Biochemistry Eighth Edition Textbook by Berg et al.

Biochemistry
EIGHTH
EDITION
Jeremy M. Berg
Tymoczko
Gregory J. Gatto, Jr.
John L.
Lubert Stryer
Publisher: Kate Ahr Parker
Senior Acquisitions Editor: Lauren Schultz
Developmental Editor: Irene Pech
Editorial Assistants: Shannon Moloney and Nandini
Ahuja Senior Project Editor: Denise Showers with
Sherrill Redd Manuscript Editors: Irene Vartanoff and
Mercy Heston
Cover and Interior Design: Vicki Tomaselli
Illustrations: Jeremy Berg with Network Graphics, Gregory J.
Gatto, Jr. Illustration Coordinator: Janice Donnola
Photo Editor: Christine Buese
Photo Researcher: Jacquelyn Wong
Production Coordinator: Paul Rohloff
Executive Media Editor: Amanda Dunning
Media Editor: Donna Brodman
Executive Marketing Manager: Sandy Lindelof
Composition: Aptara®, Inc.
Printing and Binding: RR Donnelley
Library of Congress Control Number: 2014950359
Gregory J. Gatto, Jr., is an employee of GlaxoSmithKline (GSK), which has not
supported or funded this work in any way. Any views expressed herein do not
necessarily represent the views of GSK.
ISBN-13: 978-1-4641-2610-9
ISBN-10: 1-4641-2610-0
©2015, 2012, 2007, 2002 by W. H. Freeman and Company; © 1995, 1988, 1981,
1975 by Lubert Stryer
All rights reserved
Printed in the United States of America
First printing
W. H. Freeman and Company
41 Madison Avenue
New York, NY 10010
www.whfreeman.com
To our teachers and our students
Professor of Computational and Systems Biology
and Pittsburgh Foundation Professor and Director
of the Institute for Personalized Medicine. He
JEREMY M. BERG received his B.S. and M.S.
served as President of the American Society for
degrees in Chemistry from Stanford (where he did Biochemistry and Molecular Biology from 2011–
research with Keith Hodgson and Lubert Stryer)
2013. He is a Fellow of the American Association
and his Ph.D. in Chemistry from Harvard with
for the Advancement of Science and a member of
Richard Holm. He then completed a postdoctoral
the Institute of Medicine of the National Academy
fellowship with Carl Pabo in Biophysics at Johns
of Sciences. He received the American Chemical
Hopkins University School of Medicine. He was an Society Award in Pure Chemistry (1994) and the
Assistant Professor in the Department of
Eli Lilly Award for Fundamental Research in
Chemistry at Johns Hopkins from 1986 to 1990.
Biological Chemistry (1995), was named Maryland
He then moved to Johns Hopkins University
Outstanding Young Scientist of the Year (1995),
School of Medicine as Professor and Director of
received the Harrison Howe Award (1997), and
the Department of Biophysics and Biophysical
received public service awards from the
Chemistry, where he remained until 2003. He then Biophysical Society, the American Society for
became Director of the National Institute of
Biochemistry and Molecular Biology, the American
General Medical Sciences at the National
Chemical Society, and the American Society for
Institutes of Health. In 2011, he moved to the
Cell Biology. He also received numerous teaching
University of Pittsburgh where he is now
ABOUT THE AUTHORS
awards, including the W. Barry Wood Teaching
Award (selected by medical students), the
Graduate Student Teaching Award, and the
Professor’s Teaching Award for the Preclinical
Sciences. He is coauthor, with Stephen J.
Lippard, of the textbook Principles of Bioinorganic
Chemistry.
JOHN L. TYMOCZKO is Towsley Professor
of Biology at Carleton College, where he has
taught since 1976. He currently teaches
Biochemistry, Biochemistry Laboratory,
Oncogenes and the
Molecular Biology of Cancer, and Exercise
Biochemistry and coteaches an introductory
course, Energy Flow in Biological Systems.
Professor Tymoczko received his B.A. from the
University of Chicago in 1970 and his Ph.D. in
Biochemistry from the University of Chicago with
Shutsung Liao at the Ben May Institute for
Cancer Research. He then had a postdoctoral
position with Hewson Swift of the Department of
Biology at the University of Chicago. The focus
of his research has been on steroid recep tors,
ribonucleoprotein particles, and proteolytic
processing enzymes.
GREGORY J. GATTO, JR., received his A.B.
degree in Chemistry from Princeton University,
iv
PREFACE
where he worked with Martin F. Semmelhack and
was awarded the Everett S. Wallis Prize in
Organic Chemistry. In 2003, he received his
M.D. and Ph.D. degrees from the Johns Hopkins
University School of Medicine, where he studied
the structural biology of peroxisomal targeting
signal recognition with Jeremy M. Berg and
received the Michael A. Shanoff Young
Investigator Research Award. He completed a
postdoctoral fellowship in 2006 with Christopher
T. Walsh at Harvard Medical School, where he
studied the biosynthesis of the macrolide
immunosuppres sants. He is currently a Senior
Scientific Investigator in the Heart Failure
Discovery Performance Unit at GlaxoSmithKline.
LUBERT STRYER is Winzer Professor of Cell
Biology, Emeritus, in the School of Medicine and
Professor of Neurobiology, Emeritus, at Stanford
University, where he has been on the faculty
since 1976. He received his M.D. from Harvard
Medical School. Professor Stryer has received
many awards for his research on the interplay of
light and life, including the Eli Lilly Award for
Fundamental Research in Biological Chemistry,
the Distinguished Inventors Award of the
Intellectual Property Owners’ Association, and
election to the National Academy of Sciences
and the American Philosophical Society. He was
awarded the National Medal of Science in 2006.
The publication of his first edition of Biochemistry
in 1975 transformed the teaching of biochemistry.
to help students see main points without the
distraction of excess detail.
• Physiological relevance. It has always been
our goal to help students connect biochemistry
to their own lives on a variety of scales.
Pathways and pro cesses are presented in a
physiological context so
For several generations of students and
teachers,
Biochemistry has been an invaluable resource,
pre senting the concepts and details of molecular
structure, metabolism, and laboratory techniques
in a streamlined and engaging way.
Biochemistry’s success in helping students learn
the subject for the first time is built on a number of
hallmark features:
• Clear writing and simple illustrations. The lan
guage of biochemistry is made as accessible as
possi ble for students learning the subject for the
first time. To complement the straightforward
language and organization of concepts in the
text, figures illustrate a single concept at a time
students can see how biochemistry works in the
body and under different conditions, and Clinical
Application sections in every chapter show
students how the concepts they are studying
impact human health. The eighth edition
includes a number of new Clinical Application
sections based on recent dis coveries in
biochemistry and health. (For a full list, see p.
xi)
• Evolutionary perspective. Discussions of
evolution are woven into the narrative of the text,
just as evolu tion shapes every pathway and
molecular structure described in the text.
Molecular Evolution sections highlight important
milestones in the evolution of life as a way to
provide context for the processes and molecules
being discussed. (For a full list, see p. x)
• Problem-solving practice. Every chapter of
Biochemistry provides numerous opportunities
for students to practice problem-solving skills
and apply the concepts described in the text.
End-of-chapter
interpretation
structures. All
0%
problems ask stu
molecular structures
1.0
dents to draw
in the book, with few
conclusions
from
exceptions, have
0.9
50%
data taken from real been selected and
0.8
research papers;
rendered by Jeremy
0.7
AB
and chapter
Berg and Gregory
integration problems Gatto to emphasize
require students to the aspect of
connect concepts
structure most impor
from across
tant to the topic at
chapters. Further
hand. Students are
0%
problem-solving
introduced to
problems are divided
practice is pro vided realistic renderings
into three categories
online, on the
of molecules through
to address different
Biochemistry
a molecular model
problem-solving
LaunchPad. (For
“primer” in the
skills: Mechanism
100% 50%
more details on
appendices to
prob lems ask
LaunchPad
Chapters 1 and 2 so
students to suggest
100%
resources, see p.
they are wellor describe a
viii)
equipped to
chemical
recognize and
• A variety of
mechanism; Data
interpret
molecular
Light aerobic effort
the structures throughout the
book. Figure legends
Maximal aerobic effort
direct students explicitly to the key features of a
Figure 27.12 An idealized representation of fuels use as a function of
model, and often include PDB numbers so the
aerobic exercise intensity. (A) With increased exercise intensity, the
reader can access the file used in generating
use of fats as fuels falls as the utilization of glucose increases. (B)
the structure from the Protein Data Bank
The respiratory quotient (RQ) measures the alteration in fuel use.
website (www.pdb.org). Students
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Figure 9.48 Single molecule motion. (A) A trace of the position of a single dimeric myosin V molecule as it moves across
a surface coated with actin filaments. (B) A model of how the dimeric molecule moves in discrete steps with an average
size of 74 6 5 nm. [Data from A. Yildiz et al., Science 300(5628)2061–2065, 2003.]
can explore molecular structures further online
through the Living Figures, in which they can
rotate 3D models
of molecules and view
alternative renderings.
In this revision of Biochemistry, we focused on
build ing on the strengths of the previous editions
to present biochemistry in an even more clear and
streamlined manner, as well as incorporating
exciting new advances from the field. Throughout
the book, we have updated explanations of basic
concepts and bolstered them with examples from
new research. Some new topics that we present in
the eighth edition include:
• Environmental factors that influence
human biochemistry (Chapter 1)
• Genome editing (Chapter 5)
• Horizontal gene transfer events that may explain
unex pected branches of the evolutionary tree
(Chapter 6)
• Penicillin irreversibly inactivating a key enzyme
in bacterial cell-wall synthesis (Chapter 8)
• Scientists watching single molecules of
myosin move (Chapter 9)
• Glycosylation functions in nutrient
sensing (Chapter 11)
• The structure of a SNARE complex (Chapter
12) • The mechanism of ABC transporters
(Chapter 13) • The structure of the gap junction
(Chapter 13)
• The structural basis for activation of the badrenergic receptor (Chapter 14)
• Excessive fructose consumption can lead to
patho logical conditions (Chapter 16)
• Alterations in the glycolytic pathway by cancer
cells (Chapter 16)
• Regulation of mitochondrial ATP
synthase (Chapter 18)
• Control of chloroplast ATP synthase (Chapter
19) • Activation of rubisco by rubisco activase
(Chapter 20)
Figure 12.39 SNARE complexes initiate membrane fusion. The SNARE protein synaptobrevin
(yellow) from one membrane forms a tight four-helical bundle with the corresponding SNARE
proteins syntaxin-1 (blue) and SNAP25 (red) from a second membrane. The complex brings
the membranes close together, initiating the fusion event. [Drawn from 1SFC.pdb.]
• The structural details of ligand binding by
TLRs (Chapter 34)
• The role of the pentose phosphate pathway in
rapid cell growth (Chapter 20)
• Biochemical characteristics of muscle fiber
types (Chapter 21)
• Alteration of fatty acid metabolism in tumor
cells (Chapter 22)
• Biochemical basis of neurological
symptoms of phenylketonuria (Chapter
24)
• Ribonucleotide reductase as a
chemotherapeutic target (Chapter 25)
Preface vii
• The role of excess choline in the
development of heart disease (Chapter 26)
• Cycling of the LDL receptor is regulated (Chapter
26)
• The role of ceramide metabolism in
stimulating tumor growth (Chapter 26)
• The extraordinary power of DNA repair
systems illustrated by Deinococcus
radiodurans (Chapter 28)
All of the new media resources for Biochemistry
will be available in our new system.
www.macmillanhighered.com/launchpad/berg8e
LaunchPad is a dynamic, fully integrated learning
environment that brings together all of our teaching
and learning resources in one place. It also
contains the fully interactive e-Book and other
newly updated resources
for students and
instructors, including the following:
• NEW Case Studies are a series of biochemistry
case studies you can integrate into your course.
Each case study gives students practice in
working with
Figure 34.3 Recognition of a PAMP by a Toll-like receptor. The structure
of TLR3 bound to its PAMP, a fragment of double-stranded RNA, as
seen from
data, developing critical thinking skills,
connecting topics, and applying knowledge to
real scenarios. We also provide instructional
guidance with each case study (with
suggestions on how to use the case in the
classroom) and aligned assessment questions
for quizzes and exams.
• Newly Updated Clicker Questions allow
instruc tors to integrate active learning in the
classroom and to assess students’
understanding of key concepts during lectures.
Available in Microsoft Word and PowerPoint
students have learned in the book to
(PPT).
novel medical situ ations. Students read
clinical case studies and use basic
• Newly Updated Lecture PowerPoints have
biochemistry concepts to solve the
been developed to minimize preparation time for
medical mysteries, applying and
new users of the book. These files offer
reinforcing what they learn in lecture and
suggested lectures
from the book.
including key illustrations and summaries
• Hundreds of self-graded practice prob
that instructors can adapt to their teaching
lems allow students to test their
styles.
understanding of concepts explained in the
• Updated Layered PPTs deconstruct key
text, with immedi ate feedback.
concepts, sequences, and processes
• The Metabolic Map helps students
from the textbook images, allowing
under stand the principles and applications
instructors to pres ent complex ideas
of the core metabolic pathways. Students
step-by-step.
can work through guided tutorials with
• Updated Textbook Images and Tables
embedded
are offered as high-resolution JPEG files.
assessment
questions,
or explore the
Each image has been fully optimized to
Metabolic
Map
on
their
own using the
increase type sizes and adjust color
dragging
and
zooming
functionality
of the
saturation. These images have been
map.
tested in a large lecture hall to ensure
maximum clarity and visibility.
• Jmol tutorials by Jeffrey Cohlberg, California
the side (top) and from above (bottom). Notice that the PAMP induces
• The Clinical Companion, by Gregory
receptor dimerization by binding the surfaces on the side of each of the
Raner, The University of North Carolina at extracellular domains. [Drawn from 3CIY.pdb].
Greensboro and Douglas Root, University
of North Texas, applies concepts that
State University at Long Beach, teach students
viii
how to create models of proteins in Jmol based
on data from the Protein Data Bank. By working
through the tutorial and answering assessment
ques tions at the end of each exercise, students
learn to use this important database and fully
realize the relationships between the structure
and function of enzymes.
• Living figures allow students to explore protein
structure in 3-D. Students can zoom and rotate
the “live” structures to get a better
understanding of their three-dimensional nature
and can experiment with different display styles
(space-filling, ball-and stick, ribbon, backbone)
by means of a user-friendly interface.
• Concept-based tutorials by Neil D. Clarke help
students build an intuitive understanding of
some of the more difficult concepts covered in
the textbook.
• Animated techniques help students grasp
experi mental techniques used for exploring
genes and proteins.
• NEW animations show students biochemical
pro cesses in motion. The eighth edition
includes many new animations.
• Online end-of-chapter questions are
assignable and self-graded multiple-choice
versions of the
end-of-chapter questions in the book, giving
stu dents a way to practice applying chapter
content in an online environment.
• Flashcards are an interactive tool that
allows students to study key terms from
the book.
• LearningCurve is a self-assessment tool that
helps students evaluate their progress.
Students can test their understanding by taking
an online multiple choice quiz provided for each
chapter, as well as a general chemistry review.
Updated Student Companion
[1-4641-8803-3]
For each chapter of the textbook, the Student
Companion includes:
• Chapter Learning Objectives and Summary
• Self-Assessment Problems, including multiple
choice, short-answer, matching questions, and
chal lenge problems, and their answers
• Expanded Solutions to end-of-chapter
problems in the textbook
ix
MOLECULAR EVOLUTION
This icon signals the start of the many discussions that highlight
protein commonalities or other molecular evolutionary insights.
Only L amino acids make up proteins (p. 29)
Why this set of 20 amino acids? (p. 35)
Sickle-cell trait and malaria (p. 206)
Additional human globin genes (p. 208)
Catalytic triads in hydrolytic enzymes (p. 258)
Major classes of peptide-cleaving enzymes (p.
260) Common catalytic core in type II restriction
enzymes (p. 275)
P-loop NTPase domains (p. 280)
Conserved catalytic core in protein kinases (p.
298) Why do different human blood types
exist? (p. 331) Archaeal membranes (p. 346)
Ion pumps (p. 370)
P-type ATPases (p. 374)
ATP-binding cassettes (p. 374)
Sequence comparisons of Na1 and Ca21 channels (p.
382) Small G proteins (p. 414)
Metabolism in the RNA world (p. 444)
Why is glucose a prominent fuel? (p. 451)
NAD1 binding sites in dehydrogenases (p. 465)
Isozymic forms of lactate dehydrogenase (p.
487) Evolution of glycolysis and
gluconeogenesis (p. 487) The a-ketoglutarate
dehydrogenase complex (p. 505) Domains of
succinyl CoA synthetase (p. 507) Evolution of
the citric acid cycle (p. 516)
Mitochondrial evolution (p. 525)
Conserved structure of cytochrome c (p. 541)
Common features of ATP synthase and G proteins (p.
548) Pigs lack uncoupling protein 1 (UCP-1) and
brown fat (p. 556)
Related uncoupling proteins (p. 556)
Chloroplast evolution (p. 568)
Evolutionary origins of photosynthesis (p.
584) Evolution of the C4 pathway (p. 601)
The relationship of the Calvin cycle and the
pentose phosphate pathway (p. 610)
Increasing sophistication of glycogen
phosphorylase regulation (p. 629)
Glycogen synthase is homologous to
glycogen phosphorylase (p. 631)
A recurring motif in the activation of carboxyl
groups (p. 649)
Prokaryotic counterparts of the ubiquitin pathway and
the proteasome (p. 686)
A family of pyridoxal-dependent enzymes (p.
692) Evolution of the urea cycle (p. 696)
The P-loop NTPase domain in nitrogenase (p. 716)
Conserved amino acids in transaminases determine
amino acid chirality (p. 721)
Feedback inhibition (p. 731)
Recurring steps in purine ring synthesis (p.
749) Ribonucleotide reductases (p. 755)
Increase in urate levels during primate evolution (p.
761) Deinococcus radiodurans illustrates the power
of DNA repair systems (p. 828)
DNA polymerases (p. 829)
Thymine and the fidelity of the genetic message (p.
849) Sigma factors in bacterial transcription (p. 865)
Similarities in transcription between archaea and
eukaryotes (p. 876)
Evolution of spliceosome-catalyzed splicing (p.
888) Classes of aminoacyl-tRNA synthetases
(p. 901) Composition of the primordial ribosome
(p. 903) Homologous G proteins (p. 908)
A family of proteins with common ligand-binding
domains (p. 930)
The independent evolution of DNA-binding sites
of regulatory proteins (p. 931)
Key principles of gene regulation are similar in bacteria
and archaea (p. 937)
CpG islands (p. 949)
Iron-response elements (p. 955)
miRNAs in gene evolution (p. 957)
The odorant-receptor family (p. 963)
Photoreceptor evolution (p. 973)
The immunoglobulin fold (p. 988)
Relationship of tubulin to prokaryotic proteins (p. 1023)
x
CLINICAL APPLICATIONS
This icon signals the start of a clinical application in the text. Additional,
briefer clinical correlations appear in the text as appropriate.
Osteogenesis imperfecta (p. 46)
Protein-misfolding diseases (p. 56)
Protein modification and scurvy (p. 57)
Antigen/antibody detection with ELISA (p.
82) Synthetic peptides as drugs (p. 92)
PCR in diagnostics and forensics (p.142)
Gene therapy (p. 164)
Aptamers in biotechnology and medicine (p.
187) Functional magnetic resonance imaging
(p. 193) 2,3-BPG and fetal hemoglobin (p.
201)
Carbon monoxide poisoning (p. 201)
Sickle-cell anemia (p. 205)
Thalassemia (p. 207)
Aldehyde dehydrogenase deficiency (p.
228) Action of penicillin (p. 239)
Protease inhibitors (p. 263)
Carbonic anhydrase and osteopetrosis (p.
264) Isozymes as a sign of tissue damage
(p. 293)
Trypsin inhibitor helps prevent pancreatic damage (p.
302) Emphysema (p. 303)
Blood clotting involves a cascade of zymogen
activations (p. 303)
Vitamin K (p. 306)
Antithrombin and hemorrhage (p. 307)
Hemophilia (p.308)
Monitoring changes in glycosylated hemoglobin (p.
321) Erythropoietin (p. 327)
Hurler disease (p. 327)
Mucins (p. 329)
Blood groups (p. 331)
I-cell disease (p. 332)
Influenza virus binding (p. 335)
Clinical applications of liposomes (p. 349)
Aspirin and ibuprofen (p. 353)
Digitalis and congestive heart failure (p.
373) Multidrug resistance (p. 374)
Long QT syndrome (p. 388)
Signal-transduction pathways and cancer (p.
416) Monoclonal antibodies as anticancer drugs
(p. 416) Protein kinase inhibitors as anticancer
drugs (p. 417) G-proteins, cholera and
whooping cough (p. 417) Vitamins (p. 438)
Triose phosphate isomerase deficiency (p.
454) Excessive fructose consumption (p.
466)
Lactose intolerance (p. 467)
Galactosemia (p. 468)
Aerobic glycolysis and cancer (p. 474)
Phosphatase deficiency (p. 512)
Defects in the citric acid cycle and the
development of cancer (p. 513)
Beriberi and mercury poisoning (p. 515)
Frataxin mutations cause Friedreich’s ataxia (p. 531)
Reactive oxygen species (ROS) are implicated in a
variety of diseases (p. 539)
ROS may be important in signal transduction (p.
540) IF1 overexpression and cancer (p. 554)
Brown adipose tissue (p. 555)
Mild uncouplers sought as drugs (p.557)
Mitochondrial diseases (p. 557)
Glucose 6-phosphate dehydrogenase deficiency
causes drug-induced hemolytic anemia (p. 610)
Glucose 6-phosphate dehydrogenase deficiency
protects against malaria (p. 612)
Developing drugs for type 2 diabetes (p.
636) Glycogen-storage diseases (p. 637)
Chanarin-Dorfman syndrome (p. 648)
Carnitine deficiency (p. 650)
Zellweger syndrome (p. 657)
Diabetic ketosis (p. 659)
Ketogenic diets to treat epilepsy (p. 660)
Some fatty acids may contribute to pathological
conditions (p. 661)
The use of fatty acid synthase inhibitors as
drugs (p. 667)
Effects of aspirin on signaling pathways (p. 669)
Diseases resulting from defects in transporters of
amino acids (p. 682)
Diseases resulting from defects in E3 proteins (p.
685) Drugs target the ubiquitin-proteasome system
(p.687) Using proteasome inhibitors to treat
tuberculosis (p. 687) Blood levels of
aminotransferases indicate liver damage (p. 691)
Inherited defects of the urea cycle
(hyperammonemia) (p. 697)
Alcaptonuria, maple syrup urine disease,
and phenylketonuria (p. 705)
xi
High homocysteine levels and vascular disease (p.
726) Inherited disorders of porphyrin metabolism (p.
737) Anticancer drugs that block the synthesis of
thymidylate (p. 757)
Ribonucleotide reductase is a target for cancer
therapy (p. 759)
Adenosine deaminase and severe combined
immunodefi ciency (p. 760)
Gout (p. 761)
Lesch–Nyhan syndrome (p. 761)
Folic acid and spina bifida (p. 762)
Enzyme activation in some cancers to generate
phospho choline (p. 770)
Excess choline and heart disease (p. 771)
Gangliosides and cholera (p. 773)
Second messengers derived from sphingolipids and
diabetes (p. 773)
Respiratory distress syndrome and Tay–Sachs
disease (p. 774) Ceramide metabolism stimulates
tumor growth (p. 775) Phosphatidic acid phosphatase
and lipodystrophy (p. 776) Hypercholesterolemia and
atherosclerosis (p. 784) Mutations in the LDL receptor
(p. 785)
LDL receptor cycling is regulated (p. 787)
The role of HDL in protecting against arteriosclerosis
(p. 787) Clinical management of cholesterol levels (p.
788) Bile salts are derivatives of cholesterol (p. 789)
The cytochrome P450 system is protective (p. 791) A
new protease inhibitor also inhibits a cytochrome P450
enzyme (p. 792)
Aromatase inhibitors in the treatment of breast and
ovarian cancer (p. 794)
Rickets and vitamin D (p. 795)
Caloric homeostasis is a means of regulating body
weight (p. 802)
The brain plays a key role in caloric homeostasis (p.
804) Diabetes is a common metabolic disease often
resulting from obesity (p. 807)
Exercise beneficially alters the biochemistry of cells (p.
813) Food intake and starvation induce metabolic
changes (p. 816) Ethanol alters energy metabolism in
the liver (p. 819) Antibiotics that target DNA gyrase (p.
839)
Blocking telomerase to treat cancer (p. 845)
Huntington disease (p. 850)
Defective repair of DNA and cancer (p. 850)
Detection of carcinogens (Ames test) (p.
852) Translocations can result in
diseases (p. 855)
Antibiotic inhibitors of transcription (p. 869)
Burkitt lymphoma and B-cell leukemia (p.
876) Diseases of defective RNA splicing (p.
884)
Vanishing white matter disease (p. 913)
Antibiotics that inhibit protein synthesis (p.
914) Diphtheria (p. 914)
Ricin, a lethal protein-synthesis inhibitor (p.
915) Induced pluripotent stem cells (p. 947)
Anabolic steroids (p. 951)
Color blindness (p. 974)
The use of capsaicin in pain management (p.
978) Immune-system suppressants (p. 994)
MHC and transplantation rejection (p.
1002) AIDS (p. 1003)
Autoimmune diseases (p. 1005)
Immune system and cancer (p. 1005)
Vaccines (p. 1006)
Charcot-Marie-Tooth disease (p. 1022)
Taxol (p. 1023)
xii
task.
ACKNOWLEDGMENTS
Writing a popular textbook is both a challenge and
an honor. Our goal is to convey to our students
our enthu siasm and understanding of a discipline
to which we are devoted. They are our inspiration.
Consequently, not a word was written or an
illustration constructed with out the knowledge that
bright, engaged students would immediately
detect vagueness and ambiguity. We also thank
our colleagues who supported, advised, instruct
ed, and simply bore with us during this arduous
Paul Adams
University of Arkansas,
Fayetteville Kevin Ahern
Oregon State University
Zulfiqar Ahmad
A.T. Still University of Health
Sciences Young-Hoon An
Wayne State University
Richard Amasino
University of Wisconsin
Kenneth Balazovich
University of Michigan
Donald Beitz
Iowa State University
Matthew Berezuk
Azusa Pacific University
Melanie Berkmen
Suffolk University
Steven Berry
University of Minnesota,
Duluth Loren Bertocci
Marian University
Mrinal Bhattacharjee
Long Island University
Elizabeth Blinstrup-Good
University of Illinois
Brian Bothner
Montana State University
Mark Braiman
Syracuse University
We are grateful to our colleagues throughout the
world who patiently answered our questions and
shared their insights into recent developments.
We also especially thank those who served as
review ers for this new edition. Their thoughtful
comments, suggestions, and encouragement
have been of immense help to us in maintaining
the excellence of the preceding editions. These
reviewers are:
David Brown
Florida Gulf Coast
University Donald Burden
Middle Tennessee State
University Nicholas Burgis
Eastern Washington
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We have been working with the people
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xiv
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children, especially Timothy and Mark Gatto.
Without their support, comfort, and understanding,
this endeavor could never have been undertaken,
let alone successfully completed.
xv
BRIEF CONTENTS
3 Exploring Proteins and Proteomes 65
4 DNA, RNA, and the Flow of Genetic Information 105 5
Exploring Genes and Genomes 135
6 Exploring Evolution and Bioinformatics 169 7
Hemoglobin: Portrait of a Protein in Action 191 8
Enzymes: Basic Concepts and Kinetics 215 9 Catalytic
Strategies 251
10 Regulatory Strategies 285
11 Carbohydrates 315
12 Lipids and Cell Membranes 341
13 Membrane Channels and Pumps 367
14 Signal-Transduction Pathways 397
Part II TRANSDUCING AND STORING ENERGY
15 Metabolism: Basic Concepts and Design 423 16
Glycolysis and Gluconeogenesis 449
17 The Citric Acid Cycle 495
18 Oxidative Phosphorylation 523
19 The Light Reactions of Photosynthesis 565 20 The
Calvin Cycle and the Pentose Phosphate Pathway 589
21 Glycogen Metabolism 617
22 Fatty Acid Metabolism 643
23 Protein Turnover and Amino Acid Catabolism 681
Part III SYNTHESIZING THE MOLECULES OF LIFE
24 The Biosynthesis of Amino Acids 713
25 Nucleotide Biosynthesis 743
26 The Biosynthesis of Membrane Lipids and
Steroids 767
27 The Integration of Metabolism 801
28 DNA Replication, Repair, and Recombination 827 29
RNA Synthesis and Processing 859
30 Protein Synthesis 893
31 The Control of Gene Expression in Prokaryotes 925 32
The Control of Gene Expression in Eukaryotes 941
Part IV RESPONDING TO ENVIRONMENTAL
CHANGES
33 Sensory Systems 961
34 The Immune System 981
35 Molecular Motors 1011
36 Drug Development 1033
CONTENTS
Preface v Part I THE MOLECULAR DESIGN OF LIFE
1 CHAPTER 1 Biochemistry: An Evolving Science 1
CHAPTER 1 Biochemistry: An Evolving Science 1
1.1 Biochemical Unity Underlies Biological Diversity 1 1.2
Part I THE MOLECULAR DESIGN OF LIFE 1
Biochemistry: An Evolving Science 1
2 Protein Composition and Structure 27
DNA Illustrates the Interplay Between Form and Function 4
DNA is constructed from four building blocks 4 Two single
strands of DNA combine to form a double helix 5 DNA
structure explains heredity and the storage of information
5
1.3 Concepts from Chemistry Explain the Properties of
Biological Molecules 6 The formation of the DNA double helix
as a key example 6 The double helix can form from its
component strands 6 Covalent and noncovalent bonds are
CHAPTER 3 Exploring Proteins and Proteomes 65
important for the structure and stability of biological
molecules 6 The double helix is an expression of the rules of CHAPTER 3 Exploring Proteins and Proteomes 65
chemistry 9 The laws of thermodynamics govern the behavior
of biochemical systems 10 Heat is released in the formation
The proteome is the functional representation of
of the double helix 12 Acid–base reactions are central in
the genome 66 3.1 The Purification of Proteins Is an
many biochemical processes 13 Acid–base reactions can
Essential First Step in Understanding Their Function 66
disrupt the double helix 14 Buffers regulate pH in organisms
The assay: How do we recognize the protein that we are
and in the laboratory 15 1.4 The Genomic Revolution Is
looking for? 67 Proteins must be released from the cell to
Transforming Biochemistry, Medicine, and Other Fields 17
Genome sequencing has transformed biochemistry and
other fields 17 Environmental factors influence human
biochemistry 20 Genome sequences encode proteins and
patterns
of expression 21
APPENDIX: Visualizing Molecular Structures I:
Small Molecules 22
CHAPTER 2 Protein Composition and Structure 27
CHAPTER 2 Protein Composition and Structure 27
2.1 Proteins Are Built from a Repertoire of 20 Amino Acids 29
2.2 Primary Structure: Amino Acids Are Linked by Peptide
be purified 67 Proteins can be purified according to
solubility, size, charge, and binding affinity 68 Proteins
can be separated by gel electrophoresis and displayed
71 A protein purification scheme can be quantitatively
evaluated 75 Ultracentrifugation is valuable for separating
biomolecules and determining their masses 76 Protein
purification can be made easier with the
use of recombinant DNA technology 78 3.2 Immunology
Provides Important Techniques with Which to Investigate
Proteins 79
Antibodies to specific proteins can be generated 79
Monoclonal antibodies with virtually any desired
specificity can be readily prepared 80 Proteins can be
detected and quantified by using an enzyme-linked
immunosorbent assay 82 Western blotting permits the
detection of proteins separated by gel electrophoresis 83
Fluorescent markers make the visualization of proteins in
the cell possible 84
Contents xvii
Bonds to Form Polypeptide Chains 35
Proteins have unique amino acid sequences specified by
genes 37 Polypeptide chains are flexible yet
conformationally restricted 38
2.3 Secondary Structure: Polypeptide Chains Can Fold into
Regular Structures Such As the Alpha Helix, the Beta Sheet,
and Turns and Loops 40
The alpha helix is a coiled structure stabilized by
intrachain hydrogen bonds 40 Beta sheets are stabilized
by hydrogen bonding between polypeptide strands 42
3.3 Mass Spectrometry Is a Powerful Technique for the
Identification of Peptides and Proteins 85 Peptides can be
sequenced by mass spectrometry 87 Proteins can be
specifically cleaved into small peptides to facilitate analysis
88 Genomic and proteomic methods are complementary 89
The amino acid sequence of a protein provides valuable
information 90 Individual proteins can be identified by mass
spectrometry 91 3.4 Peptides Can Be Synthesized by
Automated Solid-Phase Methods 92
Polypeptide chains can change direction by making
reverse turns and loops 44 Fibrous proteins provide
structural support for cells and tissues 44
2.4 Tertiary Structure: Water-Soluble Proteins Fold into
3.5 Three-Dimensional Protein Structure Can Be Determined
by X-ray Crystallography and NMR Spectroscopy 95 X-ray
crystallography reveals three-dimensional
structure in atomic detail 95 Nuclear magnetic resonance
spectroscopy can reveal the structures of proteins in
Compact Structures with Nonpolar Cores 46
2.5 Quaternary Structure: Polypeptide Chains Can Assemble
into Multisubunit Structures 48
2.6 The Amino Acid Sequence of a Protein Determines Its
Three-Dimensional Structure 49 Amino acids have different
propensities for
forming a helices, b sheets, and turns 51 Protein folding is
a highly cooperative process 52 Proteins fold by progressive
stabilization of intermediates rather than by random search
53 Prediction of three-dimensional structure from sequence
remains a great challenge 54 Some proteins are inherently
unstructured and can exist in multiple conformations 55
Protein misfolding and aggregation are associated with
some neurological diseases 56 Protein modification and
cleavage confer new capabilities 57 APPENDIX: Visualizing
Molecular Structures II: Proteins 61
solution 97
CHAPTER 4 DNA, RNA, and the Flow of
CHAPTER 4 DNA, RNA, and the Flow of
Genetic Information 105 Genetic Information 105
4.1 A Nucleic Acid Consists of Four Kinds of Bases Linked to
a Sugar–Phosphate Backbone 106 RNA and DNA differ in the
sugar component and one of the bases 106 Nucleotides are
the monomeric units of nucleic acids 107 DNA molecules are
very long and have directionality 108 4.2 A Pair of Nucleic
Acid Strands with
Complementary Sequences Can Form a
Double-Helical Structure 109 The double helix is stabilized by
genomic DNA 147 Complementary DNA prepared from
mRNA can be expressed in host cells 149 Proteins with
new functions can be created through directed changes
in DNA 150 Recombinant methods enable the exploration
of the functional effects of disease-causing mutations 152
hydrogen bonds and van der Waals interactions 109 DNA
can assume a variety of structural forms 111 Z-DNA is a
5.3 Complete Genomes Have Been Sequenced and Analyzed
left-handed double helix in which
backbone phosphates zigzag 112 Some DNA molecules 152 The genomes of organisms ranging from bacteria to
are circular and supercoiled 113 Single-stranded nucleic multicellular eukaryotes have been sequenced 153 The
sequence of the human genome has been completed 154
acids can adopt elaborate
structures 113 4.3 The Double Helix Facilitates the
Accurate Transmission of Hereditary Information 114
Differences in DNA density established the validity of the
semiconservative replication hypothesis 115 The double
helix can be reversibly melted 116
4.4 DNA Is Replicated by Polymerases That Take Instructions
from Templates 117 DNA polymerase catalyzes
phosphodiester
bridge formation 117 The genes of some viruses are
made of RNA 118 4.5 Gene Expression Is the Transformation
of DNA Information into Functional Molecules 119 Several
kinds of RNA play key roles in gene expression 119
xviii Contents
All cellular RNA is synthesized by RNA polymerases 120
RNA polymerases take instructions from DNA
templates 121 Transcription begins near promoter sites
and ends at terminator sites 122 Transfer RNAs are the
adaptor molecules in protein synthesis 123
4.6 Amino Acids Are Encoded by Groups of Three Bases
Starting from a Fixed Point 124 Major features of the genetic
code 125 Messenger RNA contains start and stop signals for
protein synthesis 126 The genetic code is nearly universal
126 4.7 Most Eukaryotic Genes Are Mosaics of Introns and
Exons 127 RNA processing generates mature RNA 127 Many
Next-generation sequencing methods enable the rapid
determination of a complete genome sequence 155
Comparative genomics has become a powerful
research tool 156 5.4 Eukaryotic Genes Can Be Quantitated
and Manipulated with Considerable Precision 157 Geneexpression levels can be comprehensively
examined 157 New genes inserted into eukaryotic cells
can be
efficiently expressed 159 Transgenic animals harbor and
express genes introduced into their germ lines 160 Gene
disruption and genome editing provide clues to gene
function and opportunities for new therapies 160 RNA
interference provides an additional tool for
disrupting gene expression 162 Tumor-inducing plasmids
can be used to introduce new genes into plant cells 163
Human gene therapy holds great promise for medicine
164
CHAPTER 6 Exploring Evolution and
CHAPTER 6 Exploring Evolution and
Bioinformatics 169 Bioinformatics 169
exons encode protein domains 128
CHAPTER 5 Exploring Genes and Genomes 135 CHAPTER 5
Exploring Genes and Genomes 135
5.1 The Exploration of Genes Relies on Key Tools 136
Restriction enzymes split DNA into specific
fragments 137 Restriction fragments can be separated by
gel
electrophoresis and visualized 137 DNA can be
sequenced by controlled termination of replication 138
DNA probes and genes can be synthesized by
automated solid-phase methods 139 Selected DNA
sequences can be greatly amplified
by the polymerase chain reaction 141 PCR is a powerful
technique in medical diagnostics, forensics, and studies
of molecular evolution 142 The tools for recombinant DNA
technology have been used to identify disease-causing
mutations 143
5.2 Recombinant DNA Technology Has
Revolutionized All Aspects of Biology 143 Restriction
enzymes and DNA ligase are key tools in forming
recombinant DNA molecules 143 Plasmids and l phage
are choice vectors for DNA cloning in bacteria 144
Bacterial and yeast artificial chromosomes 147 Specific
genes can be cloned from digests of
6.1 Homologs Are Descended from a Common Ancestor 170
6.2 Statistical Analysis of Sequence Alignments Can Detect
Homology 171 The statistical significance of alignments can
be
estimated by shuffling 173 Distant evolutionary
relationships can be detected
through the use of substitution matrices 174 Databases
can be searched to identify homologous
sequences 177
6.3 Examination of Three-Dimensional Structure Enhances Our
Understanding of Evolutionary Relationships 177 Tertiary
structure is more conserved than primary
structure 178 Knowledge of three-dimensional structures
can aid in the evaluation of sequence alignments 179
Repeated motifs can be detected by aligning sequences
with themselves 180 Convergent evolution illustrates
common solutions to biochemical challenges 181
Comparison of RNA sequences can be a source of
insight into RNA secondary structures 182
6.4 Evolutionary Trees Can Be Constructed on the Basis of
Sequence Information 183 Horizontal gene transfer events
may explain unexpected branches of the evolutionary tree
184
6.5 Modern Techniques Make the Experimental Exploration of
Evolution Possible 185 Ancient DNA can sometimes be
amplified and
sequenced 185 Molecular evolution can be examined
experimentally 185
Contents xix
CHAPTER 7 Hemoglobin: Portrait of a Protein
Enzymes: Basic Concepts and
CHAPTER 7 Hemoglobin: Portrait of a Protein
in Action 191
in Action 191
7.1 Myoglobin and Hemoglobin Bind Oxygen at Iron Atoms in
Heme 192 Changes in heme electronic structure upon oxygen
binding are the basis for functional imaging studies 193 The
structure of myoglobin prevents the release of
reactive oxygen species 194 Human hemoglobin is an
assembly of four myoglobin like subunits 195
7.2 Hemoglobin Binds Oxygen Cooperatively 195 Oxygen
binding markedly changes the quaternary
structure of hemoglobin 197 Hemoglobin cooperativity can
be potentially explained by several models 198 Structural
changes at the heme groups are transmitted to the a1b1–
a2b2 interface 200 2,3-Bisphosphoglycerate in red cells is
crucial in
determining the oxygen affinity of hemoglobin 200 Carbon
monoxide can disrupt oxygen transport by hemoglobin 201
7.3 Hydrogen Ions and Carbon Dioxide Promote the Release of
Oxygen: The Bohr Effect 202 7.4 Mutations in Genes Encoding
Hemoglobin Subunits Can Result in Disease 204 Sickle-cell
anemia results from the aggregation of
mutated deoxyhemoglobin molecules 205 Thalassemia is
caused by an imbalanced production of hemoglobin
chains 207 The accumulation of free alpha-hemoglobin
chains is prevented 207 Additional globins are encoded in
the human genome 208
APPENDIX: Binding Models Can Be
Formulated in Quantitative Terms: The Hill
Plot and the
Concerted Model 210
The formation of an enzyme–substrate complex is the first
step in enzymatic catalysis 222 The active sites of
enzymes have some common
features 223 The binding energy between enzyme and
substrate is important for catalysis 225
8.4 The Michaelis–Menten Model Accounts for the Kinetic
Properties of Many Enzymes 225 Kinetics is the study of
reaction rates 225 The steady-state assumption facilitates a
description of enzyme kinetics 226 Variations in KM can have
physiological consequences 228 KM and Vmax values can be
determined by several means 228
KM and Vmax values are important enzyme
characteristics 229 kcat/KM is a measure of catalytic
efficiency 230 Most biochemical reactions include multiple
substrates 231 Allosteric enzymes do not obey Michaelis–
Menten
kinetics 233 8.5 Enzymes Can Be Inhibited by Specific
Molecules 234 The different types of reversible inhibitors are
kinetically distinguishable 235 Irreversible inhibitors can be
used to map the active site 237 Penicillin irreversibly
inactivates a key enzyme in
bacterial cell-wall synthesis 239 Transition-state analogs
are potent inhibitors of
enzymes 240 Catalytic antibodies demonstrate the
importance
of selective binding of the transition state to enzymatic
activity 241
8.6 Enzymes Can Be Studied One Molecule at a Time 242
APPENDIX: Enzymes are Classified on the Basis of the
Types of Reactions That They Catalyze 245
CHAPTER 8
Enzymes: Basic Concepts and CHAPTER 8
Kinetics 215
CHAPTER 9
Catalytic Strategies
251
Formation of the Transition State 221
Kinetics 215
8.1 Enzymes are Powerful and Highly Specific Catalysts 216
Many enzymes require cofactors for activity 217 Enzymes
can transform energy from one form
into another 217
8.2 Gibbs Free Energy Is a Useful Thermodynamic Function for
Understanding Enzymes 218 The free-energy change provides
information about the spontaneity but not the rate of a
reaction 218 The standard free-energy change of a reaction
is related to the equilibrium constant 219 Enzymes alter only
the reaction rate and not the reaction equilibrium 220
8.3 Enzymes Accelerate Reactions by Facilitating the
CHAPTER 9 Catalytic Strategies 251
A few basic catalytic principles are used by
many enzymes 252 9.1 Proteases Facilitate a Fundamentally
Difficult Reaction 253 Chymotrypsin possesses a highly
reactive serine
residue 253 Chymotrypsin action proceeds in two steps
linked by a covalently bound intermediate 254 Serine is
part of a catalytic triad that also includes
histidine and aspartate 255 Catalytic triads are found in
other hydrolytic enzymes 258 The catalytic triad has been
dissected by site-directed mutagenesis 260 Cysteine,
aspartyl, and metalloproteases are other
major classes of peptide-cleaving enzymes 260 Protease
inhibitors are important drugs 263
xx Contents
9.2 Carbonic Anhydrases Make a Fast
Reaction Faster 264 Carbonic anhydrase contains a bound
zinc ion essential for catalytic activity 265 Catalysis entails
zinc activation of a water molecule 265 A proton shuttle
facilitates rapid regeneration of the active form of the
enzyme 267
9.3 Restriction Enzymes Catalyze Highly
Specific DNA-Cleavage Reactions 269 Cleavage is by in-line
displacement of 39-oxygen
from phosphorus by magnesium-activated water 269
Restriction enzymes require magnesium for catalytic
activity 271 The complete catalytic apparatus is assembled
only within complexes of cognate DNA molecules,
ensuring specificity 272 Host-cell DNA is protected by the
addition of methyl groups to specific bases 274 Type II
restriction enzymes have a catalytic core in common and
are probably related by horizontal
gene transfer 275
9.4 Myosins Harness Changes in Enzyme
Conformation to Couple ATP Hydrolysis to
Mechanical Work 275 ATP hydrolysis proceeds by the attack
of water on the gamma-phosphoryl group 276 Formation
of the transition state for ATP hydrolysis is associated with
a substantial conformational change 277 The altered
conformation of myosin persists for a
substantial period of time 278 Scientists can watch single
molecules of myosin move 279 Myosins are a family of
enzymes containing
Cleavage 299 Chymotrypsinogen is activated by specific
cleavage of a single peptide bond 299 Proteolytic activation
of chymotrypsinogen leads
to the formation of a substrate-binding site 300 The
generation of trypsin from trypsinogen leads
to the activation of other zymogens 301 Some proteolytic
enzymes have specific inhibitors 302 Blood clotting is
accomplished by a cascade of
zymogen activations 303 Prothrombin requires a vitamin Kdependent
modification for activation 304 Fibrinogen is converted by
thrombin into a fibrin clot 304 Vitamin K is required for the
formation of
g-carboxyglutamate 306 The clotting process must be
precisely regulated 307 Hemophilia revealed an early step
in clotting 308
CHAPTER 11 Carbohydrates 315CHAPTER 11 Carbohydrates
315
11.1 Monosaccharides Are the Simplest
Carbohydrates 316 Many common sugars exist in cyclic forms
318 Pyranose and furanose rings can assume different
conformations 320 Glucose is a reducing sugar 321
Monosaccharides are joined to alcohols and amines
through glycosidic bonds 322 Phosphorylated sugars are
key intermediates in energy generation and biosyntheses
322
11.2 Monosaccharides Are Linked to Form
Complex Carbohydrates 323 Sucrose, lactose, and maltose are
P-loop structures 280
CHAPTER 10 Regulatory Strategies 285 CHAPTER 10
Regulatory Strategies 285
10.1 Aspartate Transcarbamoylase Is Allosterically Inhibited by
the End Product of Its Pathway 286 Allosterically regulated
enzymes do not follow
Michaelis–Menten kinetics 287 ATCase consists of
separable catalytic and regulatory subunits 287 Allosteric
interactions in ATCase are mediated by large changes in
quaternary structure 288 Allosteric regulators modulate the
T-to-R
equilibrium 291
10.2 Isozymes Provide a Means of Regulation Specific to
Distinct Tissues and Developmental Stages 292 10.3 Covalent
Modification Is a Means of
Regulating Enzyme Activity 293 Kinases and phosphatases
control the extent of
protein phosphorylation 294 Phosphorylation is a highly
effective means of
regulating the activities of target proteins 296
Cyclic AMP activates protein kinase A by altering
the quaternary structure 297 ATP and the target protein
bind to a deep cleft in the catalytic subunit of protein
kinase A 298
10.4 Many Enzymes Are Activated by Specific Proteolytic
the common
disaccharides 323 Glycogen and starch are storage forms
of glucose 324 Cellulose, a structural component of plants,
is made of chains of glucose 324
11.3 Carbohydrates Can Be Linked to Proteins to Form
Glycoproteins 325 Carbohydrates can be linked to proteins
through
asparagine (N-linked) or through serine or threonine (Olinked) residues 326 The glycoprotein erythropoietin is a
vital hormone 327 Glycosylation functions in nutrient
sensing 327 Proteoglycans, composed of polysaccharides
and
protein, have important structural roles 327 Proteoglycans
are important components of cartilage 328 Mucins are
glycoprotein components of mucus 329 Protein
glycosylation takes place in the lumen of the
endoplasmic reticulum and in the Golgi complex 330
Specific enzymes are responsible for oligosaccharide
assembly 331 Blood groups are based on protein
glycosylation
patterns 331 Errors in glycosylation can result in
pathological
conditions 332 Oligosaccharides can be “sequenced” 332
11.4 Lectins Are Specific Carbohydrate-Binding Proteins 333
Lectins promote interactions between cells 334 Lectins are
organized into different classes 334 Influenza virus binds to
sialic acid residues 335
CHAPTER 12 Lipids and Cell Membranes 341 CHAPTER 12
Lipids and Cell Membranes 341
Many common features underlie the diversity of
biological membranes 342 12.1 Fatty Acids Are Key
Constituents of Lipids 342 Fatty acid names are based on their
parent
hydrocarbons 342 Fatty acids vary in chain length and
degree of
unsaturation 343 12.2 There Are Three Common Types of
Membrane Lipids 344 Phospholipids are the major class of
membrane lipids 344 Membrane lipids can include
carbohydrate moieties 345 Cholesterol is a lipid based on a
steroid nucleus 346 Archaeal membranes are built from
ether lipids with
branched chains 346 A membrane lipid is an amphipathic
molecule containing a hydrophilic and a hydrophobic
moiety 347
12.3 Phospholipids and Glycolipids Readily Form Bimolecular
Sheets in Aqueous Media 348 Lipid vesicles can be formed
from phospholipids 348 Lipid bilayers are highly impermeable
to ions and most polar molecules 349
12.4 Proteins Carry Out Most Membrane
Processes 350 Proteins associate with the lipid bilayer in a
variety of ways 351 Proteins interact with membranes in a
variety of ways 351 Some proteins associate with
membranes through
covalently attached hydrophobic groups 354
Transmembrane helices can be accurately predicted from
amino acid sequences 354
12.5 Lipids and Many Membrane Proteins Diffuse Rapidly in
the Plane of the Membrane 356 The fluid mosaic model allows
lateral movement but not rotation through the membrane 357
Membrane fluidity is controlled by fatty acid
composition and cholesterol content 357 Lipid rafts are
highly dynamic complexes formed
between cholesterol and specific lipids 358 All biological
membranes are asymmetric 358
Contents xxi
12.6 Eukaryotic Cells Contain Compartments Bounded by
Internal Membranes 359
CHAPTER 13 Membrane Channels and Pumps 367CHAPTER
13 Membrane Channels and Pumps 367
ATPases couple phosphorylation and
conformational changes to pump calcium ions
across membranes 370 Digitalis specifically inhibits the
Na1–K1 pump
by blocking its dephosphorylation 373 P-type ATPases are
evolutionarily conserved and
play a wide range of roles 374 Multidrug resistance
highlights a family of membrane pumps with ATP-binding
cassette domains 374
13.3 Lactose Permease Is an Archetype of
Secondary Transporters That Use One
Concentration Gradient to Power the Formation of Another 376
13.4 Specific Channels Can Rapidly Transport Ions Across
Membranes 378
Action potentials are mediated by transient changes in Na1
and K1 permeability 378 Patch-clamp conductance
measurements reveal the activities of single channels 379
The structure of a potassium ion channel is an archetype
for many ion-channel structures 379 The structure of the
potassium ion channel reveals the basis of ion specificity
380 The structure of the potassium ion channel explains its
rapid rate of transport 383 Voltage gating requires
substantial conformational
changes in specific ion-channel domains 383 A channel
can be inactivated by occlusion of the pore: the ball-andchain model 384 The acetylcholine receptor is an
archetype for
ligand-gated ion channels 385 Action potentials integrate
the activities of several ion channels working in concert
387 Disruption of ion channels by mutations or chemicals
can be potentially life-threatening 388
13.5 Gap Junctions Allow Ions and Small Molecules to Flow
Between Communicating Cells 389 13.6 Specific Channels
Increase the Permeability of Some Membranes to Water 390
xxii Contents
CHAPTER 14 Signal-Transduction Pathways 397 CHAPTER 14
Si l T d i P h Signal-Transduction Pathways 397
Signal transduction depends on molecular circuits 398 14.1
Heterotrimeric G Proteins Transmit Signals and Reset
Themselves 399
Ligand binding to 7TM receptors leads to the activation of
heterotrimeric G proteins 400 Activated G proteins transmit
signals by binding to other proteins 402 Cyclic AMP
stimulates the phosphorylation of many target proteins by
activating protein kinase A 403 G proteins spontaneously
reset themselves through GTP hydrolysis 403 Some 7TM
receptors activate the phosphoinositide cascade 404
Calcium ion is a widely used second messenger 405
Calcium ion often activates the regulatory protein
calmodulin 407 14.2 Insulin Signaling: Phosphorylation
Cascades Are Central to Many Signal-Transduction Processes
The expression of transporters largely defines the
407
metabolic activities of a given cell type 368
The insulin receptor is a dimer that closes around a bound
13.1 The Transport of Molecules Across a
insulin molecule 408 Insulin binding results in the crossMembrane May Be Active or Passive 368 Many molecules
phosphorylation and activation of the insulin receptor 408
require protein transporters to cross membranes 368 Free
The activated insulin-receptor kinase initiates a kinase
energy stored in concentration gradients can be quantified
cascade 409 Insulin signaling is terminated by the action of
369
phosphatases 411 14.3 EGF Signaling: Signal-Transduction
13.2 Two Families of Membrane Proteins Use ATP Hydrolysis Pathways Are Poised to Respond 411
to Pump Ions and Molecules Across Membranes 370 P-type
EGF binding results in the dimerization of the EGF
receptor 411 The EGF receptor undergoes phosphorylation
of its carboxyl-terminal tail 413 EGF signaling leads to the
activation of Ras, a small G protein 413 Activated Ras
initiates a protein kinase cascade 414 EGF signaling is
terminated by protein phosphatases and the intrinsic
GTPase activity of Ras 414
14.4 Many Elements Recur with Variation in Different SignalTransduction Pathways 415 14.5 Defects in Signal-Transduction
Pathways Can Lead to Cancer and Other Diseases 416
Monoclonal antibodies can be used to inhibit signal
transduction pathways activated in tumors 416 Protein
kinase inhibitors can be effective anticancer drugs 417
Cholera and whooping cough are the result of altered Gprotein activity 417
Glucose is generated from dietary carbohydrates 450
Glucose is an important fuel for most organisms 451
16.1 Glycolysis Is an Energy-Conversion Pathway in Many
Organisms 451 Hexokinase traps glucose in the cell and
begins
glycolysis 451 Fructose 1,6-bisphosphate is generated from
glucose 6-phosphate 453 The six-carbon sugar is cleaved
into two three-carbon fragments 454 Mechanism: Triose
phosphate isomerase salvages a three-carbon fragment
455 The oxidation of an aldehyde to an acid powers the
formation of a compound with high phosphoryl-transfer
potential 457 Mechanism: Phosphorylation is coupled to the
oxidation of glyceraldehyde 3-phosphate by a thioester
intermediate 458 ATP is formed by phosphoryl transfer
from
1,3-bisphosphoglycerate 459 Additional ATP is generated
with the formation of pyruvate 460 Two ATP molecules are
formed in the conversion of glucose into pyruvate 461
Part II TRANSDUCING AND STORING ENERGY
NAD1 is regenerated from the metabolism of pyruvate 462
Fermentations provide usable energy in the absence of
oxygen 464 The binding site for NAD1 is similar in many
dehydrogenases 465 Fructose is converted into glycolytic
intermediates by fructokinase 465 Excessive fructose
consumption can lead to pathological conditions 466
Galactose is converted into glucose 6-phosphate 466 Many
adults are intolerant of milk because they are deficient in
lactase 467 Galactose is highly toxic if the transferase is
missing 468
CHAPTER 15 Metabolism: Basic Concepts
CHAPTER 15 Metabolism: Basic Concepts and Design 423
and Design 423
15.1 Metabolism Is Composed of Many Coupled,
Interconnecting Reactions 424 Metabolism consists of energyyielding and energy requiring reactions 424
16.2 The Glycolytic Pathway Is Tightly Controlled 469
Glycolysis in muscle is regulated to meet the need for ATP
A thermodynamically unfavorable reaction can be driven
469 The regulation of glycolysis in the liver illustrates the
by a favorable reaction 425
biochemical versatility of the liver 472 A family of
transporters enables glucose to enter and leave animal
15.2 ATP Is the Universal Currency of Free
cells 473 Aerobic glycolysis is a property of rapidly growing
Energy in Biological Systems 426 ATP hydrolysis is exergonic
cells 474 Cancer and endurance training affect glycolysis in
426 ATP hydrolysis drives metabolism by shifting the
a similar fashion 476
equilibrium of coupled reactions 427 The high phosphoryl
potential of ATP results from structural differences
16.3 Glucose Can Be Synthesized from
between ATP and its hydrolysis products 429 Phosphoryl- Noncarbohydrate Precursors 476 Gluconeogenesis is not a
transfer potential is an important form of cellular energy
reversal of glycolysis 478 The conversion of pyruvate into
transformation 430
phosphoenolpyruvate begins with the formation of
15.3 The Oxidation of Carbon Fuels Is an
Important Source of Cellular Energy 432 Compounds with high
phosphoryl-transfer potential can couple carbon oxidation
to ATP synthesis 432 Ion gradients across membranes
provide an important form of cellular energy that can be
coupled to
ATP synthesis 433 Phosphates play a prominent role in
biochemical
processes 434 Energy from foodstuffs is extracted in three
stages 434
oxaloacetate 478 Oxaloacetate is shuttled into the
cytoplasm and
converted into phosphoenolpyruvate 480 The conversion of
fructose 1,6-bisphosphate into
fructose 6-phosphate and orthophosphate is an
irreversible step 480 The generation of free glucose is an
important
control point 481 Six high-transfer-potential phosphoryl
groups are spent in synthesizing glucose from pyruvate
481
16.4 Gluconeogenesis and Glycolysis Are
15.4 Metabolic Pathways Contain Many
Recurring Motifs 435 Activated carriers exemplify the modular Reciprocally Regulated 482 Energy charge determines
design and economy of metabolism 435 Many activated
whether glycolysis or
carriers are derived from vitamins 438 Key reactions are
gluconeogenesis will be most active 482 The balance
between glycolysis and gluconeogenesis in the liver is
reiterated throughout metabolism 440 Metabolic processes
sensitive to blood-glucose concentration 483 Substrate
are regulated in three principal ways 442 Aspects of
cycles amplify metabolic signals and
metabolism may have evolved from an
RNA world 444 CHAPTER 16 Glycolysis and produce heat 485 Lactate and alanine formed by
contracting muscle are used by other organs 485
Glycolysis and gluconeogenesis are evolutionarily
intertwined 487
Gluconeogenesis
449CHAPTER
Gluconeogenesis 449
16
Glycolysis
and
CHAPTER 17 The Citric Acid Cycle 495 CHAPTER 17 The
xxiv Contents
Citric Acid Cycle 495
The citric acid cycle harvests high-energy electrons 496
17.1 The Pyruvate Dehydrogenase Complex Links Glycolysis to
the Citric Acid Cycle 497
Mechanism: The synthesis of acetyl coenzyme A from
pyruvate requires three enzymes and five coenzymes 498
Contents xxiii
Flexible linkages allow lipoamide to move between
different active sites 500
17.2 The Citric Acid Cycle Oxidizes
Two-Carbon Units 501 Citrate synthase forms citrate from
The high-potential electrons of NADH enter the
respiratory chain at NADH-Q oxidoreductase 532 Ubiquinol
is the entry point for electrons from
FADH2 of flavoproteins 533 Electrons flow from ubiquinol to
cytochrome c through Q-cytochrome c oxidoreductase 533
The Q cycle funnels electrons from a two-electron carrier
to a one-electron carrier and pumps protons 535
Cytochrome c oxidase catalyzes the reduction of
molecular oxygen to water 535 Toxic derivatives of
molecular oxygen such as superoxide radicals are
scavenged by protective enzymes 538 Electrons can be
transferred between groups that are not in contact 540 The
conformation of cytochrome c has remained
essentially constant for more than a billion years 541 18.4 A
oxaloacetate and acetyl coenzyme A 502 Mechanism: The
Proton Gradient Powers the Synthesis of ATP 541
mechanism of citrate synthase prevents undesirable
ATP synthase is composed of a proton-conducting unit and
reactions 502 Citrate is isomerized into isocitrate 504
a catalytic unit 543 Proton flow through ATP synthase leads
Isocitrate is oxidized and decarboxylated to alpha
to the release of tightly bound ATP: The binding-change
ketoglutarate 504 Succinyl coenzyme A is formed by the
mechanism 544 Rotational catalysis is the world’s smallest
oxidative
molecular motor 546 Proton flow around the c ring powers
decarboxylation of alpha-ketoglutarate 505 A compound
ATP synthesis 546 ATP synthase and G proteins have
with high phosphoryl-transfer potential is generated from
several common features 548
succinyl coenzyme A 505 Mechanism: Succinyl coenzyme
18.5 Many Shuttles Allow Movement Across Mitochondrial
A synthetase transforms types of biochemical energy 506
Oxaloacetate is regenerated by the oxidation of succinate Membranes 549 Electrons from cytoplasmic NADH enter
mitochondria by shuttles 549 The entry of ADP into
507 The citric acid cycle produces high-transfer-potential
mitochondria is coupled to the exit of ATP by ATP-ADP
electrons, ATP, and CO2 508
translocase 550 Mitochondrial transporters for metabolites
17.3 Entry to the Citric Acid Cycle and Metabolism Through It have a
Are Controlled 510 The pyruvate dehydrogenase complex is
common tripartite structure 551 18.6 The Regulation of
regulated
Cellular Respiration Is Governed Primarily by the Need for ATP
allosterically and by reversible phosphorylation 511 The 552 The complete oxidation of glucose yields about
citric acid cycle is controlled at several points 512 Defects in
30 molecules of ATP 552 The rate of oxidative
the citric acid cycle contribute to the
phosphorylation is determined by the need for ATP 553
development of cancer 513
ATP synthase can be regulated 554 Regulated uncoupling
17.4 The Citric Acid Cycle Is a Source of
Biosynthetic Precursors 514 The citric acid cycle must be
capable of being rapidly replenished 514 The disruption of
pyruvate metabolism is the cause of beriberi and poisoning
by mercury and arsenic 515 The citric acid cycle may have
evolved from preexisting pathways 516
leads to the generation of heat 554 Oxidative
phosphorylation can be inhibited at many stages 556
Mitochondrial diseases are being discovered 557
Mitochondria play a key role in apoptosis 557 Power
transmission by proton gradients is a central motif of
17.5 The Glyoxylate Cycle Enables Plants and Bacteria to Grow
bioenergetics 558
on Acetate 516
CHAPTER 19 The Light Reactions of
CHAPTER 18 Oxidative Phosphorylation 523CHAPTER 18
Oxidative Phosphorylation 523
18.1 Eukaryotic Oxidative Phosphorylation Takes Place in
Mitochondria 524 Mitochondria are bounded by a double
membrane 524 Mitochondria are the result of an
endosymbiotic event 525 18.2 Oxidative Phosphorylation
Depends on Electron Transfer 526 The electron-transfer
potential of an electron is
measured as redox potential 526 A 1.14-volt potential
difference between NADH and molecular oxygen drives
electron transport through the chain and favors the
formation of a proton gradient 528
18.3 The Respiratory Chain Consists of Four Complexes: Three
Proton Pumps and a Physical Link to the Citric Acid Cycle 529
Iron–sulfur clusters are common components of
the electron transport chain 531
CHAPTER 19 The Light Reactions of
Photosynthesis 565 Photosynthesis 565
Photosynthesis converts light energy into chemical energy
566 19.1 Photosynthesis Takes Place in Chloroplasts 567 The
primary events of photosynthesis take place in thylakoid
membranes 567
Chloroplasts arose from an endosymbiotic event 568 19.2
Light Absorption by Chlorophyll Induces Electron Transfer 568
A special pair of chlorophylls initiate charge separation 569
Cyclic electron flow reduces the cytochrome of the reaction
center 572
19.3 Two Photosystems Generate a Proton Gradient and
Crassulacean acid metabolism permits growth in arid
ecosystems 601
NADPH in Oxygenic Photosynthesis 572 Photosystem II
transfers electrons from water to
plastoquinone and generates a proton gradient 572
Cytochrome bf links photosystem II to photosystem I 575
Photosystem I uses light energy to generate reduced
ferredoxin, a powerful reductant 575 Ferredoxin–NADP1
reductase converts NADP1 into NADPH 576
20.3 The Pentose Phosphate Pathway Generates NADPH and
Synthesizes Five-Carbon Sugars 601 Two molecules of NADPH
are generated in the
conversion of glucose 6-phosphate into ribulose
5-phosphate 602 The pentose phosphate pathway and
glycolysis
are linked by transketolase and transaldolase
19.4 A Proton Gradient across the Thylakoid Membrane Drives
602 Mechanism: Transketolase and transaldolase stabilize
ATP Synthesis 578 The ATP synthase of chloroplasts closely
carbanionic intermediates by different mechanisms 605
resembles those of mitochondria and prokaryotes 578 The
20.4 The Metabolism of Glucose 6-Phosphate by the Pentose
activity of chloroplast ATP synthase is regulated 579 Cyclic
electron flow through photosystem I leads to the production of Phosphate Pathway Is Coordinated with Glycolysis 607
ATP instead of NADPH 580 The absorption of eight photons
The rate of the pentose phosphate pathway is controlled
yields one O2, two NADPH, and three ATP molecules 581
by the level of NADP1 607 The flow of glucose 6-phosphate
depends on the need for NADPH, ribose 5-phosphate, and
19.5 Accessory Pigments Funnel Energy into Reaction Centers
ATP 608 The pentose phosphate pathway is required for
581 Resonance energy transfer allows energy to move from
rapid cell growth 610 Through the looking-glass: The
the site of initial absorbance to the reaction center 582 The
Calvin cycle and the pentose phosphate pathway are
components of photosynthesis are highly organized 583 Many
mirror images 610
herbicides inhibit the light reactions of
20.5 Glucose 6-Phosphate Dehydrogenase Plays a Key Role in
photosynthesis 584 19.6 The Ability to Convert Light into
Protection Against Reactive Oxygen Species 610 Glucose 6Chemical Energy Is Ancient 584
phosphate dehydrogenase deficiency
Artificial photosynthetic systems may provide clean,
causes a drug-induced hemolytic anemia 610 A deficiency
of glucose 6-phosphate dehydrogenase confers an
evolutionary advantage in some circumstances 612
Contents xxv
renewable energy 585
Muscle phosphorylase is regulated by the intracellular
energy charge 625 Biochemical characteristics of muscle
fiber types differ 625 Phosphorylation promotes the
conversion of
phosphorylase b to phosphorylase a 626 Phosphorylase
kinase is activated by phosphorylation and calcium ions
626
CHAPTER 20 The Calvin Cycle and the
CHAPTER 20 The Calvin Cycle and the
Pentose Phosphate Pathway 589 Pentose Phosphate
Pathway 589
20.1 The Calvin Cycle Synthesizes Hexoses from Carbon
Dioxide and Water 590 Carbon dioxide reacts with ribulose 1,5bisphosphate to form two molecules of 3-phosphoglycerate
591 Rubisco activity depends on magnesium and carbamate
592 Rubisco activase is essential for rubisco activity 593
Rubisco also catalyzes a wasteful oxygenase reaction:
Catalytic imperfection 593 Hexose phosphates are made from
phosphoglycerate, and ribulose 1,5-bisphosphate is
regenerated 594 Three ATP and two NADPH molecules are
used to bring carbon dioxide to the level of a hexose 597
Starch and sucrose are the major carbohydrate stores in
plants 597
20.2 The Activity of the Calvin Cycle Depends on
Environmental Conditions 598
Rubisco is activated by light-driven changes in
proton and magnesium ion concentrations 598 Thioredoxin
plays a key role in regulating the
Calvin cycle 599 The C4 pathway of tropical plants
accelerates
photosynthesis by concentrating carbon dioxide 599
CHAPTER 21 Glycogen Metabolism 617
21.3 Epinephrine and Glucagon Signal the Need for Glycogen
Breakdown 627 G proteins transmit the signal for the initiation
of
glycogen breakdown 627 Glycogen breakdown must be
rapidly turned off when necessary 629 The regulation of
glycogen phosphorylase became more sophisticated as
the enzyme evolved 629
21.4 Glycogen Is Synthesized and Degraded by Different
Pathways 630 UDP-glucose is an activated form of glucose
630 Glycogen synthase catalyzes the transfer of glucose from
UDP-glucose to a growing chain 630 A branching enzyme
forms a-1,6 linkages 631 Glycogen synthase is the key
regulatory enzyme in glycogen synthesis 632 Glycogen is an
efficient storage form of glucose 632
21.5 Glycogen Breakdown and Synthesis Are Reciprocally
Regulated 632 Protein phosphatase 1 reverses the regulatory
effects of kinases on glycogen metabolism 633 Insulin
stimulates glycogen synthesis by inactivating glycogen
synthase kinase 635 Glycogen metabolism in the liver
regulates the
blood-glucose level 635 A biochemical understanding of
glycogen-storage
diseases is possible 637
CHAPTER 22 Fatty Acid Metabolism
Glycogen metabolism is the regulated release and
storage of glucose 618
Glycogen
CHAPTER 21
Metabolism
617
21.1 Glycogen Breakdown Requires the Interplay of Several
Enzymes 619 Phosphorylase catalyzes the phosphorolytic
cleavage of glycogen to release glucose 1-phosphate 619
643
Mechanism: Pyridoxal phosphate participates in the
phosphorolytic cleavage of glycogen 620 A debranching
enzyme also is needed for the breakdown of glycogen 621
Phosphoglucomutase converts glucose 1-phosphate into
glucose 6-phosphate 622 The liver contains glucose 6phosphatase, a hydrolytic enzyme absent from muscle 622
21.2 Phosphorylase Is Regulated by Allosteric Interactions and
Reversible Phosphorylation 623 Liver phosphorylase produces
glucose for use by
other tissues 623
CHAPTER 22
22.6 Acetyl CoA Carboxylase Plays a Key Role in Controlling
Fatty Acid Metabolism 670 Acetyl CoA carboxylase is
regulated by conditions in the cell 671 Acetyl CoA
carboxylase is regulated by a variety of hormones 671
CHAPTER 23 Protein Turnover and
CHAPTER 23 Protein Turnover and
Fatty Acid Metabolism Amino Acid Catabolism 681 Amino Acid Catabolism 681
643 23.1 Proteins are Degraded to Amino Acids 682 The digestion of
Fatty acid degradation and synthesis mirror each other in
their chemical reactions 644
22.1 Triacylglycerols Are Highly Concentrated Energy Stores
645 Dietary lipids are digested by pancreatic lipases 645
Dietary lipids are transported in chylomicrons 646
22.2 The Use of Fatty Acids as Fuel Requires Three Stages of
Processing 647 Triacylglycerols are hydrolyzed by hormone
stimulated lipases 647 Free fatty acids and glycerol are
released
into the blood 648 Fatty acids are linked to coenzyme A
before they are oxidized 648 Carnitine carries long-chain
activated fatty acids into the mitochondrial matrix 649
Acetyl CoA, NADH, and FADH2 are generated in each
round of fatty acid oxidation 650
xxvi Contents
The complete oxidation of palmitate yields
106 molecules of ATP 652 22.3 Unsaturated and Odd-Chain
Fatty Acids Require Additional Steps for Degradation 652 An
isomerase and a reductase are required for the
oxidation of unsaturated fatty acids 652 Odd-chain fatty
acids yield propionyl CoA in the final thiolysis step 654
Vitamin B12 contains a corrin ring and a cobalt atom 654
Mechanism: Methylmalonyl CoA mutase catalyzes a
rearrangement to form succinyl CoA 655 Fatty acids are
also oxidized in peroxisomes 656 Ketone bodies are
formed from acetyl CoA when fat breakdown predominates
657 Ketone bodies are a major fuel in some tissues 658
Animals cannot convert fatty acids into glucose 660 Some
fatty acids may contribute to the development of
pathological conditions 661
22.4 Fatty Acids Are Synthesized by Fatty Acid Synthase 661
Fatty acids are synthesized and degraded by different
pathways 661 The formation of malonyl CoA is the committed
step in fatty acid synthesis 662 Intermediates in fatty acid
synthesis are attached to an acyl carrier protein 662 Fatty
acid synthesis consists of a series of condensation, reduction,
dehydration, and reduction reactions 662 Fatty acids are
synthesized by a multifunctional enzyme complex in animals
664 The synthesis of palmitate requires 8 molecules of acetyl
CoA, 14 molecules of NADPH, and 7 molecules of ATP 666
Citrate carries acetyl groups from mitochondria to the
cytoplasm for fatty acid synthesis 666 Several sources supply
NADPH for fatty acid synthesis 667 Fatty acid metabolism is
altered in tumor cells 667 22.5 The Elongation and Unsaturation
of Fatty Acids are Accomplished by Accessory Enzyme Systems
668 Membrane-bound enzymes generate unsaturated fatty
acids 668 Eicosanoid hormones are derived from
polyunsaturated fatty acids 669 Variations on a theme:
Polyketide and nonribosomal peptide synthetases resemble
fatty acid synthase 670
dietary proteins begins in the stomach and is completed in
the intestine 682 Cellular proteins are degraded at different
rates 682
23.2 Protein Turnover Is Tightly Regulated 683 Ubiquitin tags
proteins for destruction 683 The proteasome digests the
ubiquitin-tagged proteins 685 The ubiquitin pathway and the
proteasome have
prokaryotic counterparts 686 Protein degradation can be
used to regulate biological function 687
23.3 The First Step in Amino Acid Degradation Is the Removal
of Nitrogen 687 Alpha-amino groups are converted into
ammonium ions by the oxidative deamination of glutamate
687 Mechanism: Pyridoxal phosphate forms Schiff-base
intermediates in aminotransferases 689 Aspartate
aminotransferase is an archetypal pyridoxal dependent
transaminase 690 Blood levels of aminotransferases serve a
diagnostic function 691 Pyridoxal phosphate enzymes
catalyze a wide array of reactions 691 Serine and threonine
can be directly deaminated 692 Peripheral tissues transport
nitrogen to the liver 692 23.4 Ammonium Ion Is Converted into
Urea in Most Terrestrial Vertebrates 693 The urea cycle begins
with the formation of carbamoyl phosphate 693 Carbamoyl
phosphate synthetase is the key regulatory enzyme for urea
synthesis 694 Carbamoyl phosphate reacts with ornithine to
begin the urea cycle 694 The urea cycle is linked to
gluconeogenesis 696 Urea-cycle enzymes are evolutionarily
related to
enzymes in other metabolic pathways 696 Inherited
defects of the urea cycle cause
hyperammonemia and can lead to brain damage 697 Urea
is not the only means of disposing of excess nitrogen 698 23.5
Carbon Atoms of Degraded Amino Acids Emerge as Major
Metabolic Intermediates 698 Pyruvate is an entry point into
metabolism for a
number of amino acids 699 Oxaloacetate is an entry point
into metabolism for aspartate and asparagine 700 Alphaketoglutarate is an entry point into metabolism for fivecarbon amino acids 700 Succinyl coenzyme A is a point of
entry for several nonpolar amino acids 701 Methionine
degradation requires the formation of a key methyl donor,
S-adenosylmethionine 701 The branched-chain amino
acids yield acetyl CoA, acetoacetate, or propionyl CoA 701
Oxygenases are required for the degradation of
aromatic amino acids 703
23.6 Inborn Errors of Metabolism Can Disrupt Amino Acid
Degradation 705 Phenylketonuria is one of the most common
metabolic disorders 706 Determining the basis of the
neurological symptoms of phenylketonuria is an active area
of research 706
Part III SYNTHESIZING THE MOLECULES OF LIFE
CHAPTER 24 The Biosynthesis of Amino Acids 713 CHAPTER
24 The Biosynthesis of Amino Acids 713
Amino acid synthesis requires solutions to three key
biochemical problems 714
24.1 Nitrogen Fixation: Microorganisms Use ATP and a
Powerful Reductant to Reduce Atmospheric Nitrogen to
Ammonia 714
The iron–molybdenum cofactor of nitrogenase binds and
reduces atmospheric nitrogen 715 Ammonium ion is
assimilated into an amino acid
through glutamate and glutamine 717
24.2 Amino Acids Are Made from Intermediates of the Citric
Acid Cycle and Other Major Pathways 719 Human beings can
synthesize some amino acids but must obtain others from
their diet 719 Aspartate, alanine, and glutamate are formed by
the addition of an amino group to an alpha-ketoacid 720 A
common step determines the chirality of all
amino acids 721 The formation of asparagine from
aspartate requires an adenylated intermediate 721
Glutamate is the precursor of glutamine, proline, and
arginine 722 3-Phosphoglycerate is the precursor of serine,
cysteine, and glycine 722 Tetrahydrofolate carries
activated one-carbon units at several oxidation levels 723
S-Adenosylmethionine is the major donor of methyl groups
724 Cysteine is synthesized from serine and homocysteine
726 High homocysteine levels correlate with vascular
disease 726 Shikimate and chorismate are intermediates in
the biosynthesis of aromatic amino acids 727 Tryptophan
synthase illustrates substrate channeling in enzymatic
catalysis 729
24.3 Feedback Inhibition Regulates Amino Acid Biosynthesis
730 Branched pathways require sophisticated regulation 731
The sensitivity of glutamine synthetase to allosteric regulation
is altered by covalent modification 732
24.4 Amino Acids Are Precursors of Many
Biomolecules 734 Glutathione, a gamma-glutamyl peptide,
serves as a sulfhydryl buffer and an antioxidant 734 Nitric
oxide, a short-lived signal molecule, is formed from
arginine 735 Porphyrins are synthesized from glycine and
succinyl coenzyme A 736 Porphyrins accumulate in some
a pyrimidine nucleotide and is converted into
uridylate 746 Nucleotide mono-, di-, and triphosphates are
interconvertible 747 CTP is formed by amination of UTP 747
Salvage pathways recycle pyrimidine bases 748
25.2 Purine Bases Can Be Synthesized de Novo or Recycled by
Salvage Pathways 748 The purine ring system is assembled on
ribose
phosphate 749 The purine ring is assembled by successive
steps
of activation by phosphorylation followed by
displacement 749 AMP and GMP are formed from IMP 751
Enzymes of the purine synthesis pathway associate with
one another in vivo 752 Salvage pathways economize
intracellular energy
expenditure 752
25.3 Deoxyribonucleotides Are Synthesized by the Reduction of
Ribonucleotides Through a Radical Mechanism 753
Mechanism: A tyrosyl radical is critical to the action of
ribonucleotide reductase 753 Stable radicals other than
tyrosyl radical are employed by other ribonucleotide
reductases 755 Thymidylate is formed by the methylation
of
deoxyuridylate 755 Dihydrofolate reductase catalyzes the
regeneration of tetrahydrofolate, a one-carbon carrier 756
Several valuable anticancer drugs block the synthesis of
thymidylate 757
25.4 Key Steps in Nucleotide Biosynthesis Are Regulated by
Feedback Inhibition 758 Pyrimidine biosynthesis is regulated
by aspartate
transcarbamoylase 758 The synthesis of purine nucleotides
is controlled by feedback inhibition at several sites 758
The synthesis of deoxyribonucleotides is controlled by the
regulation of ribonucleotide reductase 759
25.5 Disruptions in Nucleotide Metabolism
Can Cause Pathological Conditions 760 The loss of adenosine
deaminase activity results in
severe combined immunodeficiency 760 Gout is induced
by high serum levels of urate 761 Lesch–Nyhan syndrome
is a dramatic consequence of mutations in a salvagepathway enzyme 761 Folic acid deficiency promotes birth
defects such as spina bifida 762
xxviii Contents
CHAPTER 26 The Biosynthesis of Membrane
CHAPTER 26 The Biosynthesis of Membrane Lipids and
inherited disorders of porphyrin metabolism 737
CHAPTER 25 Nucleotide Biosynthesis 743 CHAPTER 25
Steroids 767
Lipids and Steroids 767
Nucleotide Biosynthesis 743
Nucleotides can be synthesized by de novo or
salvage pathways 744
26.1 Phosphatidate Is a Common Intermediate in the
Contents xxvii Synthesis of Phospholipids and
Triacylglycerols 768 The synthesis of phospholipids requires
25.1 The Pyrimidine Ring Is Assembled de Novo or Recovered
by Salvage Pathways 744 Bicarbonate and other oxygenated
carbon compounds are activated by phosphorylation 745 The
side chain of glutamine can be hydrolyzed to
generate ammonia 745 Intermediates can move between
active sites by
channeling 745 Orotate acquires a ribose ring from PRPP
to form
an activated intermediate 769 Some phospholipids are
synthesized from an activated alcohol 770
Phosphatidylcholine is an abundant phospholipid 770
Excess choline is implicated in the development of heart
disease 771 Base-exchange reactions can generate
phospholipids 771 Sphingolipids are synthesized from
ceramide 772 Gangliosides are carbohydrate-rich
sphingolipids that contain acidic sugars 772 Sphingolipids
confer diversity on lipid structure and function 773
Respiratory distress syndrome and Tay–Sachs disease
result from the disruption of lipid metabolism 774 Ceramide
metabolism stimulates tumor growth 774 Phosphatidic acid
phosphatase is a key regulatory
enzyme in lipid metabolism 775
26.2 Cholesterol Is Synthesized from Acetyl Coenzyme A in
Three Stages 776 The synthesis of mevalonate, which is
activated as isopentenyl pyrophosphate, initiates the
synthesis
of cholesterol 776 Squalene (C30) is synthesized from six
molecules of isopentenyl pyrophosphate (C5) 777
Squalene cyclizes to form cholesterol 778
26.3 The Complex Regulation of Cholesterol Biosynthesis Takes
Place at Several Levels 779 Lipoproteins transport cholesterol
and triacylglycerols throughout the organism 782 Low-density
lipoproteins play a central role in
cholesterol metabolism 784 The absence of the LDL
receptor leads to
hypercholesterolemia and atherosclerosis 784 Mutations in
the LDL receptor prevent LDL release and result in
receptor destruction 785 Cycling of the LDL receptor is
regulated 787 HDL appears to protect against
atherosclerosis 787 The clinical management of cholesterol
levels can be understood at a biochemical level 788
26.4 Important Derivatives of Cholesterol Include Bile Salts and
Steroid Hormones 788 Letters identify the steroid rings and
numbers identify the carbon atoms 790 Steroids are
hydroxylated by cytochrome P450
monooxygenases that use NADPH and O2 790 The
cytochrome P450 system is widespread and
performs a protective function 791 Pregnenolone, a
precursor of many other steroids,
is formed from cholesterol by cleavage of its side chain 792
Progesterone and corticosteroids are synthesized from
pregnenolone 792 Androgens and estrogens are
synthesized from
pregnenolone 792 Vitamin D is derived from cholesterol by
the ring
Beneficially Alters the Biochemistry of Cells 813
Mitochondrial biogenesis is stimulated by muscular activity
813 Fuel choice during exercise is determined by the
intensity and duration of activity 813 27.5 Food Intake and
Starvation Induce Metabolic Changes 816
The starved–fed cycle is the physiological response to a
fast 816 Metabolic adaptations in prolonged starvation
minimize protein degradation 818
27.6 Ethanol Alters Energy Metabolism in the Liver 819 Ethanol
metabolism leads to an excess of NADH 820 Excess ethanol
consumption disrupts vitamin
metabolism 821
CHAPTER 28 DNA Replication, Repair,
CHAPTER 28 DNA Replication, Repair,
and Recombination 827 and Recombination 827
28.1 DNA Replication Proceeds by the
Polymerization of Deoxyribonucleoside
Triphosphates Along a Template 828
DNA polymerases require a template and a primer 829 All
DNA polymerases have structural features in
common 829 Two bound metal ions participate in the
polymerase reaction 829 The specificity of replication is
dictated by
complementarity of shape between bases 830 An RNA
primer synthesized by primase enables
DNA synthesis to begin 831 One strand of DNA is made
continuously, whereas the other strand is synthesized in
fragments 831 DNA ligase joins ends of DNA in duplex
regions 832 The separation of DNA strands requires
specific
helicases and ATP hydrolysis 832
28.2 DNA Unwinding and Supercoiling Are Controlled by
splitting activity of light 794 Topoisomerases 833 The linking number of DNA, a topological
property, determines the degree of supercoiling 835
CHAPTER 27 The Integration of Metabolism 801 CHAPTER Topoisomerases prepare the double helix for
unwinding 836 Type I topoisomerases relax supercoiled
27 The Integration of Metabolism 801
structures 836 Type II topoisomerases can introduce
negative
supercoils through coupling to ATP hydrolysis 837 28.3
27.1 Caloric Homeostasis Is a Means of Regulating Body
Weight 802 27.2 The Brain Plays a Key Role in Caloric
DNA Replication Is Highly Coordinated 839 DNA replication
Homeostasis 804 Signals from the gastrointestinal tract induce requires highly processive
feelings of satiety 804 Leptin and insulin regulate longpolymerases 839 The leading and lagging strands are
term control over caloric homeostasis 805 Leptin is one of
synthesized in a coordinated fashion 840 DNA replication
several hormones secreted by
in Escherichia coli begins at a
adipose tissue 806 Leptin resistance may be a contributing
unique site 842 DNA synthesis in eukaryotes is initiated at
factor to
multiple sites 843 Telomeres are unique structures at the
obesity 806 Dieting is used to combat obesity 807 27.3
ends of linear chromosomes 844 Telomeres are replicated
Diabetes Is a Common Metabolic Disease Often Resulting from
by telomerase, a specialized polymerase that carries its
own RNA template 845
Obesity 807 Insulin initiates a complex signal-transduction
pathway in muscle 808 Metabolic syndrome often precedes 28.4 Many Types of DNA Damage Can Be
type 2 diabetes 809 Excess fatty acids in muscle modify Repaired 845 Errors can arise in DNA replication 846 Bases
metabolism 810 Insulin resistance in muscle facilitates
can be damaged by oxidizing agents, alkylating agents,
pancreatic failure 810 Metabolic derangements in type 1
and light 846 DNA damage can be detected and repaired
by a variety of systems 847 The presence of thymine
diabetes result from
instead of uracil in DNA permits the repair of deaminated
insulin insufficiency and glucagon excess 812 27.4 Exercise
cytosine 849 Some genetic diseases are caused by the
expansion of repeats of three nucleotides 850 Many
cancers are caused by the defective repair of DNA 850
Many potential carcinogens can be detected by their
mutagenic action on bacteria 852
28.5 DNA Recombination Plays Important Roles in Replication,
Repair, and Other Processes 852 RecA can initiate
recombination by promoting
strand invasion 853 Some recombination reactions
proceed through
Holliday-junction intermediates 854
to Both Mechanism and Evolution 886
xxx Contents
CHAPTER 30 Protein Synthesis 893 CHAPTER 30 P iS hrotein
Synthesis 893
30.1 Protein Synthesis Requires the Translation of Nucleotide
Sequences into Amino Acid Sequences 894 The synthesis of
Contents xxix long proteins requires a low error frequency 894 Transfer
RNA molecules have a common design 895 Some transfer
RNA molecules recognize more than one codon because of
wobble in base-pairing 897
CHAPTER 29 RNA S h i d P RNA Synthesis and Processing
30.2 Aminoacyl Transfer RNA Synthetases
Read the Genetic Code 898 Amino acids are first activated by
adenylation 898 Aminoacyl-tRNA synthetases have highly
RNA synthesis comprises three stages: Initiation,
discriminating amino acid activation sites 899 Proofreading
elongation, and termination 860 29.1 RNA Polymerases
by aminoacyl-tRNA synthetases
Catalyze Transcription 861 RNA chains are formed de novo
increases the fidelity of protein synthesis 900 Synthetases
and grow in the
recognize various features of transfer
59-to-39 direction 862 RNA polymerases backtrack and
RNA molecules 901 Aminoacyl-tRNA synthetases can be
divided into two classes 901
correct errors 863 RNA polymerase binds to promoter sites
on the
30.3 The Ribosome Is the Site of Protein Synthesis 902
DNA template to initiate transcription 864 Sigma subunits
Ribosomal RNAs (5S, 16S, and 23S rRNA) play a central
of RNA polymerase recognize
role in protein synthesis 903 Ribosomes have three tRNAbinding sites that bridge the 30s and 50s subunits 905 The
promoter sites 865 RNA polymerases must unwind the
start signal is usually AUG preceded by several bases that
template
pair with 16S rRNA 905 Bacterial protein synthesis is
double helix for transcription to take place 865 Elongation
initiated by
takes place at transcription bubbles that move along the
formylmethionyl transfer RNA 906 Formylmethionyl-tRNAf
DNA template 866 Sequences within the newly transcribed
is placed in the P site of the ribosome in the formation of
RNA signal termination 866 Some messenger RNAs
the 70S initiation complex 907
directly sense metabolite
Elongation factors deliver aminoacyl-tRNA to the ribosome
concentrations 867 The rho protein helps to terminate the
907 Peptidyl transferase catalyzes peptide-bond synthesis
transcription of some genes 868 Some antibiotics inhibit
908 The formation of a peptide bond is followed by the
transcription 869 Precursors of transfer and ribosomal RNA
GTP driven translocation of tRNAs and mRNA 909 Protein
are cleaved and chemically modified after transcription in
synthesis is terminated by release factors that read stop
prokaryotes 870
codons 910
29.2 Transcription in Eukaryotes Is Highly Regulated 871 Three
30.4 Eukaryotic Protein Synthesis Differs from Bacterial Protein
types of RNA polymerase synthesize RNA in eukaryotic
Synthesis Primarily in Translation Initiation 911 Mutations in
cells 872 Three common elements can be found in the
RNA polymerase II promoter region 874 The TFIID protein initiation factor 2 cause a curious
pathological condition 913 30.5 A Variety of Antibiotics and
complex initiates the assembly of the active transcription
Toxins
Can Inhibit Protein Synthesis 913
complex 874 Multiple transcription factors interact with
Some antibiotics inhibit protein synthesis 914 Diphtheria
eukaryotic promoters 875 Enhancer sequences can
stimulate transcription at
toxin blocks protein synthesis in
start sites thousands of bases away 876
eukaryotes by inhibiting translocation 914 Ricin fatally
29.3 The Transcription Products of Eukaryotic Polymerases Are modifies 28S ribosomal RNA 915 30.6 Ribosomes Bound to the
859CHAPTER 29 RNA Synthesis and Processing 859
Processed 876 RNA polymerase I produces three ribosomal
Endoplasmic Reticulum Manufacture Secretory and Membrane
Proteins 915 Protein synthesis begins on ribosomes that are
RNAs 877 RNA polymerase III produces transfer RNA 877
The product of RNA polymerase II, the pre-mRNA transcript, free in the cytoplasm 916 Signal sequences mark proteins for
translocation
acquires a 59 cap and a 39 poly(A) tail 878 Small regulatory
across the endoplasmic reticulum membrane 916
RNAs are cleaved from larger precursors 879 RNA editing
changes the proteins encoded by mRNA 879 Sequences at
the ends of introns specify splice sites in mRNA precursors
880 Splicing consists of two sequential transesterification
Transport vesicles carry cargo proteins to their final
reactions 881 Small nuclear RNAs in spliceosomes catalyze
the
splicing of mRNA precursors 882 Transcription and
processing of mRNA are coupled 883 Mutations that affect
pre-mRNA splicing cause disease 884 Most human predestination 918
mRNAS can be spliced in alternative
ways to yield different proteins 885
29.4 The Discovery of Catalytic RNA was Revealing in Regard
CHAPTER 31 The Control of Gene Expression
CHAPTER 31 The Control of Gene Expression in
Prokaryotes 925
Controlled at Posttranscriptional Levels 954 Genes associated
in Prokaryotes 925
31.1 Many DNA-Binding Proteins Recognize Specific DNA
Sequences 926 The helix-turn-helix motif is common to many
prokaryotic DNA-binding proteins 927 31.2 Prokaryotic
with iron metabolism are translationally regulated in
animals 954 Small RNAs regulate the expression of many
eukaryotic genes 956
Part IV RESPONDING TO ENVIRONMENTAL
DNA-Binding Proteins Bind Specifically to Regulatory Sites in
Operons 927 An operon consists of regulatory elements and
CHANGES
protein-encoding genes 928 The lac repressor protein in
the absence of lactose
CHAPTER 33 Sensory Systems 961 CHAPTER 33 Sensory
binds to the operator and blocks transcription 929 Ligand
Systems 961
binding can induce structural changes in
regulatory proteins 930 The operon is a common regulatory
unit in prokaryotes 930 Transcription can be stimulated by 33.1 A Wide Variety of Organic Compounds Are Detected by
proteins that contact RNA polymerase 931
Olfaction 962 Olfaction is mediated by an enormous family of
seven-transmembrane-helix receptors 962 Odorants are
31.3 Regulatory Circuits Can Result in Switching Between
Patterns of Gene Expression 932 The l repressor regulates its decoded by a combinatorial mechanism 964 33.2 Taste Is a
own expression 932 A circuit based on the l repressor and Cro Combination of Senses That Function by Different Mechanisms
forms
966 Sequencing of the human genome led to the discovery of
a genetic switch 933 Many prokaryotic cells release
a large family of 7TM bitter receptors 967 A heterodimeric
chemical signals that regulate gene expression in other
7TM receptor responds to sweet
cells 933 Biofilms are complex communities of prokaryotes
compounds 968 Umami, the taste of glutamate and
934
aspartate, is mediated by a heterodimeric receptor related
31.4 Gene Expression Can Be Controlled at Posttranscriptional
to the sweet receptor 969 Salty tastes are detected
primarily by the passage of sodium ions through channels
Levels 935 Attenuation is a prokaryotic mechanism for
regulating transcription through the modulation of nascent
969 Sour tastes arise from the effects of hydrogen ions
(acids) on channels 969
RNA secondary structure 935 CHAPTER 32 The Control of
33.3 Photoreceptor Molecules in the Eye Detect Visible Light
970 Rhodopsin, a specialized 7TM receptor, absorbs
Gene Expression
CHAPTER 32 The Control of Gene Expression in
Eukaryotes 941
in Eukaryotes 941
visible light 970 Light absorption induces a specific
isomerization of bound 11-cis-retinal 971 Light-induced
lowering of the calcium level coordinates recovery 972
Color vision is mediated by three cone receptors that are
homologs of rhodopsin 973 Rearrangements in the genes
for the green and red pigments lead to “color blindness”
974
33.4 Hearing Depends on the Speedy Detection of Mechanical
32.1 Eukaryotic DNA Is Organized into Chromatin 943 Stimuli 975 Hair cells use a connected bundle of stereocilia to
Nucleosomes are complexes of DNA and histones 943 DNA
detect tiny motions 975
Contents xxxi
wraps around histone octamers to form
nucleosomes 943 32.2 Transcription Factors Bind DNA and
Regulate Transcription Initiation 945
A range of DNA-binding structures are employed by
eukaryotic DNA-binding proteins 945 Activation domains
interact with other proteins 946 Multiple transcription
factors interact with eukaryotic regulatory regions 946
Enhancers can stimulate transcription in specific
cell types 946 Induced pluripotent stem cells can be
generated by introducing four transcription factors into
differentiated cells 947 32.3 The Control of Gene
Expression Can Require Chromatin Remodeling 948
Mechanosensory channels have been identified in
Drosophila and vertebrates 976
33.5 Touch Includes the Sensing of Pressure, Temperature, and
Other Factors 977 Studies of capsaicin reveal a receptor for
sensing
high temperatures and other painful stimuli 977 CHAPTER 34
The Immune System 981CHAPTER 34 The Immune System
981
The methylation of DNA can alter patterns of gene
expression 949 Steroids and related hydrophobic
molecules pass through membranes and bind to DNAbinding receptors 949 Nuclear hormone receptors regulate
transcription by recruiting coactivators to the transcription
complex 950 Steroid-hormone receptors are targets for
drugs 951 Chromatin structure is modulated through
covalent modifications of histone tails 952 Histone
deacetylases contribute to transcriptional
repression 953
32.4 Eukaryotic Gene Expression Can Be
Innate immunity is an evolutionarily ancient defense
system 982 The adaptive immune system responds by
using the principles of evolution 984
34.1 Antibodies Possess Distinct Antigen-Binding and Effector
Units 985
34.2 Antibodies Bind Specific Molecules Through
Hypervariable Loops 988 The immunoglobulin fold consists of
a beta-sandwich framework with hypervariable loops 988 Xray analyses have revealed how antibodies bind
antigens 989 Large antigens bind antibodies with
numerous
CHAPTER 36 Drug Development 1033
interactions 990
34.3 Diversity Is Generated by Gene
Rearrangements 991 J (joining) genes and D (diversity) genes
increase
antibody diversity 991 More than 108 antibodies can be
formed by
combinatorial association and somatic mutation 992 The
oligomerization of antibodies expressed on the surfaces of
immature B cells triggers antibody secretion 993 Different
classes of antibodies are formed by the
hopping of VH genes 994 34.4 Major-Histocompatibility-
Complex Proteins Present Peptide Antigens on Cell Surfaces for
Recognition by T-Cell Receptors 995
Peptides presented by MHC proteins occupy a deep
groove flanked by alpha helices 996 T-cell receptors are
antibody-like proteins containing variable and constant
regions 998 CD8 on cytotoxic T cells acts in concert with Tcell receptors 998 Helper T cells stimulate cells that
display foreign
peptides bound to class II MHC proteins 1000 Helper T
cells rely on the T-cell receptor and CD4 to recognize
foreign peptides on antigen-presenting cells 1000 MHC
proteins are highly diverse 1002 Human immunodeficiency
viruses subvert the immune system by destroying helper T
cells 1003
34.5 The Immune System Contributes to the
Prevention and the Development of
Human Diseases 1004 T cells are subjected to positive and
negative selection in the thymus 1004 Autoimmune
diseases result from the generation of immune responses
against self-antigens 1005
xxxii Contents
36.1 The Development of Drugs Presents Huge Challenges 1034
Drug candidates must be potent and selective
modulators of their targets 1035
Drugs must have suitable properties to reach
their targets 1036 Toxicity can limit drug effectiveness 1040
36.2 Drug Candidates Can Be Discovered by Serendipity,
Screening, or Design 1041 Serendipitous observations can
drive drug
development 1041 Natural products are a valuable source
of drugs and drug leads 1043 Screening libraries of
synthetic compounds expands the opportunity for
identification of drug leads 1044 Drugs can be designed on
the basis of three-dimensional structural information about
their targets 1046
36.3 Analyses of Genomes Hold Great Promise for Drug
Discovery 1048 Potential targets can be identified in the
human
proteome 1048 Animal models can be developed to test the
validity of potential drug targets 1049 Potential targets can
be identified in the genomes of pathogens 1050 Genetic
differences influence individual responses to drugs 1050
36.4 The Clinical Development of Drugs Proceeds Through
Several Phases 1051 Clinical trials are time consuming and
expensive 1052 The evolution of drug resistance can limit
the utility of drugs for infectious agents and cancer 1053
Answers to Problems A1 Selected Readings B1 Index C1
The immune system plays a role in cancer prevention 1005
Vaccines are a powerful means to prevent and
eradicate disease 1006 CHAPTER 35 Molecular Motors
1011 CHAPTER 35 Molecular Motors 1011
35.1 Most Molecular-Motor Proteins Are Members of the PLoop NTPase Superfamily 1012 Molecular motors are generally
oligomeric proteins with an ATPase core and an extended
structure 1012 ATP binding and hydrolysis induce changes in
the conformation and binding affinity of motor proteins 1014
35.2 Myosins Move Along Actin Filaments 1016 Actin is a polar,
self-assembling, dynamic polymer 1016 Myosin head
domains bind to actin filaments 1018 Motions of single motor
proteins can be directly
observed 1018 Phosphate release triggers the myosin
power stroke 1019 Muscle is a complex of myosin and actin
1019 The length of the lever arm determines motor velocity
1022
35.3 Kinesin and Dynein Move Along Microtubules 1022
Microtubules are hollow cylindrical polymers 1022 Kinesin
motion is highly processive 1024
35.4 A Rotary Motor Drives Bacterial Motion 1026 Bacteria swim
by rotating their flagella 1026 Proton flow drives bacterial
flagellar rotation 1026 Bacterial chemotaxis depends on
reversal of the
direction of flagellar rotation 1028 CHAPTER 36 Drug
Biochemistry: An Evolving Science
1
CHAPTER
C
H
O
H2 C
HN
OC
CH
2
+
H+
C
O
—
C
H
O
H2 C
HN
OC
CH
C
2
O
H
Chemistry in action.Human activities require energy. The
interconversionof different forms of energy requires large biochemical
machines comprising many thousands of atoms such as the complex
shown above. Yet, the functions of these elaborate assemblies depend on
simple chemical processes such as the protonationand deprotonationof
the carboxylic acid groups shown on the right. The photograph is of
Nobel Prize winners Peter Agre , M.D., and Carol Greider , Ph.D., who
used, respectively, biochemical techniques to reveal key mechanisms of
how water is transported into and out of cells, and how chromosomes are
replicated faithfully. [Keith Weller for Johns Hopkins Medicine.]
B
iochemistry is the study of the chemistry of
life processes. Since the
the great unity of all living things at the
biochemical level.
1.1 Biochemical Unity Underlies Biological
discovery that biological molecules such as urea Diversity
could be synthesized from nonliving components
in 1828, scientists have explored the chemistry of The biological world is magnificently diverse. The
animal kingdom is rich with species ranging from
life with great intensity. Through these
nearly microscopic insects to elephants and
investigations, many of the most fundamental
whales. The plant kingdom includes species as
mysteries of how living things function at a
small and relatively
biochemical level have now been solved.
However, much remains to be investigated. As is
often the case, each discovery raises at least as O U T L I N E
many new questions as it answers. Furthermore,
1.1 Biochemical Unity Underlies Biological Diversity
we are now in an age of unprecedented
opportunity for the application of our tremendous 1.2 DNA Illustrates the Interplay BetweenForm and Function
knowledge of biochemistry to problems in
1.3 Concepts from Chemistry Explain the Properties of
medicine, dentistry, agriculture, forensics,
Biological
Molecules
anthropology, envi ronmental sciences, alternative
energy, and many other fields. We begin our
1.4 The Genomic Revolution Is Transforming Biochemistry,
journey into biochemistry with one of the most
Medicine, and Other Fields
startling discoveries of the past century: namely,
simple as algae and as large and from cells suggested that these
CHAPTER 1 Biochemistry: An Evolving Science
complex as giant sequoias. This
diverse organisms might have
diversity extends further when we more in common than is apparent
descend into the microscopic
from their outward appearance.
world. Organisms such as
With the development of
protozoa, yeast, and bacteria are biochemistry, this suggestion has
present with great diversity in
been tremendously supported and
water, in soil, and on or within
expanded. At the biochemical level,
larger organisms. Some organisms all organisms have many common
can survive and even thrive in
features (Figure 1.1).
seemingly hostile environments
As mentioned earlier, biochemistry
such as hot springs and glaciers. is the study of the chemistry of life
The development of the
processes. These processes entail
microscope revealed a key unifying the interplay of two different
feature that underlies this diversity. classes of molecules: large
CH2OH
Large organisms are built up of
molecules such as proteins and
O
cells, resembling, to some extent, nucleic acids, referred to as
single-celled microscopic
biological macromolecules, and
OH
CH2OH
organisms. The construction of ani low-molecular-weight molecules
mals, plants, and microorganisms such as glu
HO C H
referred to as metabolites, things. For example,
built from the same set of
HO
that are chemically
deoxyribonucleic acid
20 building blocks in all
OH
transformed in biological (DNA) stores genetic
organisms. Furthermore,
OH
processes.
information in all cellular proteins that play similar
Glucose
Members of both these organisms. Proteins, the roles in different
classes of molecules are macromol ecules that are organisms often have very
CH2OH
key participants in most similar three dimensional
Glycerol
common, with minor
cose and glycerol,
biological processes, are structures (Figure 1.1).
variations, to all living
2
Sulfolobus archaea Arabidopsis thaliana Homo sapiens
1
FIGURE 1.1 Biological diversity and similarity. The shape of a key molecule in gene
regulation (the TATA-box-binding protein) is similar in three very different organisms that are
separated from one another by billions of years of evolution. [(Left) Eye of Science/Science
Source; (middle) Holt Studios/Photo Researchers; (right) Time Life Pictures/Getty Images.]
lc
r
a
u
m
u
n
s
m
a
i
de
s
u
c
n
ip
o
o
m
a
r
h
ti
s
c
a
iM
E
ll
w
e
i
e
s
3
m
o
r
iD
s
s
o
ht
n
gr
o
f
H
s
c
r
C
Oxygen
atmosphere
forming
o
s
r
i
g
1.1 Unity and
Diversity
n
c
a
i
n
a
e
gr
b
o
s
n
M
4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 Billions of years
FIGURE 1.2 A possible time line for biochemical evolution. Selected key events are indicated. Note
that life on Earth began approximately 3.5 billion years ago, whereas human beings emerged quite
recently.
Key metabolic processes also are common to many organisms. For
example, the set of chemical transformations that converts glucose
and oxy gen into carbon dioxide and water is essentially identical in
simple bacteria such as Escherichia coli (E. coli) and human beings.
Even processes that appear to be quite distinct often have common
features at the biochemical level. Remarkably, the biochemical
processes by which plants capture light energy and convert it into
more-useful forms are strikingly similar to steps used in animals to
capture energy released from the breakdown of glucose.
These observations overwhelmingly suggest that all living things
on Earth have a common ancestor and that modern organisms have
evolved from this ancestor into their present forms. Geological and
biochemical find ings support a time line for this evolutionary path
(Figure 1.2). On the basis of their biochemical characteristics, the
diverse organisms of the modern world can be divided into three
fundamental groups called domains: Eukarya (eukaryotes), Bacteria,
and Archaea . Domain Eukarya comprises all multicel lular
organisms, including human beings as well as many microscopic
unicel lular organisms such as yeast. The defining characteristic of
eukaryotes is the presence of a well-defined nucleus within each
cell. Unicellular organisms such as bacteria, which lack a nucleus,
are referred to as prokaryotes . The pro karyotes were reclassified as
two separate domains in response to Carl Woese’s discovery in
1977 that certain bacteria-like organisms are biochemi cally quite
distinct from other previously characterized bacterial species. These
organisms, now recognized as having diverged from bacteria early in
evolution, are the archaea . Evolutionary
paths from a common ancestor to modern
organisms can
BACTERIA EUKARYA ARCHAEA
be deduced on the basis of biochemical information. One
s
s
s
such path is shown in Figure 1.3.
e
u
m
u
c
c
u
b
c
y
i
Much of this book will explore the chemical reactions
a
a
r
o
o
l
l
i
m
e
c
l
t
g
h
o
o
e
c
r
c
s
o
and the associated biological macromolecules and metab
i
n
n
a
a
e
r
u
a
o
o
l
b
h
e
a
l
h
i
c
o
olites that are found in biological processes common to all
h
h
t
m
l
m
c
a
l
c
c
c
e
o
a
a
r
a
a
s
e
E
S
B
H
S
Z
M
A
H
organisms. The unity of life at the biochemical level
makes this approach possible. At the same time, different
organisms have specific needs, depending on the particu
lar biological niche in which they evolved and live. By
comparing and contrasting details of particular biochemi
cal pathways in different organisms, we can learn how
biological challenges are solved at the biochemical level.
In most cases, these challenges are addressed by the adap
tation of existing macromolecules to new roles rather than
by the evolution of entirely new ones.
Biochemistry has been greatly enriched by our for many of the advances in biochemistry and many
ability to examine the three-dimensional structures of other fields, extending to the present.
biological macromolecules in great detail. Some of The structure of DNA powerfully illustrates a basic
these structures
principle common to all biological macromolecules:
the intimate relation between structure and function.
FIGURE 1.3 The tree of life. A possible evolutionary path from a common
The remarkable properties of this chemical
ancestor approximately 3.5 billion years ago at the bottom of the tree to
substance allow it to function as a very efficient and
organisms found in the modern world at the top.
4
robust vehicle for storing information. We start with
an examination of the covalent structure of DNA and
CHAPTER 1 Biochemistry: An Evolving Science
its exten
are simple and elegant, whereas others are
incredibly complicated. In any case, these structures sion into three dimensions.
provide an essential framework for understanding
function. We begin our exploration of the interplay
DNA is constructed from four building blocks
between structure and function with the genetic
DNA is a linear polymer made up of four different
material, DNA.
types of monomers. It has a fixed backbone from
which protrude variable substituents , referred to as
bases (Figure 1.4). The backbone is built of
1.2 DNA Illustrates the Interplay Between
repeating sugar–phosphate units. The sugars are
Form and Function
molecules of deoxyribose from which DNA receives
its name. Each sugar is connected to two phosphate
A fundamental biochemical feature common to all groups through different linkages. Moreover, each
cellular organisms is the use of DNA for the storage sugar is oriented in the same way, and so each DNA
of genetic information. The discovery that DNA plays strand has directionality, with one end distinguishable
this central role was first made in studies of bacteria from the other. Joined to each deoxyribose is one of
in the 1940s. This discovery was followed by a
four possible bases: adenine (A), cyto sine (C),
compelling proposal for the three-dimensional
guanine (G), and thymine (T).
structure of DNA in 1953, an event that set the stage
NH2
N
NH2
H
N
N
H
N
H
NN
O
N
N
H
N
O
O
NH
H CH3 N
H
NH2
O
NH
Adenine (A) Cytosine (C) Guanine (G) Thymine (T)
These bases are connected to the sugar components in the DNA
backbone through the bonds shown in black in Figure 1.4. All four
bases are planar but differ significantly in other respects. Thus, each
monomer of DNA consists of a sugar–phosphate unit and one of four
bases attached to the sugar. These bases can be arranged in any
order along a strand of DNA.
base1 base2 base3
OO
O
FIGURE 1.4 Covalent structure of
O
DNA. Each unit of the polymeric
structure
O
OO
OO
is composed of a sugar
(deoxyribose), a
phosphate, and a variable base that
O
P
P
P
OO
protrudes from the sugar–phosphate backbone.
O O –– –
OO
Sugar Phosphate
Two single strands of DNA combine to form a double helix
Most DNA molecules consist of not one but two
strands (Figure 1.5). In 1953, James Watson and
Francis Crick deduced the arrangement of these
strands and proposed a three-dimensional
structure for DNA molecules. This structure is a
carbon or carbon–nitrogen bonds that define the
double helix composed of two inter
struc
tures of the bases themselves. Such weak
twined strands arranged such that the sugar–
bonds
are crucial to biochemical systems; they
phosphate backbone lies on the outside and the
are
weak
enough to be reversibly broken in
bases on the inside. The key to this structure is
biochemical
processes, yet they are strong
that the bases form specific base pairs ( bp ) held
enough,
particularly
when many form simultane
together by hydrogen bonds (Section 1.3):
ously,
to
help
stabilize
specific structures such as
adenine pairs with thymine (A–T) and guanine
the
double
helix.
pairs with cytosine (G–C), as shown in Figure 1.6.
Hydrogen bonds are much weaker than covalent
FIGURE 1.5 The double helix. The double-helical structure of DNA
bonds such as the carbon–
proposed by Watson and Crick. The sugar–phosphate backbones of
5
the two chains are shown in red and blue, and the bases are shown in
green, purple, orange, and yellow. The two strands are antiparallel,
running in opposite directions with respect to the axis of the double
helix, as indicated by the arrows.
1.2 DNA: Form and Function
H
N
H
O
N
N
N
H
N
NHN
H
CH3
N
O
N
N
H
N
O
N
N
N
O
N
H
H
Guanine (G) Cytosine (C)
Adenine (A) Thymine (T)
shape (Figure 1.6) and thus fit equally well
into the center of the double-helical
(A – T), and guanine with cytosine (G – C). The dashed green
structure of any sequence. Without any
lines represent hydrogen bonds.
constraints, the sequence of bases along a
DNA strand can act as an efficient means
of storing information. Indeed, the
DNA structure explains heredity and the storage of sequence of bases along DNA strands is
information
how genetic information is stored.
The structure proposed by Watson and
Crick has two properties of central
importance to the role of DNA as the
hereditary material. First, the structure is
compatible with any sequence of bases.
While the bases are distinct in structure,
the base pairs have essentially the same
FIGURE 1.6 Watson–Crick base pairs. Adenine pairs with thymine
G
C
A
sequences and protein activities
base-pairing, one strand C
C
The DNA
of the
molecules within cells. the
Newly
sequence
ribonucleic that carry out Second,
sequence of synthesized
determines acid (RNA) most of the because of bases along strands
C
A
G
the
sequence along G
wrote: “It has not specific pairing
the
other
strand.
GG
escaped our
completely
Crick so coyly
As
Watson
and
C
determines the
notice that the
T
T
C
T
A
T
A
A
G
we have postulated immediately suggests a possible copying mechanism for
T
the genetic material.” Thus, if the DNA double helix is separated into two
G
C
1.3 Concepts from Chemistry Explain the
Properties of Biological Molecules
single strands, each strand can act as a template
for the generation of its partner strand through
specific base-pair formation (Figure 1.7). The three We have seen how a chemical insight into the
dimensional structure of DNA beautifully illustrates hydrogen-bonding capabili ties of the bases of DNA
the close connection between molecular form and led to a deep understanding of a fundamental
function.
biological process. To lay the groundwork for the
rest of the book, we begin our study of
FIGURE 1.7 DNA replication. If a DNA molecule is separated into two
strands, each strand can act as the template for the generation of its partner biochemistry by examining selected concepts from
strand.
chemistry and showing how these concepts apply
6
to biological systems. The concepts include the
CHAPTER 1 Biochemistry:
types of chemical bonds; the structure of water, the
An Evolving Science
solvent in which most biochemical processes take
place; the First and Second Laws of
Thermodynamics; and the principles of acid–base
chemistry.
The formation of the DNA double helix as a key example
We will use these concepts to examine an
archetypical biochemical process— namely, the
formation of a DNA double helix from its two
component strands. The process is but one of
many examples that could have been chosen to
illustrate these topics. Keep in mind that, although
the specific discussion is about DNA and doublehelix formation, the concepts consid
ered are quite general and will apply to many other
classes of molecules and processes that will be
discussed in the remainder of the book. In the
course of these discussions, we will touch on the
properties of water and the concepts of pK a and
buffers that are of great importance to many
aspects of biochemistry.
The double helix can form from its component strands
G
CG
C
GATTAAT
CTAATTA
GATTAAT
CGATTAAT and
The discovery that DNA from natural sources
exists in a double-helical form with Watson–Crick
base pairs suggested, but did not prove, that such
double helices would form spontaneously outside
biological systems. Suppose that two short strands
of DNA were chemically synthesized to
have complementary sequences so that they could,
in principle, form a double helix with Watson–Crick
base pairs. Two such sequences are
molecules C
in solution can be
sequences are mixed, a What forces cause the
examined by a variety of double helix with T
two strands of DNA to
techniques. In isolation, Watson–Crick base pairs bind to each other?
T
does form (Figure 1.8).
each sequence exists
This reaction pro
almost exclusively as a T
single-stranded molecule. ceeds nearly to
A
completion.
A
A
However, when the two
several factors: the types of interactions and bonds
FIGURE 1.8 Formation of a double helix. When two DNA strands with
in biochemical systems and the ener getic favorability
appropriate, complementary sequences are mixed, they spontaneously assemble
of the reaction. We must also consider the influence
to form a double helix.
To analyze this binding reaction, we must consider of the solution conditions—in particular, the
consequences of acid– base reactions.
Covalent and noncovalentbonds are important for the structure and stability
of biological molecules
Atoms interact with one another through chemical bonds. These
bonds include the covalent bonds that define the structure of
molecules as well as a variety of noncovalent bonds that are of great
importance to biochemistry.
Covalent bonds. The strongest bonds are covalent bonds, such as the
bonds that hold the atoms together within the individual bases shown
on page 4. A covalent bond is formed by the sharing of a pair of
electrons between adjacent atoms. A typical carbon–carbon (C}C)
covalent bond has
a bond length of 1.54 Å and bond energy of 355 kJ covalent bonding can be written. For example,
mol 1 (85 kcal mol 1 ). Because covalent bonds are adenine can be written in two nearly equivalent ways
so strong, considerable energy must be expended to called resonance structures.
7
break them. More than one electron pair can be
1.3 Chemical Concepts
shared between two atoms to form a multiple
covalent bond. For example, three of the bases in
Figure 1.6 include carbon–oxygen (C“O) double
Distance and energy units
bonds. These bonds are even stronger than C}C
Interatomic distances and bond lengths are usually measured in angstrom (Å) units:
single bonds, with energies near 730 kJ mol 1
1 Å 5 10210 m 5 1028 cm 5 0.1 nm
(175 kcal mol 1 ) and are somewhat shorter.
Several energy units are in common use. One joule (J) is the amount of energy
For some molecules, more than one pattern of
required to move 1 meter against a force of 1 newton.
required to raise the
NH2
temperature of 1
gram of water 1
5
degree
A kilojoule (kJ) is
Celsius. A kilocalorie
One joule is equal to
1000 joules. One
(kcal) is 1000
0.239 cal.
calorie is
calories.
the amount of energy N
NH2
5
H
NN
H
NN
N
4N
N
H
4
H
stability than does a molecule without multiple
resonance structures.
These adenine structures depict alternative
arrangements of single and double bonds that Noncovalent bonds. Noncovalent bonds are
are possible within the same structural
weaker than covalent bonds but are crucial for
framework. Resonance structures are shown
biochemical processes such as the formation of a
connected by a double-headed arrow. Adenine’s double helix. Four fundamental noncovalent
true structure is a composite of its two resonance bond types are ionic interactions, hydrogen
structures. The composite structure is
bonds, van der Waals interactions, and
manifested in the bond lengths such as that for hydrophobic interactions . They differ in
the bond joining carbon atoms C-4 and C-5. The geometry, strength, and specificity. Furthermore,
observed bond length of 1.40 Å is between that these bonds are affected in vastly different ways
expected for a C}C single bond (1.54 Å) and a
by the presence of water. Let us consider the
C“C double bond (1.34 Å). A molecule that can characteristics of each type:
be written as several resonance structures of
approximately equal energies has greater
1. Ionic Interactions . A charged group on one
q1 q2 r
molecule can attract an oppo sitely charged
group on the same or another molecule. The
energy of an ionic interaction (sometimes called 8
CHAPTER 1 Biochemistry: An Evolving Science
an electrostatic interaction) is given by the
2. Hydrogen Bonds . These interactions are
Coulomb energy:
largely ionic interactions, with partial charges on
nearby atoms attracting one another. Hydrogen
E 5 kq1q2/Dr
bonds are responsible for specific base-pair
where E is the energy, q1 and q2 are the charges formation in the DNA double helix. The hydrogen
on the two atoms (in units of the electronic
atom in a hydrogen bond is partially shared by
charge), r is the distance between the two atoms two electronega
(in ang stroms), D is the dielectric constant
tive atoms such as nitrogen or oxygen. The
(which decreases the strength of the Coulomb
hydrogen-bond donor is the group
depending on the intervening solvent or
medium), and k is a pro portionality constant (k 5
1389, for energies in units of kilojoules per mole,
or 332 for energies in kilocalories per mole).
By convention, an attractive interaction has a
negative energy. The ionic interaction between
two ions bearing single opposite charges sepa
rated by 3 Å in water (which has a dielectric
constant of 80) has an energy of 2 5.8 kJ mol 1
( 2 1.4 kcal mol 1 ). Note how important the
dielectric constant of the medium is. For the
same ions separated by 3 Å in a nonpolar
solvent such as hexane (which has a dielectric
constant of 2), the energy of this interaction is 2
232 kJ mol 1 ( 2 55 kcal mol 1 ).
Hydrogen bond donor
that includes both the atom to
atom itself, whereas the
Hydrogen
which
the
hydrogen
atom
is
more
hydrogen-bond acceptor is
bond acceptor
tightly linked and the hydrogen
NHN
develops a partial positive charge ( d ). Thus, the
+ − −
hydrogen atom with a partial positive charge can
interact with an atom having a partial negative
NHO
charge ( d ) through an ionic interaction.
Hydrogen bonds are much weaker than covalent
OHN
bonds. They have ener gies ranging from 4 to 20 kJ
OHO
mol 1 (from 1 to 5 kcal mol 1 ). Hydrogen bonds are
FIGURE 1.9 Hydrogen bonds. Hydrogen bonds are depicted by dashed green
also somewhat longer than covalent bonds; their
lines. The positions of the partial charges (d and d ) are shown.
bond lengths (measured from the hydrogen atom)
the atom less tightly linked to the hydrogen atom
range from 1.5 Å to 2.6 Å; hence, a distance ranging
(Figure 1.9). The electro negative atom to which the from 2.4 Å to 3.5 Å separates the two nonhydrogen
hydrogen atom is covalently bonded pulls elec tron atoms
density away from the hydrogen atom, which thus
approximately straight, such that
in a hydrogen bond.
Hydrogen
bond donor
The strongest hydrogen bonds the hydrogen-bond donor, the
hydrogen atom, and the
have a tendency to be
0.9 Å 2.0 Å
Hydrogen-bond acceptor
t
c
a
r
t
t
A
N H O 180°
y
gr
e
n
E
n
o
i
s
lu
p
e
R
n
o
i
0
van der Waals
respect to one
fluctuates with time.
another. Hydrogen- At any instant, the
bonding interactions charge distribution is
are responsible for not perfectly
many of the
symmetric. This
properties of water transient asymmetry
that make it such a in the electronic
special solvent, as charge about an
will be described
atom acts through
contact distance Distance
shortly.
ionic interactions to
induce a
hydrogen-bond
3. van der Waals
complementary
acceptor lie along a Interactions . The
asymmetry in the
straight line. This
basis of a van der
tendency toward lin Waals interaction is electron distribution
within its neighboring
earity can be
that the distribution
atoms. The atom and
important for
of electronic charge
its neighbors then
orienting interacting around an atom
attract one another.
molecules with
This attraction
increases as two
atoms come closer
to each other, until
they are separated distances shorter
repulsive forces
the two atoms
by the van der Waals than the van der
become dominant
overlap.
contact distance
Waals contact
because the outer
(Figure 1.10). At
distance, very strong electron clouds of
FIGURE 1.10 Energy of a van der Waals interaction as two atoms approach each hydrogen bonds hold the structure together; similar
other. The energy is most favorable at the van der Waals contact distance.
interactions link molecules in
Owing to electron–electron repulsion, the energy rises rapidly as the distance
liquid water and account for many of the properties of
between the atoms becomes shorter than the contact distance.
water. In the liquid state, approximately one in four of
the hydrogen bonds present in ice are broken. The
polar nature of water is responsible for its high
dielectric constant of 80. Molecules in aqueous
solution interact with water mole
cules through the formation of hydrogen bonds and
through ionic interactions. These interactions make
water a versatile solvent, able to readily dissolve
many species, especially polar and charged
Electric
dipole
compounds that can participate in these interactions.
Energies associated with van der Waals interactions
are quite small; typical interactions contribute from 2 The hydrophobic effect. A final fundamental
to 4 kJ mol 1 (from 0.5 to 1 kcal mol 1 ) per atom
interaction called the hydrophobic effect is a
pair. When the surfaces of two large molecules come manifestation of the proper ties of water. Some
together, however, a large number of atoms are in molecules (termed nonpolar molecules ) cannot
van der Waals contact, and the net effect, summed participate in hydrogen bonding or ionic interac
over many atom pairs, can be substantial.
9
We will cover the fourth noncovalent interaction, the 1.3 Chemical Concepts
hydrophobic inter action, after we examine the
characteristics of water; these characteristics are
essential to understanding the hydrophobic
interaction.
Properties of water. Water is the solvent in which
most biochemical reac tions take place, and its
properties are essential to the formation of macro
molecular structures and the progress of chemical
reactions. Two properties of water are especially
relevant:
1. Water is a polar molecule . The water molecule is
bent, not linear, and so
O
–
the distribution of charge is asymmetric. The oxygen
nucleus draws elec
HH+
trons away from the two hydrogen nuclei, which
leaves the region around
tions. The interactions of nonpolar molecules with
each hydrogen atom with a net positive charge. The water molecules are not as favorable as are
water molecule is thus an electrically polar structure. interactions between the water molecules
themselves. The water molecules in contact with
2. Water is highly cohesive . Water molecules
interact strongly with one another through hydrogen these nonpolar molecules form “cages” around
bonds. These interactions are apparent in
FIGURE 1.11 Structure of ice. Hydrogen bonds (shown as dashed green lines)
are formed between water molecules to produce a highly ordered and open
the structure of ice (Figure 1.11). Networks of
structure.
them, becoming more well ordered than water molecules free in
solution. However, when two such nonpolar molecules come together,
some of the water molecules are released, allowing them to interact
freely with bulk water (Figure 1.12). The release of water from such
cages is favorable for reasons to be considered shortly. The result is
that nonpolar molecules show an increased tendency to associate
with one another in water compared with other, less polar and less
self-associating, solvents. This tendency is called the hydrophobic
effect and the associated interactions are called hydrophobic
interactions .
The double helix is an expression of the rules of chemistry
Let us now see how these four noncovalent interactions work together
in driving the association of two strands of DNA to form a double helix.
First, each phosphate group in a DNA strand carries a negative
charge. These negatively charged groups interact unfavorably with
one another over dis
tances. Thus, unfavorable ionic interactions take place when two
strands of Nonpolar
molecule
molecule
Nonpolar molecule
aggregation of nonpolar groups in water leads to
the release of water molecules, initially
interacting with the nonpolar surface, into bulk
water. The release of water molecules into
solution makes the aggregation of nonpolar
groups favorable.
Nonpolar molecule
Nonpolar
FIGURE 1.12 The hydrophobic effect. The
DNA come together. These phosphate groups are far apart in the
double helix with distances greater than 10 Å, but many such
interactions take place (Figure 1.13). Thus, ionic interactions oppose
the formation of the double helix. The strength of these repulsive ionic
interactions is dimin
ished by the high dielectric constant of water and the presence of ionic
species such as Na or Mg 2 ions in solution. These positively charged
species interact with the phosphate groups and partly neutralize their
negative charges.
Second, as already noted, hydrogen bonds are important in
determining the formation of specific base pairs in the double helix.
However, in single stranded DNA, the hydrogen-bond donors and
acceptors are exposed to solution and can form hydrogen bonds
with water molecules.
FIGURE 1.13 Ionic
(the phosphorus atom being O
shown in purple) that bears H
a negative charge. The
H
C
interactions in DNA. Each O
unit within the double helix
includes a phosphate group
H
+
O
O H
H H OH
HN
C
H
O
H
N
unfavorable interactions of one phosphate with several others are shown by red
lines. These repulsive interactions oppose the formation of a double helix.
FIGURE 1.14 Base stacking. In the DNA double helix, adjacent base pairs are
stacked nearly on top of one another, and so many atoms in each base pair are
separated by their van der Waals contact distance. The central base pair is
shown in dark blue and the two adjacent base pairs in light blue. Several van
der Waals contacts are shown in red.
van der Waals
contacts
When two single strands come together, these
hydrogen bonds with water are broken and new
hydrogen bonds between the bases are formed.
Because the number of hydrogen bonds broken is
the same as the number formed, these hydrogen
bonds do not contribute substantially to driving the
overall process of double-helix formation. However, favorability of base stacking. More-complete base
they contribute greatly to the specificity of binding. stacking moves the nonpolar surfaces of the bases
Suppose two bases that cannot form Watson–Crick out of water into contact with each other.
base pairs are brought together. Hydrogen bonds
The principles of double-helix formation between two
with water must be bro
strands of DNA apply to many other biochemical
ken as the bases come into contact. Because the
processes. Many weak interactions con tribute to the
bases are not complemen tary in structure, not all of overall energetics of the process, some favorably
and some unfavorably. Furthermore, surface
these bonds can be simultaneously replaced by
hydrogen bonds between the bases. Thus, the
complementarity is a key feature: when
formation of a double helix between
complementary surfaces meet, hydrogen-bond
noncomplementary sequences is disfavored.
donors align with hydrogen bond acceptors and
nonpolar surfaces come together to maximize van
Third, within a double helix, the base pairs are
parallel and stacked nearly on top of one another. der Waals interactions and minimize nonpolar
surface area exposed to the aque ous environment.
The typical separation between the planes of
The properties of water play a major role in
adjacent base pairs is 3.4 Å, and the distances
determining the importance of these interactions.
between the most closely approaching atoms are
approximately 3.6 Å. This separation distance cor
The laws of thermodynamics govern the behavior of
responds nicely to the van der Waals contact
distance (Figure 1.14). Bases tend to stack even in biochemical systems
single-stranded DNA molecules. However, the base We can look at the formation of the double helix from
stacking and associated van der Waals interactions a different perspec tive by examining the laws of
are nearly optimal in a double-helical structure.
thermodynamics. These laws are general
Fourth, the hydrophobic effect also contributes to the
10
principles that apply to all physical (and biological) system plus that of its sur roundings always
processes. They are of great importance because increases. For example, the release of water from
they determine the conditions under which spe cific nonpolar surfaces responsible for the hydrophobic
processes can or cannot take place. We will consider effect is favorable because water molecules free in
these laws from a general perspective first and then solution are more disordered than they are when
apply the principles that we have devel oped to the they are associated with nonpolar surfaces. At first
formation of the double helix.
glance, the Second Law appears to contradict much
The laws of thermodynamics distinguish between a common experience, particularly about biological sys
system and its surroundings. A system refers to the tems. Many biological processes, such as the
matter within a defined region of space. The matter generation of a leaf from car bon dioxide gas and
in the rest of the universe is called the surroundings. other nutrients, clearly increase the level of order
and hence decrease entropy. Entropy may be
The First Law of Thermodynamics states that the
decreased locally in the formation of such ordered
total energy of a system and its surroundings is
constant . In other words, the energy content of the structures only if the entropy of other parts of the
universe is increased by an equal or greater
uni
verse is constant; energy can be neither created nor amount. The local decrease in entropy is often
accomplished by a release of heat, which increases
destroyed. Energy can take different forms,
however. Heat, for example, is one form of energy. the entropy of the surroundings.
We can analyze this process in quantitative terms.
Heat is a manifestation of the kinetic energy
First, consider the system. The entropy ( S ) of the
associated with the random motion of molecules.
system may change in the course of a chemical
Alternatively, energy can be present as potential
energy —energy that will be released on the
reaction by an amount DSsystem . If heat flows from
occurrence of some process. Consider, for example, the system to its surroundings, then the heat
a ball held at the top of a tower. The ball has
content, often referred to as the enthalpy ( H ) ,
considerable potential energy because, when it is
of the system will be reduced by an amount
released, the ball will develop kinetic energy
DHsystem . To apply the Second Law, we must
associated with its motion as it falls. Within chemical determine the change in entropy of the surround
systems, potential energy is related to the likelihood ings. If heat flows from the system to the
that atoms can react with one another. For instance, surroundings, then the entropy of the surroundings
a mixture of gasoline and oxy gen has a large
will increase. The precise change in the entropy of
potential energy because these molecules may react the surroundings depends on the temperature; the
to form carbon dioxide and water and release
change in entropy is greater when heat is added to
energy as heat. The First Law requires that any
relatively cold surroundings than when heat is added
energy released in the formation of chemical bonds to surroundings at high temperatures that are
must be used to break other bonds, released as heat already in a high degree of disorder. To be even
or light, or stored in some other form.
more specific, the change in the entropy of the sur
Another important thermodynamic concept is that of roundings will be proportional to the amount of heat
entropy, a measure of the degree of randomness or transferred from the system and inversely
disorder in a system. The Second Law of
proportional to the temperature ( T ) of the surround
Thermodynamics states that the total entropy of a
ings. In biological systems, T [in kelvins (K), absolute
temperature] is
if and only if
11
¢Ssystem . ¢HsystemyT (6) Rearranging gives
1.3 Chemical Concepts
12
TDSsystem . DH or, in other words, entropy will
increase if and only if
CHAPTER 1 Biochemistry: An Evolving Science
usually assumed to be constant. Thus, a change in
the entropy of the surroundings is given by
¢G 5 ¢Hsystem 2 T¢Ssystem , 0 (7)
¢Ssurroundings 5 2¢HsystemyT (1) The total entropy Thus, the free-energy change must be negative for
a process to take place spontaneously. There is
change is given by the expression
negative free-energy change when and only when
the overall entropy of the universe is increased .
Again, the free energy represents a single term that
takes into account both the entropy of the system
and the entropy of the surroundings.
Heat is released in the formation of the double helix
¢Stotal 5 ¢Ssystem 1 ¢Ssurroundings (2)
Let us see how the principles of thermodynamics
apply
to the formation of the double helix (Figure
Substituting equation 1 into equation 2 yields
1.15). Suppose solutions containing each of the two
¢Stotal 5 ¢Ssystem 2 ¢HsystemyT (3) Multiplying by single strands are mixed. Before the double helix
forms, each of the single strands is free to translate
2T gives
and rotate in solution, whereas each matched pair of
strands in the double helix must move together.
2T¢Stotal 5 ¢Hsystem 2 T¢Ssystem (4)
Furthermore, the free single strands exist in more
The function 2TDS has units of energy and is
referred to as free energy or Gibbs free energy, after conformations than possible when bound together in
Josiah Willard Gibbs, who developed this function in a double helix. Thus, the formation of a double helix
from two single strands appears to result in an
1878:
increase in order for the system, that is, a decrease
in the entropy of the system.
¢G 5 ¢Hsystem 2 T¢Ssystem (5)
On the basis of this analysis, we expect that the
The free-energy change, DG , will be used
double helix cannot form without violating the
throughout this book to describe the energetics of Second Law of Thermodynamics unless heat is
biochemical reactions. The Gibbs free energy is
released to increase the entropy of the
essentially an accounting tool that keeps track of
surroundings. Experimentally, we can measure the
both the entropy of the system (directly) and the
heat released by allowing the solutions containing
entropy of the surroundings (in the form of heat
the two single strands to come together within a
released from the system).
water bath, which here corresponds to the surround
Recall that the Second Law of Thermodynamics
ings. We then determine how much heat must be
states that, for a process to take place, the entropy absorbed by the water bath or released from it to
of the universe must increase. Examination of
maintain it at a constant temperature. This
equation 3 shows that the total entropy will increase experiment
reveals that approximatelresult
quite large,
a substantial y 250 kJ mol reveals that 2 250 kJ mol
amount of 1 (60 kcal
the change 1 , consistent
heat is
mol 1 ). This in enthalpy with our
released— experimentalfor the
namely,
process is
C
T
A
A
C
T
A
A
T
T
A
GCT
AAT
AATT
A
C
T
A
GATT
TTA
AAT
A
CG
C
GATT
GCT
GAT
T
AA
T
AATT
C
A
G
G
CG
T
T
expectation that significant heat would have to
released to the
surroundings for the
process
be
not to violate the
roundings to
Second Law. We
ensure that the
see in quanti
entropy of the
tative terms how
uni verse
order within a
increases. We
Acid–base reactions
system can be
will encounter
increased by
are central in many
this general
releasing
theme again and
sufficient heat to
again throughout
the sur
this book.
AG
T
A
A
T
C
G
A
T
A
TA
T
A
A
T
T
GC
A
T
G
C
TAA
A
T
TA
T
C
A
T
T
C
G
C
GATT
AAT
T
Mixing
TTAA
A
CG
GCT
AATT
C
GATT
G
C
T
A
T
T
A
A
A
T
T
A
A
T
C
G
A
A
T
GCT
C
G
A
T
T
A
A
T
G
C
T
A
TTA
C
G
T
TTA
TAA
biochemical
processes
Throughou t our
considerati on of the
formation
AATT
AAT
TAA
C
G
A
T
T
TTAA A
A
A
of the double
helix, we have
dealt only with
the
C
A
AATT
GATT
noncovalent
bonds that are
formed or
broken in this
bonds. A
the formation particularly
and cleavage important
of covalent
class of
T
T
A
A
GCT
process. Many
biochemical
processes entail
A
A
GATT
CG
C
G
A
T
T
G
C
T
A
CG
A
T
AAT
AAT
C
GAT
T
AA
T
C
reactions
reactions .
promi nent in
biochemistry
is acid – base
T
C
T
A
A
T
T
A
T
A
A
T
A
A
T
T
T
GC
AAT
A
A
T
T
A
A
G
C
T
C
A
T
GCT A
G
are added to
Throughout the which the
In acid and basemolecules or
book, we will
addition or
reactions,
removed from encounter many removal of
hydrogen ions them.
processes in
atoms is
processes
crucial, such by which
C
as the meta carbohydrat
TA
bolic
es are
hydrogen
release
energy for
other uses.
Thus,
degraded to
TT
understanding of
a thorough
the basic princi
reactions is
essential.
ples of these
written as H , cor
A hydrogen ion, often
responds to a
proton. In fact,
hydrogen ions
C
G
G
A
T
T
A
TTA
T
TA
A
T
A
A
T
T
A
A
A
T
GC
A
T
T
ATT A
Reacting
TTA
TAA
T
TAA
GA
G
C
T
C
G
A
GC
A
C
G
AC
C
TTAA
A
TT
A
G
CG
G
A
A
T
T
TT
T
A
TTAA
TTAA A
A
TAA
TAG
C
T
A
C
GCT
TAA
G
T
AATT
TC
A
GATT
GCT C
AATT
T
AAT
A
GAT
A
A
GCT
C
T
exist in solution bound to water molecules, thus
T
A
AAT
GATT
T
T
C
A
T
G
AATT
forming what are known as hydronium ions, H 3 O .
A
A
A
G
T
T
G
C
T
C
T
C
A
AAT
A
A
G
For simplicity, we will continue to write H ,
A
T
T
A
T
A
but we should keep in mind that H is short
hand for the actual species present. The
concentration of hydrogen ions in solu tion is
expressed as the pH. Specifically, the pH of a
solution is defined as
FIGURE 1.15 Double-helix formation and entropy. When solutions containing
DNA strands with complementary sequences are mixed, the strands react to
form double helices. This process results in a loss of entropy from the system,
indicating that heat must be released to the surroundings to prevent a violation
of the Second Law of Thermodynamics.
pH 5 2log[H1]
where [H ] is in units of molarity. Thus, pH 7.0 refers to a solution
for which 2 log[H ] 5 7.0, and so log[H ] 5 2 7.0 and [H ] 5 10 log[H
] 5 10 7.0 5 1.0 3 10 7 M.
The pH also indirectly expresses the concentration of hydroxide
ions, [OH ], in solution. To see how, we must realize that water
molecules can dissociate to form H and OH ions in an equilibrium
process.
H2O Δ H1 1 OH2
The equilibrium constant ( K ) for the dissociation of water is
defined as K 5 [H1][OH2]/[H2O]
13
14
The concentration of water, [H 2 O], in pure water is
55.5 M, and this concentration is constant under
most conditions. Thus, we can define a new
constant, KW :
CHAPTER 1 Biochemistry: An Evolving Science
KW 5 K[H2O] 5 [H1][OH2]
K[H2O] 5 1.8 3 10216 3 55.5
5 1.0 3 10214
Because KW 5 [H ][OH ] 5 1.0 3 10 14 , we can
calculate [OH2] 5 10214/[H1] and [H1] 5 10214/[OH2]
1.0
n
i
and has a value of K 5 1.8 3 10 16 . Note that an
equilibrium constant does not formally have units.
Nonetheless, the value of the equilibrium constant
given assumes that particular units are used for
concentration (sometimes referred to a standard
states); in this case and in many others, units of
molar
ity (M) are assumed.
With these relations in hand, we can easily calculate
the concentration of hydroxide ions in an aqueous
solution, given the pH. For example, at pH 5 7.0, we
know that [H ] 5 10 7 M and so [OH ] 5 10 14 /10 7 5
10 7 M. In acidic solutions, the concentration of
hydrogen ions is higher than 10 7 and, hence, the pH
is below 7. For example, in 0.1 M HCl, [H ] 5 10 1 M
and so pH 5 1.0 and [OH ] 5 10 14 /10 1 5 10 13 M.
Acid–base reactions can disrupt the double helix
The reaction that we have been considering
between two strands of DNA to
and treat it with a
the first additions of ate into its
0.8 0.6 0.4 0.2 0
solution of
base are made, the component single
concentrated base pH rises, but the
strands. As the pH
7 8 9 10 11 pH
(i.e., with a high
concentration of the continues to rise
concentration of OH double-helical DNA from 9 to 10, this
form a double helix ). As the base is
does not change
dissociation
takes place readily at added, we monitor significantly.
becomes essentially
pH 7.0. Suppose that the pH and the
However, as the pH complete. Why do
we take the solution fraction of DNA in
approaches 9, the the two strands
containing the
double-helical form DNA double helix
double-helical DNA (Figure 1.16). When begins to dissoci
in DNA base pairs to remove certain protons. The
FIGURE 1.16 DNA denaturation by the addition of a base. The addition of a
base to a solution of double-helical DNA initially at pH 7 causes the double
most susceptible proton is the one bound to the N-1
helix to separate
nitrogen atom in a guanine base.
dissociate? The hydroxide ions can react with bases
e
s
lbu
o
e
l
d
u
c
e
lo
m
f
o
n
o
it
c
a
r
F
m
r
o
f
l
a
c
il
e
h-
into single strands. The process is
half
O
complete at slightly above pH 9.
N
NH
−
O
NH
N
+ pKa = 9.7 +
H
H
N N Guanine (G)
NH2
NN
NH2
Proton dissociation for a substance HA (such as that bound to N-1
on gua nine) has an equilibrium constant defined by the expression
Ka 5 [H1][A2]y[HA]
The susceptibility of a proton to removal by reaction with a base
is often described by its pKa value:
pKa 5 2log(Ka)
When the pH is equal to the p Ka , we have
pH 5 pKa
and so
2log[H1] 5 2log([H1][A2]y[HA])
and
[H1] 5 [H1][A2]y[HA]
Dividing by [H ] reveals that
1 5 [A2]y[HA]
and so
[A2] 5 [HA]
Thus, when the pH equals the p Ka , the
concentration of the deprotonated form of the group
or molecule is equal to the concentration of the
proton ated form; the deprotonation process is
halfway to completion.
The p Ka for the proton on N-1 of guanine is typically
9.7. When the pH approaches this value, the proton
on N-1 is lost (Figure 1.16). Because this proton
participates in an important hydrogen bond, its loss
substantially destabilizes the DNA double helix. The
DNA double helix is also destabi
lized by low pH. Below pH 5, some of the hydrogen
bond acceptors that participate in base-pairing
become protonated. In their protonated forms, these
bases can no longer form hydrogen bonds and the
double helix sepa rates. Thus, acid–base reactions
that remove or donate protons at specific positions
on the DNA bases can disrupt the double helix.
Buffers regulate pH in organisms and in the laboratory
These observations about DNA reveal that a
12
significant change in pH can disrupt molecular
structure. The same is true for many other biological the amount of acid added. In contrast, when acid is
added to a buffered solu tion, the pH drops more
macromolecules; changes in pH can protonate or
gradually. Buffers also mitigate the pH increase
deprotonate key groups, potentially disrupting
structures and initiating harmful reactions. Thus,
systems have evolved to mitigate changes in pH in
biological systems. Solutions that resist such
changes are called buffers . Specifically, when acid
is added to an unbuffered aqueous solution, the pH
drops in proportion to
10
caused by the addition of base and changes in pH
caused by dilution. 8
Compare the result of adding a 1 M solution of the
strong acid HCl drop by drop to pure water with
adding it to a solution containing 100 mM of the 6
buffer sodium acetate (Na CH 3 COO ; Figure
1.17). The process of
2
drops of acid. However, for the sodium acetate
solution, the pH first falls rapidly from its initial value
near 10, then changes more gradually until the
H
p
0
gradually adding known amounts of reagent to a
pH reaches 3.5, and then falls more rapidly again.
solution with which the 4
Why does the pH decrease so gradually in the
reagent reacts while monitoring the results is called a middle of the titration? The answer is that,
titration . For pure water, the pH drops from 7 to
15
close to 2 on the addition of the first few
1.3 Chemical Concepts
when hydrogen ions are added to this solution, they
react with acetate ions to form acetic acid. This
reaction consumes some of the added hydrogen
ions so that the pH does not drop. Hydrogen ions
continue reacting with acetate ions until essentially
all of the acetate ion is converted into acetic acid.
After this point, added protons remain free in solution
and the pH begins to fall sharply again.
0.1 M Na+CH3COO−
Gradual pH change
Water
FIGURE 1.17 Buffer action. The addition of a strong acid, 1 M HCl, to pure
water results in an immediate drop in pH to near 2. In contrast, the addition of
the acid to a 0.1 M sodium acetate (Na CH3COO ) solution results in a much
more gradual change in pH until the pH drops below 3.5.
6050403020100 Number of drops
equation to our titration of sodium
acetate. The p Ka of acetic acid is
4.75. We can calculate the ratio of
the concentration of acetate ion to
the concentration of acetic acid as
a function of pH by using the
Henderson–Hasselbalch equation,
slightly rearranged.
16
CHAPTER 1 Biochemistry: An Evolving Science
12
10
60504030201000%
We can analyze the effect of the
buffer in quantitative terms. The
equi librium constant for the
deprotonation of an acid is
[Acetate ion]y[Acetic acid] 5
[A2]y[HA] 5 10pH2pKa
At pH 9, this ratio is 10 9 4.75 5 10
4.25 5 17,800; very little acetic acid
8
percentage
has been formed. At pH 4.75 (when
Ka 5 [H1][A2]y[HA]
6
the pH equals the p Ka ), the ratio is
Taking logarithms of both sides
10 4.75 4.75 5 10 0 5 1. At pH 3, the
4
yields
ratio is 10 3 4.75 5 10 1.25 5 0.02;
1
2
2
log(Ka) 5 log([H ]) 1 log([A ]y[HA]) almost all of the acetate ion has
been converted into acetic acid.
Recalling the definitions of p Ka
0
We can fol
and pH and rearranging gives pH low the conversion of acetate ion
Number of drops
5 pKa 1 log([A2]y[HA])
into acetic acid over the entire
100%
This expression is referred to as titration (Figure 1.18). The graph
shows that the region of relatively
the Henderson – Hasselbalch
constant pH
equation . We can apply the
From this discussion, we see that a buffer functions
FIGURE 1.18 Buffer protonation. When acid is added to sodium acetate, the
added hydrogen ions are used to convert acetate ion into acetic acid. Because best close to the p K value of its acid component.
a
the proton concentration does not increase significantly, the pH remains
Physiological
pH
is
typically
about 7.4. An important
relatively constant until all of the acetate has been converted into acetic acid.
buffer
in
biological
systems
is
based on phosphoric
corresponds precisely to the region in which acetate
acid (H 3 PO 4 ). The acid can be deprotonated in
ion is being protonated to form acetic acid.
three steps to form a phosphate ion.
Acetic acid
H
p
H
H H
PO43
H2PO4 HPO42 H3PO4 pKa 2.12 pKa pKa 12.67
7.21
At about pH 7.4, inorganic phosphate exists primarily as a nearly equal
mixture of H2PO42 and HPO422. Thus, phosphate solutions function
as effective buffers near pH 7.4. The concentration of inorganic
phosphate in blood is typically approximately 1 mM, providing a useful
buffer against processes that produce either acid or base. We can
examine this utility in quantitative terms with the use of the
Henderson–Hasselbalch equation. What concentration of acid must
be added to change the pH of 1 mM phos phate buffer from 7.4 to 7.3?
Without buffer, this change in [H ] corre
sponds to a change of 10 7.3 2 10 7.4 M 5 (5.0 3 10 8 2 4.0 3 10 8 )
M 5 1.0 3 10 8 M. Let us now consider what happens to the buffer
com ponents. At pH 7.4,
[HPO422]y[H2PO42] 5 107.427.21 5 100.19 5 1.55
The total concentration of phosphate, [HPO422] 1 [H2PO42], is
1 mM, Thus,
[HPO422] 5 (1.55/2.55) 3 1 mM 5 0.608 mM
and
[H2PO42] 5 (1/2.55) 3 1 mM 5 0.392 mM
GGAGAAGT
At pH 7.3,
CTGCCGTTACTGCCCTGTGGGGCAAGGTGAACG
[HPO42 2]y[H2PO42] 5 107.327.21 5 100.09 5 1.23
TGGA . . .
and so
is a part of one of the genes that encodes
hemoglobin, the oxygen carrier in our blood. This
[HPO422] 5 (1.23y2.23) 5 0.552 mM
gene is found on the end of chromosome 9 of our 24
and
distinct chromosomes. If we were to include the
complete sequence of our entire genome, this
[H2PO42] 5 (1y2.23) 5 0.448 mM
chapter would run to more than 500,000 pages. The
22
Thus, (0.608 2 0.552) 5 0.056 mM HPO4 is
sequenc
converted into H2PO42, consuming 0.056 mM 5 5.6 ing of our genome is truly a landmark in human
3 10 5 M [H ]. Thus, the buffer increases the
history. This sequence contains a vast amount of
amount of acid required to produce a drop in pH from information, some of which we can now extract and
7.4 to 7.3 by a factor of 5.6 3 10 5y1.0 3 10 8 5
interpret, but much of which we are only beginning to
5600 compared with pure water.
understand. For example, some human diseases
have been linked to particular variations in genomic
sequence. Sickle-cell anemia, discussed in detail in
Chapter 7, is caused by a single base change of an
1.4 The Genomic Revolution Is Transforming
A (noted in boldface type in the
Biochemistry, Medicine, and Other Fields
17
1.4 The Genomic Revolution
Watson and Crick’s discovery of the structure of
18
DNA suggested the hypothesis that hereditary
information is stored as a sequence of bases along CHAPTER 1 Biochemistry: An Evolving Science
preceding sequence) to a T. We will encounter many
long strands of DNA. This remarkable insight
other examples of dis eases that have been linked to
provided an entirely new way of thinking about
specific DNA sequence changes. Determining the
biology. However, at the time that it was made,
first human genome sequences was a great
Watson and Crick’s discovery, though full of
challenge. It required the efforts of large teams of
potential, remained to be confirmed and many
geneticists, molecular biologists, bio chemists, and
features needed to be elucidated. How is the
computer scientists, as well as billions of dollars,
sequence information read and translated into
because there was no previous framework for
action? What are the sequences of naturally
aligning the sequences of various DNA fragments.
occurring DNA molecules and how can such
sequences be experimentally determined? Through One human genome sequence can serve as a
advances in bio chemistry and related sciences, we reference for other sequences. The availability of
such reference sequences enables much more
now have essentially complete answers to these
rapid characterization of partial or complete
questions. Indeed, in the past decade or so,
genomes from other indi viduals. As we will discuss
scientists have deter
mined the complete genome sequences of hundreds in Chapter 5, arrays containing millions of target
single-stranded DNA molecules with sequences from
of different organ isms, including simple
microorganisms, plants, animals of varying degrees the reference genome and known or potential
of complexity, and human beings. Comparisons of variants are powerful tools. These arrays can
these genome sequences, with the use of methods be exposed to mixtures of DNA fragments for a
introduced in Chapter 6, have been sources of many particular individual and those single-stranded
targets that bind to their complementary strands can
insights that have transformed biochemistry. In
be determined. This allows many positions within the
addition to its experimental and clinical aspects,
genome of the indi vidual to be simulaneously
biochemistry has now become an information
interrogated.
science .
Genome sequencing has transformed biochemistry
and other fields
The sequencing of a human genome was a daunting
task because it contains approximately 3 billion (3 3
10 9 ) base pairs. For example, the sequence
ACATTTGCTTCTGACACAACTGTGTTCACTAGCA
ACCTC
AAACAGACACCATGGTGCATCTGACTCCTG A
Methods for sequencing DNA have also been
d
e
c
n
e
u
q
e
s
e
m
o
n
e
g
n
a
m
u
h
r
e
p
t
s
o
C
$100M
improv
sequencing rate and decreases in
cost (Figure 1.19). The availability
of such powerful sequencing
technology is transforming many
$100K
fields, including medicine, dentistry,
microbiology, pharmacology, and
$10K
ecology, although a great deal
$1K
remains to be done to improve the
200620052004200320022001 20112010200920082007
accuracy and precision of the
20132012
interpretation of these large
Year
genomic and related data sets.
ing rapidly, driven by a deep
understanding of the biochemistry Each person has a unique
sequence of DNA base pairs. How
of DNA replication and other
different are we from one another
processes. This has resulted in
both dramatic increases in the DNA at the
$1M
$10M
reveals that, on average, each pair of individuals has
a different base in one position per 200 bases; that
is, the difference is approximately 0.5%. This
variation between individuals who are not closely
Human Genome Research Institute. www.genome.gov/sequencingcosts]
related is
quite substantial compared with
genomic level? Examination of genomic variation
differences in popu
lations. The average difference between two people within one ethnic
group is greater than the difference between the averages of two
differ ent ethnic groups.
The significance of much of this genetic variation is not understood.
As noted earlier, variation in a single base within the genome can lead
to a disease such as sickle-cell anemia. Scientists have now identified
the genetic variations associated with hundreds of diseases for which
the cause can be traced to a single gene. For other diseases and
traits, we know that variation in many different genes contributes in
significant and often complex ways. Many of the most prevalent
human ailments such as heart disease are linked to variations in many
genes. Furthermore, in most cases, the presence of a particular
variation or set of variations does not inevita bly result in the onset of
a disease but, instead, leads to a predisposition to the development
of the disease.
Our own genes are not the only ones that can contribute to health
and disease. Our bodies, including our skin, mouth, digestive tract,
genito urinary tract, respiratory tract, and other areas, contain large
number of microorganisms. These complex communities have been
characterized through powerful methods that allow DNA isolated
from these biologi cal samples to be sequenced without any previous
knowledge of the
organisms present. Many of these They appear to play roles in health
organisms had not previously been and in diseases such as obesity
discovered because they can only and dental caries (Figure 1.20).
grow as part of complex
In addition to the implications for Nasal
communities and thus cannot be understanding human health and
isolated through standard
microbiological techniques.
Remarkably, it appears that we are Gastrointestinal Urogenital
outnumbered in our own bodies!
Each of us contains approximately
Skin
ten times more microbial cells than
human cells and these microbial
cells include many more genes
than do our own genomes. These
microbiomes differ from site to site, Oral
from one person to another and
over time in the same individual.
disease, the genome sequence is a source of deep of different individuals and populations, we can
learn a great deal about human history. On the
insight into other aspects of human biology and
culture. For example, by comparing the sequences basis of such analysis, a compelling case can be
FIGURE 1.19 Decreasing costs of DNA sequencing. Through the Human
Genome Project, the cost of DNA sequencing dropped steadily. With the
advent of new methods, these costs dropped dramatically and are now
approaching $1000 for a complete human genome sequence. [National
made that the human species originated in Africa, evolutionary and functional relatives in the genomes
and the occurrence and even the timing of
of bacteria. Because many studies that are possible
important migrations of groups of human beings can in model organisms are difficult or unethical to
be dem
conduct in human beings, these discoveries have
onstrated (Figure 1.21). Finally, comparisons of the many practical implica tions. Comparative genomics
has become a powerful science, linking evolu
human genome with the genomes of other
organisms are confirming the tremendous unity that tion and biochemistry.
exists at the level of biochemistry and are revealing
1.20 The human microbiome. Microorganisms cover the human body.
key steps in the course of evolution from relatively FIGURE
Examination of the microbial communities using DNA sequencing methods
simple, single-celled organisms to complex,
revealed many previously uncharacterized species. The Venn diagrams
multicellular organisms such as human beings. For represent populations of related species as determined by DNA sequence
example, many genes that are key to the function of comparisons. The populations present on different body surfaces are largely
distinct. [Adapted from www.nature. com/nature/journal/v486/n7402/fig_tab/
the human brain and nervous system have
nature11234_F1.html]
50,000–60,000
years ago
46,000–50,000
years ago
15,000–19,000 years ago
150,000
years ago
20,000–30,000
coastal
route
years ago 15,000 years ago 40,000 years ago
12,500 years ago
FIGURE 1.21 Human migrations supported by DNA sequence comparisons. Modern human beings
originated in Africa, migrated first to Asia, and then to Europe, Australia, and North and South
America. [Adapted from S. Oppenheimer, “Out-of-Africa, the peopling of continents and islands: tracing
uniparental gene trees across the map.” Philos. Trans. R. Soc. Lond. B. Biol. Sci. 367(1590):770–784]
20
CHAPTER 1 Biochemistry: An Evolving Science
element. Despite the fact that the most important
essential dietary factors have been
Environmental factors influence human biochemistry
Although our genetic makeup (and that of our
microbiomes ) is an impor tant factor that contributes
to disease susceptibility and to other traits, factors in
a person’s environment also are significant. What are
these envi ronmental factors? Perhaps the most
obvious are chemicals that we eat or are exposed to
in some other way. The adage “you are what you
eat” has considerable validity; it applies both to
substances that we ingest in sig nificant quantities
and to those that we ingest in only trace amounts.
Throughout our study of biochemistry, we will
encounter vitamins and trace elements and their
derivatives that play crucial roles in many pro cesses.
In many cases, the roles of these chemicals were
known for some time, new roles for them continue to
first revealed through investigation of deficiency
disorders observed in people who do not take in a be discovered. A healthful diet requires a balance of
major food
sufficient quantity of a particular vitamin or trace
Vitamins
and minerals
19
FruitsGrains
Fats
Carbohydrates
Dairy
groups. In addition to providing
vitamins and trace elements,
food provides calories in the form FIGURE 1.22 Nutrition. Proper health depends
of sub stances that can be
Protein
broken down to release energy in which food, particularly rich
that drives other biochemical
foods such as meat, was scarce.
processes. Proteins, fats, and With the development of
carbohydrates provide the
agriculture and modern
building blocks used to construct economies, rich foods are now
the molecules of life (Figure
plentiful in parts of the world.
1.22). Finally, it is possible to get Some of the most prevalent
too much of a good thing.
diseases in the so-called
Human beings evolved under
developed world, such as
circumstances
Vegetables Protein
Just as vitamin deficiencies and genetic diseases
have revealed funda mental principles of
biochemistry and biology, investigations of variations
choosemyplate.gov]
in behavior and their linkage to genetic and
heart disease and diabetes, can be attributed to the biochemical factors are potential sources of great
large quantities of fats and carbohydrates present in insight into mechanisms within the brain. For
modern diets. We are now developing a deeper
example, studies of drug addiction have revealed
understanding of the biochemical consequences of neural circuits and biochemical pathways that
these diets and the inter play between diet and
greatly influence aspects of behavior. Unraveling the
genetic factors.
inter play between biology and behavior is one of the
Chemicals are only one important class of
great challenges in modern science, and
environmental factors. Our behaviors also have
biochemistry is providing some of the most important
biochemical consequences. Through physical activ concepts and tools for this endeavor.
ity, we consume the calories that we take in,
Genome sequences encode proteins and patterns of expression
ensuring an appropriate bal ance between food
The structure of DNA revealed how information is
intake and energy expenditure. Activities ranging
from exercise to emotional responses such as fear stored in the base sequence along a DNA strand.
But what information is stored and how is it
and love may activate specific biochemical
expressed? The most fundamental role of DNA is to
pathways, leading to changes in levels of gene
encode the sequences of proteins. Like DNA,
expression, the release of hormones, and other
consequences. Furthermore, the interplay between proteins are linear polymers. However, proteins
differ from DNA in two important ways. First, proteins
biochemistry and behavior is bidirectional. Just as
our biochemistry is affected by our behavior, so, too, are built from 20 building blocks, called amino acids,
rather than just four, as in DNA. The chemical
our behavior is affected, although certainly not
completely determined, by our genetic makeup and complex
ity provided by this variety of build
other aspects of our biochemistry. Genetic factors
associated with a range of behavioral characteristics 21
1.4 The Genomic Revolution
have been at least tentatively identified.
a solution of double-helical
molecules. A similar
spontaneous folding
process gives proteins their
three-dimensional structure.
A bal ance of hydrogen
bonding, van der
on an appropriate combination of food groups (fruits, vegetables, protein,
grains, dairy) (left) to provide an optimal mix of biochemicals (carbohydrates,
proteins, fats, vitamins, and minerals) (right). [Adapted from www.
ing blocks enables proteins 1 2 3
to per form a wide range of
Amino acid sequence 1
functions. Second, proteins
spontaneously fold into
elaborate three-dimensional
structures, determined only
by their amino acid
sequences (Figure 1.23).
We have explored in depth
123
how solutions containing
two appropri ate strands of Amino acid sequence 2
DNA come together to form
Waals interactions, and hydrophobic interactions
overcomes the entropy lost in going from an
The fundamental unit of hereditary information, the
gene, is becom ing increasingly difficult to precisely
define as our knowledge of the com plexities of
genetics and genomics increases. The genes that
are simplest to define encode the sequences of
proteins. For these protein-encoding genes, a block
of DNA bases encodes the amino acid sequence of
a spe cific protein molecule. A set of three bases
along the DNA strand, called a codon, determines
the identity of one amino acid within the protein
sequence. The set of rules that links the DNA
sequence to the encoded protein sequence is
called the genetic code . One of the biggest
surprises from the sequencing of the human
genome is the small number of pro tein-encoding
genes. Before the genome-sequencing project
began, the consensus view was that the human
genome would include approximately 100,000
protein-encoding genes. The current analysis
suggests that the actual number is between 20,000
and 25,000. We shall use an estimate of 21,000
throughout this book. However, additional
mechanisms allow many genes to encode more
than one protein. For example, the genetic
information in some genes is translated in more
than one way to produce a set of proteins that differ
from one another in parts of their amino acid
sequences. In other cases, proteins are modified
after they have been syn thesized through the
addition of accessory chemical groups. Through
these indirect mechanisms, much more complexity
is encoded in our genomes than would be
expected from the number of protein-encoding
genes alone.
On the basis of current knowledge, the proteinencoding regions account for only about 3% of the
human genome. What is the function of the rest of
the DNA? Some of it contains information that
regulates the expression of specific genes (i.e., the
production of specific proteins) in particular cell
types and physiological conditions. Essentially
every human
FIGURE 1.23 Protein folding. Proteins are linear polymers of amino acids that
fold into elaborate structures. The sequence of amino acids determines
the three-dimensional structure. Thus, amino acid sequence 1 gives rise only
to a protein with the shape depicted in blue, not the shape depicted in red.
22
CHAPTER 1 Biochemistry: An Evolving Science
cell contains the same DNA genome, yet cell types
differ considerably in the proteins that they
produce. For example, hemoglobin is expressed
only in precursors of red blood cells, even though
the genes for hemoglobin are present in essentially
every cell. Specific sets of genes are expressed in
response to hormones, even though these genes
are not expressed in the same cell in the absence
of the hormones. The control regions that regulate
such differences account for only a small amount of
the remainder of our genomes. The truth is that we
do not yet understand all of the function of much of
the remainder of the DNA. Some of it is sometimes
referred to as “junk”—stretches of DNA that were
inserted at some stage of evolution and have
remained. In some cases, this DNA may, in fact,
serve important functions. In others, it may serve
no function but, because it does not cause
significant harm, it has remained.
APPENDIX: Visualizing Molecular Structures I: Small Molecules
The authors of a biochemistry
textbook face the problem of
trying to present threedimensional molecules in the two Z X C
W
W
ZW
of biomolecules
dimensions available on the printed page. The
ZX≡≡
interplay between the three-dimensional structures Y X
frequently
and their biological functions will be Y
discussed extensively throughout Fischer
this book. Toward this end, we will projection
use representations that, although of necessity are
rendered in two dimensions, emphasize the threedimensional struc tures of molecules.
Stereochemical Renderings
Most of the chemical formulas in this book are drawn to
depict the geometric arrangement of atoms, crucial to
chemical bonding and reactivity, as accurately as possi
ble. For example, the carbon atom of methane is tetra
hedral, with H–C–H angles of 109.5 degrees, whereas
the carbon atom in formaldehyde has bond angles of
120 degrees.
Y
Stereochemical
rendering
carbon are represented by horizontal and vertical lines
from the substituent atoms to the carbon atom, which is
assumed to be at the center of the cross. By convention,
the horizontal bonds are assumed to project out of the
page toward the viewer, whereas the vertical bonds are
assumed to project behind the page away from the
viewer.
Molecular Models for Small Molecules
For depicting the molecular architecture of small
molecules in more detail, two types of models will often
be used: space filling and ball and stick. These models
show structures at
In a Fischer projection, the bonds to the central
HH
C
H H
C
H H
the atomic level.
O
Methane Formaldehyde
space-filling models are the most
realistic. The size and position of
an atom in a space
1. Space-Filling Models . The
shown in Figure 1.24.
To illustrate the correct stereochemistry about tetra
hedral carbon atoms, wedges will be used to depict the
direction of a bond into or out of the plane of the page. A
solid wedge with the broad end away from the carbon
atom denotes a bond coming toward the viewer out of
the plane. A dashed wedge, with its broad end at the
carbon atom, represents a bond going away from the
viewer behind the plane of the page. The remaining two
bonds are depicted as straight lines.
2. Ball-and-Stick Models . Ball-and-stick models are not
as realistic as space-filling models, because the atoms
are depicted as spheres of radii smaller than their van
der Waals radii. However, the bonding arrangement is
easier to see because the
bonds are explicitly
represented as sticks. In an illustration, the taper of a
stick, representing parallax, tells which of a pair of
bonded atoms is closer to the reader. A ball-and-stick
Fischer Projections
model reveals a complex structure more clearly than a
Although representative of the actual structure of a
com pound, stereochemical structures are often difficult space filling model does. Ball-and-stick models of
several simple molecules are shown in Figure 1.24.
to draw quickly. An alternative, less-representative
23
method of depicting structures with tetrahedral carbon
Problems
centers relies on the use of Fischer projections .
filling model are determined by its bonding properties
and van der Waals radius, or contact distance. A van der Molecular models for depicting large molecules will be
Waals radius describes how closely two atoms can discussed in the appendix to Chapter 2.
approach each other when they are not linked by a
covalent bond. The colors of the model are set by
Water Acetate Formamide Cysteine SH
convention.
Carbon, black Hydrogen, white Nitrogen, blue
Oxygen, red Sulfur, yellow Phosphorus, purple
Space-filling models of several simple molecules are
FIGURE 1.24 Molecular representations. Structural formulas (bottom), ball-and- molecules are shown. Black 5 carbon, red 5 oxygen, white 5 hydrogen, yellow
5 sulfur, blue 5 nitrogen.
stick models (top), and space-filling representations (middle) of selected
+H N
3
O
H3C C
H
− H2N C
H
O
H2O
O
O
OC−
KEY TERMS
biological macromolecule (p
. 2) metabolite (p. 2)
deoxyribonucleic acid (DNA) (p.
2) protein (p. 2)
Eukarya (p. 3)
Bacteria (p. 3)
Archaea (p. 3)
eukaryote (p. 3)
prokaryote (p. 3)
double helix (p. 5)
covalent bond (p. 5)
resonance structure (p.
7) ionic interaction (p.
7)
hydrogen bond (p. 8)
van der Waals interaction (p.
8) hydrophobic effect (p. 9)
hydrophobic interaction (p. 9)
entropy (p. 11)
enthalpy (p. 11)
free energy (Gibbs free energy) (p.
12) pH (p. 13)
p Ka value (p. 14)
buffer (p. 15)
predisposition (p. 18)
microbiome (p. 19)
amino acid (p. 21)
genetic code (p. 21)
PROBLEMS
1. Donors and acceptors . Identify the hydrogen-bond
donors and acceptors in each of the four bases on page
4.
2. Resonance structures . The structure of an amino
acid, tyro sine, is shown here. Draw an alternative
resonance structure.
O H
H
H
H
H
H CH2 C
+H N COO−
3
24
CHAPTER 1 Biochemistry: An Evolving Science
(b) DH 5 2 84 kJ mol 1 ( 2 20 kcal mol 1 ),
3. It takes all types . What types of noncovalent bonds
hold together the following solids?
( a ) Table salt ( NaCl ), which contains Na and Cl
ions. (b) Graphite (C), which consists of sheets of
covalently bonded carbon atoms.
4. Don’t break the law . Given the following values for
the changes in enthalpy ( DH ) and entropy ( DS ), which
of the following processes can take place at 298 K
without violat
ing the Second Law of
Thermodynamics? (a)
DH 5 2 84 kJ mol 1 ( 2 20
kcal mol 1 ) ,
DS 5 1 125 J mol 1 K 1 ( 1 30 cal mol 1 K
1)
DS 5 2 125 J mol 1 K 1 ( 2 30 cal mol 1 K 1 )
(c) DH 5 1 84 kJ mol 1 ( 1 20 kcal mol 1 ),
DS 5 1 125 J mol 1 K 1 ( 1 30 cal mol 1 K 1 )
(d) DH 5 1 84 kJ mol 1 ( 1 20 kcal mol 1 ),
DS 5 2 125 J mol 1 K 1 ( 2 30 cal mol 1 K 1 )
5. Double-helix-formation entropy . For double-helix
forma tion, D G can be measured to be 2 54 kJ mol 1 (
2 13 kcal mol 1 ) at pH 7.0 in 1 M NaCl at 25 8 C (298
K). The heat released indicates an enthalpy change of
2 251 kJ mol 1 ( 2 60 kcal mol 1 ) . For this process,
calculate the entropy change for the system and the
entropy change for the surroundings.
6. Find the pH . What are the pH values for the
following solutions?
(a) 0.1 M HCl
(b) 0.1 M NaOH
(c) 0.05 M HCl
(d) 0.05 M NaOH
7. A weak acid . What is the pH of a 0.1 M solution of
acetic acid ( p Ka 5 4. 75 ) ?
(Hint: Let x be the concentration of H ions released
from acetic acid when it dissociates. The solutions to a
quadratic equation of the form ax2 1 bx 1 c = 0 are x 5
(2b 6 2b2 2 4ac)y2a.)
8. Substituent effects . What is the pH of a 0.1 M
solution of chloroacetic acid ( ClCH 2 COOH, pKa 5 2 .
86)? 9. Water in water . Given a density of 1 g/ml and
a molecu lar weight of 18 g/mol, calculate the
concentration of water in water.
10. Basic fact . What is the pH of a 0.1 M solution of
ethylamine, given that the p Ka of ethylammonium ion (
CH 3 CH 2 NH 3+ ) is 10.70?
11. Comparison . A solution is prepared by adding 0.01
M acetic acid and 0.01 M ethylamine to water and
adjusting the pH to 7.4. What is the ratio of acetate to
acetic acid?
What is the ratio of ethylamine to
ethylammonium ion?
12. Concentrate . Acetic acid is added to water until the
pH value reaches 4.0. What is the total concentration of
the added acetic acid?
13. Dilution . 100 mL of a solution of hydrochloric acid
with pH 5.0 is diluted to 1 L . What is the pH of the diluted
solution?
14. Buffer dilution . 100 mL of a 0.1 mM buffer solution
made from acetic acid and sodium acetate with pH 5.0 is
diluted to 1 L . What is the pH of the diluted solution?
17. What’s the ratio? An acid with a p Ka of 8.0 is
present in a solution with a pH of 6.0. What is the ratio
of the proton ated to the deprotonated form of the acid?
18. Phosphate buffer . What is the ratio of the
concentra tions of H2PO4 and HPO42 at (a) pH 7.0; (b)
pH 7.5; (c) pH 8.0?
19. Neutralization of phosphate . Given that phosphoric
acid (H 3 PO 4 ) can give up three protons with different
pK a values, sketch a plot of pH as a function of added
drops of sodium hydroxide solution, starting with a
solution of phosphoric acid at pH 1.0.
20. Buffer capacity . Two solutions of sodium acetate are
prepared, one with a concentration of 0.1 M and the other
with a concentration of 0.01 M . Calculate the pH values
when the following concentrations of HCl have been
added to each of these solutions: 0.0025 M , 0.005 M ,
0.01 M , and 0.05 M .
21. Buffer preparation . You wish to prepare a buffer con
sisting of acetic acid and sodium acetate with a total
acetic acid plus acetate concentration of 250 mM and a
pH of 5.0. What concentrations of acetic acid and
sodium acetate should you use? Assuming you wish to
make 2 liters of this buffer, how many moles of acetic
acid and sodium acetate will you need? How many
grams of each will you need (molecular weights: acetic
acid 60 . 05 g mol 1 , sodium acetate, 82 . 03 g mol 1 )?
22. An alternative approach . When you go to prepare
the buffer described in Problem 21, you discover that
your laboratory is out of sodium acetate, but you do have
sodium hydroxide. How much (in moles and grams)
acetic acid and sodium hydroxide do you need to make
the buffer?
23. Another alternative . Your friend from another
labora tory was out of acetic acid, so tries to prepare
the buffer in Problem 21 by dissolving 41.02 g of
sodium acetate in water, carefully adding 180.0 ml of 1
M HCl , and adding more water to reach a total volume
of 2 liters . What is the total concentration of acetate
plus acetic acid in the solu tion? Will this solution have
pH 5.0? Will it be identical with the desired buffer? If
not, how will it differ?
24. Blood substitute . As noted in this chapter, blood
con tains a total concentration of phosphate of
approximately 1 mM and typically has a pH of 7.4.
You wish to make 100 liters of phosphate buffer with a
pH of 7.4 from NaH 2 PO 4 (molecular weight, 119 . 98 g
(molec
mol 1 ) and Na 2 HPO 4
ular weight, 141 . 96 g
1
mol ) . How much of each (in grams) do you need?
15. Find the pKa . For an acid HA, the concentrations of
HA and A are 0.075 and 0.025, respectively, at pH 6.0. 25. A potential problem . You wish to make a buffer
with pH 7.0. You combine 0.060 grams of acetic acid
What is the p Ka value for HA?
16. pH indicator . A dye that is an acid and that appears and 14.59 grams of sodium acetate and add water to
as different colors in its protonated and deprotonated yield a total vol ume of 1 liter . What is the pH? Will this
forms can be used as a pH indicator. Suppose that you be the useful pH 7.0 buffer you seek?
have a 0.001 M solution of a dye with a p Ka of 7.2. From 26. Charge! Suppose two phosphate groups in DNA
the color, the concentration of the protonated form is (each with a charge of 2 1) are separated by 12 Å.
found to be 0.0002 M . Assume that the remainder of What is the energy of the ionic interaction between
the dye is in the deprotonated form. What is the pH of these two phos phates assuming a dielectric constant of
80? Repeat the calculation assuming a dielectric
the solution?
constant of 2.
27. Vive la différence . On average, estimate how many
base differences there are between two human beings.
25
Problems
28. Epigenomics . The human body contains many
distinct cell types yet almost all human cells contain the
same genome with 21,000 genes. The distinct cell types
are pri marily due to differences in gene expression.
Assume that one set of 1000 genes is expressed in all
cell types and that the remaining 20,000 genes can be
divided into sets of 1000 genes that are either all
expressed or all not expressed in a given cell type. How
many different cell types are possible if each cell type
expresses 10 sets of these genes? Note that the number
of combinations of n objects into m sets is given by n !/
(m!(n-m)!) where n! = 1*2*
…*(n 2 1)*n.
29. Predispositions in populations . Assume that 10% of
the members of a population will get a particular disease
over the course of their lifetime. Genomic studies reveal
that 5% of the population have sequences in their
genomes such that their probability of getting the
disease over the course of their lifetimes is 50%. What
is the average lifetime risk of this disease for the
remaining 95% of the population with
out these sequences?
Protein Composition and
Structure
N
Leu
Tyr
Gln
Leu
Glu
Asn
Tyr
C
2
within this sequence can fold into regular structures (the secondary structure),
such as the a-helix. Entire chains fold into well-defined structures (the tertiary
structure)—in this case, a single insulin molecule. Such structures assemble with
other chains to form arrays such as the complex of six insulin molecules shown
at the far right (the quaternary structure). These arrays can often be induced to
form well-defined crystals (photograph at left), which allows a determination of
these structures in detail. [Photograph from Christo Nanev.]
CHAPTER
Leu
Insulin is a protein hormone, crucial for maintaining blood sugar at appropriate
levels. (Below) Chains of amino acids in a specific sequence (the primary
Glu
structure) define a protein such as insulin. Amino acids close to one another
Secondary structure
Tertiary structure
Quarternary structure
Primary structure
dimensional structure formed by hydrogen bonds
between amino acids near one another is called
secondary structure, whereas tertiary structure is
roteins are the most versatile macromolecules formed by long-range interactions between amino
acids. Protein function depends directly on this three
dimensional structure (Figure 2.1). Thus, proteins are
in living systems and
the embodiment of the transition from the oneserve crucial functions in essentially all biological
dimensional world of sequences to the threeprocesses. They func tion as catalysts, transport and dimensional world of molecules capable of diverse
store other molecules such as oxygen, pro vide
activities . Many proteins also display
mechanical support and immune protection, generate O U T L I N E
movement, transmit nerve impulses, and control
growth and differentiation. Indeed, much of this book 2.1 Proteins Are Built from a Repertoire of 20 Amino Acids
will focus on understanding what proteins do and
2.2 Primary Structure: Amino Acids Are Linked by Peptide Bonds to
how they perform these functions.
Form Polypeptide Chains
Several key properties enable proteins to participate
2.3 Secondary Structure: Polypeptide Chains Can Fold into Regular
in a wide range of functions.
P
Structures Such As the Alpha
Helix, the Beta Sheet, and Turns and Loops
1. Proteins are linear polymers built of monomer
units called amino acids, which are linked end to end. 2.4 Tertiary Structure: Water-Soluble Proteins Fold into Compact
The sequence of linked amino acids is called the
Structures with Nonpolar Cores
primary structure. Remarkably, proteins
2.5 Quaternary Structure: Polypeptide Chains Can Assemble into
spontaneously fold up into three-dimensional
Multisubunit Structures
structures that are determined by the sequence of
2.6 The Amino Acid Sequence of a Protein Determines Its Three
amino acids in the protein polymer. ThreeDimensional Structure
27
DNA
FIGURE 2.1 Structure dictates function. A protein component of the DNA
2. Proteins contain a wide range of functional groups
. These functional groups include alcohols, thiols,
thioethers, carbox ylic acids, carboxamides, and a
variety of basic groups. Most of these groups are
chemically reactive. When combined in various
sequences, this array of functional groups accounts
for the broad spectrum of protein function. For
instance, their reactive properties are essential to
the function of enzymes, the proteins that catalyze
specific chemical reactions in bio logical systems
(Chapters 8 through 10).
3. Proteins can interact with one another and with
other bio logical macromolecules to form complex
assemblies . The proteins within these assemblies
can act synergistically to generate capabilities that
individual proteins may lack. Examples of these
assemblies include macromolecular machines that
repli cate DNA, transmit signals within cells, and
enable muscle cells to contract (Figure 2.2).
replication machinery surrounds a section
quaternary structure, in which the functional protein is (A)
composed of several distinct polypeptide chains.
of DNA double helix depicted as a
cylinder. The protein, which consists of
two identical subunits (shown in red and
yellow), acts as a clamp that allows large
segments of DNA to be copied without
the replication
machinery dissociating from the DNA.
Myofibrils
[Drawn from 2POL.pdb.]
Single muscle fiber (cell) Plasma
membrane
Single
myofibril
Nucleus
Sarcomere
I band A band I band
Z line
(B)
(C)
FIGURE 2.2
A complex
protein
assembly. (A) A
single muscle cell
contains
multiple
myofibrils, each
Thick
of which is
filaments
comprised of
numerous repeats Z line
of a
complex protein
assembly known
as the
sarcomere. (B) The banding pattern of a
sarcomere, evident by electron microscopy,
is caused by (C) the interdigitation of
filaments made up of many individual
proteins. [(B) Courtesy of Dr. Hugh Huxley.]
H zone
Thin
filaments
4. Some proteins are quite rigid, whereas others
display considerable flexibility . Rigid units can
function as structural elements in the cytoskeleton
(the internal scaffolding within cells) or in connective
tissue. Proteins with some flexibility may act as
hinges, springs, or levers. In addition, conformational
changes within proteins enable the regulated
assembly of larger protein complexes as well as the
transmission of information within and between cells
Iron
2.1 Proteins Are Built from a Repertoire of 20
Amino Acids
Amino acids are the building blocks of proteins. An amino acid consists of a central carbon atom, called
the carbon, linked to an amino group, a carboxylic
acid group, a hydrogen atom, and a distinctive R
group. The R group is often referred to as the side
chain . With four different groups connected to the
tetrahe
dral a -carbon atom, a -amino acids are chiral: they
may exist in one or the other of two mirror-image
forms, called the L isomer and the D isomer (Figure
2.4).
Only L amino acids are constituents of proteins . For
almost all amino acids, the L isomer has S (rather
than R ) absolute configuration (Figure 2.5). What is
the basis for the preference for L amino acids? The
answer has been lost
to evolutionary history. It is possible that the
preference for L over D amino acids was a
consequence of a chance selection. However, there
is evidence that L amino acids are slightly more
soluble than a racemic mixture of D and L amino
29
2.1 Amino Acids
FIGURE 2.3 Flexibility and
function. On binding iron, the protein lactoferrin undergoes a substantial change
in conformation that allows other molecules to distinguish between the iron-free
and the iron-bound forms. [Drawn from 1LFH.pdb and 1LFG.pdb.]
Notation for distinguishing stereoisomers The four different substituents of an
asymmetric carbon atom are assigned a priority according to atomic number. The
lowest-priority substituent, often hydrogen, is pointed away from the viewer. The
configuration about the carbon atom is called S (from the Latin sinister, “left”) if the
progression from the highest to the lowest priority is counterclockwise. The
configuration is called R (from the Latin rectus, “right”) if the progression is
acids, which tend to form
crystals. This small solubility
clockwise.
difference could have been
(also called zwitterions ). In the
amplified over time so that the L dipolar form, the amino group is
isomer became dominant in
protonated
solution. Amino acids in
R
solution at neutral pH exist
predominantly as dipolar ions H (4)
(3)
(1) (2) Cα
HRRHC C
α
COO−
NH3+
α
(}NH3 ) and the carboxyl group is deprotonated
(}COO ). The ionization state of an amino acid varies
with pH (Figure 2.6). In acid solution (e.g., pH 1), the
amino group is protonated (}NH3 ) and the carboxyl
group is not dissociated (}COOH). As the pH is
raised, the carboxylic acid is the first group to give
up a proton, inasmuch as its p Ka is near 2. The
dipolar form persists until the pH approaches 9,
when the protonated amino group loses a proton.
NH3+ NH3+
COO− COO−
L isomer D isomer
FIGURE 2.4 The L and D isomers of amino acids. The letter R refers to the side
chain. The L and D isomers are mirror images of each other.
FIGURE 2.5 Only L amino acids are found in proteins. Almost all L amino
acids have an S absolute configuration. The counterclockwise direction of the
arrow from highest- to lowest-priority substituents indicates that the chiral
center is of the S configuration.
30
CHAPTER 2 Protein Composition and Structure
RH
FIGURE 2.6 Ionization state as a
function of pH. The
acids is altered by a change in R
HC
ionization state of amino
pH.
H+
H+
The zwitterionic form predominates near physiological pH.
RHC
C
H
+ COOH + +H N H N COO– H N COO– H+
3
3
2
o
it
a
r
t
n
e
c
n
o
C
Zwitterionic form
Both groups
protonated
Both groups deprotonated
n
0 2 4 6810 12 14 pH
Twenty kinds of side chains varying in size, shape, charge,
hydrogen bonding capacity, hydrophobic character, and chemical
reactivity are com monly found in proteins. Indeed, all proteins in all
species—bacterial, archaeal, and eukaryotic—are constructed from
the same set of 20 amino acids with only a few exceptions. This
fundamental alphabet for the con struction of proteins is several
billion years old. The remarkable range of functions mediated by
proteins results from the diversity and versatility of these 20 building
blocks. Understanding how this alphabet is used to create the
intricate three-dimensional structures that enable proteins to carry out
so many biological processes is an exciting area of biochemistry and
one that we will return to in Section 2.6.
Although there are many ways to classify amino acids, we will
sort these molecules into four groups, on the basis of the general
chemical characteris tics of their R groups:
1. Hydrophobic amino acids with nonpolar R groups
2. Polar amino acids with neutral R groups but the charge is not
evenly distributed
3. Positively charged amino acids with R groups that have a positive
charge at physiological pH
4. Negatively charged amino acids with R groups that have a
negative charge at physiological pH
Hydrophobic amino acids. The simplest amino acid is glycine, which
Glycine
(Gly,
G)
Alanine
(Ala, A)
has a single hydrogen atom as its side chain. With two hydrogen
atoms bonded to the a -carbon atom, glycine is unique in being achiral
. Alanine, the next simplest amino acid, has a methyl group (}CH 3 )
as its side chain (Figure 2.7).
C
H
H
H
H CH2
C
CH3
H3C CH3
Valine
(Val,
V)
Proline
(Pro,
P)
2C
Leucine
(Leu, L)
CH3
H
+H N
3
COO–
+H N
3
COO–
C
H2C
H2COO– H
2C
CH3 C
H
+ COO– H N C
3
+H N
3
(Gly, G)
(Ala, A)
H2
H2C CH2 N+ COO–
Alanine
C
+ COO– H N C
3
H
Proline (Pro, P)
CH3
CH2
+H N
3
C
+H N
3
CH3
COO–
H
Glycine
HC
CH
COO–
H
CH
H
N+
COO–
CH3
H CH3 C
Valine
(Val, V)
H CH3 C
H
CH2
H
+H N
3
Leucine (Leu, L)
COO–
C
Isoleucine
(Ile, I)
H3C
CH3
H
H C CH *
2
H
3
Methionine (Met,
M)
S
H2C
Tryptophan (Trp,
W)
H
H
H
Phenylalanine
(Phe, F)
C
+H N
3
C H COO–
+H N
3
C
+H N
3
CH2
H
C
CH2
H
H
HNH
COO–
CH2
H
+H N COO–
3
CH3
C
H CH3
CH2
CH2
H
C
CH3 S
+H N COO– HC
3
COO–
+H N
3
Isoleucine (Ile, I)
CH
HC
C
CH2
HC
CH
COO–
HN
HC
CH
HC
CH2 C
CH
H
H
H
CH
CC
+ COO– H N C
3
Phenylalanine
CH
H
H
CH2
C
(Phe, F)
representation (see the Appendix to Chapter 1).
FIGURE 2.7 Structures of hydrophobic amino
acids. For each amino acid, a ball-and-stick model The additional chiral center in isoleucine is indicated
(top) shows the arrangement of atoms and bonds in by an asterisk.
space. A stereochemically realistic formula (middle) + COO– H3N C
shows the geometric arrangement of bonds around
atoms, and a Fischer projection (bottom) shows all H
Tryptophan
bonds as being perpendicular for a simplified
31
Methionine (Met, M)
(Trp, W)
Tyrosine (Tyr, Y)
Asparagine (Asn,
N)
Glutamine (Gln, Q)
O
HO
CH
H
Threonine (Thr, T)
Serine (Ser, S)
CH3
H
COO–
H
C
*
H2N
OH
H
H
C
NH2
+H N
3
+H COO–
OC O CH
H 2
3N
H
OH
H
C
H
COO–
H
(Ser, S) Threonine (Thr, T)
C
HC
CH
(Cys, C)
C
CH
+H N
3
HCH2
+H N COO–
3
O NH2 C
O NH2
C
HC
C
CH2
2
+H N COO–
3
C
H
Serine
HO
CH3
C
+ COO– H N
3
Cysteine
H
CH
+H N COO–
3
OH
CH2
+H N
3
H2C
H
H
Asparagine
CH2
CH2
+ COO– H N C
3
CH2
+ COO– H N C
3
Tyrosine (Tyr, Y)
CH
COO–
(Asn, N)
Glutamine (Gln, Q)
H
SH
HCH2
+H N COO–
3
SH
CH2
+ COO– H N
3
C
H
Cysteine
(Cys, C)
FIGURE 2.8 Structures of the polar amino acids. The additional chiral center
in threonine is indicated by an asterisk.
HN
tend to cluster together rather than contact water.
The three-dimensional structures of water
soluble proteins are stabilized by this tendency of
hydrophobic groups to come together, which is
called the hydrophobic effect (p. 9). The different
sizes and shapes of these hydrocarbon side chains
enable them to pack together to form compact
structures with little empty space. Proline also has an
aliphatic side chain, but it differs from other
members of the set of 20 in that its side chain is
bonded to both the nitrogen and the a -carbon atoms,
yielding a pyrrolidine ring. Proline markedly
influences protein architecture because its cyclic
struc
ture makes it more conformationally restricted than
the other amino acids. Two amino acids with
relatively simple aromatic side chains are part of the
fundamental repertoire. Phenylalanine, as its name
indicates, contains a phe nyl ring attached in place of
one of the hydrogen atoms of alanine. Tryptophan
has an indole group joined to a methylene (}CH 2})
group; the indole group comprises two fused rings
containing an NH group. Phenylalanine is purely
hydrophobic, whereas tryptophan is less so because
of its NH group.
Larger hydrocarbon side chains are found in valine,
leucine, and isoleucine . Methionine contains a
Polar amino acids. Six amino acids are polar but
largely aliphatic side chain that includes a thioether uncharged. Three amino acids, serine, threonine,
(}S}) group. The side chain of isoleucine includes an and tyrosine, contain hydroxyl groups (}OH) attached
additional chiral center; only the isomer shown in
to a hydrophobic side chain (Figure 2.8). Serine can
Figure 2.7 is found in proteins. The larger aliphatic be thought of as a version
side chains are especially hydrophobic; that is, they
of alanine with a hydroxyl group attached, threonine resembles valine with a Indole
32
hydroxyl
isoleucine,
group in place con tains an
of one of
additional
valine’s
asymmetric
methyl
center; again,
groups, and only one
tyrosine is a isomer is
version of
present in
phenylalanine proteins.
with the
In addition,
hydroxyl
the set
group
includes
replacing a asparagine NH3
hydrogen
and glutamine H2C
atom on the , two amino +
aromatic ring.acids that con Arginine (Arg, R)
The hydroxyl tain a terminal
group makes carboxamide .
these amino The side
acids much chain of
more
glutamine is
hydrophilic
one
(water loving) methylene
and reactive group longer
than their
than that of
hydrophobic asparagine.
Lysine
analogs.
H2N
(Lys, K)
Threonine,
like
H
N
+
C
HN
NH2
H
C
Histidine (His, H)
Cysteine is
structurally
similar to ser
ine but
contains a
sulfhydryl, or
thiol (}SH),
group in place
of the
hydroxyl
(}OH) group.
The sulfhydryl
group is
H2C
H2C
H
C
CH2 CH2
H
much more reactive.
Pairs of sulfhydryl
+
C
CH2 CH2
N
H
C
H
C
particularly impor tant in
stabilizing some proteins,
NH3+
as will be discussed
shortly.
CH2
NH
CH2
+H N COO–
3
Positively charged amino CH2
+H N COO–
acids. We turn
3
groups may come
together to form disul fide H3N COO–
bonds, which are
+
H2N NH2 C
H
N
HC
now to amino acids hydrophilic. Lysine CH CH
2
2
and
arginine
have
with complete posi
N
long
CH
tive charges that
CH
CH
2
2
render them highly
C
H
amino
group
and
arginine
side chains that terminate
by a guanidinium group. Lysine
with groups that are
(Lys, K)
Histidine contains an
positively charged at
+ COO– H N C
3
imidazole
neutral pH. Lysine is
+ COO– H N C
H
3
capped by a primary
CH2
Arginine
(Arg, R)
+ COO– H N C
3
H
Histidine
(His, H)
group, an aromatic ring that also can be
positively charged (Figure 2.9).
(Figure 2.10). Histidine is often found in the active
value
near
6,
the
imidazole
group
can
sites of enzymes, where NH2
With a p Ka
the imidazole ring can bind and release protons in
be uncharged or posi tively charged near neutral the course of enzymatic +
N
pH, depending on its local environment
HH
FIGURE 2.9 Positively charged amino acids lysine, arginine, and histidine.
reactions.
C
H2N NH2
C
NC
C
H
H
are charged derivatives of
called aspartate and glutamate to
This set of amino acids contains asparagine and glutamine (Figure emphasize that, at physiological
two with acidic side chains:
2.8), with a carboxylic acid in
Guanidinium
aspartic acid and glutamic acid
place of a carboxamide. Aspartic Imidazole
(Figure 2.11). These amino acids acid and glutamic acid are often
present in the acid form
Negatively charged amino acids.
HH
pH, their side chains usually lack a proton that is
and hence are negatively charged. Nonetheless, these side chains can accept
N
N
HC
HC
protons in some proteins, often with functionally important consequences.
+
CH
CH
Seven of the 20 amino acids have readily ionizable side chains. These
H+
N
N
C
C
H
7 amino acids are able to donate or accept protons to facilitate reactions as
CH2
CH2
well as to form ionic bonds. Table 2.1 gives equilibria and typical p Ka val ues for ionization of the side chains of
tyrosine, cysteine, arginine, lysine,
H
H
H+
C
bind or release protons near physiological pH.
C
NCOH
NCOH
FIGURE 2.10 Histidine ionization. Histidine can
33
Aspartate (Asp, D)
Glutamate (Glu, E)
TABLE 2.1 Typical pKa values of ionizable
groups in proteins
Group Acid Base Typical pKa*
O
O
CHO
O
O
Aspartic acid
Terminal a-carboxyl group 3.1
C–O
C–
Glutamic acid 4.1 C H
O
O–
–
CO
O
N
O
H
O
C
Histidine 6.0 C
H2
C
H
CH2
+H N COO–
3
+H N COO–
3
+
C
CH2
H
N
+
N
H
H
NH
N
H
Terminal a-amino group 8.0 H
OO–
C
H
OO–
N
H
H
Cysteine 8.3 CH2
C
CH2
S–
S
CH2
H
Tyrosine 10.9
+H COO–
3N C
H
Glutamate
H
Aspartate
+H COO–
O–
N
O
H
3N C
NH
+
(Glu, E)
Lysine 10.8 H
(Asp, D)
H
H
FIGURE 2.11 Negatively charged
H
H
amino acids.
N
N
H
+
H
H
Arginine 12.5
N
H
C
NH
N
H
C
NH
*pKa values depend on temperature, ionic strength, and the microenvironment of the ionizable group
histidine, and aspartic and glutamic acids in proteins. Two other
groups in proteins—the terminal a -amino group and the terminal a carboxyl group—can be ionized, and typical p Ka values for these
groups also are included in Table 2.1.
Amino acids are often designated by either a three-letter
abbreviation or a one-letter symbol (Table 2.2). The abbreviations for
amino acids are the first three letters of their names, except for
asparagine (Asn), glutamine (Gln), isoleucine (Ile), and tryptophan
(Trp). The symbols for many amino acids are the first letters of their
names (e.g., G for glycine and L for leucine);
TABLE 2.2 Abbreviations for amino acids
Three-letter
One-letter
abbreviation
abbreviation
Amino acid
Amino acid
Three-letter
abbreviation
One-letter
abbreviation
Alanine Ala A Methionine Met M Arginine Arg R Phenylalanine Phe F Asparagine
Asn N Proline Pro P Aspartic acid Asp D Serine Ser S Cysteine Cys C Threonine
Thr T Glutamine Gln Q Tryptophan Trp W
Glutamic acid Glu E Tyrosine Tyr Y Glycine Gly G Valine Val V Histidine His H
Asparagine or
Isoleucine Ile I aspartic acid Asx B Leucine Leu L Glutamine or
Lysine Lys K glutamic acid Glx Z
34
H
H
C
C
NH
2C
C
O
H
2C
H
OH
N
H
X
2C
NH
C
C
2
CH
2 CO
FIGURE 2.12
Undesirable
C
cyclization of serine would form
a strained, four-membered ring
2
Homoserine
H
H
O
HX+
HOH
O
Serine
X
H CHO
form a stable, five-membered
ring, potentially resulting in
peptide-bond cleavage. The
C
N
H
C
O
HX+
and is thus
disfavored. X
can be an
amino group
from a
neighboring
amino acid or
another
potential
leaving group.
the other symbols have been agreed on by convention. These
abbreviations and symbols are an integral part of the vocabulary of
biochemists.
How did this particular set of amino acids become the building
blocks of proteins? First, as a set, they are diverse: their structural
and chemical properties span a wide range, endowing proteins with
the versatility to assume many functional roles. Second, many of
these amino
acids were probably available from prebiotic reactions; that is, from
reac tions that took place before the origin of life. Finally, other
possible amino acids may have simply been too reactive. For
example, amino acids such as homoserine and homocysteine tend
to form five-membered cyclic forms that limit their use in proteins;
the alternative amino acids that are found in proteins—serine and
cysteine—do not readily cyclize, because the rings in their cyclic
forms are too small (Figure 2.12).
2.2 Primary Structure: Amino Acids Are Linked by
Peptide Bonds to Form Polypeptide Chains
Proteins are linear polymers formed by linking the a -carboxyl group
of one amino acid to the a -amino group of another amino acid. This
type of linkage is called a peptide bond or an amide bond . The
formation of a dipeptide from two amino acids is accompanied by the
loss of a water molecule (Figure 2.13). The equilibrium of this reaction
lies on the side of hydrolysis rather than synthesis under most
conditions. Hence, the biosynthesis of peptide bonds requires an input
of free energy. Nonetheless, peptide bonds are quite stable kinetically
because the rate of hydrolysis is extremely slow; the lifetime of a
reactivity in amino cyclization.
acids. Some amino Homoserine can
cyclize to
acids are
unsuitable for
35
proteins because 2.2 Primary
of undesirable
Structure
peptide bond in aqueous solution in the absence of a catalyst
approaches 1000 years.
A series of amino acids joined by peptide bonds form a polypeptide
chain, and each amino acid unit in a polypeptide is called a residue.
A poly peptide chain has directionality because its ends are different :
an a -amino group
H R1 C
O
C
OC
–
+
+H N
3
+H N
3
H R2 C
C
O
O
C
O–
+H N
3
H R1 C
O
+ H2O
–
C
H
N
O
R2 H
Peptide bond
FIGURE 2.13 Peptide-bond formation. The linking of two amino acids is accompanied by the loss of
a molecule of water.
36
CH3
CHAPTER 2 Protein Composition and
HC CH3
Structure
O
OH
O
H
+H
3N C
C
H
NC
HH
O
HH
C
N
H
H2C H2C
H
H
C
N
C C H2CH
O
C
N
H
COC
–
O
Tyr Gly Gly Phe Leu
Amino
terminal
residue
Carboxyl
terminal residue
theory of matter.
Kilodalton (kDa)
Kilodalton (kDa)
A unit of mass equal to 1000 daltonsA unit
of mass equal to 1000 daltons
FIGURE 2.14 Amino acid sequences have direction. This illustration of the
pentapeptide Tyr-Gly-Gly-Phe-Leu (YGGFL) shows the sequence from the
amino terminus to the carboxyl terminus. This pentapeptide, Leu-enkephalin, is
an opioid peptide that modulates the perception of pain. The reverse
pentapeptide, Leu-Phe-Gly-Gly-Tyr (LFGGY), is a different molecule and has
no such effects.
is present at one end and an a -carboxyl group at the
other. The amino end is taken to be the beginning
of a polypeptide chain; by convention, the sequence
of amino acids in a polypeptide chain is written
starting with the amino-terminal residue. Thus, in the
polypeptide Tyr-Gly-Gly-Phe
Leu (YGGFL), tyrosine is the amino-terminal (Nterminal) residue and leucine is the carboxylterminal (C-terminal) residue (Figure 2.14). Leu PheGly-Gly-Tyr (LFGGY) is a different polypeptide, with
Dalton
different chemical properties.
Dalton
A unit of mass very nearly equal to
A polypeptide chain consists of a regularly repeating
that of a
part,
called the main chain or backbone, and a
A unit of mass very nearly equal to that
of a hydrogen atom. Named after John variable part, comprising the distinctive side chains
(Figure 2.15). The polypeptide backbone is rich in
Dalton
hydrogen atom. Named after John Dalton hydrogen- bonding potential. Each residue contains
(1766–1844), who developed the atomic
a carbonyl group (C “ O), which is a good hydrogen(1766–1844), who developed the atomic theory of matter.
bond acceptor, and, with the exception of proline, an
consists of more than 27,000 amino acids.
NH group, which is a good hydrogen-bond donor.
Polypeptide chains made of small numbers of amino
These groups interact with each other and with
acids are called oligopeptides or simply peptides .
functional groups from side chains to stabilize
particular structures, as will be discussed in Section The mean molecular weight of an amino acid
2.3.
residue is about 110 g mol 1 , and so the molecular
Most natural polypeptide chains contain between 50 weights of most proteins are between 5500 and
and 2000 amino acid residues and are commonly
220,000 g mol 1 . We can also refer to the mass of a
referred to as proteins . The largest single
protein, which is expressed in units of
polypeptide known is the muscle protein titin , which
CCO
NH
daltons; with a
a mass of proteins,
O
C
H
HR1
N
one dalton molecular 50,000
the linear
HR3
O
N
C
H
is equal to weight of daltons, or polypeptide
C
N
C
one atomic 50,000 g 50 kDa
chain is
HR5
H
1
C
mass
unit.
(kilodaltons
C
mol has
H
O
C
H
C
A protein
). In some
N
H
O
common cross-links are disul
R2
cross-linked. The most
R4
cysteine residues (Figure 2.16). The resulting unit of
FIGURE 2.15 Components of a polypeptide chain. A polypeptide chain
two linked cysteines is called cystine . Extracellular
consists of a constant backbone (shown in black) and variable side chains
(shown in green).
proteins
fide bonds, formed by the oxidation of a pair of
often have several disulfide
OH
bonds, whereas intracel
lular proteins usually lack
cross-links derived from other C
OH
them. Rarely, nondisulfide
side chains are present
C
N
H
in proteins. For example, collagen fibers in connective
C
N
H2C
C
tissue are strengthened in this way, as are fibrin blood
S
H
clots (Section 10.4).
H2C
H
sequences
S
Cysteine
+ 2 e– +2 H
Proteins have unique amino acid
specified by genes
(Figure 2.17).
biochemistry because
it showed C
H
S
In 1953, Frederick
CH2
Sanger determined H
the amino acid
This work is a
sequence of insulin, a
landmark in
protein hormone
for the first time that a protein has a
precisely defined amino N
Reduction
+ Oxidation
H
C
CH2 C
N
S
C
OH
by peptide bonds. This
O H Cysteine
accomplishment stimulated Cystine
other
scientists to carry out sequence studies of a wideknown. The striking
variety of proteins. Currently, the complete amino FIGURE 2.16 Cross-links. The formation of a disulfide bond from two
acid sequences of more than 2,000,000 proteins are cysteine residues is an oxidation reaction.
acid sequence consisting
only of L amino acids linked
fact is that each protein has a unique, precisely defined amino acid
sequence . The amino acid sequence of a protein is referred to as its
primary structure .
SS
A chain
Gly-Ile-Val-Glu-Gln-Cys-Cys-Ala-Ser-Val-Cys-Ser-Leu-Tyr-Gln-Leu-Glu-Asn-TyrCys-Asn 5 10 15 21
SS
S
S
B chain
Phe-Val-Asn-Gln-His-Leu-Cys-Gly-Ser-His-Leu-Val-Glu-Ala-Leu-Tyr-Leu-Val-Cys-Gly-Glu-Arg-Gly-Phe-Phe-Tyr-Thr-ProLys-Ala 5 10 15 20 25 30
FIGURE 2.17 Amino acid sequence of bovine insulin.
A series of incisive studies in the late 1950s and early 1960s
revealed that the amino acid sequences of proteins are determined
by the nucleotide sequences of genes. The sequence of nucleotides
in DNA specifies a com plementary sequence of nucleotides in RNA,
which in turn specifies the amino acid sequence of a protein. In
particular, each of the 20 amino acids of the repertoire is encoded by
one or more specific sequences of three nucleotides (Section 4.6).
Knowing amino acid sequences is important for several reasons.
First, knowledge of the sequence of a protein is usually essential to
elucidating its function (e.g., the catalytic mechanism of an enzyme).
In fact, proteins with novel properties can be generated by varying the
sequence of known
proteins. Second, amino acid sequences
determine the three-dimensional structures of proteins. The amino
acid sequence is the link between the genetic message in DNA and
the three-dimensional structure that performs a protein’s biological
function. Analyses of relations between amino acid sequences and
three-dimensional structures of proteins are uncovering the rules that
govern the folding of polypeptide chains. Third, alterations in amino
acid sequence can lead to abnormal protein function and disease.
Severe and sometimes fatal diseases, such as sickle-cell anemia
(Chapter 7) and cystic fibrosis, can result from a change in a single
amino acid within a protein. Fourth, the sequence of a protein reveals
much about its evolution ary history (Chapter 6). Proteins resemble
one another in amino acid sequence only if they have a common
ancestor. Consequently, molecular events in evolution can be traced
from amino acid sequences; molecular paleontology is a flourishing
area of research.
atom and CO group of the first amino acid and the
NH group and a -carbon atom of the second amino
acid. The nature of
38
CHAPTER 2 Protein Composition and Structure
H
Polypeptide chains are flexible yet conformationally restricted
Examination of the geometry of the protein
the chemical bonding within a peptide accounts for
backbone reveals several important features. First, the bond’s planarity. The bond resonates between a
the peptide bond is essentially planar (Figure 2.18). single bond and a double bond. Because of this
Thus, for a pair of amino acids linked by a peptide
partial double-bond character, rotation about this
bond, six atoms lie in the same plane: the a -carbon bond is prevented and thus the conforma
backbone is constrained.
Cα
N
C
αC
H
tion of the peptide
C
C
O
FIGURE 2.18 Peptide bonds are planar. In a pair of linked amino acids, six
O
N
C
atoms (C , C, O, N, H, and C ) lie in a plane. Side chains are shown as green
balls.
Peptide-bond resonance structures
H
C
C
N+
C
O–
The partial double-bond character is also expressed
in the length of the bond between the CO and the
NH groups. As shown in Figure 2.19, the C}N
distance in a peptide bond is typically 1.32 Å, which
is between the values expected for a C}N single
37
bond (1.49 Å) and a C “ N
Cα
to form tightly trans over cis common cis
packed globularcan be
peptide bonds
structures.
explained by the are X}Pro
fact that steric linkages. Such
Two
bonds show
configurations clashes
are possible for between groups less preference
for the trans
a planar peptide
configuration
bond. In the
because the
trans
nitrogen of
configuration,
H
proline is
the two a bonded to two
carbon atoms
tetrahedral
are on opposite
N
carbon atoms,
sides
of
the
1.0 Å
peptide bond. attached to the limiting the
Cα
1.45 Å
In the cis
a -carbon atoms steric
1.51 Å
differences
configuration, hinder the
double bond
(1.27 Å). Finally, these groups
C
formation of the between the
trans and cis
are on the same cis
the peptide
1.32 Å
forms (Figure
side of the pep configuration
bond is
tide bond.
uncharged,
but do not arise 2.21).
Almost all
allowing
In contrast with
in the trans
peptide bonds inconfiguration
polymers of
the peptide
proteins are
amino acids
(Figure 2.20). bond, the bonds
trans . This
linked by
By far the most between the
peptide bonds preference for
1.24 Å O
two adjacent rigid peptide units can rotate
about these bonds, taking on various
amino group and the a -carbon atom and orientations. This freedom of rotation about
between the a -carbon atom and the
two
carbonyl group are pure single bonds. The
FIGURE 2.19 Typical bond lengths within a peptide unit. The peptide unit is
shown in the trans configuration.
bonds of each amino acid allows proteins to fold in
many different ways . The rotations about these
bonds can be specified by
Trans Cis
FIGURE 2.20 Trans and cis peptide bonds. The trans form is strongly favored because of steric
clashes, indicated by the orange semicircles, that arise in the cis form.
39
2.2 Primary Structure
Trans Cis
FIGURE 2.21 Trans and cis X–Pro bonds. The energies of these forms are similar to one
another because steric clashes, indicated by the orange semicircles, arise in both forms.
(B)
(C)
RH
RH
OH
(A)
C
C
C
C
N
C
N
C
N
HH
O
HR
O
= +85°
= −80°
fold into well defined structures is remarkable
thermodynamically. An unfolded polymer exists as a
acid in a polypeptide can be adjusted by rotation about two single bonds. (A)
Phi ( ) is the angle of rotation about the bond between the nitrogen and the a- random coil: each copy of an unfolded polymer will
carbon atoms, whereas psi ( ) is the angle of rotation about the bond between thehave a differ ent conformation, yielding a mixture of
a-carbon and the carbonyl carbon atoms. (B) A view down the bond between many possible conformations. The favorable
the nitrogen and the a-carbon atoms, showing how is measured. (C) A view entropy associated with a mixture of many
down the bond between the a-carbon and the carbonyl carbon atoms, showing
conformations opposes folding and must be
how is measured.
overcome by interactions favoring the folded form.
Thus, highly flexible polymers with a large number of
possible con formations do not fold into unique
torsion angles (Figure 2.22). The angle of rotation
structures. The rigidity of the peptide unit and the
about the bond between the nitrogen and the a carbon atoms is called phi ( ). The angle of rotation restricted set of allowed f and c angles limits the
number of structures accessible to the unfolded form
about the bond between the a -carbon and the
sufficiently to allow protein folding to take place.
carbonyl carbon atoms is called psi ( ) .
A clockwise rotation about either bond as viewed
from the nitrogen atom toward the a -carbon atom or
from the a -carbon atom toward the carbonyl group
corresponds to a positive value. The and angles
determine the path of the polypeptide chain.
Are all combinations of and possible?
Gopalasamudram Ramachandran recognized that
many combinations are forbidden because of steric
collisions between atoms. The allowed values can be
visualized on a two-dimensional plot called a
Ramachandran plot (Figure 2.23). Three
quarters of the possible ( , ) combinations are
excluded simply by local steric clashes. Steric
exclusion, the fact that two atoms cannot be in the
same place at the same time, can be a powerful
organizing principle.
Torsion angle
The ability of biological polymers such as proteins to A measure of the rotation about a bond, usually taken to lie between 2180 and 1180
FIGURE 2.22 Rotation about bonds in a polypeptide. The structure of each amino
degrees. Torsion angles are sometimes called dihedral angles.
+180
40
CHAPTER 2 Protein Composition and Structure
120
60
0
−60
−120
( = 90°, = −90°)
−180
−60
−120 −180
600
120
+180
Disfavore
d
2.3 Secondary Structure: Polypeptide Chains Can
Fold into Regular Structures Such As the Alpha
Helix, the Beta Sheet, and Turns and Loops
Can a polypeptide chain fold into a regularly
repeating structure? In 1951, Linus Pauling and
Robert Corey proposed two periodic structures called
the helix (alpha helix) and the pleated sheet (beta
pleated sheet). Subsequently, other structures such
as the turn and omega ( V ) loop were identified.
Although not periodic, these common turn or loop
structures are well defined and contribute with a
helices and b sheets to form the final protein
structure. Alpha helices, b strands, and turns are
formed by a regu
lar pattern of hydrogen bonds between the peptide
N}H and C “ O groups of amino acids that are near
one another in the linear sequence . Such folded
segments are called secondary structure .
The alpha helix is a coiled structure stabilized by intrachain
hydrogen bonds
In evaluating potential structures, Pauling and Corey
considered which con formations of peptides were
sterically allowed and which most fully exploited the
hydrogen-bonding capacity of the backbone NH and
CO groups. The first of their proposed structures,
the helix, is a rodlike structure (Figure 2.24). A
tightly coiled backbone forms the inner part of the rod
and the side chains extend outward in a helical
array. The a helix is stabilized by hydrogen bonds
between the NH and CO groups of the main chain. In
par ticular, the CO group of each amino acid forms a
hydrogen bond with the NH group of the amino acid
that is situated four residues ahead in the sequence
(Figure 2.25). Thus, except for amino acids near the
ends of an a helix, all the main-chain CO and NH
groups are hydrogen bonded. Each resi due is
Screw sense
related to the next one by a rise, also called
Screw sense
Describes the direction in which a helical translation, of 1.5 Å along the helix axis and a
Describes the direction in which a helical structure rotates with respect to its axis. If, rotation of 100 degrees, which gives 3.6 amino acid
structure rotates with respect to its axis. resi dues per turn of helix. Thus, amino acids spaced
If, viewed down the axis of a helix,
three and four apart in the sequence are spatially
the chain turns
quite close to one another in an a helix. In contrast,
viewed down the axis of a helix, the chain turns in a clockwise direction, it has a
amino acids spaced two apart in the sequence are
right-handed
in a clockwise direction, it has a right- situated on opposite sides of the helix and so are
handed screw sense. If the turning unlikely to make contact. The pitch of the a helix is
is counterclockwise,
the length of one complete turn along the helix axis
screw sense. If the turning is counterclockwise, the screw sense is left-handed.
and is equal to the product of the rise (1.5 Å) and the
the screw sense is left-handed.
FIGURE 2.23 A Ramachandran plot showing the number of residues per turn (3.6), or 5.4 Å. The
values of and . Not all and values are possible without collisions between screw
atoms. The most favorable regions are shown in dark green; borderline regions
are shown in light green. The structure on the right is disfavored because of
steric clashes.
(A)
(B) (C) (D)
handed
+180 120
60
0
41
2.3 Secondary Structure Left-
−60
FIGURE 2.24 Structure of the a helix. (A) A ribbon depiction shows the a-
carbon atoms and side chains (green). (B) A side view of a ball-and-stick
version depicts the hydrogen −120
helix (very rare)
bonds (dashed lines) between NH and CO groups. (C) An end view shows the
coiled backbone as the inside of the helix and the side chains (green)
projecting outward. (D) A
helix (common)
−120 −180
Right-handed
space-filling view of part interior core of the helix. −60
C shows the tightly packed
600
120
−180
+180
FIGURE 2.26 Ramachandran plot for
O
HO
Ri+2
H
i+4 H H H
R
Ri
N
C
H
O
Ri+1
helices. Both right- and left-handed helices lie in
regions of allowed conformations in the
HO
C
N
C
C
N
C
C
N
HO
OH
HR
Ri+5 H
i+3H
C
C
N
C
C
N
(A) (B)
FIGURE 2.25 Hydrogen-bonding scheme for an a helix. In the a helix, the CO
C
C
However, all a helicesare rightRamachandr
essentially in proteins handed.
an plot.
are in a helices (Figure 2.28). Indeed, about 25% of
all soluble proteins are composed of a helices
sense of an a helix can be right-handed (clockwise) connected by loops and turns of the polypeptide
or left-handed (counter clockwise). The
chain. Single a helices are usually less than 45 Å
Ramachandran plot reveals that both the rightlong. Many proteins that span biological membranes
handed and the left-handed helices are among
also contain a helices.
allowed conformations (Figure 2.26). However, righthanded helices are energetically more favorable
because there is less steric clash between the side
chains and the backbone. Essentially all helices
found in proteins are right-handed. In schematic
representations
of proteins, a helices are depicted as twisted ribbons
or rods (Figure 2.27). Not all amino acids can be
readily accommodated in an a helix. Branching at
the b -carbon atom, as in valine, threonine, and
isoleucine, tends to destabilize a helices because of
steric clashes. Serine, aspartate, and asparagine
FIGURE 2.27 Schematic views of a helices. (A) A ribbon depiction. (B) A
also tend to disrupt a helices because their side
cylindrical depiction.
chains contain hydrogen-bond donors or acceptors
in close proximity to the main chain, where they
compete for main-chain NH and CO groups. Proline
also is a helix breaker because it lacks an NH group
and because its ring structure prevents it from
assuming the value to fit into an a helix. The a helical content of proteins ranges widely, from none
to almost 100%. For example, about 75% of the
residues in ferritin, a protein that helps store iron,
FIGURE 2.28 A largely a-helical protein. Ferritin, an iron-storage protein, is
group of residue i forms a hydrogen bond with the NH group of residue i 1 4.
built from a bundle of a helices. [Drawn from 1AEW.pdb.]
is composed of two or
Pauling and Corey
proposed another periodic more polypeptide chains
called strands. A b
structural motif, which
−60
they named the pleated strand is almost fully
sheet ( b because it was extended rather than
−120
the second structure that being tightly coiled as in
the a helix. A range of
they elucidated, the a
extended structures are
helix having been the
−120 −180
first). The b pleated sheet sterically allowed (Figure
2.29).
Beta strands
(or, more simply, the b
Beta sheets are stabilized by
The distance between
sheet) differs markedly
hydrogen bonding between
adjacent
amino acids
from the rodlike a helix. It
polypeptide strands
along a b strand is approxi
600
distance of 1.5 Å
opposite directions
+180
120
along
an
a
helix.
chains of adjacent (Figure 2.30).
mately 3.5 Å, in
The
side
amino acids point in
contrast with a
0
+180
120
60
−180
−60
FIGURE 2.29 Ramachandran plot for b
strands. The red area shows the sterically
allowed conformations of extended,
b-strand-like structures.
7Å
FIGURE 2.30 Structure of a b strand. The side chains (green) are alternately above and
below the plane of the strand.
A b sheet is formed by linking two or more b strands lying next to one
another through hydrogen bonds. Adjacent strands in a b sheet can run in
opposite directions (antiparallel b sheet) or in the same direction (parallel b
sheet). In the antiparallel arrangement, the NH group and the CO group of
each amino acid are respectively hydrogen bonded to the CO group and the
NH group of a partner on the adjacent chain (Figure 2.31). In the parallel
arrangement, the hydrogen-bonding scheme is slightly more complicated.
For each amino acid, the NH group is hydrogen bonded to the CO group of
one amino acid on the adjacent strand, whereas the CO group is hydrogen
bonded to the NH group on the amino acid two residues farther along the
chain (Figure 2.32). Many strands, typically 4 or 5 but as many as 10 or
more, can come together in b sheets. Such b sheets can be purely antiparal
lel, purely parallel, or mixed (Figure 2.33).
FIGURE 2.31 An antiparallel b sheet. Adjacent b strands run in opposite directions, as
indicated by the arrows. Hydrogen bonds between NH and CO groups connect each amino
acid to a single amino acid on an adjacent strand, stabilizing the structure.
42
2.3 Secondary Structure
FIGURE 2.32 A parallel b sheet. Adjacent b strands run in the same direction, as indicated
by the arrows. Hydrogen bonds connect each amino acid on one strand with two different
amino acids on the adjacent strand.
FIGURE 2.33 Structure of a mixed b sheet. The arrows indicate directionality of each strand.
In schematic representations, b strands are usually depicted by broad
arrows pointing in the direction of the carboxyl-terminal end to indicate the
type of b sheet formed—parallel or antiparallel. More structurally diverse
than a helices, b sheets can be almost flat but most adopt a somewhat
twisted shape (Figure 2.34). The b sheet is an important structural element
in many proteins. For example, fatty acid-binding proteins, important for
lipid metabolism, are built almost entirely from b sheets (Figure 2.35).
(A) (B)
FIGURE 2.35 A protein rich in b
FIGURE 2.34 A schematic twisted b sheet. (A) A schematic model. (B) The
schematic view rotated by 90 degrees to illustrate the twist more clearly.
sheets. The structure of a fatty acid binding protein. [Drawn from 1FTP.pdb.]
44
CHAPTER 2 Protein Composition and Structure
i + 1i + 2
i+3
i
FIGURE 2.36 Structure of a reverse turn. The CO group of residue i of the
polypeptide chain is hydrogen bonded to the NH group of residue i 1 3 to
stabilize the turn.
FIGURE 2.37 Loops on a protein surface. A part of an antibody molecule
has surface loops (shown in red) that mediate interactions with other
molecules. [Drawn from 7FAB.pdb.]
Polypeptide chains can change direction by making reverse
Most proteins have compact, globular shapes
owing to reversals in the direction of their
polypeptide chains. Many of these reversals are
accom plished by a common structural element
called the reverse turn (also known as the turn or
hairpin turn ) , illustrated in Figure 2.36. In many
reverse turns, the CO group of residue i of a
polypeptide is hydrogen bonded to the NH group
of residue i 1 3. This interaction stabilizes abrupt
changes in direction of the polypeptide chain. In
other cases, more-elaborate structures are
responsible for chain reversals. These structures
are called loops or sometimes loops (omega
loops) to suggest their overall shape. Unlike a
helices and b strands, loops do not have regular,
periodic structures. Nonetheless, loop structures
are often rigid and well defined (Figure 2.37).
Turns and loops invariably lie on the surfaces of
proteins and thus often participate in interactions
between proteins and other molecules.
Fibrous proteins provide structural support for cells and
tissues
Special types of helices are present in the two
proteins a-keratin and collagen. These proteins
form long fibers that serve a structural role. a Keratin, which is an essential component of wool,
hair, and skin, con sists of two right-handed a
helices intertwined to form a type of left-handed
superhelix called an -helical coiled coil . a -Keratin
is a member of a superfam ily of proteins referred
to as coiled-coil proteins (Figure 2.38). In these
proteins, two or more a helices can entwine to
form a very stable structure, which can have a
length of 1000 Å (100 nm, or 0.1 m m) or more.
There are approximately 60 members of this
family in humans, including intermediate filaments,
pro teins that contribute to the cell cytoskeleton
(internal scaffolding in a cell), and the muscle
proteins myosin and tropomyosin (Section 35.2).
Members of this family are characterized by a
central region of 300 amino acids that contains
imperfect repeats of a sequence of seven amino
acids called a heptad repeat . The two helices in
a -keratin associate with each other by weak
interactions such as van der Waals forces and
ionic interactions. The left-handed supercoil alters
the two right-handed a helices such that there are
3.5 residues per turn instead of 3.6. Thus, the
pattern of side-chain interactions can be repeated
every seven residues, forming the heptad repeats.
Two helices with such repeats are able to interact
with one another if the repeats are complementary
(Figure 2.39). For example, the repeating residues
may be hydrophobic, allow ing van der Waals
interactions, or have opposite charge, allowing
ionic interac tions. In addition, the two helices may
be linked by disulfide bonds formed by
neighboring cysteine residues. The bonding of the
helices accounts for the physical properties of
wool, an example of an a -keratin. Wool is
extensible and can be stretched to nearly twice its
length because the a helices stretch, breaking
(A)
FIGURE 2.38 An a-helical coiled coil. (A) Space-filling model. (B) Ribbon
diagram. The two helices wind around one another to form a superhelix.
Such structures are found in many proteins, including keratin in hair, quills,
claws, and horns. [Drawn from 1C1G.pdb.]
(B)
Leu
the weak interactions
strand are absent.
between neighboring
Instead, the helix is
helices. However, the
stabilized by steric
covalent disulfide bonds repulsion of the pyrrolidine
resist breakage and return rings of the proline and
Leu
the fiber to its original
hydroxyproline resi dues
state once the stretching (Figure 2.41). The
force is released. The
pyrrolidine rings keep out
number of disulfide bond of each other’s way when
Leu
cross-links further defines the polypeptide chain
the fiber’s properties. Hair assumes its helical form,
and wool, having fewer
which has about three resi
cross-links, are flex
dues per turn. Three
strands wind around one Leu
ible. Horns, claws, and
another to form a
hooves, having more
superhelical cable that is
cross-links, are much
harder. A different type stabilized by hydrogen
bonds between strands.
of helix is present in
The hydrogen
collagen, the most
abundant protein
of mammals. Collagen is Leucine (Leu) residue
the main fibrous
component of skin, bone,
tendon, cartilage, and
teeth. This extracellular
protein is a rod-shaped
molecule, about 3000 Å
long and only 15 Å in
diameter. It contains three Leu
helical polypeptide
chains, each nearly 1000
residues long. Glycine
appears at every third
residue in the amino acid Leu
sequence, and the
sequence glycine-prolinehydroxyproline recurs
frequently (Figure 2.40).
Leu
Hydroxyproline is a
derivative of proline that C C N N
has a hydroxyl group in
place of one of the
hydrogen atoms on the
pyrrolidine ring. The
collagen helix has
properties different from
those of the a helix.
Hydrogen bonds within a
Gly
bonds form between the peptide NH groups of
glycine residues and the CO groups of residues on
the other chains. The hydroxyl groups of hydroxypro
line residues also participate in hydrogen bonding.
ProPro
13
FIGURE 2.39 Heptad repeats in a coiled-coil protein. Every seventh residue in glycine be present at every third position on each
each helix is leucine. The two helices are held together by van der Waals
strand (Figure 2.42A). The only residue that can fit in
interactions
an interior position is glycine . The amino acid
primarily between the leucine residues. [Drawn from 2ZTA.pdb.]
residue on either side of glycine is located on the
outside of the cable, where there is room for the
bulky rings of proline and hydroxyproline residues
(Figure 2.42B).
Pro
-Gly-Pro-Met-Gly-Pro-Ser-Gly-Pro-Arg 22
-Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Ala-Hyp 31
-Gly-Pro-Gln-Gly-Phe-Gln-Gly-Pro-Hyp 40
-Gly-Glu-Hyp-Gly-Glu-Hyp-Gly-Ala-Ser 49
-Gly-Pro-Met-Gly-Pro-Arg-Gly-Pro-Hyp 58
-Gly-Pro-Hyp-Gly-Lys-Asn-Gly-Asp-Asp
ProGly
FIGURE 2.41 Conformation of a single strand of a collagen triple helix.
FIGURE 2.40 Amino acid sequence of a part of a collagen chain. Every
The inside of the triple-stranded helical cable is very third residue is a glycine. Proline and hydroxyproline (Hyp) also are abundant.
crowded and accounts for the requirement that
(A)
(B)
G
G
FIGURE 2.42 Structure of the protein collagen. (A) Space
filling model of collagen. Each strand is shown in a different
G
color. (B) Cross section of a model of collagen. Each strand
is hydrogen bonded to the other two strands. The a-carbon
atom of a glycine residue is identified by the letter G. Every
third residue must be glycine because there is no space in
the center of the helix. Notice that the pyrrolidine rings of the
proline residues are on the outside.
Myoglobin is an extremely compact molecule . Its
CHAPTER 2 Protein Composition and Structure
overall dimensions are 45 3 35 3 25 Å, an order of
The importance of the positioning of glycine inside magnitude less than if it were fully stretched out
the triple helix is illustrated in the disorder
(Figure 2.43). About 70% of the main chain is folded
osteogenesis imperfecta, also known as brittle bone into eight a heli
disease. In this condition, which can vary from mild toces, and much of the rest of the chain forms turns
very
and loops between helices. The folding of the main
severe, other amino acids replace the internal glycinechain of myoglobin, like that of most other pro teins,
residue. This replace ment leads to a delayed and
is complex and devoid of symmetry. The overall
improper folding of collagen. The most serious
course of the poly peptide chain of a protein is
symptom is severe bone fragility. Defective collagen referred to as its tertiary structure . A unifying
in the eyes causes the whites of the eyes to have a principle emerges from the distribution of side chains.
blue tint (blue sclera).
Strikingly, the inte rior consists almost entirely of
nonpolar residues such as leucine, valine,
methionine, and phenylalanine (Figure 2.44).
Charged residues such as aspartate, glutamate,
2.4 Tertiary Structure: Water-Soluble Proteins
Fold into Compact Structures with Nonpolar Cores lysine, and arginine are absent from the inside of
myo globin. The only polar residues inside are two
Let us now examine how amino acids are grouped histidine residues, which play critical roles in binding
iron and oxygen. The outside of myoglobin, on the
together in a complete protein. X-ray
crystallographic and nuclear magnetic resonance
(NMR) studies (Section 3.5) have revealed the
detailed three-dimensional structures of thousands
of proteins. We begin here with an examination of
myoglobin,
the first protein to be seen in atomic detail.
Myoglobin, the oxygen storage protein in muscle, is
a single polypeptide chain of 153 amino acids
(Chapter 7). The capacity of myoglobin to bind
oxygen depends on the presence of heme, a
nonpolypeptide prosthetic (helper) group consisting
of protoporphyrin IX and a central iron atom.
46
45
(B) Heme group
(A)
Heme group
Iron atom
FIGURE 2.43 Three-dimensional structure of myoglobin. (A) A ribbon diagram
shows that the protein consists largely of a helices. (B) A space-filling model in
the same orientation shows how tightly packed the folded protein is. Notice that
the heme group is nestled into a crevice in the compact protein with only an
edge exposed. One helix is blue to allow comparison of the two structural
depictions. [Drawn from 1A6N.pdb.]
model of myoglobin with hydrophobic
amino acids shown in yellow, charged
amino acids shown in blue, and others
shown in white. Notice that the surface of
the molecule has many charged amino
acids, as well as some hydrophobic amino
acids. (B) In this cross-sectional view,
notice that mostly hydrophobic amino acids
are found on the inside of the structure,
whereas the charged amino acids are
found on the protein surface. [Drawn from
1MBD.pdb.]
other hand, consists of both polar and nonpolar residues. The space-filling
model shows that there is very little empty space inside.
This contrasting distribution of polar and nonpolar residues reveals a
key facet of protein architecture. In an aqueous environment, protein
folding is driven by the strong tendency of hydrophobic residues to be
excluded from water. Recall that a system is more thermodynamically
stable when hydrophobic groups are clustered rather than extended
into the aqueous surroundings (p. 9). The polypeptide chain therefore folds
so that its hydrophobic side chains are buried and its polar, charged chains
are on the surface . Many a helices and b strands are amphipathic; that is,
the a helix or b strand has a hydrophobic face, which points into the
protein interior, and a more polar face, which points into solution. The
fate of the main chain accompanying the
“the exceptions that prove the rule” because they
hydrophobic side chains is important, too. An
have the reverse distribution of hydrophobic and
unpaired peptide NH or CO group markedly prefers hydrophilic amino acids. For example, consider
water to a nonpolar milieu. The secret of burying a porins, proteins found in the outer membranes of
segment of main chain in a hydrophobic
many bacteria (Figure 2.45). Membranes are built
environment is to pair all the NH and CO groups by largely of hydropho
hydrogen bonding. This pairing is neatly
bic alkane chains (Section 12.2). Thus, porins are
accomplished in an a helix or b sheet. Van der
cov ered on the outside largely with hydrophobic
Waals interactions between tightly packed hydrocar residues that interact with the neighboring alkane
bon side chains also contribute to the stability of pro chains. In contrast, the center of the protein
contains many charged and polar amino acids that
teins. We can now understand why the set of 20
surround a water-filled chan nel running through the
amino acids contains several that differ subtly in
size and shape. They provide a palette from which middle of the protein. Thus,
FIGURE 2.45 “Inside out” amino acid distribution in porin. The outside of
to choose to fill the interior of a protein neatly and
porin (which contacts hydrophobic groups
thereby maxi mize van der Waals interactions, which
in membranes) is covered largely with hydrophobic residues, whereas the
require inti mate contact.
center includes a water-filled channel lined with charged and polar amino
Some proteins that span biological membranes are acids. [Drawn from 1PRN.pdb.]
hydrophobic environments, they
are “inside out” relative to proteins
that function in aqueous solution.
Water-filled
hydrophilic channel
Largely hydrophobic exterior
because porins function in
47