Tools and Techniques The seventh edition of Biochemistry offers three chapters that present the tools and techniques of biochemistry:"Exploring Proteins and Proteomes"(Chapter 3), "Exploring Genes and Genomes"(Chapter 5),and"Exploring Evolution and Bioinformatics"(Chapter 6).Additional experimental techniques are presented throughout the book,as appropriate. Exploring Proteins and Proteomes (Chapter 3) Mutagenesis techniques(p.156) Protein purification(p.66) Next-generation sequencing(p.160) Differential centrifugation(p.67) Quantitative PCR(p.161) Salting out(p.68) Examining expression levels(DNA microarrays)(p.162) Dialysis (p.69) Introducing genes into eukaryotes(p.163) Gel-filtration chromatography(p.69) Transgenic animals(p.164) Ion-exchange chromatography(p.69) Gene disruption(p.164) Affinity chromatography (p.70) Gene disruption by RNA interference(p.165) High-pressure liquid chromatography(p.71) Tumor-inducing plasmids(p.166) Gel electrophoresis(p.71) Isoelectric focusing(p.73) Exploring Genes (other chapters) Two-dimensional electrophoresis(p.74) Density-gradient equilibrium sedimentation(p.119) Qualitative and quantitative evaluation of protein Chromatin immunoprecipitation(ChIP)(p.945) purification(p.75) Ultracentrifugation(p.76) Edman degradation(p.80) Exploring Evolution and Bioinformatics Protein sequencing(p.82) (Chapter 6) Production of polyclonal antibodies(p.86) Sequence-comparison methods(p.174) Production of monoclonal antibodies (p.86) Sequence-alignment methods(p.176) Enzyme-linked immunoabsorbent assay(ELISA)(p.88) Estimating the statistical significance of alignments Western blotting(p.89) (by shuffling)(p.177) Fluorescence microscopy(p.89) Substitution matrices(p.178) Green fluorescent protein as a marker(p.89) Performing a BLAST database search(p.181) Immunoelectron microscopy(p.91) Sequence templates(p.184) MALDI-TOF mass spectrometry (p.91) Detecting repeated motifs(p.184) Tandem mass spectrometry(p.93) Mapping secondary structures through RNA sequence Proteomic analysis by mass spectrometry(p.94) comparisons(p.186) Automated solid-phase peptide synthesis(p.95) Construction of evolutionary trees(p.187) X-ray crystallography(p.98) Combinatorial chemistry(p.188) Nuclear magnetic resonance spectroscopy(p.101) Molecular evolution in the laboratory (p.189) NOESY spectroscopy(p.102) Other Techniques Exploring Proteins (other chapters) Functional magnetic resonance imaging(fMRI)(p.197) Basis of fluorescence in green fluorescent protein(p.58) Sequencing of carbohydrates by using MALDI-TOF mass Using irreversible inhibitors to map the active site(p.241) spectroscopy(p.336) Enzyme studies with catalytic antibodies(p.243) The use of liposomes to investigate membrane Single-molecule studies(p.246) permeability(p.353) The use of hydropathy plots to locate transmembrane Exploring Genes and Genomes (Chapter 5) helices(p.360) Restriction-enzyme analysis(p.141) Fluorescence recovery after photobleaching(FRAP)for measuring Southern and northern blotting techniques(p.142) lateral diffusion in membranes(p.361) Sanger dideoxy method of DNA sequencing(p.143) Patch-clamp technique for measuring channel activity(p.383) Solid-phase synthesis of nucleic acids(p.144) Measurement of redox potential(p.528) Polymerase chain reaction(PCR)(p.145) Recombinant DNA technology(p.148) kAnimated Techniques DNA cloning in bacteria(p.149) Animated explanations of experimental techniques used for exploring Creating cDNA libraries(p.154) genes and proteins are available at www.whfreeman.com/berg7e. xii
xii Exploring Proteins and Proteomes (Chapter 3) Protein purification (p. 66) Differential centrifugation (p. 67) Salting out (p. 68) Dialysis (p. 69) Gel-filtration chromatography (p. 69) Ion-exchange chromatography (p. 69) Affinity chromatography (p. 70) High-pressure liquid chromatography (p. 71) Gel electrophoresis (p. 71) Isoelectric focusing (p. 73) Two-dimensional electrophoresis (p. 74) Qualitative and quantitative evaluation of protein purification (p. 75) Ultracentrifugation (p. 76) Edman degradation (p. 80) Protein sequencing (p. 82) Production of polyclonal antibodies (p. 86) Production of monoclonal antibodies (p. 86) Enzyme-linked immunoabsorbent assay (ELISA) (p. 88) Western blotting (p. 89) Fluorescence microscopy (p. 89) Green fluorescent protein as a marker (p. 89) Immunoelectron microscopy (p. 91) MALDI-TOF mass spectrometry (p. 91) Tandem mass spectrometry (p. 93) Proteomic analysis by mass spectrometry (p. 94) Automated solid-phase peptide synthesis (p. 95) X-ray crystallography (p. 98) Nuclear magnetic resonance spectroscopy (p. 101) NOESY spectroscopy (p. 102) Exploring Proteins (other chapters) Basis of fluorescence in green fluorescent protein (p. 58) Using irreversible inhibitors to map the active site (p. 241) Enzyme studies with catalytic antibodies (p. 243) Single-molecule studies (p. 246) Exploring Genes and Genomes (Chapter 5) Restriction-enzyme analysis (p. 141) Southern and northern blotting techniques (p. 142) Sanger dideoxy method of DNA sequencing (p. 143) Solid-phase synthesis of nucleic acids (p. 144) Polymerase chain reaction (PCR) (p. 145) Recombinant DNA technology (p. 148) DNA cloning in bacteria (p. 149) Creating cDNA libraries (p. 154) Mutagenesis techniques (p. 156) Next-generation sequencing (p. 160) Quantitative PCR (p. 161) Examining expression levels (DNA microarrays) (p. 162) Introducing genes into eukaryotes (p. 163) Transgenic animals (p. 164) Gene disruption (p. 164) Gene disruption by RNA interference (p. 165) Tumor-inducing plasmids (p. 166) Exploring Genes (other chapters) Density-gradient equilibrium sedimentation (p. 119) Chromatin immunoprecipitation (ChIP) (p. 945) Exploring Evolution and Bioinformatics (Chapter 6) Sequence-comparison methods (p. 174) Sequence-alignment methods (p. 176) Estimating the statistical significance of alignments (by shuffling) (p. 177) Substitution matrices (p. 178) Performing a BLAST database search (p. 181) Sequence templates (p. 184) Detecting repeated motifs (p. 184) Mapping secondary structures through RNA sequence comparisons (p. 186) Construction of evolutionary trees (p. 187) Combinatorial chemistry (p. 188) Molecular evolution in the laboratory (p. 189) Other Techniques Functional magnetic resonance imaging (fMRI) (p. 197) Sequencing of carbohydrates by using MALDI-TOF mass spectroscopy (p. 336) The use of liposomes to investigate membrane permeability (p. 353) The use of hydropathy plots to locate transmembrane helices (p. 360) Fluorescence recovery after photobleaching (FRAP) for measuring lateral diffusion in membranes (p. 361) Patch-clamp technique for measuring channel activity (p. 383) Measurement of redox potential (p. 528) Animated Techniques Animated explanations of experimental techniques used for exploring genes and proteins are available at www.whfreeman.com/berg7e. The seventh edition of Biochemistry offers three chapters that present the tools and techniques of biochemistry: “Exploring Proteins and Proteomes” (Chapter 3), “Exploring Genes and Genomes” (Chapter 5), and “Exploring Evolution and Bioinformatics” (Chapter 6). Additional experimental techniques are presented throughout the book, as appropriate. Tools and Techniques
Acknowledgments Thanks go first and foremost to our students.Not We thank Susan J.Baserga and Erica A.Champion a word was written or an illustration constructed of the Yale University School of Medicine for their without the knowledge that bright,engaged students outstanding contributions in the sixth edition's revi- would immediately detect vagueness and ambiguity. sion of Chapter 29.We also especially thank those We also thank our colleagues who supported,advised, who served as reviewers for this new edition.Their instructed,and simply bore with us during this arduous thoughtful comments,suggestions,and encourage- task.We are also grateful to our colleagues through- ment have been of immense help to us in maintain- out the world who patiently answered our questions ing the excellence of the preceding editions.These and shared their insights into recent developments. reviewers are: Fareed Aboul-Ela Paul Hager M.Kazem Mostafapour Louisiana State University East Carolina University University of Michigan,Dearborn Paul Adams Frans Huijing Duarte Mota de Freitas University of Arkansas,Fayetteville University of Miami Loyola University of Chicago Kevin Ahern Nitin Jain Stephen Munroe Oregon State University University of Tennessee Marquette University Edward Behrman Gerwald Jogl Xiaping Pan Ohio State University Brown University East Carolina University Donald Beitz Kelly Johanson Scott Pattison lowa State University Xavier University of Louisiana Ball State University Sanford Bernstein Todd Johnson Stefan Paula San Diego State University Weber State University Northern Kentucky University Martin Brock Michael Kalafatis David Pendergrass Easter Kentucky University Cleveland State University University of Kansas W.Malcom Byrnes Mark Kearly Reuben Peters Howard University College of Medicine Florida State University lowa State University C.Britt Carlson Sung-Kun Kim Wendy Pogozelski Brookdale Community College Baylor University State University of New York,Geneseo Geraldine Prody Graham Carpenter Roger Koeppe Vanderbilt University University of Arkansas,Fayetteville Western Washington University Jun Chung Dmitry Kolpashchikov Greg Raner Louisiana State University University of Central Florida University of North Carolina,Greensboro Michael Cusanovich Joshua Rausch John Koontz Elmhurst College University of Arizona University of Tennessee David Daleke Glen Legge Tanea Reed Eastern Kentucky University Indiana University University of Houston, Lori Robins Margaret Daugherty University Park California Polytechnic University,San Luis Colorado College John Stephen Lodmell Obispo Dan Davis University of Montana Douglas Root University of Arkansas,Fayetteville Timothy Logan University of North Texas Mary Farwell Florida State University Theresa Salerno East Carolina University Michael Massiah Minnesota State University,Mankato Brent Feske Oklahoma State University Scott Samuels Armstrong Atlantic University Diana McGill University of Montana,Missoula Wilson Francisco Northern Kentucky University Benjamin Sandler Arizona State University Michael Mendenhall Oklahoma State University Masaya Fujita University of Kentucky Joel Schildbach University of Houston,University Park David Merkler Johns Hopkins University Peter Gegenheimer University of South Florida Hua Shi University of Kansas Gary Merrill State University of New York,University John Goers Oregon State University at Albany California Polytechnic University,San Debra Moriarity Kerry Smith Luis Obispo University of Alabama,Huntsville Clemson University Neena Grover Patricia Moroney Robert Stach Colorado College Louisiana State University University of Michigan,Flint xiii
xiii Thanks go first and foremost to our students. Not a word was written or an illustration constructed without the knowledge that bright, engaged students would immediately detect vagueness and ambiguity. We also thank our colleagues who supported, advised, instructed, and simply bore with us during this arduous task. We are also grateful to our colleagues throughout the world who patiently answered our questions and shared their insights into recent developments. We thank Susan J. Baserga and Erica A. Champion of the Yale University School of Medicine for their outstanding contributions in the sixth edition’s revision of Chapter 29. We also especially thank those who served as reviewers 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: Acknowledgments Fareed Aboul-Ela Louisiana State University Paul Adams University of Arkansas, Fayetteville Kevin Ahern Oregon State University Edward Behrman Ohio State University Donald Beitz Iowa State University Sanford Bernstein San Diego State University Martin Brock Eastern Kentucky University W. Malcom Byrnes Howard University College of Medicine C. Britt Carlson Brookdale Community College Graham Carpenter Vanderbilt University Jun Chung Louisiana State University Michael Cusanovich University of Arizona David Daleke Indiana University Margaret Daugherty Colorado College Dan Davis University of Arkansas, Fayetteville Mary Farwell East Carolina University Brent Feske Armstrong Atlantic University Wilson Francisco Arizona State University Masaya Fujita University of Houston, University Park Peter Gegenheimer University of Kansas John Goers California Polytechnic University, San Luis Obispo Neena Grover Colorado College Paul Hager East Carolina University Frans Huijing University of Miami Nitin Jain University of Tennessee Gerwald Jogl Brown University Kelly Johanson Xavier University of Louisiana Todd Johnson Weber State University Michael Kalafatis Cleveland State University Mark Kearly Florida State University Sung-Kun Kim Baylor University Roger Koeppe University of Arkansas, Fayetteville Dmitry Kolpashchikov University of Central Florida John Koontz University of Tennessee Glen Legge University of Houston, University Park John Stephen Lodmell University of Montana Timothy Logan Florida State University Michael Massiah Oklahoma State University Diana McGill Northern Kentucky University Michael Mendenhall University of Kentucky David Merkler University of South Florida Gary Merrill Oregon State University Debra Moriarity University of Alabama, Huntsville Patricia Moroney Louisiana State University M. Kazem Mostafapour University of Michigan, Dearborn Duarte Mota de Freitas Loyola University of Chicago Stephen Munroe Marquette University Xiaping Pan East Carolina University Scott Pattison Ball State University Stefan Paula Northern Kentucky University David Pendergrass University of Kansas Reuben Peters Iowa State University Wendy Pogozelski State University of New York, Geneseo Geraldine Prody Western Washington University Greg Raner University of North Carolina, Greensboro Joshua Rausch Elmhurst College Tanea Reed Eastern Kentucky University Lori Robins California Polytechnic University, San Luis Obispo Douglas Root University of North Texas Theresa Salerno Minnesota State University, Mankato Scott Samuels University of Montana, Missoula Benjamin Sandler Oklahoma State University Joel Schildbach Johns Hopkins University Hua Shi State University of New York, University at Albany Kerry Smith Clemson University Robert Stach University of Michigan, Flint
Scott Stagg Liang Tong Xuemin Wang Florida State University Columbia University University of Missouri,St.Louis Wesley Stites Kenneth Traxler Kevin Williams University of Arkansas,Fayetteville Bemidji State University Western Kentucky University Paul Straight Peter Van Der Geer Warren Williams Texas A&M University San Diego State University University of British Columbia Gerald Stubbs Nagarajan Vasumathi Shiyong Wu Vanderbilt University Jacksonville State University Ohio University Takita Felder Sumter Stefan Vetter Laura Zapanta Winthrop University Florida Atlantic University University of Pittsburgh Jeremy Thorner Edward Walker University of California,Berkeley Weber State University Three of us have had the pleasure of working with the Coordinator,deftly directed the rendering of new illustra- folks at W.H.Freeman and Company on a number of tions.Paul Rohloff,Production Coordinator,made sure projects,whereas one of us is new to the Freeman fam- that the significant difficulties of scheduling,composi- ily.Our experiences have always been delightful and tion,and manufacturing were smoothly overcome. rewarding.Writing and producing the seventh edition Andrea Gawrylewski,Patrick Shriner,Marni Rolfes, of Biochemistry was no exception.The Freeman team and Rohit Phillip did a wonderful job in their manage- has a knack for undertaking stressful,but exhilarating, ment of the media program.Amanda Dunning ably projects and reducing the stress without reducing the coordinated the print supplemants plan.Special thanks exhilaration and a remarkable ability to coax without also to editorial assistant Anna Bristow.Debbie Clare, ever nagging.We have many people to thank for this Associate Director of Marketing,enthusiastically experience.First,we would like to acknowledge the introduced this newest edition of Biochemistry to the encouragement,patience,excellent advice,and good academic world.We are deeply appreciative of the sales humor of Kate Ahr Parker,Publisher.Her enthusi- staff for their enthusiastic support.Without them,all of asm is source of energy for all of us.Lisa Samols is our our excitement and enthusiasm would ultimately come wonderful developmental editor.Her insight,patience, to naught.Finally,we owe a deep debt of gratitude to and understanding contributed immensely to the suc- Elizabeth Widdicombe,President of W.H.Freeman cess of this project.Beth Howe and Erica Champion and Company.Her vision for science textbooks and assisted Lisa by developing several chapters,and we her skill at gathering exceptional personnel make are grateful to them for their help.Georgia Lee Hadler, working with W.H.Freeman and Company a true Senior Project Editor,managed the flow of the entire pleasure. project,from copyediting through bound book,with Thanks also to our many colleagues at our own insti- her usual admirable efficiency.Patricia Zimmerman tutions as well as throughout the country who patiently and Nancy Brooks,our manuscript editors,enhanced answered our questions and encouraged us on our quest. the literary consistency and clarity of the text.Vicki Finally,we owe a debt of gratitude to our families- Tomaselli,Design Manager,produced a design and our wives,Wendie Berg,Alison Unger,and Megan layout that makes the book exciting and eye-catching Williams,and our children,Alex,Corey,and Monica Berg, while maintaining the link to past editions.Photo Janina and Nicholas Tymoczko,and Timothy and Mark Editor Christine Beuse and Photo Researcher Jacalyn Gatto.Without their support,comfort,and understand- Wong found the photographs that we hope make ing,this endeavor could never have been undertaken, the text more inviting.Janice Donnola,Illustration let alone successfully completed. xiv
xiv Scott Stagg Florida State University Wesley Stites University of Arkansas, Fayetteville Paul Straight Texas A&M University Gerald Stubbs Vanderbilt University Takita Felder Sumter Winthrop University Jeremy Thorner University of California, Berkeley Liang Tong Columbia University Kenneth Traxler Bemidji State University Peter Van Der Geer San Diego State University Nagarajan Vasumathi Jacksonville State University Stefan Vetter Florida Atlantic University Edward Walker Weber State University Xuemin Wang University of Missouri, St. Louis Kevin Williams Western Kentucky University Warren Williams University of British Columbia Shiyong Wu Ohio University Laura Zapanta University of Pittsburgh Three of us have had the pleasure of working with the folks at W. H. Freeman and Company on a number of projects, whereas one of us is new to the Freeman family. Our experiences have always been delightful and rewarding. Writing and producing the seventh edition of Biochemistry was no exception. The Freeman team has a knack for undertaking stressful, but exhilarating, projects and reducing the stress without reducing the exhilaration and a remarkable ability to coax without ever nagging. We have many people to thank for this experience. First, we would like to acknowledge the encouragement, patience, excellent advice, and good humor of Kate Ahr Parker, Publisher. Her enthusiasm is source of energy for all of us. Lisa Samols is our wonderful developmental editor. Her insight, patience, and understanding contributed immensely to the success of this project. Beth Howe and Erica Champion assisted Lisa by developing several chapters, and we are grateful to them for their help. Georgia Lee Hadler, Senior Project Editor, managed the flow of the entire project, from copyediting through bound book, with her usual admirable efficiency. Patricia Zimmerman and Nancy Brooks, our manuscript editors, enhanced the literary consistency and clarity of the text. Vicki Tomaselli, Design Manager, produced a design and layout that makes the book exciting and eye-catching while maintaining the link to past editions. Photo Editor Christine Beuse and Photo Researcher Jacalyn Wong found the photographs that we hope make the text more inviting. Janice Donnola, Illustration Coordinator, deftly directed the rendering of new illustrations. Paul Rohloff, Production Coordinator, made sure that the significant difficulties of scheduling, composition, and manufacturing were smoothly overcome. Andrea Gawrylewski, Patrick Shriner, Marni Rolfes, and Rohit Phillip did a wonderful job in their management of the media program. Amanda Dunning ably coordinated the print supplemants plan. Special thanks also to editorial assistant Anna Bristow. Debbie Clare, Associate Director of Marketing, enthusiastically introduced this newest edition of Biochemistry to the academic world. We are deeply appreciative of the sales staff for their enthusiastic support. Without them, all of our excitement and enthusiasm would ultimately come to naught. Finally, we owe a deep debt of gratitude to Elizabeth Widdicombe, President of W. H. Freeman and Company. Her vision for science textbooks and her skill at gathering exceptional personnel make working with W. H. Freeman and Company a true pleasure. Thanks also to our many colleagues at our own institutions as well as throughout the country who patiently answered our questions and encouraged us on our quest. Finally, we owe a debt of gratitude to our families— our wives, Wendie Berg, Alison Unger, and Megan Williams, and our children, Alex, Corey, and Monica Berg, Janina and Nicholas Tymoczko, and Timothy and Mark Gatto. Without their support, comfort, and understanding, this endeavor could never have been undertaken, let alone successfully completed
BRIEF CONTENTS CONTENTS Part I THE MOLECULAR DESIGN OF LIFE Preface 1 Biochemistry:An Evolving Science 1 2 Protein Composition and Structure 25 Part I THE MOLECULAR DESIGN OF LIFE 3 Exploring Proteins and Proteomes 65 4 DNA,RNA,and the Flow of Genetic Information 109 Chapter 1 Biochemistry:An Evolving Science 5 Exploring Genes and Genomes 139 1.1 Biochemical Unity Underlies 6 Exploring Evolution and Bioinformatics 173 Biological Diversity 7 Hemoglobin:Portrait of a Protein in Action 195 1.2 DNA Illustrates the Interplay Between 8 Enzymes:Basic Concepts and Kinetics 219 Form and Function 9 Catalytic Strategies 253 DNA is constructed from four building blocks 10 Regulatory Strategies 289 Two single strands of DNA combine to form a 11 Carbohydrates 319 double helix 12 Lipids and Cell Membranes 345 DNA structure explains heredity and the storage of information 13 Membrane Channels and Pumps 371 1.3 Concepts from Chemistry Explain the 14 Signal-Transduction Pathways 401 Properties of Biological Molecules The double helix can form from its component strands 6 Part II TRANSDUCING AND STORING ENERGY Covalent and noncovalent bonds are important for the 15 Metabolism:Basic Concepts and Design 427 structure and stability of biological molecules 16 Glycolysis and Gluconeogenesis 453 The double helix is an expression of the rules 17 The Citric Acid Cycle 497 of chemistry 10 18 Oxidative Phosphorylation 525 The laws of thermodynamics govern the behavior of biochemical systems 11 19 The Light Reactions of Photosynthesis 565 Heat is released in the formation of the double helix 12 20 The Calvin Cycle and the Pentose Phosphate Pathway 589 Acid-base reactions are central in many biochemical processes 13 21 Glycogen Metabolism 615 Acid-base reactions can disrupt the double helix 14 22 Fatty Acid Metabolism 639 Buffers regulate pH in organisms and in the laboratory 15 23 Protein Turnover and Amino Acid Catabolism 673 1.4 The Genomic Revolution Is Transforming Biochemistry and Medicine Part IlI SYNTHESIZING THE MOLECULES OF LIFE The sequencing of the human genome is a landmark 24 The Biosynthesis of Amino Acids 705 in human history 17 25 Nucleotide Biosynthesis 735 Genome sequences encode proteins and patterns of expression 18 26 The Biosynthesis of Membrane Lipids and Steroids 759 Individuality depends on the interplay between genes and environment 19 27 The Integration of Metabolism 791 APPENDIX:Visualizing Molecular Structures I: 28 DNA Replication,Repair,and Recombination 819 Small Molecules 21 29 RNA Synthesis and Processing 851 30 Protein Synthesis 887 31 The Control of Gene Expression in Prokaryotes 921 Chapter 2 Protein Composition and Structure 25 32 The Control of Gene Expression in Eukaryotes 937 2.1 Proteins Are Built from a Repertoire of 20 Amino Acids 27 Part IV RESPONDING TO ENVIRONMENTAL CHANGES 2.2 Primary Structure:Amino Acids Are Linked by 33 Sensory Systems 957 Peptide Bonds to Form Polypeptide Chains 33 34 The Immune System 977 Proteins have unique amino acid sequences specified 35 Molecular Motors 1007 by genes 35 36 Drug Development 1029 Polypeptide chains are flexible yet conformationally restricted 36
Part I THE MOLECULAR DESIGN OF LIFE 1 Biochemistry: An Evolving Science 1 2 Protein Composition and Structure 25 3 Exploring Proteins and Proteomes 65 4 DNA, RNA, and the Flow of Genetic Information 109 5 Exploring Genes and Genomes 139 6 Exploring Evolution and Bioinformatics 173 7 Hemoglobin: Portrait of a Protein in Action 195 8 Enzymes: Basic Concepts and Kinetics 219 9 Catalytic Strategies 253 10 Regulatory Strategies 289 11 Carbohydrates 319 12 Lipids and Cell Membranes 345 13 Membrane Channels and Pumps 371 14 Signal-Transduction Pathways 401 Part II TRANSDUCING AND STORING ENERGY 15 Metabolism: Basic Concepts and Design 427 16 Glycolysis and Gluconeogenesis 453 17 The Citric Acid Cycle 497 18 Oxidative Phosphorylation 525 19 The Light Reactions of Photosynthesis 565 20 The Calvin Cycle and the Pentose Phosphate Pathway 589 21 Glycogen Metabolism 615 22 Fatty Acid Metabolism 639 23 Protein Turnover and Amino Acid Catabolism 673 Part III SYNTHESIZING THE MOLECULES OF LIFE 24 The Biosynthesis of Amino Acids 705 25 Nucleotide Biosynthesis 735 26 The Biosynthesis of Membrane Lipids and Steroids 759 27 The Integration of Metabolism 791 28 DNA Replication, Repair, and Recombination 819 29 RNA Synthesis and Processing 851 30 Protein Synthesis 887 31 The Control of Gene Expression in Prokaryotes 921 32 The Control of Gene Expression in Eukaryotes 937 Part IV RESPONDING TO ENVIRONMENTAL CHANGES 33 Sensory Systems 957 34 The Immune System 977 35 Molecular Motors 1007 36 Drug Development 1029 Preface v Part I THE MOLECULAR DESIGN OF LIFE Chapter 1 Biochemistry: An Evolving Science 1 1.1 Biochemical Unity Underlies Biological Diversity 1 1.2 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 double helix can form from its component strands 6 Covalent and noncovalent bonds are important for the structure and stability of biological molecules 7 The double helix is an expression of the rules of chemistry 10 The laws of thermodynamics govern the behavior of biochemical systems 11 Heat is released in the formation of the double helix 12 Acid–base reactions are central in many biochemical processes 13 Acid–base reactions can disrupt the double helix 14 Buffers regulate pH in organisms and in the laboratory 15 1.4 The Genomic Revolution Is Transforming Biochemistry and Medicine 17 The sequencing of the human genome is a landmark in human history 17 Genome sequences encode proteins and patterns of expression 18 Individuality depends on the interplay between genes and environment 19 APPENDIX: Visualizing Molecular Structures I: Small Molecules 21 Chapter 2 Protein Composition and Structure 25 2.1 Proteins Are Built from a Repertoire of 20 Amino Acids 27 2.2 Primary Structure: Amino Acids Are Linked by Peptide Bonds to Form Polypeptide Chains 33 Proteins have unique amino acid sequences specified by genes 35 Polypeptide chains are flexible yet conformationally restricted 36 BRIEF CONTENTS CONTENTS
xvi Contents 2.3 Secondary Structure:Polypeptide Chains Can 3.2 Amino Acid Sequences of Proteins Can Fold into Regular Structures Such As the Alpha Be Determined Experimentally 79 Helix,the Beta Sheet,and Turns and Loops 38 Peptide sequences can be determined by automated The alpha helix is a coiled structure stabilized Edman degradation 80 by intrachain hydrogen bonds 38 Proteins can be specifically cleaved into small Beta sheets are stabilized by hydrogen bonding between peptides to facilitate analysis 82 polypeptide strands 40 Genomic and proteomic methods are complementary 84 Polypeptide chains can change direction by 3.3 Immunology Provides Important Techniques making reverse turns and loops 42 with Which to Investigate Proteins 84 Fibrous proteins provide structural support for 84 cells and tissues 43 Antibodies to specific proteins can be generated Monoclonal antibodies with virtually any 2.4 Tertiary Structure:Water-Soluble Proteins desired specificity can be readily prepared 86 Fold into Compact Structures with Proteins can be detected and quantified by using an Nonpolar Cores 45 enzyme-linked immunosorbent assay 88 2.5 Quaternary Structure:Polypeptide Chains Western blotting permits the detection of Can Assemble into Multisubunit Structures 48 proteins separated by gel electrophoresis 89 2.6 The Amino Acid Sequence of a Protein Fluorescent markers make the visualization of Determines Its Three-Dimensional Structure 49 proteins in the cell possible 90 Amino acids have different propensities for 3.4 Mass Spectrometry Is a Powerful Technique forming alpha helices,beta sheets,and beta turns 50 for the ldentification of Peptides and Proteins 91 Protein folding is a highly cooperative process 52 The mass of a protein can be precisely determined Proteins fold by progressive stabilization of by mass spectrometry 91 intermediates rather than by random search 52 Peptides can be sequenced by mass spectrometry 93 Prediction of three-dimensional structure from Individual proteins can be identified by sequence remains a great challenge 54 mass spectrometry 94 Some proteins are inherently unstructured and 3.5 Peptides Can Be Synthesized by can exist in multiple conformations 54 Automated Solid-Phase Methods 95 Protein misfolding and aggregation are associated 3.6 Three-Dimensional Protein Structure with some neurological diseases 55 Can Be Determined by X-ray Crystallography Protein modification and cleavage confer and NMR Spectroscopy 98 new capabilities 57 APPENDIX:Visualizing Molecular Structures II:Proteins 60 X-ray crystallography reveals three-dimensional structure in atomic detail 98 Nuclear magnetic resonance spectroscopy can reveal the structures of proteins in solution 101 Chapter 3 Exploring Proteins and Proteomes 65 The proteome is the functional representation of Chapter 4 DNA,RNA,and the Flow of the genome 66 Information 109 3.1 The Purification of Proteins Is an Essential 4.1 A Nucleic Acid Consists of Four Kinds of First Step in Understanding Their Function 66 Bases Linked to a Sugar-Phosphate Backbone 110 The assay:How do we recognize the protein that we are looking for? 67 RNA and DNA differ in the sugar component and one of the bases 110 Proteins must be released from the cell to be purified 67 Nucleotides are the monomeric units of nucleic acids 111 Proteins can be purified according to solubility,size, 113 charge,and binding affinity 68 DNA molecules are very long Proteins can be separated by gel electrophoresis and 4.2 A Pair of Nucleic Acid Chains with displayed 71 Complementary Sequences Can Form a A protein purification scheme can be quantitatively Double-Helical Structure 113 evaluated 75 The double helix is stabilized by hydrogen bonds and Ultracentrifugation is valuable for separating van der Waals interactions 113 biomolecules and determining their masses 76 DNA can assume a variety of structural forms 115 Protein purification can be made easier with the use Z-DNA is a left-handed double helix in which of recombinant DNA technology 78 backbone phosphates zigzag 116
xvi Contents 2.3 Secondary Structure: Polypeptide Chains Can Fold into Regular Structures Such As the Alpha Helix, the Beta Sheet, and Turns and Loops 38 The alpha helix is a coiled structure stabilized by intrachain hydrogen bonds 38 Beta sheets are stabilized by hydrogen bonding between polypeptide strands 40 Polypeptide chains can change direction by making reverse turns and loops 42 Fibrous proteins provide structural support for cells and tissues 43 2.4 Tertiary Structure: Water-Soluble Proteins Fold into Compact Structures with Nonpolar Cores 45 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 alpha helices, beta sheets, and beta turns 50 Protein folding is a highly cooperative process 52 Proteins fold by progressive stabilization of intermediates rather than by random search 52 Prediction of three-dimensional structure from sequence remains a great challenge 54 Some proteins are inherently unstructured and can exist in multiple conformations 54 Protein misfolding and aggregation are associated with some neurological diseases 55 Protein modification and cleavage confer new capabilities 57 APPENDIX: Visualizing Molecular Structures II: Proteins 60 Chapter 3 Exploring Proteins and Proteomes 65 The proteome is the functional representation of the genome 66 3.1 The Purification of Proteins Is an Essential First Step in Understanding Their Function 66 The assay: How do we recognize the protein that we are looking for? 67 Proteins must be released from the cell to 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 Amino Acid Sequences of Proteins Can Be Determined Experimentally 79 Peptide sequences can be determined by automated Edman degradation 80 Proteins can be specifically cleaved into small peptides to facilitate analysis 82 Genomic and proteomic methods are complementary 84 3.3 Immunology Provides Important Techniques with Which to Investigate Proteins 84 Antibodies to specific proteins can be generated 84 Monoclonal antibodies with virtually any desired specificity can be readily prepared 86 Proteins can be detected and quantified by using an enzyme-linked immunosorbent assay 88 Western blotting permits the detection of proteins separated by gel electrophoresis 89 Fluorescent markers make the visualization of proteins in the cell possible 90 3.4 Mass Spectrometry Is a Powerful Technique for the Identification of Peptides and Proteins 91 The mass of a protein can be precisely determined by mass spectrometry 91 Peptides can be sequenced by mass spectrometry 93 Individual proteins can be identified by mass spectrometry 94 3.5 Peptides Can Be Synthesized by Automated Solid-Phase Methods 95 3.6 Three-Dimensional Protein Structure Can Be Determined by X-ray Crystallography and NMR Spectroscopy 98 X-ray crystallography reveals three-dimensional structure in atomic detail 98 Nuclear magnetic resonance spectroscopy can reveal the structures of proteins in solution 101 Chapter 4 DNA, RNA, and the Flow of Information 109 4.1 A Nucleic Acid Consists of Four Kinds of Bases Linked to a Sugar–Phosphate Backbone 110 RNA and DNA differ in the sugar component and one of the bases 110 Nucleotides are the monomeric units of nucleic acids 111 DNA molecules are very long 113 4.2 A Pair of Nucleic Acid Chains with Complementary Sequences Can Form a Double-Helical Structure 113 The double helix is stabilized by hydrogen bonds and van der Waals interactions 113 DNA can assume a variety of structural forms 115 Z-DNA is a left-handed double helix in which backbone phosphates zigzag 116