《生物化学原理》英文版（一）：1

Lehninger Principles of Biochemistry Fourth Edition David L. Nelson (U. of Wisconsin–Madison) Michael M. Cox (U. of Wisconsin–Madison) 1. The Foundations of Biochemistry 1.1 Cellular Foundations 1.2 Chemical Foundations 1.3 Physical Foundations 1.4 Genetic Foundations 1.5 Evolutionary Foundations Distilled and reorganized from Chapters 1–3 of the previous edition, this overview provides a refresher on the cellular, chemical, physical, genetic, and evolutionary background to biochemistry, while orienting students toward what is unique about biochemistry. PART I. STRUCTURE AND CATALYSIS 2. Water 2.1 Weak Interactions in Aqueous Systems 2.2 Ionization of Water, Weak Acids, and Weak Bases 2.3 Buffering against pH Changes in Biological Systems 2.4 Water as a Reactant 2.5 The Fitness of the Aqueous Environment for Living Organisms Includes new coverage of the concept of protein-bound water, illustrated with molecular graphics. 3. Amino Acids, Peptides, and Proteins 3.1 Amino Acids 3.2 Peptides and Proteins 3.3 Working with Proteins 3.4 The Covalent Structure of Proteins 3.5 Protein Sequences and Evolution Adds important new material on genomics and proteomics and their implications for the study of protein structure, function, and evolution. 4. The Three-Dimensional Structure of Proteins 4.1 Overview of Protein Structure 4.2 Protein Secondary Structure 4.3 Protein Tertiary and Quaternary Structures 4.4 Protein Denaturation and Folding Adds a new box on scurvy. 5. Protein Function 5.1 Reversible Binding of a Protein to a Ligand: Oxygen-Binding Proteins 5.2 Complementary Interactions between Proteins and Ligands: The Immune System and Immunoglobulins 5.3 Protein Interactions Modulated by Chemical Energy: Actin, Myosin, and Molecular Motors Adds a new box on carbon monoxide poisoning 6. Enzymes 6.1 An Introduction to Enzymes 6.2 How Enzymes Work 6.3 Enzyme Kinetics as An Approach to Understanding Mechanism 6.4 Examples of Enzymatic Reactions 6.5 Regulatory Enzymes Offers a revised presentation of the mechanism of chymotrypsin (the first reaction mechanism in the book), featuring a two-page figure that takes students through this particular mechanism, while serving as a step-by-step guide to interpreting any reaction mechanism Features new coverage of the mechanism for lysozyme including the controversial aspects of the mechanism and currently favored resolution based on work published in 2001. 7. Carbohydrates and Glycobiology 7.1 Monosaccharides and Disaccharides 7.2 Polysaccharides 7.3 Glycoconjugates: Proteoglycans, Glycoproteins, and Glycolipids 7.4 Carbohydrates as Informational Molecules: The Sugar Code 7.5 Working with Carbohydrates Includes new section on polysaccharide conformations. A striking new discussion of the "sugar code" looks at polysaccharides as informational molecules, with detailed discussions of lectins, selectins, and oligosaccharide-bearing hormones. Features new material on structural heteropolysaccharides and proteoglycans Covers recent techniques for carbohydrate analysis. 8. Nucleotides and Nucleic Acids 8.1 Some Basics 8.2 Nucleic Acid Structure 8.3 Nucleic Acid Chemistry 8.4 Other Functions of Nucleotides 9. DNA-Based Information Technologies 9.1 DNA Cloning: The Basics 9.2 From Genes to Genomes 9.3 From Genomes to Proteomes 9.4 Genome Alterations and New Products of Biotechnology Introduces the human genome. Biochemical insights derived from the human genome are integrated throughout the text. Tracking the emergence of genomics and proteomics, this chapter establishes DNA technology as a core topic and a path to understanding metabolism, signaling, and other topics covered in the middle chapters of this edition. Includes up-to-date coverage of microarrays, protein chips, comparative genomics, and techniques in cloning and analysis. 10. Lipids 10.1 Storage Lipids 10.2 Structural Lipids in Membranes 10.3 Lipids as Signals, Cofactors, and Pigments 10.4 Working with Lipids Integrates new topics specific to chloroplasts and archaebacteria Adds material on lipids as signal molecules. 11. Biological Membranes and Transport 11.1 The Composition and Architecture of Membranes 11.2 Membrane Dynamics 11.3 Solute Transport across Membranes Includes a description of membrane rafts and microdomains within membranes, and a new box on the use of atomic force microscopy to visualize them. Looks at the role of caveolins in the formation of membrane caveolae Covers the investigation of hop diffusion of membrane lipids using FRAP (fluorescence recovery after photobleaching) Adds new details to the discussion of the mechanism of Ca 2 - ATPase (SERCA pump), revealed by the recently available high-resolution view of its structure Explores new facets of the mechanisms of the K+ selectivity filter, brought to light by recent high-resolution structures of the K+ channel Illuminates the structure, role, and mechanism of aquaporins with important new details Describes ABC transporters, with particular attention to the multidrug transporter (MDR1) Includes the newly solved structure of the lactose transporter of E. coli. 12. Biosignaling 12.1 Molecular Mechanisms of Signal Transduction 12.2 Gated Ion Channels 12.3 Receptor Enzymes 12.4 G Protein-Coupled Receptors and Second Messengers 12.5 Multivalent Scaffold Proteins and Membrane Rafts 12.6 Signaling in Microorganisms and Plants 12.7 Sensory Transduction in Vision, Olfaction, and Gustation 12.8 Regulation of Transcription by Steroid Hormones 12.9 Regulation of the Cell Cycle by Protein Kinases 12.10 Oncogenes, Tumor Suppressor Genes, and Programmed Cell Death Updates the previous edition's groundbreaking chapter to chart the continuing rapid development of signaling research Includes discussion on general mechanisms for activation of protein kinases in cascades Now covers the roles of membrane rafts and caveolae in signaling pathways, including the activities of AKAPs (A Kinase Anchoring Proteins) and other scaffold proteins Examines the nature and conservation of families of multivalent protein binding modules, which combine to create many discrete signaling pathways Adds a new discussion of signaling in plants and bacteria, with comparison to mammalian signaling pathways Features a new box on visualizing biochemistry with fluorescence resonance energy transfer (FRET) with green fluorescent protein (GFP) PART II: BIOENERGETICS AND METABOLISM 13. Principles of Bioenergetics 13.1 Bioenergetics and Thermodynamics 13.2 Phosphoryl Group Transfers and ATP 13.3 Biological Oxidation-Reduction Reactions Examines the increasing awareness of the multiple roles of polyphosphate Adds a new discussion of niacin deficiency and pellagra. 14. Glycolysis, Gluconeogenesis, and the Pentose Phosphate Pathway 14.1 Glycolysis 14.2 Feeder Pathways for Glycolysis 14.3 Fates of Pyruvate under Anaerobic Conditions: Fermentation 14.4 Gluconeogenesis 14.5 Pentose Phosphate Pathway of Glucose Oxidation Now covers gluconeogenesis immediately after glycolysis, discussing their relatedness, differences, and coordination and setting up the completely new chapter on metabolic regulation that follows Adds coverage of the mechanisms of phosphohexose isomerase and aldolase Revises the presentation of the mechanism of glyceraldehyde 3-phosphate dehydrogenase. New Chapter 15. Principles of Metabolic Regulation, Illustrated with Glucose and Glycogen Metabolism 15.1 The Metabolism of Glycogen in Animals 15.2 Regulation of Metabolic Pathways 15.3 Coordinated Regulation of Glycolysis and Gluconeogenesis 15.4 Coordinated Regulation of Glycogen Synthesis and Breakdown 15.5 Analysis of Metabolic Control Brings together the concepts and principles of metabolic regulation in one chapter Concludes with the latest conceptual approaches to the regulation of metabolism, including metabolic control analysis and contemporary methods for studying and predicting the flux through metabolic pathways 16. The Citric Acid Cycle 16.1 Production of Acetyl-CoA (Activated Acetate) 16.2 Reactions of the Citric Acid Cycle 16.3 Regulation of the Citric Acid Cycle 16.4 The Glyoxylate Cycle Expands and updates the presentation of the mechanism for pyruvate carboxylase. Adds coverage of the mechanisms of isocitrate dehydrogenase and citrate synthase. 17. Fatty Acid Catabolism 17.1 Digestion, Mobilization, and Transport of Fats 17.2 Oxidation of Fatty Acids 17.3 Ketone Bodies Updates coverage of trifunctional protein New section on the role of perilipin phosphorylation in the control of fat mobilization New discussion of the role of acetyl-CoA in the integration of fatty acid oxidation and synthesis Updates coverage of the medical consequences of genetic defects in fatty acyl–CoA dehydrogenases Takes a fresh look at medical issues related to peroxisomes 18. Amino Acid Oxidation and the Production of Urea 18.1 Metabolic Fates of Amino Groups 18.2 Nitrogen Excretion and the Urea Cycle 18.3 Pathways of Amino Acid Degradation Integrates the latest on regulation of reactions throughout the chapter, with new material on genetic defects in urea cycle enzymes, and updated information on the regulatory function of N-acetylglutamate synthase. Reorganizes coverage of amino acid degradation to focus on the big picture Adds new material on the relative importance of several degradative pathways Includes a new description of the interplay of the pyridoxal phosphate and tetrahydrofolate cofactors in serine and glycine metabolism 19. Oxidative Phosphorylation and Photophosphorylation Oxidative Phosporylation 19.1 Electron-Transfer Reactions in Mitochondria 19.2 ATP Synthesis 19.3 Regulation of Oxidative Phosphorylation 19.4 Mitochondrial Genes: Their Origin and the Effects of Mutations 19.5 The Role of Mitochondria in Apoptosis and Oxidative Stress Photosynthesis: Harvesting Light Energy 19.6 General Features of Photophosphorylation 19.7 Light Absorption 19.8 The Central Photochemical Event: Light-Driven Electron Flow 19.9 ATP Synthesis by Photophosphorylation Adds a prominent new section on the roles of mitochondria in apoptosis and oxidative stress Now covers the role of IF1 in the inhibition of ATP synthase during ischemia Includes revelatory details on the light-dependent pathways of electron transfer in photosynthesis, based on newly available molecular structures 20. Carbohydrate Biosynthesis in Plants and Bacteria 20.1 Photosynthetic Carbohydrate Synthesis 20.2 Photorespiration and the C 4 and CAM Pathways 20.3 Biosynthesis of Starch and Sucrose 20.4 Synthesis of Cell Wall Polysaccharides: Plant Cellulose and Bacterial Peptidoglycan 20.5 Integration of Carbohydrate Metabolism in the Plant Cell Reorganizes the coverage of photosynthesis and the C 4 and CAM pathways Adds a major new section on the synthesis of cellulose and bacterial peptidoglycan 21. Lipid Biosynthesis 21.1 Biosynthesis of Fatty Acids and Eicosanoids 21.2 Biosynthesis of Triacylglycerols 21.3 Biosynthesis of Membrane Phospholipids 21.4 Biosynthesis of Cholesterol, Steroids, and Isoprenoids Features an important new section on glyceroneogenesis and the triacylglycerol cycle between adipose tissue and liver, including their roles in fatty acid metabolism (especially during starvation) and the emergence of thiazolidinediones as regulators of glyceroneogenesis in the treatment of type II diabetes Includes a timely new discussion on the regulation of cholesterol metabolism at the genetic level, with consideration of sterol regulatory element-binding proteins (SREBPs). 22. Biosynthesis of Amino Acids, Nucleotides, and Related Molecules 22.1 Overview of Nitrogen Metabolism 22.2 Biosynthesis of Amino Acids 22.3 Molecules Derived from Amino Acids 22.4 Biosynthesis and Degradation of Nucleotides Adds material on the regulation of nitrogen metabolism at the level of transcription Significantly expands coverage of synthesis and degradation of heme 23. Integration and Hormonal Regulation of Mammalian Metabolism 23.1 Tissue-Specific Metabolism: The Division of Labor 23.2 Hormonal Regulation of Fuel Metabolism 23.3 Long Term Regulation of Body Mass 23.4 Hormones: Diverse Structures for Diverse Functions Reorganized presentation leads students through the complex interactions of integrated metabolism step by step Features extensively revised coverage of insulin and glucagon metabolism that includes the integration of carbohydrate and fat metabolism New discussion of the role of AMP-dependent protein kinase in metabolic integration Updates coverage of the fast-moving field of obesity, regulation of body mass, and the leptin and adiponectin regulatory systems Adds a discussion of Ghrelin and PYY3-36 as regulators of short-term eating behavior Covers the effects of diet on the regulation of gene expression, considering the role of peroxisome proliferator-activated receptors (PPARs) PART III. INFORMATION PATHWAYS 24. Genes and Chromosomes 24.1 Chromosomal Elements 24.2 DNA Supercoiling 24.3 The Structure of Chromosomes Integrates important new material on the structure of chromosomes, including the roles of SMC proteins and cohesins, the features of chromosomal DNA, and the organization of genes in DNA 25. DNA Metabolism 25.1 DNA Replication 25.2 DNA Repair 25.3 DNA Recombination Adds a section on the "replication factories" of bacterial DNA Includes latest perspectives on DNA recombination and repair 26. RNA Metabolism 26.1 DNA-Dependent Synthesis of RNA 26.2 RNA Processing 26.3 RNA-Dependent Synthesis of RNA and DNA Updates coverage on mechanisms of mRNA processing Adds a subsection on the 5' cap of eukaryotic mRNAs Adds important new information about the structure of bacterial RNA polymerase and its mechanism of action. 27. Protein Metabolism 27.1 The Genetic Code 27.2 Protein Synthesis 27.3 Protein Targeting and Degradation Includes a presentation and analysis of the long-awaited structure of the ribosome- -one of the most important updates in this new edition Adds a new box on the evolutionary significance of ribozyme-catalyzed peptide synthesis. 28. Regulation of Gene Expression 28.1 Principles of Gene Regulation 28.2 Regulation of Gene Expression in Prokaryotes 28.3 Regulation of Gene Expression in Eukaryotes Adds a new section on RNA interference (RNAi), including the medical potential of gene silencing. chapter F ifteen to twenty billion years ago, the universe arose as a cataclysmic eruption of hot, energy-rich sub- atomic particles. Within seconds, the simplest elements (hydrogen and helium) were formed. As the universe expanded and cooled, material condensed under the in- fluence of gravity to form stars. Some stars became enormous and then exploded as supernovae, releasing the energy needed to fuse simpler atomic nuclei into the more complex elements. Thus were produced, over bil- lions of years, the Earth itself and the chemical elements found on the Earth today. About four billion years ago, life arose—simple microorganisms with the ability to ex- tract energy from organic compounds or from sunlight, which they used to make a vast array of more complex biomolecules from the simple elements and compounds on the Earth’s surface. Biochemistry asks how the remarkable properties of living organisms arise from the thousands of differ- ent lifeless biomolecules. When these molecules are iso- lated and examined individually, they conform to all the physical and chemical laws that describe the behavior of inanimate matter—as do all the processes occurring in living organisms. The study of biochemistry shows how the collections of inanimate molecules that consti- tute living organisms interact to maintain and perpetu- ate life animated solely by the physical and chemical laws that govern the nonliving universe. Yet organisms possess extraordinary attributes, properties that distinguish them from other collections of matter. What are these distinguishing features of liv- ing organisms? A high degree of chemical complexity and microscopic organization. Thousands of differ- ent molecules make up a cell’s intricate internal structures (Fig. 1–1a). Each has its characteristic sequence of subunits, its unique three-dimensional structure, and its highly specific selection of binding partners in the cell. Systems for extracting, transforming, and using energy from the environment (Fig. 1–1b), enabling organisms to build and maintain their intricate structures and to do mechanical, chemical, osmotic, and electrical work. Inanimate matter tends, rather, to decay toward a more disordered state, to come to equilibrium with its surroundings. THE FOUNDATIONS OF BIOCHEMISTRY 1.1 Cellular Foundations 3 1.2 Chemical Foundations 12 1.3 Physical Foundations 21 1.4 Genetic Foundations 28 1.5 Evolutionary Foundations 31 With the cell, biology discovered its atom ...To characterize life, it was henceforth essential to study the cell and analyze its structure: to single out the common denominators, necessary for the life of every cell; alternatively, to identify differences associated with the performance of special functions. —Fran?ois Jacob, La logique du vivant: une histoire de l’hérédité (The Logic of Life: A History of Heredity), 1970 We must, however, acknowledge, as it seems to me, that man with all his noble qualities . . . still bears in his bodily frame the indelible stamp of his lowly origin. —Charles Darwin, The Descent of Man, 1871 1 1 8885d_c01_01-46 10/27/03 7:48 AM Page 1 mac76 mac76:385_reb: A capacity for precise self-replication and self-assembly (Fig. 1–1c). A single bacterial cell placed in a sterile nutrient medium can give rise to a billion identical “daughter” cells in 24 hours. Each cell contains thousands of different molecules, some extremely complex; yet each bacterium is a faithful copy of the original, its construction directed entirely from information contained within the genetic material of the original cell. Mechanisms for sensing and responding to alterations in their surroundings, constantly adjusting to these changes by adapting their internal chemistry. Defined functions for each of their compo- nents and regulated interactions among them. This is true not only of macroscopic structures, such as leaves and stems or hearts and lungs, but also of microscopic intracellular structures and indi- vidual chemical compounds. The interplay among the chemical components of a living organism is dy- namic; changes in one component cause coordinat- ing or compensating changes in another, with the whole ensemble displaying a character beyond that of its individual parts. The collection of molecules carries out a program, the end result of which is reproduction of the program and self-perpetuation of that collection of molecules—in short, life. A history of evolutionary change. Organisms change their inherited life strategies to survive in new circumstances. The result of eons of evolution is an enormous diversity of life forms, superficially very different (Fig. 1–2) but fundamentally related through their shared ancestry. Despite these common properties, and the funda- mental unity of life they reveal, very few generalizations about living organisms are absolutely correct for every organism under every condition; there is enormous di- versity. The range of habitats in which organisms live, from hot springs to Arctic tundra, from animal intestines to college dormitories, is matched by a correspondingly wide range of specific biochemical adaptations, achieved Chapter 1 The Foundations of Biochemistry2 (a) (c) (b) FIGURE 1–1 Some characteristics of living matter. (a) Microscopic complexity and organization are apparent in this colorized thin sec- tion of vertebrate muscle tissue, viewed with the electron microscope. (b) A prairie falcon acquires nutrients by consuming a smaller bird. (c) Biological reproduction occurs with near-perfect fidelity. FIGURE 1–2 Diverse living organisms share common chemical fea- tures. Birds, beasts, plants, and soil microorganisms share with hu- mans the same basic structural units (cells) and the same kinds of macromolecules (DNA, RNA, proteins) made up of the same kinds of monomeric subunits (nucleotides, amino acids). They utilize the same pathways for synthesis of cellular components, share the same genetic code, and derive from the same evolutionary ancestors. Shown here is a detail from “The Garden of Eden,” by Jan van Kessel the Younger (1626–1679). 8885d_c01_002 11/3/03 1:38 PM Page 2 mac76 mac76:385_reb: within a common chemical framework. For the sake of clarity, in this book we sometimes risk certain general- izations, which, though not perfect, remain useful; we also frequently point out the exceptions that illuminate scientific generalizations. Biochemistry describes in molecular terms the struc- tures, mechanisms, and chemical processes shared by all organisms and provides organizing principles that underlie life in all its diverse forms, principles we refer to collectively as the molecular logic of life. Although biochemistry provides important insights and practical applications in medicine, agriculture, nutrition, and industry, its ultimate concern is with the wonder of life itself. In this introductory chapter, then, we describe (briefly!) the cellular, chemical, physical (thermody- namic), and genetic backgrounds to biochemistry and the overarching principle of evolution—the develop- ment over generations of the properties of living cells. As you read through the book, you may find it helpful to refer back to this chapter at intervals to refresh your memory of this background material. 1.1 Cellular Foundations The unity and diversity of organisms become apparent even at the cellular level. The smallest organisms consist of single cells and are microscopic. Larger, multicellular organisms contain many different types of cells, which vary in size, shape, and specialized function. Despite these obvious differences, all cells of the simplest and most complex organisms share certain fundamental properties, which can be seen at the biochemical level. Cells Are the Structural and Functional Units of All Living Organisms Cells of all kinds share certain structural features (Fig. 1–3). The plasma membrane defines the periphery of the cell, separating its contents from the surroundings. It is composed of lipid and protein molecules that form a thin, tough, pliable, hydrophobic barrier around the cell. The membrane is a barrier to the free passage of inorganic ions and most other charged or polar com- pounds. Transport proteins in the plasma membrane al- low the passage of certain ions and molecules; receptor proteins transmit signals into the cell; and membrane enzymes participate in some reaction pathways. Be- cause the individual lipids and proteins of the plasma membrane are not covalently linked, the entire struc- ture is remarkably flexible, allowing changes in the shape and size of the cell. As a cell grows, newly made lipid and protein molecules are inserted into its plasma membrane; cell division produces two cells, each with its own membrane. This growth and cell division (fission) occurs without loss of membrane integrity. The internal volume bounded by the plasma mem- brane, the cytoplasm (Fig. 1–3), is composed of an aqueous solution, the cytosol, and a variety of sus- pended particles with specific functions. The cytosol is a highly concentrated solution containing enzymes and the RNA molecules that encode them; the components (amino acids and nucleotides) from which these macro- molecules are assembled; hundreds of small organic molecules called metabolites, intermediates in biosyn- thetic and degradative pathways; coenzymes, com- pounds essential to many enzyme-catalyzed reactions; inorganic ions; and ribosomes, small particles (com- posed of protein and RNA molecules) that are the sites of protein synthesis. All cells have, for at least some part of their life, ei- ther a nucleus or a nucleoid, in which the genome— 1.1 Cellular Foundations 3 Nucleus (eukaryotes) or nucleoid (bacteria) Contains genetic material–DNA and associated proteins. Nucleus is membrane-bounded. Plasma membrane Tough, flexible lipid bilayer. Selectively permeable to polar substances. Includes membrane proteins that function in transport, in signal reception, and as enzymes. Cytoplasm Aqueous cell contents and suspended particles and organelles. Supernatant: cytosol Concentrated solution of enzymes, RNA, monomeric subunits, metabolites, inorganic ions. Pellet: particles and organelles Ribosomes, storage granules, mitochondria, chloroplasts, lysosomes, endoplasmic reticulum. centrifuge at 150,000 g FIGURE 1–3 The universal features of living cells. All cells have a nucleus or nucleoid, a plasma membrane, and cytoplasm. The cytosol is defined as that portion of the cytoplasm that remains in the super- natant after centrifugation of a cell extract at 150,000 g for 1 hour. 8885d_c01_003 12/20/03 7:03 AM Page 3 mac76 mac76:385_reb: the complete set of genes, composed of DNA—is stored and replicated. The nucleoid, in bacteria, is not sepa- rated from the cytoplasm by a membrane; the nucleus, in higher organisms, consists of nuclear material en- closed within a double membrane, the nuclear envelope. Cells with nuclear envelopes are called eukaryotes (Greek eu, “true,” and karyon, “nucleus”); those with- out nuclear envelopes—bacterial cells—are prokary- otes (Greek pro, “before”). Cellular Dimensions Are Limited by Oxygen Diffusion Most cells are microscopic, invisible to the unaided eye. Animal and plant cells are typically 5 to 100 H9262m in di- ameter, and many bacteria are only 1 to 2 H9262m long (see the inside back cover for information on units and their abbreviations). What limits the dimensions of a cell? The lower limit is probably set by the minimum number of each type of biomolecule required by the cell. The smallest cells, certain bacteria known as mycoplasmas, are 300 nm in diameter and have a volume of about 10 H1100214 mL. A single bacterial ribosome is about 20 nm in its longest dimension, so a few ribosomes take up a sub- stantial fraction of the volume in a mycoplasmal cell. The upper limit of cell size is probably set by the rate of diffusion of solute molecules in aqueous systems. For example, a bacterial cell that depends upon oxygen- consuming reactions for energy production must obtain molecular oxygen by diffusion from the surrounding medium through its plasma membrane. The cell is so small, and the ratio of its surface area to its volume is so large, that every part of its cytoplasm is easily reached by O 2 diffusing into the cell. As cell size increases, how- ever, surface-to-volume ratio decreases, until metabo- lism consumes O 2 faster than diffusion can supply it. Metabolism that requires O 2 thus becomes impossible as cell size increases beyond a certain point, placing a theoretical upper limit on the size of the cell. There Are Three Distinct Domains of Life All living organisms fall into one of three large groups (kingdoms, or domains) that define three branches of evolution from a common progenitor (Fig. 1–4). Two large groups of prokaryotes can be distinguished on bio- chemical grounds: archaebacteria (Greek arche - , “ori- gin”) and eubacteria (again, from Greek eu, “true”). Eubacteria inhabit soils, surface waters, and the tissues of other living or decaying organisms. Most of the well- studied bacteria, including Escherichia coli, are eu- bacteria. The archaebacteria, more recently discovered, are less well characterized biochemically; most inhabit extreme environments—salt lakes, hot springs, highly acidic bogs, and the ocean depths. The available evi- dence suggests that the archaebacteria and eubacteria diverged early in evolution and constitute two separate Chapter 1 The Foundations of Biochemistry4 Purple bacteria Cyanobacteria Flavobacteria Thermotoga Extreme halophiles Methanogens Extreme thermophiles Microsporidia Flagellates Plants Fungi CiliatesAnimals Archaebacteria Gram- positive bacteria Eubacteria Eukaryotes Green nonsulfur bacteria FIGURE 1–4 Phylogeny of the three domains of life. Phylogenetic relationships are often illustrated by a “family tree” of this type. The fewer the branch points between any two organisms, the closer is their evolutionary relationship. 8885d_c01_01-46 10/27/03 7:48 AM Page 4 mac76 mac76:385_reb: domains, sometimes called Archaea and Bacteria. All eu- karyotic organisms, which make up the third domain, Eukarya, evolved from the same branch that gave rise to the Archaea; archaebacteria are therefore more closely related to eukaryotes than to eubacteria. Within the domains of Archaea and Bacteria are sub- groups distinguished by the habitats in which they live. In aerobic habitats with a plentiful supply of oxygen, some resident organisms derive energy from the trans- fer of electrons from fuel molecules to oxygen. Other environments are anaerobic, virtually devoid of oxy- gen, and microorganisms adapted to these environments obtain energy by transferring electrons to nitrate (form- ing N 2 ), sulfate (forming H 2 S), or CO 2 (forming CH 4 ). Many organisms that have evolved in anaerobic envi- ronments are obligate anaerobes: they die when ex- posed to oxygen. We can classify organisms according to how they obtain the energy and carbon they need for synthesiz- ing cellular material (as summarized in Fig. 1–5). There are two broad categories based on energy sources: pho- totrophs (Greek trophe - , “nourishment”) trap and use sunlight, and chemotrophs derive their energy from oxidation of a fuel. All chemotrophs require a source of organic nutrients; they cannot fix CO 2 into organic com- pounds. The phototrophs can be further divided into those that can obtain all needed carbon from CO 2 (au- totrophs) and those that require organic nutrients (heterotrophs). No chemotroph can get its carbon atoms exclusively from CO 2 (that is, no chemotrophs are autotrophs), but the chemotrophs may be further classified according to a different criterion: whether the fuels they oxidize are inorganic (lithotrophs) or or- ganic (organotrophs). Most known organisms fall within one of these four broad categories—autotrophs or heterotrophs among the photosynthesizers, lithotrophs or organotrophs among the chemical oxidizers. The prokaryotes have several gen- eral modes of obtaining carbon and energy. Escherichia coli, for example, is a chemoorganoheterotroph; it re- quires organic compounds from its environment as fuel and as a source of carbon. Cyanobacteria are photo- lithoautotrophs; they use sunlight as an energy source and convert CO 2 into biomolecules. We humans, like E. coli, are chemoorganoheterotrophs. Escherichia coli Is the Most-Studied Prokaryotic Cell Bacterial cells share certain common structural fea- tures, but also show group-specific specializations (Fig. 1–6). E. coli is a usually harmless inhabitant of the hu- man intestinal tract. The E. coli cell is about 2 H9262m long and a little less than 1 H9262m in diameter. It has a protec- tive outer membrane and an inner plasma membrane that encloses the cytoplasm and the nucleoid. Between the inner and outer membranes is a thin but strong layer of polymers called peptidoglycans, which gives the cell its shape and rigidity. The plasma membrane and the 1.1 Cellular Foundations 5 Heterotrophs (carbon from organic compounds) Examples: ?Purple bacteria ?Green bacteria Autotrophs (carbon from CO 2 ) Examples: ?Cyanobacteria ?Plants Heterotrophs (carbon from organic compounds) Phototrophs (energy from light) Chemotrophs (energy from chemical compounds) All organisms Lithotrophs (energy from inorganic compounds) Examples: ?Sulfur bacteria ?Hydrogen bacteria Organotrophs (energy from organic compounds) Examples: ?Most prokaryotes ?All nonphototrophic eukaryotes FIGURE 1–5 Organisms can be classified according to their source of energy (sunlight or oxidizable chemical compounds) and their source of carbon for the synthesis of cellular material. 8885d_c01_005 12/20/03 7:04 AM Page 5 mac76 mac76:385_reb: layers outside it constitute the cell envelope. In the Archaea, rigidity is conferred by a different type of poly- mer (pseudopeptidoglycan). The plasma membranes of eubacteria consist of a thin bilayer of lipid molecules penetrated by proteins. Archaebacterial membranes have a similar architecture, although their lipids differ strikingly from those of the eubacteria. The cytoplasm of E. coli contains about 15,000 ribosomes, thousands of copies each of about 1,000 different enzymes, numerous metabolites and cofac- tors, and a variety of inorganic ions. The nucleoid contains a single, circular molecule of DNA, and the cytoplasm (like that of most bacteria) contains one or more smaller, circular segments of DNA called plas- mids. In nature, some plasmids confer resistance to toxins and antibiotics in the environment. In the labo- ratory, these DNA segments are especially amenable to experimental manipulation and are extremely use- ful to molecular geneticists. Most bacteria (including E. coli) lead existences as individual cells, but in some bacterial species cells tend to associate in clusters or filaments, and a few (the myxobacteria, for example) demonstrate simple social behavior. Eukaryotic Cells Have a Variety of Membranous Organelles, Which Can Be Isolated for Study Typical eukaryotic cells (Fig. 1–7) are much larger than prokaryotic cells—commonly 5 to 100 H9262m in diameter, with cell volumes a thousand to a million times larger than those of bacteria. The distinguishing characteristics of eukaryotes are the nucleus and a variety of membrane- bounded organelles with specific functions: mitochondria, endoplasmic reticulum, Golgi complexes, and lysosomes. Plant cells also contain vacuoles and chloroplasts (Fig. 1–7). Also present in the cytoplasm of many cells are granules or droplets containing stored nutrients such as starch and fat. In a major advance in biochemistry, Albert Claude, Christian de Duve, and George Palade developed meth- ods for separating organelles from the cytosol and from each other—an essential step in isolating biomolecules and larger cell components and investigating their Chapter 1 The Foundations of Biochemistry6 Ribosomes Bacterial ribosomes are smaller than eukaryotic ribosomes, but serve the same function— protein synthesis from an RNA message. Nucleoid Contains a single, simple, long circular DNA molecule. Pili Provide points of adhesion to surface of other cells. Flagella Propel cell through its surroundings. Cell envelope Structure varies with type of bacteria. Gram-negative bacteria Outer membrane; peptidoglycan layer Outer membrane Peptidoglycan layer Inner membrane Gram-positive bacteria No outer membrane; thicker peptidoglycan layer Cyanobacteria Gram-negative; tougher peptidoglycan layer; extensive internal membrane system with photosynthetic pigments Archaebacteria No outer membrane; peptidoglycan layer outside plasma membrane Peptidoglycan layer Inner membrane FIGURE 1–6 Common structural features of bacterial cells. Because of differences in the cell envelope structure, some eubacteria (gram- positive bacteria) retain Gram’s stain, and others (gram-negative bacteria) do not. E. coli is gram-negative. Cyanobacteria are also eubacteria but are distinguished by their extensive internal membrane system, in which photosynthetic pigments are localized. Although the cell envelopes of archaebacteria and gram-positive eubacteria look similar under the electron microscope, the structures of the membrane lipids and the polysaccharides of the cell envelope are distinctly dif- ferent in these organisms. 8885d_c01_006 11/3/03 1:39 PM Page 6 mac76 mac76:385_reb: 1.1 Cellular Foundations 7 Ribosomes are protein- synthesizing machines Peroxisome destroys peroxides Lysosome degrades intracellular debris Transport vesicle shuttles lipids and proteins between ER, Golgi, and plasma membrane Golgi complex processes, packages, and targets proteins to other organelles or for export Smooth endoplasmic reticulum (SER) is site of lipid synthesis and drug metabolism Nucleus contains the genes (chromatin) Ribosomes Cytoskeleton Cytoskeleton supports cell, aids in movement of organells Golgi complex Nucleolus is site of ribosomal RNA synthesis Rough endoplasmic reticulum (RER) is site of much protein synthesis Mitochondrion oxidizes fuels to produce ATP Plasma membrane separates cell from environment, regulates movement of materials into and out of cell Chloroplast harvests sunlight, produces ATP and carbohydrates Starch granule temporarily stores carbohydrate products of photosynthesis Thylakoids are site of light- driven ATP synthesis Cell wall provides shape and rigidity; protects cell from osmotic swelling Cell wall of adjacent cell Plasmodesma provides path between two plant cells Nuclear envelope segregates chromatin (DNA H11001 protein) from cytoplasm Vacuole degrades and recycles macromolecules, stores metabolites (a) Animal cell (b) Plant cell Glyoxysome contains enzymes of the glyoxylate cycle FIGURE 1–7 Eukaryotic cell structure. Schematic illustrations of the two major types of eukaryotic cell: (a) a representative animal cell and (b) a representative plant cell. Plant cells are usually 10 to 100 H9262m in diameter—larger than animal cells, which typically range from 5 to 30 H9262m. Structures labeled in red are unique to either animal or plant cells. 8885d_c01_007 1/15/04 3:28 PM Page 7 mac76 mac76:385_reb: ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? structures and functions. In a typical cell fractionation (Fig. 1–8), cells or tissues in solution are disrupted by gentle homogenization. This treatment ruptures the plasma membrane but leaves most of the organelles in- tact. The homogenate is then centrifuged; organelles such as nuclei, mitochondria, and lysosomes differ in size and therefore sediment at different rates. They also differ in specific gravity, and they “float” at different levels in a density gradient. Differential centrifugation results in a rough fraction- ation of the cytoplasmic contents, which may be further purified by isopycnic (“same density”) centrifugation. In this procedure, organelles of different buoyant densities (the result of different ratios of lipid and protein in each type of organelle) are separated on a density gradient. By carefully removing material from each region of the gra- dient and observing it with a microscope, the biochemist can establish the sedimentation position of each organelle Chapter 1 The Foundations of Biochemistry8 ? ? ? ? ? ? ? ? ? ? ? ? ? ? Centrifugation Fractionation Sample Less dense component More dense component Sucrose gradient 8765 3421 ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? Isopycnic (sucrose-density) centrifugation ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? (b) ? ? ? ? ? ? ? ? ? ? ? ? ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ?? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ▲ Low-speed centrifugation (1,000 g, 10 min) Supernatant subjected to medium-speed centrifugation (20,000 g, 20 min) Supernatant subjected to high-speed centrifugation (80,000 g, 1 h) Supernatant subjected to very high-speed centrifugation (150,000 g, 3 h) Differential centrifugation Tissue homogenization Tissue homogenate Pellet contains mitochondria, lysosomes, peroxisomes Pellet contains microsomes (fragments of ER), small vesicles Pellet contains ribosomes, large macromolecules Pellet contains whole cells, nuclei, cytoskeletons, plasma membranes Supernatant contains soluble proteins ? ? ? ? ? ?? ? ? ? ? ? ? ? ? ? ? ? ? ▲ ? ? ?? (a) ▲ ▲ ▲ ▲ ▲▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ▲ ? ? ? ?? ? ? ? ? ? ? ? ? ? ? ? ▲ ▲ ▲ ▲▲ ▲ ▲ ▲ ▲ ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? FIGURE 1–8 Subcellular fractionation of tissue. A tissue such as liver is first mechanically homogenized to break cells and disperse their contents in an aqueous buffer. The sucrose medium has an osmotic pressure similar to that in organelles, thus preventing diffusion of wa- ter into the organelles, which would swell and burst. (a) The large and small particles in the suspension can be separated by centrifugation at different speeds, or (b) particles of different density can be sepa- rated by isopycnic centrifugation. In isopycnic centrifugation, a cen- trifuge tube is filled with a solution, the density of which increases from top to bottom; a solute such as sucrose is dissolved at different concentrations to produce the density gradient. When a mixture of organelles is layered on top of the density gradient and the tube is centrifuged at high speed, individual organelles sediment until their buoyant density exactly matches that in the gradient. Each layer can be collected separately. 8885d_c01_01-46 10/27/03 7:48 AM Page 8 mac76 mac76:385_reb: and obtain purified organelles for further study. For example, these methods were used to establish that lysosomes contain degradative enzymes, mitochondria contain oxidative enzymes, and chloroplasts contain photosynthetic pigments. The isolation of an organelle en- riched in a certain enzyme is often the first step in the purification of that enzyme. The Cytoplasm Is Organized by the Cytoskeleton and Is Highly Dynamic Electron microscopy reveals several types of protein fila- ments crisscrossing the eukaryotic cell, forming an inter- locking three-dimensional meshwork, the cytoskeleton. There are three general types of cytoplasmic filaments— actin filaments, microtubules, and intermediate filaments (Fig. 1–9)—differing in width (from about 6 to 22 nm), composition, and specific function. All types provide structure and organization to the cytoplasm and shape to the cell. Actin filaments and microtubules also help to produce the motion of organelles or of the whole cell. Each type of cytoskeletal component is composed of simple protein subunits that polymerize to form fila- ments of uniform thickness. These filaments are not per- manent structures; they undergo constant disassembly into their protein subunits and reassembly into fila- ments. Their locations in cells are not rigidly fixed but may change dramatically with mitosis, cytokinesis, amoeboid motion, or changes in cell shape. The assem- bly, disassembly, and location of all types of filaments are regulated by other proteins, which serve to link or bundle the filaments or to move cytoplasmic organelles along the filaments. The picture that emerges from this brief survey of cell structure is that of a eukaryotic cell with a meshwork of structural fibers and a complex system of membrane-bounded compartments (Fig. 1–7). The fila- ments disassemble and then reassemble elsewhere. Mem- branous vesicles bud from one organelle and fuse with another. Organelles move through the cytoplasm along protein filaments, their motion powered by energy de- pendent motor proteins. The endomembrane system segregates specific metabolic processes and provides surfaces on which certain enzyme-catalyzed reactions occur. Exocytosis and endocytosis, mechanisms of transport (out of and into cells, respectively) that involve membrane fusion and fission, provide paths between the cytoplasm and surrounding medium, allowing for secre- tion of substances produced within the cell and uptake of extracellular materials. 1.1 Cellular Foundations 9 Actin stress fibers (a) Microtubules (b) Intermediate filaments (c) FIGURE 1–9 The three types of cytoskeletal filaments. The upper pan- els show epithelial cells photographed after treatment with antibodies that bind to and specifically stain (a) actin filaments bundled together to form “stress fibers,” (b) microtubules radiating from the cell center, and (c) intermediate filaments extending throughout the cytoplasm. For these experiments, antibodies that specifically recognize actin, tubu- lin, or intermediate filament proteins are covalently attached to a fluorescent compound. When the cell is viewed with a fluorescence microscope, only the stained structures are visible. The lower panels show each type of filament as visualized by (a, b) transmission or (c) scanning electron microscopy. 8885d_c01_009 12/20/03 7:04 AM Page 9 mac76 mac76:385_reb: Although complex, this organization of the cyto- plasm is far from random. The motion and the position- ing of organelles and cytoskeletal elements are under tight regulation, and at certain stages in a eukaryotic cell’s life, dramatic, finely orchestrated reorganizations, such as the events of mitosis, occur. The interactions be- tween the cytoskeleton and organelles are noncovalent, reversible, and subject to regulation in response to var- ious intracellular and extracellular signals. Cells Build Supramolecular Structures Macromolecules and their monomeric subunits differ greatly in size (Fig. 1–10). A molecule of alanine is less than 0.5 nm long. Hemoglobin, the oxygen-carrying pro- tein of erythrocytes (red blood cells), consists of nearly 600 amino acid subunits in four long chains, folded into globular shapes and associated in a structure 5.5 nm in diameter. In turn, proteins are much smaller than ribo- somes (about 20 nm in diameter), which are in turn much smaller than organelles such as mitochondria, typ- ically 1,000 nm in diameter. It is a long jump from sim- ple biomolecules to cellular structures that can be seen Chapter 1 The Foundations of Biochemistry10 Uracil Thymine -D-Ribose 2-Deoxy- -D-ribose O H OH NH 2 HOCH 2 Cytosine H HH OH H O H OH HOCH 2 H HH OH OH Adenine Guanine COO H11002 Oleate Palmitate H CH 2 OH O HO OH -D-Glucose HH H OH OH H (b) The components of nucleic acids (c) Some components of lipids (d) The parent sugar HO P O H11002 O OH Phosphoric acid N Choline H11001 CH 2 CH 2 OH CH 3 CH 3 CH 3 Glycerol CH 2 OH CHOH CH 2 OH CH 2 CH 3 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 3 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 COO H11002 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH 2 CH CH H9251 H9251 H9251 C NH 2 C C CH HC N N N H N C O C C CH C HN N N H N C O O CH CH C HN N H O CH CH C N N H C O O CH C C HN N H H 2 N CH 3 Nitrogenous bases Five-carbon sugars H 3 H11001 N H 3 H11001 N H 3 H11001 H11001 N H 3 H11001 N OC A COO H11002 COO H11002 COO H11002 COO H11002 H 3 H11001 N COO H11002 H 3 H11001 N COO H11002 COO H11002 A CH 3 OH OC A A CH 2 OH OH OC A A C A H 2 OH Alanine Serine Aspartate OC A A C A SH H 2 OH Cysteine Histidine C A OC A OH H 2 OH Tyrosine OC A A C A H 2 OH C H CH HC N NH (a) Some of the amino acids of proteins FIGURE 1–10 The organic compounds from which most cellular materials are constructed: the ABCs of biochemistry. Shown here are (a) six of the 20 amino acids from which all proteins are built (the side chains are shaded pink); (b) the five nitrogenous bases, two five- carbon sugars, and phosphoric acid from which all nucleic acids are built; (c) five components of membrane lipids; and (d) D-glucose, the parent sugar from which most carbohydrates are derived. Note that phosphoric acid is a component of both nucleic acids and membrane lipids. 8885d_c01_010 1/15/04 3:28 PM Page 10 mac76 mac76:385_reb: with the light microscope. Figure 1–11 illustrates the structural hierarchy in cellular organization. The monomeric subunits in proteins, nucleic acids, and polysaccharides are joined by covalent bonds. In supramolecular complexes, however, macromolecules are held together by noncovalent interactions—much weaker, individually, than covalent bonds. Among these noncovalent interactions are hydrogen bonds (between polar groups), ionic interactions (between charged groups), hydrophobic interactions (among nonpolar groups in aqueous solution), and van der Waals inter- actions—all of which have energies substantially smaller than those of covalent bonds (Table 1–1). The nature of these noncovalent interactions is described in Chap- ter 2. The large numbers of weak interactions between macromolecules in supramolecular complexes stabilize these assemblies, producing their unique structures. In Vitro Studies May Overlook Important Interactions among Molecules One approach to understanding a biological process is to study purified molecules in vitro (“in glass”—in the test tube), without interference from other molecules present in the intact cell—that is, in vivo (“in the liv- ing”). Although this approach has been remarkably re- vealing, we must keep in mind that the inside of a cell is quite different from the inside of a test tube. The “in- terfering” components eliminated by purification may be critical to the biological function or regulation of the molecule purified. For example, in vitro studies of pure 1.1 Cellular Foundations 11 Level 4: The cell and its organelles Level 3: Supramolecular complexes Level 2: Macromolecules Level 1: Monomeric units Nucleotides Amino acids Protein Cellulose Plasma membrane Chromosome Cell wall Sugars DNA O H11002 P H11002 OO O O CH2 NH2 H H N N H HOH H O H CH3N COO H11002 CH3 H O H OH CH 2 OH H HO OH OH H O CH 2OH H H11001 FIGURE 1–11 Structural hierarchy in the molecular organization of cells. In this plant cell, the nucleus is an organelle containing several types of supramolecular complexes, including chromosomes. Chro- mosomes consist of macromolecules of DNA and many different pro- teins. Each type of macromolecule is made up of simple subunits— DNA of nucleotides (deoxyribonucleotides), for example. *The greater the energy required for bond dissociation (breakage), the stronger the bond. TABLE 1–1 Strengths of Bonds Common in Biomolecules Bond Bond dissociation dissociation Type energy* Type energy of bond (kJ/mol) of bond (kJ/mol) Single bonds Double bonds OOH 470 CPO 712 HOH 435 CPN 615 POO 419 CPC 611 COH 414 PPO 502 NOH 389 COO 352 Triple bonds COC 348 CmC 816 SOH 339 NmN 930 CON 293 COS 260 NOO 222 SOS 214 enzymes are commonly done at very low enzyme con- centrations in thoroughly stirred aqueous solutions. In the cell, an enzyme is dissolved or suspended in a gel- like cytosol with thousands of other proteins, some of which bind to that enzyme and influence its activity. 8885d_c01_011 12/20/03 7:04 AM Page 11 mac76 mac76:385_reb: Some enzymes are parts of multienzyme complexes in which reactants are channeled from one enzyme to an- other without ever entering the bulk solvent. Diffusion is hindered in the gel-like cytosol, and the cytosolic com- position varies in different regions of the cell. In short, a given molecule may function quite differently in the cell than in vitro. A central challenge of biochemistry is to understand the influences of cellular organization and macromolecular associations on the function of individ- ual enzymes and other biomolecules—to understand function in vivo as well as in vitro. SUMMARY 1.1 Cellular Foundations ■ All cells are bounded by a plasma membrane; have a cytosol containing metabolites, coenzymes, inorganic ions, and enzymes; and have a set of genes contained within a nucleoid (prokaryotes) or nucleus (eukaryotes). ■ Phototrophs use sunlight to do work; chemotrophs oxidize fuels, passing electrons to good electron acceptors: inorganic compounds, organic compounds, or molecular oxygen. ■ Bacterial cells contain cytosol, a nucleoid, and plasmids. Eukaryotic cells have a nucleus and are multicompartmented, segregating certain processes in specific organelles, which can be separated and studied in isolation. ■ Cytoskeletal proteins assemble into long filaments that give cells shape and rigidity and serve as rails along which cellular organelles move throughout the cell. ■ Supramolecular complexes are held together by noncovalent interactions and form a hierarchy of structures, some visible with the light microscope. When individual molecules are removed from these complexes to be studied in vitro, interactions important in the living cell may be lost. 1.2 Chemical Foundations Biochemistry aims to explain biological form and func- tion in chemical terms. As we noted earlier, one of the most fruitful approaches to understanding biological phenomena has been to purify an individual chemical component, such as a protein, from a living organism and to characterize its structural and chemical charac- teristics. By the late eighteenth century, chemists had concluded that the composition of living matter is strik- ingly different from that of the inanimate world. Antoine Lavoisier (1743–1794) noted the relative chemical sim- plicity of the “mineral world” and contrasted it with the complexity of the “plant and animal worlds”; the latter, he knew, were composed of compounds rich in the ele- ments carbon, oxygen, nitrogen, and phosphorus. During the first half of the twentieth century, par- allel biochemical investigations of glucose breakdown in yeast and in animal muscle cells revealed remarkable chemical similarities in these two apparently very dif- ferent cell types; the breakdown of glucose in yeast and muscle cells involved the same ten chemical intermedi- ates. Subsequent studies of many other biochemical processes in many different organisms have confirmed the generality of this observation, neatly summarized by Jacques Monod: “What is true of E. coli is true of the elephant.” The current understanding that all organisms share a common evolutionary origin is based in part on this observed universality of chemical intermediates and transformations. Only about 30 of the more than 90 naturally occur- ring chemical elements are essential to organisms. Most of the elements in living matter have relatively low atomic numbers; only five have atomic numbers above that of selenium, 34 (Fig. 1–12). The four most abun- dant elements in living organisms, in terms of percent- age of total number of atoms, are hydrogen, oxygen, nitrogen, and carbon, which together make up more than 99% of the mass of most cells. They are the light- est elements capable of forming one, two, three, and four bonds, respectively; in general, the lightest elements Chapter 1 The Foundations of Biochemistry12 1 2 34 56 78910 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87 88 H He Li Be B C N O F Ne Na Mg Al Si P S Cl Ar K Ca Sc Ti V Cr Mn Fe Co Ni Cu Zn Ga Ge As Se Br Kr Rb Sr Y Zr Nb Mo Tc Ru Rh Pd Ag Cd In Sn Sb Te I Xe Cs Ba Hf Ta W Re Os Ir Pt Au Hg Tl Pb Bi Po At Rn Fr Ra Lanthanides Actinides Bulk elements Trace elements FIGURE 1–12 Elements essential to animal life and health. Bulk elements (shaded orange) are structural components of cells and tissues and are required in the diet in gram quantities daily. For trace elements (shaded bright yellow), the requirements are much smaller: for humans, a few milligrams per day of Fe, Cu, and Zn, even less of the others. The elemental requirements for plants and microorganisms are similar to those shown here; the ways in which they acquire these elements vary. 8885d_c01_01-46 10/27/03 7:48 AM Page 12 mac76 mac76:385_reb: form the strongest bonds. The trace elements (Fig. 1–12) represent a miniscule fraction of the weight of the hu- man body, but all are essential to life, usually because they are essential to the function of specific proteins, including enzymes. The oxygen-transporting capacity of the hemoglobin molecule, for example, is absolutely dependent on four iron ions that make up only 0.3% of its mass. Biomolecules Are Compounds of Carbon with a Variety of Functional Groups The chemistry of living organisms is organized around carbon, which accounts for more than half the dry weight of cells. Carbon can form single bonds with hy- drogen atoms, and both single and double bonds with oxygen and nitrogen atoms (Fig. 1–13). Of greatest sig- nificance in biology is the ability of carbon atoms to form very stable carbon–carbon single bonds. Each carbon atom can form single bonds with up to four other car- bon atoms. Two carbon atoms also can share two (or three) electron pairs, thus forming double (or triple) bonds. The four single bonds that can be formed by a car- bon atom are arranged tetrahedrally, with an angle of about 109.5H11034 between any two bonds (Fig. 1–14) and an average length of 0.154 nm. There is free rotation around each single bond, unless very large or highly charged groups are attached to both carbon atoms, in which case rotation may be restricted. A double bond is shorter (about 0.134 nm) and rigid and allows little rotation about its axis. Covalently linked carbon atoms in biomolecules can form linear chains, branched chains, and cyclic struc- tures. To these carbon skeletons are added groups of other atoms, called functional groups, which confer specific chemical properties on the molecule. It seems likely that the bonding versatility of carbon was a ma- jor factor in the selection of carbon compounds for the molecular machinery of cells during the origin and evo- lution of living organisms. No other chemical element can form molecules of such widely different sizes and shapes or with such a variety of functional groups. Most biomolecules can be regarded as derivatives of hydrocarbons, with hydrogen atoms replaced by a va- riety of functional groups to yield different families of organic compounds. Typical of these are alcohols, which have one or more hydroxyl groups; amines, with amino groups; aldehydes and ketones, with carbonyl groups; and carboxylic acids, with carboxyl groups (Fig. 1–15). Many biomolecules are polyfunctional, containing two or more different kinds of functional groups (Fig. 1–16), each with its own chemical characteristics and reac- tions. The chemical “personality” of a compound is de- termined by the chemistry of its functional groups and their disposition in three-dimensional space. 1.2 Chemical Foundations 13 HCH H H11001 O H11001 CO H11001 CN C C O H11001 N H11001 C H11001 CC C H11001 CCCC C C C C C C C H11001C O C C C N C N O C CC C N N OC C C C FIGURE 1–13 Versatility of carbon bonding. Carbon can form cova- lent single, double, and triple bonds (in red), particularly with other carbon atoms. Triple bonds are rare in biomolecules. FIGURE 1–14 Geometry of carbon bonding. (a) Carbon atoms have a characteristic tetrahedral arrangement of their four single bonds. (b) Carbon–carbon single bonds have freedom of rotation, as shown for the compound ethane (CH 3 OCH 3 ). (c) Double bonds are shorter and do not allow free rotation. The two doubly bonded carbons and the atoms designated A, B, X, and Y all lie in the same rigid plane. (a) (b) (c) 109.5° 109.5° C C C 120° X C C A B Y 8885d_c01_013 1/15/04 3:28 PM Page 13 mac76 mac76:385_reb: Cells Contain a Universal Set of Small Molecules Dissolved in the aqueous phase (cytosol) of all cells is a collection of 100 to 200 different small organic mole- cules (M r ~100 to ~500), the central metabolites in the major pathways occurring in nearly every cell—the metabolites and pathways that have been conserved throughout the course of evolution. (See Box 1–1 for an explanation of the various ways of referring to molecu- lar weight.) This collection of molecules includes the common amino acids, nucleotides, sugars and their phosphorylated derivatives, and a number of mono-, di-, and tricarboxylic acids. The molecules are polar or charged, water soluble, and present in micromolar to millimolar concentrations. They are trapped within the cell because the plasma membrane is impermeable to them—although specific membrane transporters can catalyze the movement of some molecules into and out Chapter 1 The Foundations of Biochemistry14 Hydroxyl ROH (alcohol) Carbonyl (aldehyde) RC O H Carbonyl (ketone) RC O R 21 Carboxyl RC O O H11002 O H11002 O H11002 O H11002 Methyl RC H H H Ethyl RC H H C H H H Ester R 1 C O OR 2 Ether R 1 OR 2 Sulfhydryl RSH Disulfide R SSR 21 Phosphoryl ROP O OH Thioester R 1 C O SR 2 Anhydride R 1 C OO CR 2 (two car- boxylic acids) O Imidazole R N CHC HN H C Guanidino RN H C N H N H H Amino RN H H Amido RC O N H H Phenyl RC HC C H H C C C H H (carboxylic acid and phosphoric acid; also called acyl phosphate) Mixed anhydride RCO O OH Phosphoanhydride R 1 O R 2 OP O P O H11002 O P O O FIGURE 1–15 Some common functional groups of biomolecules. In this figure and throughout the book, we use R to represent “any substituent.” It may be as simple as a hydrogen atom, but typically it is a carbon-containing moiety. When two or more substituents are shown in a molecule, we designate them R 1 , R 2 , and so forth. 8885d_c01_014 1/15/04 3:28 PM Page 14 mac76 mac76:385_reb: of the cell or between compartments in eukaryotic cells. The universal occurrence of the same set of compounds in living cells is a manifestation of the universality of metabolic design, reflecting the evolutionary conserva- tion of metabolic pathways that developed in the earli- est cells. There are other small biomolecules, specific to cer- tain types of cells or organisms. For example, vascular plants contain, in addition to the universal set, small molecules called secondary metabolites, which play a role specific to plant life. These metabolites include compounds that give plants their characteristic scents, and compounds such as morphine, quinine, nicotine, and caffeine that are valued for their physiological ef- fects on humans but used for other purposes by plants. The entire collection of small molecules in a given cell has been called that cell’s metabolome, in parallel with the term “genome” (defined earlier and expanded on in Section 1.4). If we knew the composition of a cell’s metabolome, we could predict which enzymes and meta- bolic pathways were active in that cell. Macromolecules Are the Major Constituents of Cells Many biological molecules are macromolecules, poly- mers of high molecular weight assembled from rela- tively simple precursors. Proteins, nucleic acids, and polysaccharides are produced by the polymerization of relatively small compounds with molecular weights of 500 or less. The number of polymerized units can range from tens to millions. Synthesis of macromolecules is a major energy-consuming activity of cells. Macromol- ecules themselves may be further assembled into supramolecular complexes, forming functional units such as ribosomes. Table 1–2 shows the major classes of biomolecules in the bacterium E. coli. 1.2 Chemical Foundations 15 SOCH 2 OCH 2 ONHOC B O OCH 2 OCH 2 ONHOC B O B O OC A H A OH O C A CH 3 A CH 3 CH 2 OOCH 3 OOC OP A O H11002 B O OOOP A B O OOOCH 2 O H N K CH B C E A NH 2 N N A H11002 OOP OH PO imidazole amino phosphoanhydride Acetyl-coenzyme A H N N C H C HC O O H methyl hydroxyl amido methyl amidothioester phosphoryl CC C H C H A A O H11002 OO FIGURE 1–16 Several common functional groups in a single biomolecule. Acetyl-coenzyme A (often abbreviated as acetyl-CoA) is a carrier of acetyl groups in some enzymatic reactions. BOX 1–1 WORKING IN BIOCHEMISTRY Molecular Weight, Molecular Mass, and Their Correct Units There are two common (and equivalent) ways to de- scribe molecular mass; both are used in this text. The first is molecular weight, or relative molecular mass, denoted M r . The molecular weight of a substance is de- fined as the ratio of the mass of a molecule of that sub- stance to one-twelfth the mass of carbon-12 ( 12 C). Since M r is a ratio, it is dimensionless—it has no asso- ciated units. The second is molecular mass, denoted m. This is simply the mass of one molecule, or the mo- lar mass divided by Avogadro’s number. The molecu- lar mass, m, is expressed in daltons (abbreviated Da). One dalton is equivalent to one-twelfth the mass of carbon-12; a kilodalton (kDa) is 1,000 daltons; a mega- dalton (MDa) is 1 million daltons. Consider, for example, a molecule with a mass 1,000 times that of water. We can say of this molecule either M r H11005 18,000 or m H11005 18,000 daltons. We can also describe it as an “18 kDa molecule.” However, the ex- pression M r H11005 18,000 daltons is incorrect. Another convenient unit for describing the mass of a single atom or molecule is the atomic mass unit (formerly amu, now commonly denoted u). One atomic mass unit (1 u) is defined as one-twelfth the mass of an atom of carbon-12. Since the experimen- tally measured mass of an atom of carbon-12 is 1.9926 H11003 10 H1100223 g, 1 u H11005 1.6606 H11003 10 H1100224 g. The atomic mass unit is convenient for describing the mass of a peak observed by mass spectrometry (see Box 3–2). 8885d_c01_015 1/15/04 3:29 PM Page 15 mac76 mac76:385_reb: Proteins, long polymers of amino acids, constitute the largest fraction (besides water) of cells. Some pro- teins have catalytic activity and function as enzymes; others serve as structural elements, signal receptors, or transporters that carry specific substances into or out of cells. Proteins are perhaps the most versatile of all biomolecules. The nucleic acids, DNA and RNA, are polymers of nucleotides. They store and transmit genetic information, and some RNA molecules have structural and catalytic roles in supramolecular complexes. The poly- saccharides, polymers of simple sugars such as glucose, have two major functions: as energy-yielding fuel stores and as extracellular structural elements with specific binding sites for particular proteins. Shorter polymers of sugars (oligosaccharides) attached to proteins or lipids at the cell surface serve as specific cellular signals. The lipids, greasy or oily hydrocarbon derivatives, serve as structural components of membranes, energy-rich fuel stores, pigments, and intracellular signals. In proteins, nucleotides, polysaccharides, and lipids, the number of monomeric subunits is very large: molecular weights in the range of 5,000 to more than 1 million for proteins, up to several billion for nucleic acids, and in the millions for polysaccharides such as starch. Individual lipid mol- ecules are much smaller (M r 750 to 1,500) and are not classified as macromolecules. However, large num- bers of lipid molecules can associate noncovalently into very large structures. Cellular membranes are built of enormous noncovalent aggregates of lipid and protein molecules. Proteins and nucleic acids are informational macromolecules: each protein and each nucleic acid has a characteristic information-rich subunit sequence. Some oligosaccharides, with six or more different sug- ars connected in branched chains, also carry informa- tion; on the outer surface of cells they serve as highly specific points of recognition in many cellular processes (as described in Chapter 7). Three-Dimensional Structure Is Described by Configuration and Conformation The covalent bonds and functional groups of a biomol- ecule are, of course, central to its function, but so also is the arrangement of the molecule’s constituent atoms in three-dimensional space—its stereochemistry. A carbon-containing compound commonly exists as stereoisomers, molecules with the same chemical bonds but different stereochemistry—that is, different configuration, the fixed spatial arrangement of atoms. Interactions between biomolecules are invariably stereo- specific, requiring specific stereochemistry in the in- teracting molecules. Figure 1–17 shows three ways to illustrate the stereo- chemical structures of simple molecules. The perspec- tive diagram specifies stereochemistry unambiguously, but bond angles and center-to-center bond lengths are better represented with ball-and-stick models. In space- Chapter 1 The Foundations of Biochemistry16 Approximate number of Percentage of different total weight molecular of cell species Water 70 1 Proteins 15 3,000 Nucleic acids DNA 1 1 RNA 6 H110223,000 Polysaccharides 3 5 Lipids 2 20 Monomeric subunits and intermediates 2 500 Inorganic ions 1 20 Molecular Components of an E. coli Cell TABLE 1–2 H 2 N#C C M O D OH HOC A H OH !H (a) (c) (b) FIGURE 1–17 Representations of molecules. Three ways to represent the structure of the amino acid alanine. (a) Structural formula in per- spective form: a solid wedge (!) represents a bond in which the atom at the wide end projects out of the plane of the paper, toward the reader; a dashed wedge (^) represents a bond extending behind the plane of the paper. (b) Ball-and-stick model, showing relative bond lengths and the bond angles. (c) Space-filling model, in which each atom is shown with its correct relative van der Waals radius. 8885d_c01_016 11/3/03 1:40 PM Page 16 mac76 mac76:385_reb: filling models, the radius of each atom is proportional to its van der Waals radius, and the contours of the model define the space occupied by the molecule (the volume of space from which atoms of other molecules are excluded). Configuration is conferred by the presence of either (1) double bonds, around which there is no freedom of rotation, or (2) chiral centers, around which substituent groups are arranged in a specific sequence. The identi- fying characteristic of configurational isomers is that they cannot be interconverted without temporarily breaking one or more covalent bonds. Figure 1–18a shows the configurations of maleic acid and its isomer, fumaric acid. These compounds are geometric, or cis- trans, isomers; they differ in the arrangement of their substituent groups with respect to the nonrotating dou- ble bond (Latin cis, “on this side”—groups on the same side of the double bond; trans, “across”—groups on op- posite sides). Maleic acid is the cis isomer and fumaric acid the trans isomer; each is a well-defined compound that can be separated from the other, and each has its own unique chemical properties. A binding site (on an enzyme, for example) that is complementary to one of these molecules would not be a suitable binding site for the other, which explains why the two compounds have distinct biological roles despite their similar chemistry. In the second type of configurational isomer, four different substituents bonded to a tetrahedral carbon atom may be arranged two different ways in space—that is, have two configurations (Fig. 1–19)—yielding two stereoisomers with similar or identical chemical proper- ties but differing in certain physical and biological prop- erties. A carbon atom with four different substituents is said to be asymmetric, and asymmetric carbons are called chiral centers (Greek chiros, “hand”; some stereoisomers are related structurally as the right hand is to the left). A molecule with only one chiral carbon can have two stereoisomers; when two or more (n) chi- ral carbons are present, there can be 2 n stereoisomers. Some stereoisomers are mirror images of each other; they are called enantiomers (Fig. 1–19). Pairs of stereoisomers that are not mirror images of each other are called diastereomers (Fig. 1–20). As Louis Pasteur first observed (Box 1–2), enan- tiomers have nearly identical chemical properties but differ in a characteristic physical property, their inter- action with plane-polarized light. In separate solutions, two enantiomers rotate the plane of plane-polarized light in opposite directions, but an equimolar solution of the two enantiomers (a racemic mixture) shows no optical rotation. Compounds without chiral centers do not rotate the plane of plane-polarized light. 1.2 Chemical Foundations 17 C G H D HOOC PC D H G COOH Maleic acid (cis) CH 3 O J C 11-cis-Retinal light C G HOOC D H PC D H G COOH Fumaric acid (trans) All-trans-Retinal H 3 CH 3 G C H 9 1210 11 H 3 CH 3 C 9 12 10 11 H 3 C C J O G H CH 3 CH 3 G D (b) (a) CH 3 CH 3 G D FIGURE 1–18 Configurations of geometric isomers. (a) Isomers such as maleic acid and fumaric acid cannot be interconverted without breaking covalent bonds, which requires the input of much energy. (b) In the vertebrate retina, the initial event in light detection is the absorption of visible light by 11-cis-retinal. The energy of the absorbed light (about 250 kJ/mol) converts 11-cis-retinal to all-trans-retinal, triggering electrical changes in the retinal cell that lead to a nerve impulse. (Note that the hydrogen atoms are omitted from the ball-and- stick models for the retinals.) 8885d_c01_017 12/20/03 7:05 AM Page 17 mac76 mac76:385_reb: Given the importance of stereochemistry in reac- tions between biomolecules (see below), biochemists must name and represent the structure of each bio- molecule so that its stereochemistry is unambiguous. For compounds with more than one chiral center, the most useful system of nomenclature is the RS system. In this system, each group attached to a chiral carbon is assigned a priority. The priorities of some common substituents are OOCH 2 H11022 OOH H11022 ONH 2 H11022 OCOOH H11022 OCHOH11022 OCH 2 OH H11022 OCH 3 H11022 OH For naming in the RS system, the chiral atom is viewed with the group of lowest priority (4 in the diagram on the next page) pointing away from the viewer. If the pri- ority of the other three groups (1 to 3) decreases in clockwise order, the configuration is (R) (Latin rectus, “right”); if in counterclockwise order, the configuration Chapter 1 The Foundations of Biochemistry18 Y Y C B (a) Mirror image of original molecule Chiral molecule: Rotated molecule cannot be superimposed on its mirror image Original molecule A Y A C X B A C B X X X A C X B X A C X B (b) Achiral molecule: Rotated molecule can be superimposed on its mirror image Mirror image of original molecule Original molecule X A C X B FIGURE 1–19 Molecular asymmetry: chiral and achiral molecules. (a) When a carbon atom has four different substituent groups (A, B, X, Y), they can be arranged in two ways that represent nonsuperim- posable mirror images of each other (enantiomers). This asymmetric carbon atom is called a chiral atom or chiral center. (b) When a tetra- hedral carbon has only three dissimilar groups (i.e., the same group occurs twice), only one configuration is possible and the molecule is symmetric, or achiral. In this case the molecule is superimposable on its mirror image: the molecule on the left can be rotated counter- clockwise (when looking down the vertical bond from A to C) to cre- ate the molecule in the mirror. Diastereomers (non–mirror images) C CH 3 CH 3 H CH X Y Enantiomers (mirror images) Enantiomers (mirror images) C CH 3 CH 3 X CY H H C CH 3 CH 3 H CY Y X H C CH 3 CH 3 H C X H FIGURE 1–20 Two types of stereoisomers. There are four different 2,3-disubstituted butanes (n H11005 2 asymmetric carbons, hence 2 n H11005 4 stereoisomers). Each is shown in a box as a perspective formula and a ball-and-stick model, which has been rotated to allow the reader to view all the groups. Some pairs of stereoisomers are mirror images of each other, or enantiomers. Other pairs are not mirror images; these are diastereomers. 8885d_c01_01-46 10/27/03 7:48 AM Page 18 mac76 mac76:385_reb: is (S) (Latin sinister, “left”). In this way each chiral car- bon is designated either (R) or (S), and the inclusion of these designations in the name of the compound pro- vides an unambiguous description of the stereochem- istry at each chiral center. Another naming system for stereoisomers, the D and L system, is described in Chapter 3. A molecule with a sin- gle chiral center (glyceraldehydes, for example) can be named unambiguously by either system. 1 4 3 2 Counterclockwise (S) 1 4 3 2 Clockwise (R) Distinct from configuration is molecular confor- mation, the spatial arrangement of substituent groups that, without breaking any bonds, are free to assume different positions in space because of the freedom of rotation about single bonds. In the simple hydrocarbon ethane, for example, there is nearly complete freedom of rotation around the COC bond. Many different, in- terconvertible conformations of ethane are possible, depending on the degree of rotation (Fig. 1–21). Two conformations are of special interest: the staggered, which is more stable than all others and thus predomi- nates, and the eclipsed, which is least stable. We cannot isolate either of these conformational forms, because CH 2 OH CH 2 OH CHO CHO HHCHO OH L-Glyceraldehyde (S)-Glyceraldehyde ≡ (2) (1)(4) (3) 1.2 Chemical Foundations 19 BOX 1–2 WORKING IN BIOCHEMISTRY Louis Pasteur and Optical Activity: In Vino, Veritas Louis Pasteur encountered the phenome- non of optical activity in 1843, during his investigation of the crystalline sediment that accumulated in wine casks (a form of tartaric acid called paratartaric acid—also called racemic acid, from Latin racemus, “bunch of grapes”). He used fine forceps to separate two types of crystals identical in shape but mirror images of each other. Both types proved to have all the chemi- cal properties of tartaric acid, but in solu- tion one type rotated polarized light to the left (levorotatory), the other to the right (dextrorotatory). Pasteur later described the experi- ment and its interpretation: In isomeric bodies, the elements and the propor- tions in which they are combined are the same, only the arrangement of the atoms is different ...We know, on the one hand, that the molecular arrange- ments of the two tartaric acids are asymmetric, and, on the other hand, that these arrangements are ab- solutely identical, excepting that they exhibit asym- metry in opposite directions. Are the atoms of the dextro acid grouped in the form of a right-handed spiral, or are they placed at the apex of an irregu- lar tetrahedron, or are they disposed according to this or that asymmetric arrangement? We do not know.* Now we do know. X-ray crystallo- graphic studies in 1951 confirmed that the levorotatory and dextrorotatory forms of tartaric acid are mirror images of each other at the molecular level and established the absolute configuration of each (Fig. 1). The same approach has been used to demonstrate that although the amino acid alanine has two stereoisomeric forms (des- ignated D and L), alanine in proteins exists exclusively in one form (the L isomer; see Chapter 3). FIGURE 1 Pasteur separated crystals of two stereoisomers of tartaric acid and showed that solutions of the separated forms rotated po- larized light to the same extent but in opposite directions. These dextrorotatory and levorotatory forms were later shown to be the (R,R) and (S,S) isomers represented here. The RS system of nomen- clature is explained in the text. Louis Pasteur 1822–1895 *From Pasteur’s lecture to the Société Chimique de Paris in 1883, quoted in DuBos, R. (1976) Louis Pasteur: Free Lance of Science, p. 95, Charles Scribner’s Sons, New York. C HOOC 1 H 2 C 3 C 4 OOH OH C HOOC 1 HO 2 C 3 C 4 O OH H (2R,3R)-Tartaric acid (2S,3S)-Tartaric acid (dextrorotatory) (levorotatory) OHH OH H 8885d_c01_019 12/20/03 7:06 AM Page 19 mac76 mac76:385_reb: they are freely interconvertible. However, when one or more of the hydrogen atoms on each carbon is replaced by a functional group that is either very large or elec- trically charged, freedom of rotation around the COC bond is hindered. This limits the number of stable con- formations of the ethane derivative. Interactions between Biomolecules Are Stereospecific Biological interactions between molecules are stereo- specific: the “fit” in such interactions must be stereo- chemically correct. The three-dimensional structure of biomolecules large and small—the combination of con- figuration and conformation—is of the utmost impor- tance in their biological interactions: reactant with enzyme, hormone with its receptor on a cell surface, antigen with its specific antibody, for example (Fig. 1–22). The study of biomolecular stereochemistry with precise physical methods is an important part of mod- ern research on cell structure and biochemical function. In living organisms, chiral molecules are usually present in only one of their chiral forms. For example, the amino acids in proteins occur only as their L iso- mers; glucose occurs only as its D isomer. (The con- ventions for naming stereoisomers of the amino acids are described in Chapter 3; those for sugars, in Chap- ter 7; the RS system, described above, is the most useful for some biomolecules.) In contrast, when a com- pound with an asymmetric carbon atom is chemically synthesized in the laboratory, the reaction usually pro- duces all possible chiral forms: a mixture of the D and L forms, for example. Living cells produce only one chiral form of biomolecules because the enzymes that syn- thesize them are also chiral. Stereospecificity, the ability to distinguish between stereoisomers, is a property of enzymes and other pro- teins and a characteristic feature of the molecular logic of living cells. If the binding site on a protein is com- plementary to one isomer of a chiral compound, it will not be complementary to the other isomer, for the same reason that a left glove does not fit a right hand. Two striking examples of the ability of biological systems to distinguish stereoisomers are shown in Figure 1–23. SUMMARY 1.2 Chemical Foundations ■ Because of its bonding versatility, carbon can produce a broad array of carbon–carbon skeletons with a variety of functional groups; these groups give biomolecules their biological and chemical personalities. ■ A nearly universal set of several hundred small molecules is found in living cells; the interconver- sions of these molecules in the central metabolic pathways have been conserved in evolution. ■ Proteins and nucleic acids are linear polymers of simple monomeric subunits; their sequences contain the information that gives each molecule its three-dimensional structure and its biological functions. Chapter 1 The Foundations of Biochemistry20 0 60 120 180 240 300 360 0 4 8 12 Potential energy (kJ/mol) Torsion angle (degrees) 12.1 kJ/mol FIGURE 1–21 Conformations. Many conformations of ethane are pos- sible because of freedom of rotation around the COC bond. In the ball-and-stick model, when the front carbon atom (as viewed by the reader) with its three attached hydrogens is rotated relative to the rear carbon atom, the potential energy of the molecule rises to a maximum in the fully eclipsed conformation (torsion angle 0H11034, 120H11034, etc.), then falls to a minimum in the fully staggered conformation (torsion angle 60H11034, 180H11034, etc.). Because the energy differences are small enough to allow rapid interconversion of the two forms (millions of times per sec- ond), the eclipsed and staggered forms cannot be separately isolated. FIGURE 1–22 Complementary fit between a macromolecule and a small molecule. A segment of RNA from the regulatory region TAR of the human immunodeficiency virus genome (gray) with a bound argin- inamide molecule (colored), representing one residue of a protein that binds to this region. The argininamide fits into a pocket on the RNA surface and is held in this orientation by several noncovalent interac- tions with the RNA. This representation of the RNA molecule is pro- duced with the computer program GRASP, which can calculate the shape of the outer surface of a macromolecule, defined either by the van der Waals radii of all the atoms in the molecule or by the “solvent exclusion volume,” into which a water molecule cannot penetrate. 8885d_c01_020 1/15/04 3:29 PM Page 20 mac76 mac76:385_reb: ■ Molecular configuration can be changed only by breaking covalent bonds. For a carbon atom with four different substituents (a chiral carbon), the substituent groups can be arranged in two different ways, generating stereoisomers with distinct properties. Only one stereoisomer is biologically active. Molecular conformation is the position of atoms in space that can be changed by rotation about single bonds, without breaking covalent bonds. ■ Interactions between biological molecules are almost invariably stereospecific: they require a complementary match between the interacting molecules. 1.3 Physical Foundations Living cells and organisms must perform work to stay alive and to reproduce themselves. The synthetic reac- tions that occur within cells, like the synthetic processes in any factory, require the input of energy. Energy is also consumed in the motion of a bacterium or an Olympic sprinter, in the flashing of a firefly or the electrical dis- charge of an eel. And the storage and expression of information require energy, without which structures rich in information inevitably become disordered and meaningless. In the course of evolution, cells have developed highly efficient mechanisms for coupling the energy obtained from sunlight or fuels to the many energy- consuming processes they must carry out. One goal of biochemistry is to understand, in quantitative and chem- ical terms, the means by which energy is extracted, channeled, and consumed in living cells. We can consider cellular energy conversions—like all other energy con- versions—in the context of the laws of thermodynamics. Living Organisms Exist in a Dynamic Steady State, Never at Equilibrium with Their Surroundings The molecules and ions contained within a living or- ganism differ in kind and in concentration from those in the organism’s surroundings. A Paramecium in a pond, a shark in the ocean, an erythrocyte in the human blood- stream, an apple tree in an orchard—all are different in composition from their surroundings and, once they have reached maturity, all (to a first approximation) maintain a constant composition in the face of con- stantly changing surroundings. Although the characteristic composition of an or- ganism changes little through time, the population of molecules within the organism is far from static. Small molecules, macromolecules, and supramolecular com- plexes are continuously synthesized and then broken down in chemical reactions that involve a constant flux of mass and energy through the system. The hemoglo- bin molecules carrying oxygen from your lungs to your brain at this moment were synthesized within the past month; by next month they will have been degraded and entirely replaced by new hemoglobin molecules. The glucose you ingested with your most recent meal is now circulating in your bloodstream; before the day is over these particular glucose molecules will have been 1.3 Physical Foundations 21 (a) H 2 C C CH 3 C CH 2 H CH 2 C O C CH 3 CH H 2 C C HC CH 3 CH 2 CH 2 C O C CH 3 CH (R)-Carvone (S)-Carvone (spearmint) (caraway) (b) C H11002 OOC CH 2 NH 3 C O N H C H C O OCH 3 H11002 OOC CH 2 C H11001 H 3 C O N H C H C O OCH 3 HC C H CH HC C CH 2 CH H11001 N HC C H CH HC C CH 2 CH H H L-Aspartyl-L-phenylalanine methyl ester (aspartame) (sweet) L-Aspartyl-D-phenylalanine methyl ester (bitter) FIGURE 1–23 Stereoisomers distinguishable by smell and taste in humans. (a) Two stereoisomers of carvone: (R)-carvone (isolated from spearmint oil) has the characteristic fragrance of spearmint; (S)-carvone (from caraway seed oil) smells like caraway. (b) Aspartame, the artificial sweetener sold under the trade name NutraSweet, is easily distinguishable by taste receptors from its bitter-tasting stereoisomer, although the two differ only in the configuration at one of the two chiral carbon atoms. 8885d_c01_021 12/20/03 7:06 AM Page 21 mac76 mac76:385_reb: converted into something else—carbon dioxide or fat, perhaps—and will have been replaced with a fresh sup- ply of glucose, so that your blood glucose concentration is more or less constant over the whole day. The amounts of hemoglobin and glucose in the blood remain nearly constant because the rate of synthesis or intake of each just balances the rate of its breakdown, consumption, or conversion into some other product. The constancy of concentration is the result of a dynamic steady state, a steady state that is far from equilibrium. Maintaining this steady state requires the constant investment of en- ergy; when the cell can no longer generate energy, it dies and begins to decay toward equilibrium with its sur- roundings. We consider below exactly what is meant by “steady state” and “equilibrium.” Organisms Transform Energy and Matter from Their Surroundings For chemical reactions occurring in solution, we can de- fine a system as all the reactants and products present, the solvent that contains them, and the immediate at- mosphere—in short, everything within a defined region of space. The system and its surroundings together con- stitute the universe. If the system exchanges neither matter nor energy with its surroundings, it is said to be isolated. If the system exchanges energy but not mat- ter with its surroundings, it is a closed system; if it ex- changes both energy and matter with its surroundings, it is an open system. A living organism is an open system; it exchanges both matter and energy with its surroundings. Living or- ganisms derive energy from their surroundings in two ways: (1) they take up chemical fuels (such as glucose) from the environment and extract energy by oxidizing them (see Box 1–3, Case 2); or (2) they absorb energy from sunlight. The first law of thermodynamics, developed from physics and chemistry but fully valid for biological sys- tems as well, describes the principle of the conservation of energy: in any physical or chemical change, the total amount of energy in the universe remains con- stant, although the form of the energy may change. Cells are consummate transducers of energy, capable of interconverting chemical, electromagnetic, mechanical, and osmotic energy with great efficiency (Fig. 1–24). The Flow of Electrons Provides Energy for Organisms Nearly all living organisms derive their energy, directly or indirectly, from the radiant energy of sunlight, which arises from thermonuclear fusion reactions carried out in the sun. Photosynthetic cells absorb light energy and use it to drive electrons from water to carbon di- oxide, forming energy-rich products such as glucose (C 6 H 12 O 6 ), starch, and sucrose and releasing O 2 into the atmosphere: light 6CO 2 H11001 6H 2 O 888n C 6 H 12 O 6 H11001 6O 2 (light-driven reduction of CO 2 ) Nonphotosynthetic cells and organisms obtain the en- ergy they need by oxidizing the energy-rich products of photosynthesis and then passing electrons to atmos- Chapter 1 The Foundations of Biochemistry22 (a) (b) (c) (d) (e) Energy transductions accomplish work Potential energy ? Nutrients in environment (complex molecules such as sugars, fats) ? Sunlight Chemical transformations within cells Cellular work: ? chemical synthesis ? mechanical work ? osmotic and electrical gradients ? light production ? genetic information transfer Heat Increased randomness (entropy) in the surroundings Metabolism produces compounds simpler than the initial fuel molecules: CO 2 , NH 3 , H 2 O, HPO 4 2H11002 Decreased randomness (entropy) in the system Simple compounds polymerize to form information-rich macromolecules: DNA, RNA, proteins FIGURE 1–24 Some energy interconversion in living organisms. Dur- ing metabolic energy transductions, the randomness of the system plus surroundings (expressed quantitatively as entropy) increases as the po- tential energy of complex nutrient molecules decreases. (a) Living or- ganisms extract energy from their surroundings; (b) convert some of it into useful forms of energy to produce work; (c) return some en- ergy to the surroundings as heat; and (d) release end-product mole- cules that are less well organized than the starting fuel, increasing the entropy of the universe. One effect of all these transformations is (e) in- creased order (decreased randomness) in the system in the form of complex macromolecules. We return to a quantitative treatment of en- tropy in Chapter 13. 8885d_c01_01-46 10/27/03 7:48 AM Page 22 mac76 mac76:385_reb: pheric O 2 to form water, carbon dioxide, and other end products, which are recycled in the environment: C 6 H 12 O 6 H11001 O 2 888n 6CO 2 H11001 6H 2 O H11001 energy (energy-yielding oxidation of glucose) Virtually all energy transductions in cells can be traced to this flow of electrons from one molecule to another, in a “downhill” flow from higher to lower electrochem- ical potential; as such, this is formally analogous to the flow of electrons in a battery-driven electric circuit. All these reactions involving electron flow are oxidation- reduction reactions: one reactant is oxidized (loses electrons) as another is reduced (gains electrons). Creating and Maintaining Order Requires Work and Energy DNA, RNA, and proteins are informational macromole- cules. In addition to using chemical energy to form the covalent bonds between the subunits in these polymers, the cell must invest energy to order the subunits in their correct sequence. It is extremely improbable that amino acids in a mixture would spontaneously condense into a single type of protein, with a unique sequence. This would represent increased order in a population of molecules; but according to the second law of thermodynamics, the tendency in nature is toward ever-greater disorder in the universe: the total entropy of the universe is continu- ally increasing. To bring about the synthesis of macro- molecules from their monomeric units, free energy must be supplied to the system (in this case, the cell). The randomness or disorder of the components of a chemical system is expressed as entropy, S (Box 1–3). Any change in randomness of the system is expressed as entropy change, H9004S, which by convention has a positive value when randomness increases. J. Willard Gibbs, who devel- oped the theory of energy changes during chemical reac- tions, showed that the free- energy content, G, of any closed system can be defined in terms of three quantities: enthalpy, H, reflecting the number and kinds of bonds; entropy, S; and the absolute temperature, T (in degrees Kelvin). The definition of free energy is G H11005 H H11002 TS. When a chemical reaction occurs at constant tempera- ture, the free-energy change, H9004G, is determined by the enthalpy change, H9004H, reflecting the kinds and num- bers of chemical bonds and noncovalent interactions broken and formed, and the entropy change, H9004S, de- scribing the change in the system’s randomness: H9004G H11005H9004H H11002 T H9004S A process tends to occur spontaneously only if H9004G is negative. Yet cell function depends largely on mole- cules, such as proteins and nucleic acids, for which the free energy of formation is positive: the molecules are less stable and more highly ordered than a mixture of their monomeric components. To carry out these ther- modynamically unfavorable, energy-requiring (ender- gonic) reactions, cells couple them to other reactions that liberate free energy (exergonic reactions), so that the overall process is exergonic: the sum of the free- energy changes is negative. The usual source of free energy in coupled biological reactions is the energy re- leased by hydrolysis of phosphoanhydride bonds such as those in adenosine triphosphate (ATP; Fig. 1–25; see also Fig. 1–15). Here, each PH22071 represents a phosphoryl group: Amino acids 888n polymer H9004G 1 is positive (endergonic) O PH22071O PH22071 888n O PH22071 H11001 PH22071 H9004G 2 is negative (exergonic) When these reactions are coupled, the sum of H9004G 1 and H9004G 2 is negative—the overall process is exergonic. By this coupling strategy, cells are able to synthesize and maintain the information-rich polymers essential to life. Energy Coupling Links Reactions in Biology The central issue in bioenergetics (the study of energy transformations in living systems) is the means by which energy from fuel metabolism or light capture is coupled to a cell’s energy-requiring reactions. In thinking about energy coupling, it is useful to consider a simple me- chanical example, as shown in Figure 1–26a. An object at the top of an inclined plane has a certain amount of potential energy as a result of its elevation. It tends to slide down the plane, losing its potential energy of po- sition as it approaches the ground. When an appropri- ate string-and-pulley device couples the falling object to another, smaller object, the spontaneous downward mo- tion of the larger can lift the smaller, accomplishing a 1.3 Physical Foundations 23 J. Willard Gibbs, 1839–1903 H11002 OP O H11002 O OP O H11002 O OP O H11002 O OCH 2 OH O H OH H HH P P P Ribose Adenine HC CH NH 2 C N C N N N FIGURE 1–25 Adenosine triphosphate (ATP). The removal of the ter- minal phosphoryl group (shaded pink) of ATP, by breakage of a phos- phoanhydride bond, is highly exergonic, and this reaction is coupled to many endergonic reactions in the cell (as in the example in Fig. 1–26b). 8885d_c01_023 1/15/04 3:30 PM Page 23 mac76 mac76:385_reb: certain amount of work. The amount of energy available to do work is the free-energy change, H9004G; this is al- ways somewhat less than the theoretical amount of en- ergy released, because some energy is dissipated as the heat of friction. The greater the elevation of the larger object, the greater the energy released (H9004G) as the ob- ject slides downward and the greater the amount of work that can be accomplished. How does this apply in chemical reactions? In closed systems, chemical reactions proceed spontaneously un- til equilibrium is reached. When a system is at equilib- rium, the rate of product formation exactly equals the rate at which product is converted to reactant. Thus there is no net change in the concentration of reactants and products; a steady state is achieved. The energy change as the system moves from its initial state to equi- librium, with no changes in temperature or pressure, is given by the free-energy change, H9004G. The magnitude of H9004G depends on the particular chemical reaction and on how far from equilibrium the system is initially. Each compound involved in a chemical reaction contains a cer- tain amount of potential energy, related to the kind and number of its bonds. In reactions that occur sponta- neously, the products have less free energy than the re- Chapter 1 The Foundations of Biochemistry24 BOX 1–3 WORKING IN BIOCHEMISTRY Entropy: The Advantages of Being Disorganized The term “entropy,” which literally means “a change within,” was first used in 1851 by Rudolf Clausius, one of the formulators of the second law of thermody- namics. A rigorous quantitative definition of entropy involves statistical and probability considerations. However, its nature can be illustrated qualitatively by three simple examples, each demonstrating one aspect of entropy. The key descriptors of entropy are ran- domness and disorder, manifested in different ways. Case 1: The Teakettle and the Randomization of Heat We know that steam generated from boiling water can do useful work. But suppose we turn off the burner under a teakettle full of water at 100 H11034C (the “sys- tem”) in the kitchen (the “surroundings”) and allow the teakettle to cool. As it cools, no work is done, but heat passes from the teakettle to the surroundings, raising the temperature of the surroundings (the kitchen) by an infinitesimally small amount until com- plete equilibrium is attained. At this point all parts of the teakettle and the kitchen are at precisely the same temperature. The free energy that was once concen- trated in the teakettle of hot water at 100 H11034C, poten- tially capable of doing work, has disappeared. Its equivalent in heat energy is still present in the teaket- tle H11001 kitchen (i.e., the “universe”) but has become completely randomized throughout. This energy is no longer available to do work because there is no tem- perature differential within the kitchen. Moreover, the increase in entropy of the kitchen (the surroundings) is irreversible. We know from everyday experience that heat never spontaneously passes back from the kitchen into the teakettle to raise the temperature of the water to 100 H11034C again. Case 2: The Oxidation of Glucose Entropy is a state not only of energy but of matter. Aerobic (heterotrophic) organisms extract free en- ergy from glucose obtained from their surroundings by oxidizing the glucose with O 2 , also obtained from the surroundings. The end products of this oxidative metabolism, CO 2 and H 2 O, are returned to the sur- roundings. In this process the surroundings undergo an increase in entropy, whereas the organism itself re- mains in a steady state and undergoes no change in its internal order. Although some entropy arises from the dissipation of heat, entropy also arises from an- other kind of disorder, illustrated by the equation for the oxidation of glucose: C 6 H 12 O 6 H11001 6O 2 O n 6CO 2 H11001 6H 2 O We can represent this schematically as The atoms contained in 1 molecule of glucose plus 6 molecules of oxygen, a total of 7 molecules, are more randomly dispersed by the oxidation reaction and are now present in a total of 12 molecules (6CO 2 H11001 6H 2 O). Whenever a chemical reaction results in an in- crease in the number of molecules—or when a solid substance is converted into liquid or gaseous products, which allow more freedom of molecular movement than solids—molecular disorder, and thus entropy, increases. Case 3: Information and Entropy The following short passage from Julius Caesar, Act IV, Scene 3, is spoken by Brutus, when he realizes that he must face Mark Antony’s army. It is an information- rich nonrandom arrangement of 125 letters of the English alphabet: 7 molecules CO 2 (a gas) H 2 O (a liquid) Glucose (a solid) O 2 (a gas) 12 molecules 8885d_c01_01-46 10/27/03 7:48 AM Page 24 mac76 mac76:385_reb: actants, thus the reaction releases free energy, which is then available to do work. Such reactions are exergonic; the decline in free energy from reactants to products is expressed as a negative value. Endergonic reactions re- quire an input of energy, and their H9004G values are posi- tive. As in mechanical processes, only part of the energy released in exergonic chemical reactions can be used to accomplish work. In living systems some energy is dissi- pated as heat or lost to increasing entropy. In living organisms, as in the mechanical example in Figure 1–26a, an exergonic reaction can be coupled to an endergonic reaction to drive otherwise unfavorable reactions. Figure 1–26b (a type of graph called a reac- tion coordinate diagram) illustrates this principle for the conversion of glucose to glucose 6-phosphate, the first step in the pathway for oxidation of glucose. The sim- plest way to produce glucose 6-phosphate would be: Reaction 1: Glucose H11001 P i O n glucose 6-phosphate (endergonic; H9004G 1 is positive) (P i is an abbreviation for inorganic phosphate, HPO 4 2H11002 . Don’t be concerned about the structure of these com- pounds now; we describe them in detail later in the book.) This reaction does not occur spontaneously; H9004G 1.3 Physical Foundations 25 There is a tide in the affairs of men, Which, taken at the flood, leads on to fortune; Omitted, all the voyage of their life Is bound in shallows and in miseries. In addition to what this passage says overtly, it has many hidden meanings. It not only reflects a complex sequence of events in the play, it also echoes the play’s ideas on conflict, ambition, and the demands of lead- ership. Permeated with Shakespeare’s understanding of human nature, it is very rich in information. However, if the 125 letters making up this quota- tion were allowed to fall into a completely random, chaotic pattern, as shown in the following box, they would have no meaning whatsoever. In this form the 125 letters contain little or no infor- mation, but they are very rich in entropy. Such con- siderations have led to the conclusion that information is a form of energy; information has been called “neg- ative entropy.” In fact, the branch of mathematics called information theory, which is basic to the programming logic of computers, is closely related to thermodynamic theory. Living organisms are highly ordered, nonran- dom structures, immensely rich in information and thus entropy-poor. a b c d e f g h i I k l m n o O r a d e f h i l m n o r a d e f h i l m n o r a d e f h i l n o r a d e f h i l n o r a d e f h i l n o a e f h i l n o a e h i n o a e i n o a e i o e i e i e e e e s t T u v w W y u s t s t s t s t s t s t s t t t t Work done raising object Loss of potential energy of position ?G > 0 ?G < 0 (b) Chemical example (a) Mechanical example ExergonicEndergonic Free energy, G Reaction 1: Glucose H11001 P i → glucose 6-phosphate Reaction 2: ATP → ADP H11001 P i Reaction 3: Glucose H11001 ATP → glucose 6-phosphate H11001 ADP ?G 1 ?G 2 ?G 3 ?G 3 = ?G 1 H11001 ?G 2 Reaction coordinate FIGURE 1–26 Energy coupling in mechanical and chemical processes. (a) The downward motion of an object releases potential energy that can do mechanical work. The potential energy made avail- able by spontaneous downward motion, an exergonic process (pink), can be coupled to the endergonic upward movement of another ob- ject (blue). (b) In reaction 1, the formation of glucose 6-phosphate from glucose and inorganic phosphate (P i ) yields a product of higher energy than the two reactants. For this endergonic reaction, H9004G is pos- itive. In reaction 2, the exergonic breakdown of adenosine triphos- phate (ATP) can drive an endergonic reaction when the two reactions are coupled. The exergonic reaction has a large, negative free-energy change (H9004G 2 ), and the endergonic reaction has a smaller, positive free- energy change (H9004G 1 ). The third reaction accomplishes the sum of re- actions 1 and 2, and the free-energy change, H9004G 3 , is the arithmetic sum of H9004G 1 and H9004G 2 . Because H9004G 3 is negative, the overall reaction is exergonic and proceeds spontaneously. 8885d_c01_025 1/15/04 3:30 PM Page 25 mac76 mac76:385_reb: is positive. A second, very exergonic reaction can occur in all cells: Reaction 2: ATP On ADP H11001 P i (exergonic; H9004G 2 is negative) These two chemical reactions share a common inter- mediate, P i , which is consumed in reaction 1 and pro- duced in reaction 2. The two reactions can therefore be coupled in the form of a third reaction, which we can write as the sum of reactions 1 and 2, with the common intermediate, P i , omitted from both sides of the equation: Reaction 3: Glucose H11001 ATP On glucose 6-phosphate H11001 ADP Because more energy is released in reaction 2 than is consumed in reaction 1, the free-energy change for re- action 3, H9004G 3 , is negative, and the synthesis of glucose 6-phosphate can therefore occur by reaction 3. The coupling of exergonic and endergonic reactions through a shared intermediate is absolutely central to the energy exchanges in living systems. As we shall see, the breakdown of ATP (reaction 2 in Fig. 1–26b) is the ex- ergonic reaction that drives many endergonic processes in cells. In fact, ATP is the major carrier of chemical energy in all cells. K eq and H9004GH11543 Are Measures of a Reaction’s Tendency to Proceed Spontaneously The tendency of a chemical reaction to go to completion can be expressed as an equilibrium constant. For the reaction aA H11001 bB 888n cC H11001 dD the equilibrium constant, K eq , is given by K eq H11005 H5007 [ [ A C e e q q ] ] a c [ [ D B e e q q ] ] b d H5007 where [A eq ] is the concentration of A, [B eq ] the concen- tration of B, and so on, when the system has reached equilibrium. A large value of K eq means the reaction tends to proceed until the reactants have been almost completely converted into the products. Gibbs showed that H9004G for any chemical reaction is a function of the standard free-energy change, H9004GH11034— a constant that is characteristic of each specific reaction—and a term that expresses the initial concen- trations of reactants and products: H9004G H11005 H9004GH11034H11001RT ln H5007 [ [ A C i i ] ] a c [ [ D B i i ] ] d b H5007 (1–1) where [A i ] is the initial concentration of A, and so forth; R is the gas constant; and T is the absolute temperature. When a reaction has reached equilibrium, no driv- ing force remains and it can do no work: H9004G H11005 0. For this special case, [A i ] H11005 [A eq ], and so on, for all reactants and products, and H11005 H5007 [ [ A C e e q q ] ] a c [ [ D B e e q q ] ] d b H5007 H11005 K eq Substituting 0 for H9004G and K eq for [C i ] c [D i ] d /[A i ] a [B i ] b in Equation 1–1, we obtain the relationship H9004GH11034H11005H11002RT ln K eq from which we see that H9004GH11034 is simply a second way (be- sides K eq ) of expressing the driving force on a reaction. Because K eq is experimentally measurable, we have a way of determining H9004GH11034, the thermodynamic constant characteristic of each reaction. The units of H9004GH11034 and H9004G are joules per mole (or calories per mole). When K eq H11022H11022 1, H9004GH11034 is large and negative; when K eq H11021H11021 1, H9004GH11034 is large and positive. From a table of experimentally determined values of ei- ther K eq or H9004GH11034, we can see at a glance which reactions tend to go to completion and which do not. One caution about the interpretation of H9004GH11034: ther- modynamic constants such as this show where the fi- nal equilibrium for a reaction lies but tell us nothing about how fast that equilibrium will be achieved. The rates of reactions are governed by the parameters of ki- netics, a topic we consider in detail in Chapter 6. Enzymes Promote Sequences of Chemical Reactions All biological macromolecules are much less thermody- namically stable than their monomeric subunits, yet they are kinetically stable: their uncatalyzed break- down occurs so slowly (over years rather than seconds) that, on a time scale that matters for the organism, these molecules are stable. Virtually every chemical reaction in a cell occurs at a significant rate only because of the presence of enzymes—biocatalysts that, like all other catalysts, greatly enhance the rate of specific chemical reactions without being consumed in the process. The path from reactant(s) to product(s) almost in- variably involves an energy barrier, called the activation barrier (Fig. 1–27), that must be surmounted for any re- action to proceed. The breaking of existing bonds and formation of new ones generally requires, first, the dis- tortion of the existing bonds, creating a transition state of higher free energy than either reactant or prod- uct. The highest point in the reaction coordinate dia- gram represents the transition state, and the difference in energy between the reactant in its ground state and in its transition state is the activation energy, H9004G ? . An enzyme catalyzes a reaction by providing a more comfortable fit for the transition state: a surface that complements the transition state in stereochemistry, po- larity, and charge. The binding of enzyme to the transi- tion state is exergonic, and the energy released by this binding reduces the activation energy for the reaction and greatly increases the reaction rate. A further contribution to catalysis occurs when two or more reactants bind to the enzyme’s surface close to each other and with stereospecific orientations that fa- [C i ] c [D i ] d H5007H5007 [A i ] a [B i ] b Chapter 1 The Foundations of Biochemistry26 8885d_c01_026 11/3/03 2:42 PM Page 26 mac76 mac76:385_reb: vor the reaction. This increases by orders of magnitude the probability of productive collisions between reac- tants. As a result of these factors and several others, discussed in Chapter 6, enzyme-catalyzed reactions commonly proceed at rates greater than 10 12 times faster than the uncatalyzed reactions. Cellular catalysts are, with a few exceptions, pro- teins. (In some cases, RNA molecules have catalytic roles, as discussed in Chapters 26 and 27.) Again with a few exceptions, each enzyme catalyzes a specific reaction, and each reaction in a cell is catalyzed by a different enzyme. Thousands of different enzymes are therefore required by each cell. The multiplicity of en- zymes, their specificity (the ability to discriminate between reactants), and their susceptibility to regula- tion give cells the capacity to lower activation barriers selectively. This selectivity is crucial for the effective regulation of cellular processes. By allowing specific re- actions to proceed at significant rates at particular times, enzymes determine how matter and energy are channeled into cellular activities. The thousands of enzyme-catalyzed chemical reac- tions in cells are functionally organized into many se- quences of consecutive reactions, called pathways, in which the product of one reaction becomes the reactant in the next. Some pathways degrade organic nutrients into simple end products in order to extract chemical energy and convert it into a form useful to the cell; to- gether these degradative, free-energy-yielding reactions are designated catabolism. Other pathways start with small precursor molecules and convert them to pro- gressively larger and more complex molecules, includ- ing proteins and nucleic acids. Such synthetic pathways, which invariably require the input of energy, are col- lectively designated anabolism. The overall network of enzyme-catalyzed pathways constitutes cellular me- tabolism. ATP is the major connecting link (the shared intermediate) between the catabolic and anabolic com- ponents of this network (shown schematically in Fig. 1–28). The pathways of enzyme-catalyzed reactions that act on the main constituents of cells—proteins, fats, sugars, and nucleic acids—are virtually identical in all living organisms. Metabolism Is Regulated to Achieve Balance and Economy Not only do living cells simultaneously synthesize thou- sands of different kinds of carbohydrate, fat, protein, and nucleic acid molecules and their simpler subunits, but they do so in the precise proportions required by 1.3 Physical Foundations 27 Activation barrier (transition state, ?) Free energy, G Reactants (A) H9004G ? cat H9004G ? uncat Products (B) H9004G B)Reaction coordinate (A FIGURE 1–27 Energy changes during a chemical reaction. An acti- vation barrier, representing the transition state, must be overcome in the conversion of reactants (A) into products (B), even though the prod- ucts are more stable than the reactants, as indicated by a large, neg- ative free-energy change (H9004G). The energy required to overcome the activation barrier is the activation energy (H9004G ? ). Enzymes catalyze re- actions by lowering the activation barrier. They bind the transition- state intermediates tightly, and the binding energy of this interaction effectively reduces the activation energy from H9004G ? uncat to H9004G ? cat . (Note that activation energy is not related to free-energy change, H9004G.) H11001 Osmotic work Stored nutrients Ingested foods Solar photons Other cellular work Complex biomolecules Mechanical work H 2 O NH 3 CO 2 ADP HPO 4 2H11002 S i m p l e p ro ducts, p r e c u r s o r s Catabolic reaction pathways (exergonic) Anabolic reaction pathways (endergonic) ATP FIGURE 1–28 The central role of ATP in metabolism. ATP is the shared chemical intermediate linking energy-releasing to energy- requiring cell processes. Its role in the cell is analogous to that of money in an economy: it is “earned/produced” in exergonic reactions and “spent/consumed” in endergonic ones. 8885d_c01_027 12/20/03 7:08 AM Page 27 mac76 mac76:385_reb: the cell under any given circumstance. For example, during rapid cell growth the precursors of proteins and nucleic acids must be made in large quantities, whereas in nongrowing cells the requirement for these precur- sors is much reduced. Key enzymes in each metabolic pathway are regulated so that each type of precursor molecule is produced in a quantity appropriate to the current requirements of the cell. Consider the pathway in E. coli that leads to the synthesis of the amino acid isoleucine, a constituent of proteins. The pathway has five steps catalyzed by five different enzymes (A through F represent the interme- diates in the pathway): If a cell begins to produce more isoleucine than is needed for protein synthesis, the unused isoleucine ac- cumulates and the increased concentration inhibits the catalytic activity of the first enzyme in the pathway, im- mediately slowing the production of isoleucine. Such feedback inhibition keeps the production and utiliza- tion of each metabolic intermediate in balance. Although the concept of discrete pathways is an im- portant tool for organizing our understanding of metab- olism, it is an oversimplification. There are thousands of metabolic intermediates in a cell, many of which are part of more than one pathway. Metabolism would be better represented as a meshwork of interconnected and in- terdependent pathways. A change in the concentration of any one metabolite would have an impact on other pathways, starting a ripple effect that would influence the flow of materials through other sectors of the cellu- lar economy. The task of understanding these complex interactions among intermediates and pathways in quan- titative terms is daunting, but new approaches, discussed in Chapter 15, have begun to offer important insights into the overall regulation of metabolism. Cells also regulate the synthesis of their own cata- lysts, the enzymes, in response to increased or dimin- ished need for a metabolic product; this is the substance of Chapter 28. The expression of genes (the translation of information in DNA to active protein in the cell) and synthesis of enzymes are other layers of metabolic con- trol in the cell. All layers must be taken into account when describing the overall control of cellular metabo- lism. Assembling the complete picture of how the cell regulates itself is a huge job that has only just begun. SUMMARY 1.3 Physical Foundations ■ Living cells are open systems, exchanging matter and energy with their surroundings, extracting and channeling energy to maintain A 1 BCDEF Threonine Isoleucine enzyme themselves in a dynamic steady state distant from equilibrium. Energy is obtained from sunlight or fuels by converting the energy from electron flow into the chemical bonds of ATP. ■ The tendency for a chemical reaction to proceed toward equilibrium can be expressed as the free-energy change, H9004G, which has two components: enthalpy change, H9004H, and entropy change, H9004S. These variables are related by the equation H9004G H11005H9004H H11002 T H9004S. ■ When H9004G of a reaction is negative, the reaction is exergonic and tends to go toward completion; when H9004G is positive, the reaction is endergonic and tends to go in the reverse direction. When two reactions can be summed to yield a third reaction, the H9004G for this overall reaction is the sum of the H9004Gs of the two separate reactions. This provides a way to couple reactions. ■ The conversion of ATP to P i and ADP is highly exergonic (large negative H9004G), and many endergonic cellular reactions are driven by coupling them, through a common intermediate, to this reaction. ■ The standard free-energy change for a reaction, H9004GH11034, is a physical constant that is related to the equilibrium constant by the equation H9004GH11034H11005H11002RT ln K eq . ■ Most exergonic cellular reactions proceed at useful rates only because enzymes are present to catalyze them. Enzymes act in part by stabilizing the transition state, reducing the activation energy, H9004G ? , and increasing the reaction rate by many orders of magnitude. The catalytic activity of enzymes in cells is regulated. ■ Metabolism is the sum of many interconnected reaction sequences that interconvert cellular metabolites. Each sequence is regulated so as to provide what the cell needs at a given time and to expend energy only when necessary. 1.4 Genetic Foundations Perhaps the most remarkable property of living cells and organisms is their ability to reproduce themselves for countless generations with nearly perfect fidelity. This continuity of inherited traits implies constancy, over mil- lions of years, in the structure of the molecules that con- tain the genetic information. Very few historical records of civilization, even those etched in copper or carved in stone (Fig. 1–29), have survived for a thousand years. But there is good evidence that the genetic instructions in living organisms have remained nearly unchanged over very much longer periods; many bacteria have nearly Chapter 1 The Foundations of Biochemistry28 8885d_c01_01-46 10/27/03 7:48 AM Page 28 mac76 mac76:385_reb: the same size, shape, and internal structure and contain the same kinds of precursor molecules and enzymes as bacteria that lived nearly four billion years ago. Among the seminal discoveries in biology in the twentieth century were the chemical nature and the three-dimensional structure of the genetic material, deoxyribonucleic acid, DNA. The sequence of the monomeric subunits, the nucleotides (strictly, deoxyri- bonucleotides, as discussed below), in this linear poly- mer encodes the instructions for forming all other cellular components and provides a template for the production of identical DNA molecules to be distributed to progeny when a cell divides. The continued existence of a biological species requires its genetic information to be maintained in a stable form, expressed accurately in the form of gene products, and reproduced with a minimum of errors. Effective storage, expression, and reproduction of the genetic message defines individual species, distinguishes them from one another, and as- sures their continuity over successive generations. Genetic Continuity Is Vested in Single DNA Molecules DNA is a long, thin organic polymer, the rare molecule that is constructed on the atomic scale in one dimen- sion (width) and the human scale in another (length: a molecule of DNA can be many centimeters long). A hu- man sperm or egg, carrying the accumulated hereditary information of billions of years of evolution, transmits this inheritance in the form of DNA molecules, in which the linear sequence of covalently linked nucleotide sub- units encodes the genetic message. Usually when we describe the properties of a chem- ical species, we describe the average behavior of a very large number of identical molecules. While it is difficult to predict the behavior of any single molecule in a col- lection of, say, a picomole (about 6 H11003 10 11 molecules) of a compound, the average behavior of the molecules is predictable because so many molecules enter into the average. Cellular DNA is a remarkable exception. The DNA that is the entire genetic material of E. coli is a single molecule containing 4.64 million nucleotide pairs. That single molecule must be replicated perfectly in every detail if an E. coli cell is to give rise to identi- cal progeny by cell division; there is no room for aver- aging in this process! The same is true for all cells. A human sperm brings to the egg that it fertilizes just one molecule of DNA in each of its 23 different chromo- somes, to combine with just one DNA molecule in each corresponding chromosome in the egg. The result of this union is very highly predictable: an embryo with all of its 35,000 genes, constructed of 3 billion nucleotide pairs, intact. An amazing chemical feat! The Structure of DNA Allows for Its Replication and Repair with Near-Perfect Fidelity The capacity of living cells to preserve their genetic ma- terial and to duplicate it for the next generation results from the structural complementarity between the two halves of the DNA molecule (Fig. 1–30). The basic unit of DNA is a linear polymer of four different monomeric subunits, deoxyribonucleotides, arranged in a precise linear sequence. It is this linear sequence that encodes the genetic information. Two of these polymeric strands are twisted about each other to form the DNA double helix, in which each deoxyribonucleotide in one strand pairs specifically with a complementary deoxyribonu- cleotide in the opposite strand. Before a cell divides, the two DNA strands separate and each serves as a template for the synthesis of a new complementary strand, gen- erating two identical double-helical molecules, one for each daughter cell. If one strand is damaged, continu- ity of information is assured by the information present in the other strand, which acts as a template for repair of the damage. The Linear Sequence in DNA Encodes Proteins with Three-Dimensional Structures The information in DNA is encoded in its linear (one- dimensional) sequence of deoxyribonucleotide sub- units, but the expression of this information results in 1.4 Genetic Foundations 29 (a) (b) FIGURE 1–29 Two ancient scripts. (a) The Prism of Sennacherib, in- scribed in about 700 B.C.E., describes in characters of the Assyrian lan- guage some historical events during the reign of King Sennacherib. The Prism contains about 20,000 characters, weighs about 50 kg, and has survived almost intact for about 2,700 years. (b) The single DNA molecule of the bacterium E. coli, seen leaking out of a disrupted cell, is hundreds of times longer than the cell itself and contains all the encoded information necessary to specify the cell’s structure and func- tions. The bacterial DNA contains about 10 million characters (nu- cleotides), weighs less than 10 H1100210 g, and has undergone only relatively minor changes during the past several million years. (The yellow spots and dark specks in this colorized electron micrograph are artifacts of the preparation.) 8885d_c01_029 12/30/03 6:34 AM Page 29 mac76 mac76:385_reb: a three-dimensional cell. This change from one to three dimensions occurs in two phases. A linear sequence of deoxyribonucleotides in DNA codes (through an inter- mediary, RNA) for the production of a protein with a corresponding linear sequence of amino acids (Fig. 1–31). The protein folds into a particular three-dimensional shape, determined by its amino acid sequence and sta- bilized primarily by noncovalent interactions. Although the final shape of the folded protein is dictated by its amino acid sequence, the folding process is aided by “molecular chaperones,” which catalyze the process by discouraging incorrect folding. The precise three- dimensional structure, or native conformation, of the protein is crucial to its function. Once in its native conformation, a protein may as- sociate noncovalently with other proteins, or with nu- cleic acids or lipids, to form supramolecular complexes such as chromosomes, ribosomes, and membranes. The individual molecules of these complexes have specific, high-affinity binding sites for each other, and within the cell they spontaneously form functional complexes. Chapter 1 The Foundations of Biochemistry30 New strand 1 Old strand 2 New strand 2 Old strand 1 C T T T T T T T T T T G G G G G G G G G G G G G C C C C C C C C C C C C C A A A A A A G Strand 2 Strand 1 Gene 1 RNA 1 RNA 2 RNA 3 Gene 2 Transcription of DNA sequence into RNA sequence Translation (on the ribosome) of RNA sequence into protein sequence and folding of protein into native conformation Formation of supramolecular complex Gene 3 Protein 1 Protein 2 Protein 3 FIGURE 1–30 Complementarity between the two strands of DNA. DNA is a linear polymer of covalently joined deoxyribonucleotides, of four types: deoxyadenylate (A), deoxyguanylate (G), deoxycytidy- late (C), and deoxythymidylate (T). Each nucleotide, with its unique three-dimensional structure, can associate very specifically but non- covalently with one other nucleotide in the complementary chain: A always associates with T, and G with C. Thus, in the double-stranded DNA molecule, the entire sequence of nucleotides in one strand is complementary to the sequence in the other. The two strands, held together by hydrogen bonds (represented here by vertical blue lines) between each pair of complementary nucleotides, twist about each other to form the DNA double helix. In DNA replication, the two strands separate and two new strands are synthesized, each with a se- quence complementary to one of the original strands. The result is two double-helical molecules, each identical to the original DNA. FIGURE 1–31 DNA to RNA to protein. Linear sequences of deoxyri- bonucleotides in DNA, arranged into units known as genes, are tran- scribed into ribonucleic acid (RNA) molecules with complementary ribonucleotide sequences. The RNA sequences are then translated into linear protein chains, which fold into their native three-dimensional shapes, often aided by molecular chaperones. Individual proteins com- monly associate with other proteins to form supramolecular com- plexes, stabilized by numerous weak interactions. 8885d_c01_01-46 10/27/03 7:48 AM Page 30 mac76 mac76:385_reb: Although protein sequences carry all necessary in- formation for the folding into their native conformation, this correct folding requires the right environment—pH, ionic strength, metal ion concentrations, and so forth. Self-assembly therefore requires both information (pro- vided by the DNA sequence) and environment (the in- terior of a living cell), and in this sense the DNA sequence alone is not enough to dictate the formation of a cell. As Rudolph Virchow, the nineteenth-century Prussian pathologist and researcher, concluded, “Omnis cellula e cellula”: every cell comes from another cell. SUMMARY 1.4 Genetic Foundations ■ Genetic information is encoded in the linear sequence of four deoxyribonucleotides in DNA. ■ The double-helical DNA molecule contains an internal template for its own replication and repair. ■ The linear sequence of amino acids in a pro- tein, which is encoded in the DNA of the gene for that protein, produces a protein’s unique three-dimensional structure. ■ Individual macromolecules with specific affinity for other macromolecules self-assemble into supramolecular complexes. 1.5 Evolutionary Foundations Nothing in biology makes sense except in the light of evolution. —Theodosius Dobzhansky, The American Biology Teacher, March 1973 Great progress in biochemistry and molecular biology during the decades since Dobzhansky made this striking generalization has amply confirmed its validity. The re- markable similarity of metabolic pathways and gene se- quences in organisms across the phyla argues strongly that all modern organisms share a common evolutionary progenitor and were derived from it by a series of small changes (mutations), each of which conferred a selective advantage to some organism in some ecological niche. Changes in the Hereditary Instructions Allow Evolution Despite the near-perfect fidelity of genetic replication, infrequent, unrepaired mistakes in the DNA replication process lead to changes in the nucleotide sequence of DNA, producing a genetic mutation (Fig. 1–32) and changing the instructions for some cellular component. Incorrectly repaired damage to one of the DNA strands has the same effect. Mutations in the DNA handed down 1.5 Evolutionary Foundations 31 Time A Mutation 2 GA A A A Mutation 1 Mutation 5 Mutation 3 Mutation 4 Mutation 6 T G A G C T A T G A C T A TGA CTAG T G A C G G G A T GA C T A T C C C A C T AG T GGA CTAG T A C A T GA C A TACTGTA CTAG FIGURE 1–32 Role of mutation in evolution. The gradual accumulation of mutations over long periods of time results in new biological species, each with a unique DNA sequence. At the top is shown a short segment of a gene in a hypothetical progenitor organism. With the passage of time, changes in nucleotide sequence (mutations, indicated here by colored boxes), occurring one nucleotide at a time, result in progeny with different DNA sequences. These mutant progeny also undergo occasional mutations, yielding their own progeny that differ by two or more nucleotides from the progenitor sequence. When two lineages have diverged so much in their genetic makeup that they can no longer interbreed, a new species has been created. 8885d_c01_031 12/20/03 7:08 AM Page 31 mac76 mac76:385_reb: to offspring—that is, mutations that are carried in the reproductive cells—may be harmful or even lethal to the organism; they may, for example, cause the synthesis of a defective enzyme that is not able to catalyze an es- sential metabolic reaction. Occasionally, however, a mu- tation better equips an organism or cell to survive in its environment. The mutant enzyme might have acquired a slightly different specificity, for example, so that it is now able to use some compound that the cell was pre- viously unable to metabolize. If a population of cells were to find itself in an environment where that com- pound was the only or the most abundant available source of fuel, the mutant cell would have a selective advantage over the other, unmutated (wild-type) cells in the population. The mutant cell and its progeny would survive and prosper in the new environment, whereas wild-type cells would starve and be eliminated. This is what Darwin meant by “survival of the fittest under se- lective pressure.” Occasionally, a whole gene is duplicated. The sec- ond copy is superfluous, and mutations in this gene will not be deleterious; it becomes a means by which the cell may evolve: by producing a new gene with a new func- tion while retaining the original gene and gene function. Seen in this light, the DNA molecules of modern or- ganisms are historical documents, records of the long journey from the earliest cells to modern organisms. The historical accounts in DNA are not complete; in the course of evolution, many mutations must have been erased or written over. But DNA molecules are the best source of biological history that we have. Several billion years of adaptive selection have re- fined cellular systems to take maximum advantage of the chemical and physical properties of the molecular raw materials for carrying out the basic energy-transforming and self-replicating activities of a living cell. Chance ge- netic variations in individuals in a population, combined with natural selection (survival and reproduction of the fittest individuals in a challenging or changing environ- ment), have resulted in the evolution of an enormous va- riety of organisms, each adapted to life in its particular ecological niche. Biomolecules First Arose by Chemical Evolution In our account thus far we have passed over the first chapter of the story of evolution: the appearance of the first living cell. Apart from their occurrence in living or- ganisms, organic compounds, including the basic bio- molecules such as amino acids and carbohydrates, are found in only trace amounts in the earth’s crust, the sea, and the atmosphere. How did the first living organisms acquire their characteristic organic building blocks? In 1922, the biochemist Aleksandr I. Oparin proposed a theory for the origin of life early in the history of Earth, postulating that the atmosphere was very different from that of today. Rich in methane, ammonia, and water, and essentially devoid of oxygen, it was a reducing atmos- phere, in contrast to the oxidizing environment of our era. In Oparin’s theory, electrical energy from lightning discharges or heat energy from volcanoes caused am- monia, methane, water vapor, and other components of the primitive atmosphere to react, forming simple or- ganic compounds. These compounds then dissolved in the ancient seas, which over many millennia became en- riched with a large variety of simple organic substances. In this warm solution (the “primordial soup”), some or- ganic molecules had a greater tendency than others to associate into larger complexes. Over millions of years, these in turn assembled spontaneously to form mem- branes and catalysts (enzymes), which came together to become precursors of the earliest cells. Oparin’s views remained speculative for many years and appeared untestable—until a surprising experiment was con- ducted using simple equipment on a desktop. Chemical Evolution Can Be Simulated in the Laboratory The classic experiment on the abiotic (nonbiological) origin of organic biomolecules was carried out in 1953 by Stanley Miller in the laboratory of Harold Urey. Miller subjected gaseous mixtures of NH 3 , CH 4 , H 2 O, and H 2 to electrical sparks produced across a pair of electrodes (to simulate lightning) for periods of a week or more, then analyzed the contents of the closed reaction ves- sel (Fig. 1–33). The gas phase of the resulting mixture contained CO and CO 2 , as well as the starting materi- als. The water phase contained a variety of organic com- pounds, including some amino acids, hydroxy acids, aldehydes, and hydrogen cyanide (HCN). This experi- ment established the possibility of abiotic production of biomolecules in relatively short times under relatively mild conditions. More refined laboratory experiments have provided good evidence that many of the chemical components of living cells, including polypeptides and RNA-like mol- ecules, can form under these conditions. Polymers of RNA can act as catalysts in biologically significant re- actions (as we discuss in Chapters 26 and 27), and RNA probably played a crucial role in prebiotic evolution, both as catalyst and as information repository. RNA or Related Precursors May Have Been the First Genes and Catalysts In modern organisms, nucleic acids encode the genetic information that specifies the structure of enzymes, and enzymes catalyze the replication and repair of nucleic acids. The mutual dependence of these two classes of biomolecules brings up the perplexing question: which came first, DNA or protein? The answer may be: neither. The discovery that RNA molecules can act as catalysts in their own forma- Chapter 1 The Foundations of Biochemistry32 8885d_c01_01-46 10/27/03 7:48 AM Page 32 mac76 mac76:385_reb: tion suggests that RNA or a similar molecule may have been the first gene and the first catalyst. According to this scenario (Fig. 1–34), one of the earliest stages of biological evolution was the chance formation, in the pri- mordial soup, of an RNA molecule that could catalyze the formation of other RNA molecules of the same se- quence—a self-replicating, self-perpetuating RNA. The concentration of a self-replicating RNA molecule would increase exponentially, as one molecule formed two, two formed four, and so on. The fidelity of self-replication was presumably less than perfect, so the process would generate variants of the RNA, some of which might be even better able to self-replicate. In the competition for nucleotides, the most efficient of the self-replicating se- quences would win, and less efficient replicators would fade from the population. The division of function between DNA (genetic information storage) and protein (catalysis) was, ac- cording to the “RNA world” hypothesis, a later devel- opment. New variants of self-replicating RNA molecules developed, with the additional ability to catalyze the condensation of amino acids into peptides. Occasion- ally, the peptide(s) thus formed would reinforce the self-replicating ability of the RNA, and the pair—RNA molecule and helping peptide—could undergo further modifications in sequence, generating even more effi- cient self-replicating systems. The recent, remarkable discovery that, in the protein-synthesizing machinery of modern cells (ribosomes), RNA molecules, not pro- teins, catalyze the formation of peptide bonds is cer- tainly consistent with the RNA world hypothesis. Some time after the evolution of this primitive protein-synthesizing system, there was a further devel- opment: DNA molecules with sequences complementary to the self-replicating RNA molecules took over the func- tion of conserving the “genetic” information, and RNA molecules evolved to play roles in protein synthesis. (We explain in Chapter 8 why DNA is a more stable molecule than RNA and thus a better repository of inheritable in- formation.) Proteins proved to be versatile catalysts and, over time, took over that function. Lipidlike compounds in the primordial soup formed relatively impermeable layers around self-replicating collections of molecules. The concentration of proteins and nucleic acids within these lipid enclosures favored the molecular interactions required in self-replication. 1.5 Evolutionary Foundations 33 Electrodes Condenser Spark gap Mixture of NH 3 , CH 4 , H 2 , and H 2 O at 80 °C FIGURE 1–33 Abiotic production of biomolecules. Spark-discharge apparatus of the type used by Miller and Urey in experiments demon- strating abiotic formation of organic compounds under primitive at- mospheric conditions. After subjection of the gaseous contents of the system to electrical sparks, products were collected by condensation. Biomolecules such as amino acids were among the products. Creation of prebiotic soup, including nucleotides, from components of Earth’s primitive atmosphere Production of short RNA molecules with random sequences Selective replication of self-duplicating catalytic RNA segments Synthesis of specific peptides, catalyzed by RNA Increasing role of peptides in RNA replication; coevolution of RNA and protein Primitive translation system develops, with RNA genome and RNA-protein catalysts Genomic RNA begins to be copied into DNA DNA genome, translated on RNA-protein complex (ribosome) with protein catalysts FIGURE 1–34 A possible “RNA world” scenario. 8885d_c01_033 12/20/03 7:09 AM Page 33 mac76 mac76:385_reb: Biological Evolution Began More Than Three and a Half Billion Years Ago Earth was formed about 4.5 billion years ago, and the first evidence of life dates to more than 3.5 billion years ago. In 1996, scientists working in Greenland found not fossil remains but chemical evidence of life from as far back as 3.85 billion years ago, forms of carbon em- bedded in rock that appear to have a distinctly bio- logical origin. Somewhere on Earth during its first billion years there arose the first simple organism, capable of replicating its own structure from a tem- plate (RNA?) that was the first genetic material. Be- cause the terrestrial atmosphere at the dawn of life was nearly devoid of oxygen, and because there were few microorganisms to scavenge organic compounds formed by natural processes, these compounds were relatively stable. Given this stability and eons of time, the improbable became inevitable: the organic com- pounds were incorporated into evolving cells to pro- duce increasingly effective self-reproducing catalysts. The process of biological evolution had begun. The First Cell Was Probably a Chemoheterotroph The earliest cells that arose in the rich mixture of or- ganic compounds, the primordial soup of prebiotic times, were almost certainly chemoheterotrophs (Fig. 1–5). The organic compounds they required were originally synthesized from components of the early atmosphere— CO, CO 2 , N 2 , CH 4 , and such—by the nonbiological ac- tions of volcanic heat and lightning. Early heterotrophs gradually acquired the ability to derive energy from compounds in their environment and to use that energy to synthesize more of their own precursor molecules, thereby becoming less dependent on outside sources. A very significant evolutionary event was the development of pigments capable of capturing the energy of light from the sun, which could be used to reduce, or “fix,” CO 2 to form more complex, organic compounds. The original electron donor for these photosynthetic processes was probably H 2 S, yielding elemental sulfur or sulfate (SO 4 2H11002 ) as the by-product, but later cells developed the enzymatic capacity to use H 2 O as the electron donor in photosynthetic reactions, eliminating O 2 as waste. Cyanobacteria are the modern descendants of these early photosynthetic oxygen-producers. Because the atmosphere of Earth in the earliest stages of biological evolution was nearly devoid of oxy- gen, the earliest cells were anaerobic. Under these conditions, chemoheterotrophs could oxidize organic compounds to CO 2 by passing electrons not to O 2 but to acceptors such as SO 4 2H11002 , yielding H 2 S as the product. With the rise of O 2 -producing photosynthetic bacteria, the atmosphere became progressively richer in oxy- gen—a powerful oxidant and deadly poison to anaer- obes. Responding to the evolutionary pressure of the “oxygen holocaust,” some lineages of microorganisms gave rise to aerobes that obtained energy by passing electrons from fuel molecules to oxygen. Because the transfer of electrons from organic molecules to O 2 re- leases a great deal of energy, aerobic organisms had an energetic advantage over their anaerobic counterparts when both competed in an environment containing oxy- gen. This advantage translated into the predominance of aerobic organisms in O 2 -rich environments. Modern bacteria inhabit almost every ecological niche in the biosphere, and there are bacteria capable of using virtually every type of organic compound as a source of carbon and energy. Photosynthetic bacteria in both fresh and marine waters trap solar energy and use it to generate carbohydrates and all other cell con- stituents, which are in turn used as food by other forms of life. The process of evolution continues—and in rap- idly reproducing bacterial cells, on a time scale that al- lows us to witness it in the laboratory. Eukaryotic Cells Evolved from Prokaryotes in Several Stages Starting about 1.5 billion years ago, the fossil record be- gins to show evidence of larger and more complex or- ganisms, probably the earliest eukaryotic cells (Fig. 1–35). Chapter 1 The Foundations of Biochemistry34 0 4,500 Formation of Earth 4,000 Formation of oceans and continents 3,500 Appearance of photosynthetic O 2 -producing cyanobacteria Appearance of photosynthetic sulfur bacteria Appearance of methanogens 2,500 Appearance of aerobic bacteria Development of O 2 -rich atmosphere 1,500 Appearance of protists, the first eukaryotes Appearance of red and green algae 1,000 Appearance of endosymbionts (mitochondria, plastids) 500 Diversification of multicellular eukaryotes (plants, fungi, animals) 3,000 2,000 Millions of years ago FIGURE 1–35 Landmarks in the evolution of life on Earth. 8885d_c01_034 11/3/03 1:44 PM Page 34 mac76 mac76:385_reb: Details of the evolutionary path from prokaryotes to eu- karyotes cannot be deduced from the fossil record alone, but morphological and biochemical comparisons of mod- ern organisms have suggested a sequence of events con- sistent with the fossil evidence. Three major changes must have occurred as prokaryotes gave rise to eukaryotes. First, as cells ac- quired more DNA, the mechanisms required to fold it compactly into discrete complexes with specific pro- teins and to divide it equally between daughter cells at cell division became more elaborate. For this, special- ized proteins were required to stabilize folded DNA and to pull the resulting DNA-protein complexes (chro- mosomes) apart during cell division. Second, as cells became larger, a system of intracellular membranes de- veloped, including a double membrane surrounding the DNA. This membrane segregated the nuclear process of RNA synthesis on a DNA template from the cytoplas- mic process of protein synthesis on ribosomes. Finally, early eukaryotic cells, which were incapable of photo- synthesis or aerobic metabolism, enveloped aerobic bac- teria or photosynthetic bacteria to form endosymbiotic associations that became permanent (Fig. 1–36). Some aerobic bacteria evolved into the mitochondria of mod- ern eukaryotes, and some photosynthetic cyanobacteria became the plastids, such as the chloroplasts of green algae, the likely ancestors of modern plant cells. Prokary- otic and eukaryotic cells are compared in Table 1–3. At some later stage of evolution, unicellular organ- isms found it advantageous to cluster together, thereby acquiring greater motility, efficiency, or reproductive success than their free-living single-celled competitors. Further evolution of such clustered organisms led to permanent associations among individual cells and eventually to specialization within the colony—to cellu- lar differentiation. The advantages of cellular specialization led to the evolution of ever more complex and highly differenti- ated organisms, in which some cells carried out the sen- sory functions, others the digestive, photosynthetic, or reproductive functions, and so forth. Many modern mul- ticellular organisms contain hundreds of different cell types, each specialized for some function that supports the entire organism. Fundamental mechanisms that evolved early have been further refined and embellished through evolution. The same basic structures and mechanisms that underlie the beating motion of cilia in Paramecium and of flagella in Chlamydomonas are employed by the highly differentiated vertebrate sperm cell. 1.5 Evolutionary Foundations 35 Anaerobic metabolism is inefficient because fuel is not completely oxidized. Ancestral anaerobic eukaryote Nucleus Aerobic metabolism is efficient because fuel is oxidized to CO 2 . Aerobic bacterium Bacterial genome Light energy is used to synthesize biomolecules from CO 2 . Photosynthetic cyanobacterium Cyanobacterial genome Aerobic eukaryote Bacterium is engulfed by ancestral eukaryote, and multiplies within it. Symbiotic system can now carry out aerobic catabolism. Some bacterial genes move to the nucleus, and the bacterial endosymbionts become mitochondria. Photosynthetic eukaryote Nonphotosynthetic eukaryote Mitochondrion Chloroplast In time, some cyanobacterial genes move to the nucleus, and endosymbionts become plastids (chloroplasts). Engulfed cyanobacterium becomes an endosymbiont and multiplies; new cell can make ATP using energy from sunlight. FIGURE 1–36 Evolution of eukaryotes through endosymbiosis. The earliest eukaryote, an anaerobe, acquired endosymbiotic purple bac- teria (yellow), which carried with them their capacity for aerobic ca- tabolism and became, over time, mitochondria. When photosynthetic cyanobacteria (green) subsequently became endosymbionts of some aerobic eukaryotes, these cells became the photosynthetic precursors of modern green algae and plants. 8885d_c01_035 12/20/03 7:09 AM Page 35 mac76 mac76:385_reb: Molecular Anatomy Reveals Evolutionary Relationships The eighteenth-century naturalist Carolus Linnaeus rec- ognized the anatomic similarities and differences among living organisms and used them to provide a framework for assessing the relatedness of species. Charles Darwin, in the nineteenth century, gave us a unifying hypothesis to explain the phylogeny of modern organisms—the ori- gin of different species from a common ancestor. Bio- chemical research in the twentieth century revealed the molecular anatomy of cells of different species—the monomeric subunit sequences and the three-dimensional structures of individual nucleic acids and proteins. Biochemists now have an enormously rich and in- creasing treasury of evidence that can be used to analyze evolutionary relationships and to refine evolu- tionary theory. The sequence of the genome (the complete genetic endowment of an organism) has been entirely determined for numerous eubacteria and for several archaebacteria; for the eukaryotic microorganisms Saccharomyces cere- visiae and Plasmodium sp.; for the plants Arabidopsis thaliana and rice; and for the multicellular animals Caenorhabditis elegans (a roundworm), Drosophila melanogaster (the fruit fly), mice, rats, and Homo sapi- ens (you) (Table 1–4). More sequences are being added to this list regularly. With such sequences in hand, detailed and quantitative comparisons among species can provide deep insight into the evolutionary process. Thus far, the molecular phylogeny derived from gene sequences is consistent with, but in many cases more precise than, the classical phylogeny based on macroscopic structures. Although organisms have continuously diverged at the level of gross anatomy, at the molecular level the basic unity of life is readily apparent; molecular structures and mechanisms are remarkably similar from the simplest to the most complex organisms. These similarities are most easily seen at the level of sequences, either the DNA se- quences that encode proteins or the protein sequences themselves. When two genes share readily detectable sequence similarities (nucleotide sequence in DNA or amino acid sequence in the proteins they encode), their sequences Chapter 1 The Foundations of Biochemistry36 TABLE 1–3 Comparison of Prokaryotic and Eukaryotic Cells Characteristic Prokaryotic cell Eukaryotic cell Size Generally small (1–10 H9262m) Generally large (5–100 H9262m) Genome DNA with nonhistone protein; DNA complexed with histone and nonhistone genome in nucleoid, not proteins in chromosomes; chromosomes in surrounded by membrane nucleus with membranous envelope Cell division Fission or budding; no mitosis Mitosis, including mitotic spindle; centrioles in many species Membrane-bounded organelles Absent Mitochondria, chloroplasts (in plants, some algae), endoplasmic reticulum, Golgi complexes, lysosomes (in animals), etc. Nutrition Absorption; some photosynthesis Absorption, ingestion; photosynthesis in some species Energy metabolism No mitochondria; oxidative Oxidative enzymes packaged in mitochondria; enzymes bound to plasma more unified pattern of oxidative metabolism membrane; great variation in metabolic pattern Cytoskeleton None Complex, with microtubules, intermediate filaments, actin filaments Intracellular movement None Cytoplasmic streaming, endocytosis, phagocytosis, mitosis, vesicle transport Source: Modified from Hickman, C.P., Roberts, L.S., & Hickman, F.M. (1990) Biology of Animals, 5th edn, p. 30, Mosby-Yearbook, Inc., St. Louis, MO. Carolus Linnaeus, 1701–1778 Charles Darwin, 1809–1882 8885d_c01_01-46 10/27/03 7:48 AM Page 36 mac76 mac76:385_reb: are said to be homologous and the proteins they encode are homologs. If two homologous genes occur in the same species, they are said to be paralogous and their protein products are paralogs. Paralogous genes are presumed to have been derived by gene duplication fol- lowed by gradual changes in the sequences of both copies (Fig. 1–37). Typically, paralogous proteins are similar not only in sequence but also in three-dimensional structure, although they commonly have acquired different func- tions during their evolution. Two homologous genes (or proteins) found in dif- ferent species are said to be orthologous, and their pro- tein products are orthologs. Orthologs are commonly found to have the same function in both organisms, and when a newly sequenced gene in one species is found to be strongly orthologous with a gene in another, this gene is presumed to encode a protein with the same function in both species. By this means, the function of gene products can be deduced from the genomic se- quence, without any biochemical characterization of the gene product. An annotated genome includes, in ad- dition to the DNA sequence itself, a description of the likely function of each gene product, deduced from com- parisons with other genomic sequences and established protein functions. In principle, by identifying the path- ways (sets of enzymes) encoded in a genome, we can deduce from the genomic sequence alone the organism’s metabolic capabilities. The sequence differences between homologous genes may be taken as a rough measure of the degree to which the two species have diverged during evo- lution—of how long ago their common evolutionary precursor gave rise to two lines with different evolu- tionary fates. The larger the number of sequence dif- ferences, the earlier the divergence in evolutionary history. One can construct a phylogeny (family tree) in which the evolutionary distance between any two species is represented by their proximity on the tree (Fig. 1–4 is an example). As evolution advances, new structures, processes, or regulatory mechanisms are acquired, reflections of the changing genomes of the evolving organisms. The genome of a simple eukaryote such as yeast should have genes related to formation of the nuclear membrane, genes not present in prokaryotes. The genome of an in- sect should contain genes that encode proteins involved in specifying the characteristic insect segmented body plan, genes not present in yeast. The genomes of all ver- tebrate animals should share genes that specify the de- velopment of a spinal column, and those of mammals should have unique genes necessary for the develop- ment of the placenta, a characteristic of mammals—and so on. Comparisons of the whole genomes of species in each phylum may lead to the identification of genes crit- ical to fundamental evolutionary changes in body plan and development. 1.5 Evolutionary Foundations 37 TABLE 1–4 Some Organisms Whose Genomes Have Been Completely Sequenced Genome size (millions Organism of nucleotide pairs) Biological interest Mycoplasma pneumoniae 0.8 Causes pneumonia Treponema pallidum 1.1 Causes syphilis Borrelia burgdorferi 1.3 Causes Lyme disease Helicobacter pylori 1.7 Causes gastric ulcers Methanococcus jannaschii 1.7 Grows at 85 H11034C! Haemophilus influenzae 1.8 Causes bacterial influenza Methanobacterium thermo- 1.8 Member of the Archaea autotrophicum Archaeoglobus fulgidus 2.2 High-temperature methanogen Synechocystis sp. 3.6 Cyanobacterium Bacillus subtilis 4.2 Common soil bacterium Escherichia coli 4.6 Some strains cause toxic shock syndrome Saccharomyces cerevisiae 12.1 Unicellular eukaryote Plasmodium falciparum 23 Causes human malaria Caenorhabditis elegans 97.1 Multicellular roundworm Anopheles gambiae 278 Malaria vector Mus musculus domesticus 2.5 H11003 10 3 Laboratory mouse Homo sapiens 2.9 H11003 10 3 Human 8885d_c01_037 12/20/03 7:09 AM Page 37 mac76 mac76:385_reb: Functional Genomics Shows the Allocations of Genes to Specific Cellular Processes When the sequence of a genome is fully determined and each gene is annotated (that is, assigned a function), molecular geneticists can group genes according to the processes (DNA synthesis, protein synthesis, generation of ATP, and so forth) in which they function and thus find what fraction of the genome is allocated to each of a cell’s activities. The largest category of genes in E. coli, A. thaliana, and H. sapiens consists of genes of as yet unknown function, which make up more than 40% of the genes in each species. The transporters that move ions and small molecules across plasma membranes take up a significant proportion of the genes in all three species, more in the bacterium and plant than in the mammal (10% of the 4,269 genes of E. coli, ~8% of the 25,706 genes of A. thaliana, and ~4% of the ~35,000 genes of H. sapiens). Genes that encode the proteins and RNA required for protein synthesis make up 3% to 4% of the E. coli genome, but in the more complex cells of A. thaliana, more genes are needed for targeting pro- teins to their final location in the cell than are needed to synthesize those proteins (about 6% and 2%, re- spectively). In general, the more complex the organism, the greater the proportion of its genome that encodes genes involved in the regulation of cellular processes and the smaller the proportion dedicated to the basic processes themselves, such as ATP generation and pro- tein synthesis. Genomic Comparisons Will Have Increasing Importance in Human Biology and Medicine The genomes of chimpanzees and humans are 99.9% identical, yet the differences between the two species are vast. The relatively few differences in genetic endowment must explain the possession of language by humans, the extraordinary athleticism of chimpanzees, and myriad other differences. Genomic comparison will allow researchers to identify candidate genes linked to divergences in the developmental pro- grams of humans and the other primates and to the emer- gence of complex functions such as language. The picture will become clearer only as more primate genomes be- come available for comparison with the human genome. Similarly, the differences in genetic endowment among humans are vanishingly small compared with the differences between humans and chimpanzees, yet these differences account for the variety among us— including differences in health and in susceptibility to chronic diseases. We have much to learn about the vari- ability in sequence among humans, and during the next decade the availability of genomic information will al- most certainly transform medical diagnosis and treat- ment. We may expect that for some genetic diseases, palliatives will be replaced by cures; and that for dis- ease susceptibilities associated with particular genetic markers, forewarning and perhaps increased preventive measures will prevail. Today’s “medical history” may be replaced by a “medical forecast.” ■ Chapter 1 The Foundations of Biochemistry38 FIGURE 1–37 Generation of genetic diversity by mutation and gene dupli- cation. 1 A mistake during replication of the genome of species A results in duplication of a gene (gene 1). The second copy is superfluous; mutations in either copy will not be deleterious as long as one good version of gene 1 is maintained. 2 As random mutations occur in one copy, the gene product changes, and in rare cases the product of the “new” gene (now gene 2) acquires a new function. Genes 1 and 2 are paralogs. 3 If species A undergoes many mutations in many genes over the course of many generations, its genome may diverge so greatly from that of the original species that it becomes a new species (species B)—that is, species A and species B cannot interbreed. Gene 1 of species A is likely to have undergone some mutations during this evolutionary period (becoming gene 1*), but it may retain enough of the original gene 1 sequence to be recognized as homologous with it, and its product may have the same function as (or similar function to) the product of gene 1. Gene 1* is an ortholog of gene 1. Species A Species B Gene duplication leads to a superfluous copy of gene 1 Mutations in gene 1 copy give rise to gene 2. Gene 2 encodes a protein with a new, different function. Mutations in many genes lead to evolution of a new species. Gene 1 Function 1 Gene 1 copy Gene 1 Function 1 Gene 2 Function 2 Homologous genes 1 and 2 are paralogs, related in sequence but encoding proteins of different function in the same species. Homologous genes 1 and 1* are orthologs, encoding proteins of the same function in different species. Gene 1* Function 1 Gene 1 Function 1 Function 1 1 2 3 8885d_c01_038 1/15/04 3:30 PM Page 38 mac76 mac76:385_reb: SUMMARY 1.5 Evolutionary Foundations ■ Occasional inheritable mutations yield an organism that is better suited for survival in an ecological niche and progeny that are preferentially selected. This process of mutation and selection is the basis for the Darwinian evolution that led from the first cell to all the organisms that now exist, and it explains the fundamental similarity of all living organisms. ■ Life originated about 3.5 billion years ago, most likely with the formation of a membrane-enclosed compartment containing a self-replicating RNA molecule. The components for the first cell were produced by the action of lightning and high temperature on simple atmospheric molecules such as CO 2 and NH 3 . ■ The catalytic and genetic roles of the early RNA genome were separated over time, with DNA becoming the genomic material and proteins the major catalytic species. ■ Eukaryotic cells acquired the capacity for photosynthesis and for oxidative phosphorylation from endosymbiotic bacteria. In multicellular organisms, differentiated cell types specialize in one or more of the functions essential to the organism’s survival. ■ Knowledge of the complete genomic nucleotide sequences of organisms from different branches of the phylogenetic tree provides insights into the evolution and function of extant organisms and offers great opportunities in human medicine. Chapter 1 Further Reading 39 Key Terms metabolite 3 nucleus 3 genome 3 eukaryote 4 prokaryote 4 archaebacteria 4 eubacteria 4 cytoskeleton 9 stereoisomers 16 configuration 16 chiral center 17 conformation 19 entropy, S 23 enthalpy, H 23 free-energy change, H9004G 23 endergonic reaction 23 exergonic reaction 23 equilibrium 24 standard free-energy change, H9004GH11034 26 activation energy, H9004G ? 26 catabolism 27 anabolism 27 metabolism 27 mutation 31 All terms are defined in the glossary. Further Reading General Fruton, J.S. (1999) Proteins, Enzymes, Genes: The Interplay of Chemistry and Biochemistry, Yale University Press, New Haven. A distinguished historian of biochemistry traces the develop- ment of this science and discusses its impact on medicine, pharmacy, and agriculture. Harold, F.M. (2001) The Way of the Cell: Molecules, Organisms, and the Order of Life, Oxford University Press, Oxford. Judson, H.F. (1996) The Eighth Day of Creation: The Makers of the Revolution in Biology, expanded edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. A highly readable and authoritative account of the rise of biochemistry and molecular biology in the twentieth century. Kornberg, A. (1987) The two cultures: chemistry and biology. Biochemistry 26, 6888–6891. The importance of applying chemical tools to biological prob- lems, described by an eminent practitioner. Monod, J. (1971) Chance and Necessity, Alfred A. Knopf, Inc., New York. [Paperback edition, Vintage Books, 1972.] Originally published (1970) as Le hasard et la nécessité, Editions du Seuil, Paris. An exploration of the philosophical implications of biological knowledge. Pace, N.R. (2001) The universal nature of biochemistry. Proc. Natl. Acad. Sci. USA 98, 805–808. A short discussion of the minimal definition of life, on Earth and elsewhere. Schr?dinger, E. (1944) What Is Life? Cambridge University Press, New York. [Reprinted (1956) in What Is Life? and Other Scientific Essays, Doubleday Anchor Books, Garden City, NY.] A thought-provoking look at life, written by a prominent physi- cal chemist. Cellular Foundations Alberts, B., Johnson, A., Bray, D., Lewis, J., Raff, M., Roberts, K., & Walter, P. (2002) Molecular Biology of the Cell, 4th edn, Garland Publishing, Inc., New York. A superb textbook on cell structure and function, covering the topics considered in this chapter, and a useful reference for many of the following chapters. Becker, W.M., Kleinsmith, L.J., & Hardin, J. (2000) The World of the Cell, 5th edn, The Benjamin/Cummings Publishing Company, Redwood City, CA. An excellent introductory textbook of cell biology. Lodish, H., Berk, A., Matsudaira, P., Kaiser, C.A., Krieger, M., Scott, M.R., Zipursky, S.L., & Darnell, J. (2003) Molecular Cell Biology, 5th edn, W. H. Freeman and Company, New York. 8885d_c01_039 1/15/04 3:30 PM Page 39 mac76 mac76:385_reb: Chapter 1 The Foundations of Biochemistry40 Like the book by Alberts and coauthors, a superb text useful for this and later chapters. Purves, W.K., Sadava, D., Orians, G.H., & Heller, H.C. (2001) Life: The Science of Biology, 6th edn, W. H. Freeman and Company, New York. Chemical Foundations Barta, N.S. & Stille, J.R. (1994) Grasping the concepts of stereochemistry. J. Chem. Educ. 71, 20–23. A clear description of the RS system for naming stereoisomers, with practical suggestions for determining and remembering the “handedness” of isomers. Brewster, J.H. (1986) Stereochemistry and the origins of life. J. Chem. Educ. 63, 667–670. An interesting and lucid discussion of the ways in which evolu- tion could have selected only one of two stereoisomers for the construction of proteins and other molecules. Kotz, J.C. & Treichel, P., Jr. (1998) Chemistry and Chemical Reactivity, Saunders College Publishing, Fort Worth, TX. An excellent, comprehensive introduction to chemistry. Vollhardt, K.P.C. & Shore, N.E. (2002) Organic Chemistry: Structure and Function, W. H. Freeman and Company, New York. Up-to-date discussions of stereochemistry, functional groups, reactivity, and the chemistry of the principal classes of biomolecules. Physical Foundations Atkins, P. W. & de Paula, J. (2001) Physical Chemistry, 7th edn, W. H. Freeman and Company, New York. Atkins, P.W. & Jones, L. (1999) Chemical Principles: The Quest for Insight, W. H. Freeman and Company, New York. Blum, H.F. (1968) Time’s Arrow and Evolution, 3rd edn, Prince- ton University Press, Princeton. An excellent discussion of the way the second law of thermody- namics has influenced biological evolution. Genetic Foundations Adams, M.D., Celniker, S.E., Holt, R.A., Evans, C.A., Gocayne, J.D., Amanatides, P.G., Scherer, S.E., Li, P.W., Hoskins, R.A., Galle, R.F., et al. (2000) The genome sequence of Drosophila melanogaster. Science 287, 2185–2195. Determination of the entire genome sequence of the fruit fly. Arabidopsis Genome Initiative. (2000) Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408, 796–815. C. elegans Sequencing Consortium. (1998) Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282, 2012–2018. Griffiths, A.J.F., Gelbart, W.M., Lewinton, R.C., & Miller, J.H. (2002) Modern Genetic Analysis: Integrating Genes and Genomes, W. H. Freeman and Company, New York. International Human Genome Sequencing Consortium. (2001) Initial sequencing and analysis of the human genome. Nature 409, 860–921. Jacob, F. (1973) The Logic of Life: A History of Heredity, Pantheon Books, Inc., New York. Originally published (1970) as La logique du vivant: une histoire de l’hérédité, Editions Gallimard, Paris. A fascinating historical and philosophical account of the route by which we came to the present molecular understanding of life. Pierce, B. (2002) Genetics: A Conceptual Approach, W. H. Freeman and Company, New York. Venter, J.C., Adams, M.D., Myers, E.W., Li, P.W., Mural, R.J., Sutton, G.G., Smith, H.O., Yandell, M., Evans, C.A., Holt, R.A., et al. (2001) The sequence of the human genome. Science 291, 1304–1351. Evolutionary Foundations Brow, J.R. & Doolittle, W.F. (1997) Archaea and the prokaryote- to-eukaryote transition. Microbiol. Mol. Biol. Rev. 61, 456–502. A very thorough discussion of the arguments for placing the Archaea on the phylogenetic branch that led to multicellular organisms. Darwin, C. (1964) On the Origin of Species: A Facsimile of the First Edition (published in 1859), Harvard University Press, Cambridge. One of the most influential scientific works ever published. de Duve, C. (1995) The beginnings of life on earth. Am. Sci. 83, 428–437. One scenario for the succession of chemical steps that led to the first living organism. de Duve, C. (1996) The birth of complex cells. Sci. Am. 274 (April), 50–57. Dyer, B.D. & Obar, R.A. (1994) Tracing the History of Eukary- otic Cells: The Enigmatic Smile, Columbia University Press, New York. Evolution of Catalytic Function. (1987) Cold Spring Harb. Symp. Quant. Biol. 52. A collection of almost 100 articles on all aspects of prebiotic and early biological evolution; probably the single best source on molecular evolution. Fenchel, T. & Finlay, B.J. (1994) The evolution of life without oxygen. Am. Sci. 82, 22–29. Discussion of the endosymbiotic hypothesis in the light of mod- ern endosymbiotic anaerobic organisms. Gesteland, R.F. & Atkins, J.F. (eds) (1993) The RNA World, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. A collection of stimulating reviews on a wide range of topics related to the RNA world scenario. Hall, B.G. (1982) Evolution on a Petri dish: the evolved H9252-galac- tosidase system as a model for studying acquisitive evolution in the laboratory. Evolutionary Biol. 15, 85–150. Knoll, A.H. (1991) End of the Proterozoic eon. Sci. Am. 265 (Oc- tober), 64–73. Discussion of the evidence that an increase in atmospheric oxy- gen led to the development of multicellular organisms, includ- ing large animals. Lazcano, A. & Miller, S.L. (1996) The origin and early evolution of life: prebiotic chemistry, the pre-RNA world, and time. Cell 85, 793–798. Brief review of developments in studies of the origin of life: primitive atmospheres, submarine vents, autotrophic versus heterotrophic origin, the RNA and pre-RNA worlds, and the time required for life to arise. Margulis, L. (1996) Archaeal-eubacterial mergers in the origin of Eukarya: phylogenetic classification of life. Proc. Natl. Acad. Sci. USA 93, 1071–1076. 8885d_c01_01-46 10/27/03 7:48 AM Page 40 mac76 mac76:385_reb: Chapter 1 Problems 41 The arguments for dividing all living creatures into five king- doms: Monera, Protoctista, Fungi, Animalia, Plantae. (Compare the Woese et al. paper below.) Margulis, L., Gould, S.J., Schwartz, K.V., & Margulis, A.R. (1998) Five Kingdoms: An Illustrated Guide to the Phyla of Life on Earth, 3rd edn, W. H. Freeman and Company, New York. Description of all major groups of organisms, beautifully illus- trated with electron micrographs and drawings. Mayr, E. (1997) This Is Biology: The Science of the Living World, Belknap Press, Cambridge, MA. A history of the development of science, with special emphasis on Darwinian evolution, by an eminent Darwin scholar. Miller, S.L. (1987) Which organic compounds could have oc- curred on the prebiotic earth? Cold Spring Harb. Symp. Quant. Biol. 52, 17–27. Summary of laboratory experiments on chemical evolution, by the person who did the original Miller-Urey experiment. Morowitz, H.J. (1992) Beginnings of Cellular Life: Metabolism Recapitulates Biogenesis, Yale University Press, New Haven. Schopf, J.W. (1992) Major Events in the History of Life, Jones and Bartlett Publishers, Boston. Smith, J.M. & Szathmáry, E. (1995) The Major Transitions in Evolution, W. H. Freeman and Company, New York. Woese, C.R., Kandler, O., & Wheelis, M.L. (1990) Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc. Natl. Acad. Sci. USA 87, 4576–4579. The arguments for dividing all living creatures into three kingdoms. (Compare the Margulis (1996) paper above.) Some problems related to the contents of the chapter follow. (In solving end-of-chapter problems, you may wish to refer to the tables on the inside of the back cover.) Each problem has a title for easy reference and discussion. 1. The Size of Cells and Their Components (a) If you were to magnify a cell 10,000 fold (typical of the magnification achieved using an electron microscope), how big would it appear? Assume you are viewing a “typical” eukaryotic cell with a cellular diameter of 50 H9262m. (b) If this cell were a muscle cell (myocyte), how many molecules of actin could it hold? (Assume the cell is spheri- cal and no other cellular components are present; actin mol- ecules are spherical, with a diameter of 3.6 nm. The volume of a sphere is 4/3 H9266r 3 .) (c) If this were a liver cell (hepatocyte) of the same di- mensions, how many mitochondria could it hold? (Assume the cell is spherical; no other cellular components are pres- ent; and the mitochondria are spherical, with a diameter of 1.5 H9262m.) (d) Glucose is the major energy-yielding nutrient for most cells. Assuming a cellular concentration of 1 mM, cal- culate how many molecules of glucose would be present in our hypothetical (and spherical) eukaryotic cell. (Avogadro’s number, the number of molecules in 1 mol of a nonionized substance, is 6.02 H11003 10 23 .) (e) Hexokinase is an important enzyme in the metabo- lism of glucose. If the concentration of hexokinase in our eukaryotic cell is 20 H9262M, how many glucose molecules are present per hexokinase molecule? 2. Components of E. coli E. coli cells are rod-shaped, about 2 H9262m long and 0.8 H9262m in diameter. The volume of a cylinder is H9266r 2 h, where h is the height of the cylinder. (a) If the average density of E. coli (mostly water) is 1.1 H11003 10 3 g/L, what is the mass of a single cell? (b) E. coli has a protective cell envelope 10 nm thick. What percentage of the total volume of the bacterium does the cell envelope occupy? (c) E. coli is capable of growing and multiplying rapidly because it contains some 15,000 spherical ribosomes (diam- eter 18 nm), which carry out protein synthesis. What per- centage of the cell volume do the ribosomes occupy? 3. Genetic Information in E. coli DNA The genetic in- formation contained in DNA consists of a linear sequence of coding units, known as codons. Each codon is a specific se- quence of three deoxyribonucleotides (three deoxyribonu- cleotide pairs in double-stranded DNA), and each codon codes for a single amino acid unit in a protein. The molecular weight of an E. coli DNA molecule is about 3.1 H11003 10 9 g/mol. The average molecular weight of a nucleotide pair is 660 g/mol, and each nucleotide pair contributes 0.34 nm to the length of DNA. (a) Calculate the length of an E. coli DNA molecule. Com- pare the length of the DNA molecule with the cell dimensions (see Problem 2). How does the DNA molecule fit into the cell? (b) Assume that the average protein in E. coli consists of a chain of 400 amino acids. What is the maximum number of proteins that can be coded by an E. coli DNA molecule? 4. The High Rate of Bacterial Metabolism Bacterial cells have a much higher rate of metabolism than animal cells. Under ideal conditions some bacteria double in size and divide every 20 min, whereas most animal cells under rapid growth conditions require 24 hours. The high rate of bacterial me- tabolism requires a high ratio of surface area to cell volume. (a) Why does surface-to-volume ratio affect the maxi- mum rate of metabolism? (b) Calculate the surface-to-volume ratio for the spher- ical bacterium Neisseria gonorrhoeae (diameter 0.5 H9262m), responsible for the disease gonorrhea. Compare it with the sur- face-to-volume ratio for a globular amoeba, a large eukaryotic cell (diameter 150 H9262m). The surface area of a sphere is 4H9266r 2 . 5. Fast Axonal Transport Neurons have long thin processes called axons, structures specialized for conducting signals throughout the organism’s nervous system. Some ax- onal processes can be as long as 2 m—for example, the ax- ons that originate in your spinal cord and terminate in the muscles of your toes. Small membrane-enclosed vesicles car- rying materials essential to axonal function move along mi- crotubules of the cytoskeleton, from the cell body to the tips of the axons. (a) If the average velocity of a vesicle is 1 H9262m/s, how long does it take a vesicle to move from a cell body in the spinal cord to the axonal tip in the toes? Problems 8885d_c01_041 1/16/04 12:35 PM Page 41 mac76 mac76:385_reb: Chapter 1 The Foundations of Biochemistry42 (b) Movement of large molecules by diffusion occurs relatively slowly in cells. (For example, hemoglobin diffuses at a rate of approximately 5 H9262m/s.) However, the diffusion of sucrose in an aqueous solution occurs at a rate ap- proaching that of fast cellular transport mechanisms (about 4 H9262m/s). What are some advantages to a cell or an organism of fast, directed transport mechanisms, compared with dif- fusion alone? 6. Vitamin C: Is the Synthetic Vitamin as Good as the Natural One? A claim put forth by some purveyors of health foods is that vitamins obtained from natural sources are more healthful than those obtained by chemical synthesis. For ex- ample, pure L-ascorbic acid (vitamin C) extracted from rose hips is better than pure L-ascorbic acid manufactured in a chemical plant. Are the vitamins from the two sources dif- ferent? Can the body distinguish a vitamin’s source? 7. Identification of Functional Groups Figures 1–15 and 1–16 show some common functional groups of biomole- cules. Because the properties and biological activities of biomolecules are largely determined by their functional groups, it is important to be able to identify them. In each of the compounds below, circle and identify by name each functional group. 8. Drug Activity and Stereochemistry The quantitative differences in biological activity between the two enantiomers of a compound are sometimes quite large. For example, the D isomer of the drug isoproterenol, used to treat mild asthma, is 50 to 80 times more effective as a bronchodilator than the L isomer. Identify the chiral center in isoproterenol. Why do the two enantiomers have such rad- ically different bioactivity? 9. Separating Biomolecules In studying a particular biomolecule (a protein, nucleic acid, carbohydrate, or lipid) in the laboratory, the biochemist first needs to separate it from other biomolecules in the sample—that is, to purify it. Specific purification techniques are described later in the text. However, by looking at the monomeric subunits of a biomolecule, you should have some ideas about the charac- teristics of the molecule that would allow you to separate it from other molecules. For example, how would you separate (a) amino acids from fatty acids and (b) nucleotides from glucose? 10. Silicon-Based Life? Silicon is in the same group of the periodic table as carbon and, like carbon, can form up to four single bonds. Many science fiction stories have been based on the premise of silicon-based life. Is this realistic? What characteristics of silicon make it less well adapted than carbon as the central organizing element for life? To answer this question, consider what you have learned about carbon’s bonding versatility, and refer to a beginning inorganic chem- istry textbook for silicon’s bonding properties. 11. Drug Action and Shape of Molecules Some years ago two drug companies marketed a drug under the trade names Dexedrine and Benzedrine. The structure of the drug is shown below. The physical properties (C, H, and N analysis, melting point, solubility, etc.) of Dexedrine and Benzedrine were identical. The recommended oral dosage of Dexedrine (which is still available) was 5 mg/day, but the recommended dosage of Benzedrine (no longer available) was twice that. Apparently it required con- siderably more Benzedrine than Dexedrine to yield the same physiological response. Explain this apparent contradiction. 12. Components of Complex Biomolecules Figure 1–10 shows the major components of complex biomolecules. For each of the three important biomolecules below (shown in their ionized forms at physiological pH), identify the constituents. (a) Guanosine triphosphate (GTP), an energy-rich nu- cleotide that serves as a precursor to RNA: P O O H11002 O P O O H11002 O O H 2 N C O NH NH 2 H H H CH OH O H N N H11002 O OP O O H11002 HH HH CCOH H H C OH H C CC O O P OH H HC C OH HO H11002 O COO H11002 COO H11002 O H11002 H H H H C OH H 3 N H11001 H 3 N H11001 NH 3 H11001 CH 3 CH 3 CH 3 CH 2 H C H H H H H C C C C C CH 2 C C O O C O OH OH OH HO NH CH 2 OH CH 2 OH Ethanolamine (a) Glycerol (b) (c) Phosphoenolpyruvate, an intermediate in glucose metabolism Threonine, an amino acid (d) Pantothenate, a vitamin D-Glucosamine (e) (f) 8885d_c01_042 1/15/04 3:31 PM Page 42 mac76 mac76:385_reb: 13. Determination of the Structure of a Biomolecule An unknown substance, X, was isolated from rabbit muscle. Its structure was determined from the following observations and experiments. Qualitative analysis showed that X was com- posed entirely of C, H, and O. A weighed sample of X was completely oxidized, and the H 2 O and CO 2 produced were measured; this quantitative analysis revealed that X contained 40.00% C, 6.71% H, and 53.29% O by weight. The molecular mass of X, determined by mass spectrometry, was 90.00 u (atomic mass units; see Box 1–1). Infrared spectroscopy showed that X contained one double bond. X dissolved read- ily in water to give an acidic solution; the solution demon- strated optical activity when tested in a polarimeter. (a) Determine the empirical and molecular formula of X. (b) Draw the possible structures of X that fit the mo- lecular formula and contain one double bond. Consider only linear or branched structures and disregard cyclic structures. Note that oxygen makes very poor bonds to itself. (c) What is the structural significance of the observed optical activity? Which structures in (b) are consistent with the observation? (d) What is the structural significance of the observa- tion that a solution of X was acidic? Which structures in (b) are consistent with the observation? (e) What is the structure of X? Is more than one struc- ture consistent with all the data? Chapter 1 Problems 43 CH 3 H11001 NP O O H11002 O H C CH 2 CH 2 OCH 2 O C O (CH 2 CH) 7 C C O C (CH 2 ) 14 CH 3 H H O (CH 2 ) 7 3 CH 2 CH 3 CH 3 HO CH 2 C H NH 2 C O N H C H H C O N H H C O N H C H C O N H C H C C S CH 3 H 2 H 2 COO H11002 H 2 CH C (b) Phosphatidylcholine, a component of many mem- branes: (c) Methionine enkephalin, the brain’s own opiate: 8885d_c01_043 1/15/04 3:31 PM Page 43 mac76 mac76:385_reb: