《生物化学原理》（英文版）BIOENERGETICS AND METABOLISM

16 Chapter 13 Principles of Bioenergetics isoalloxazine I NH H+ CH3 CH3 FADH(FMNH) FADH, (FMNH2 (fully reduced) HCOH HCOH FAD 0→P=0 0 CH O FIGURE 13-18 Structures of oxidized and reduced FAD and FMN MN consists of the structure above the dashed line on the FAD (ox- idized form). The flavin nucleotides accept two hydrogen atoms(two oHOH electrons and two protons), both of which appear in the flavin ring Flavin adenine dinucl system. When FAD or FMN accepts only one hydrogen atom, the semi- quinone, a stable free radical, forms Flavoproteins are often very complex; some have, in ad a Biological oxidation-reduction reactions can be dition to a flavin nucleotide tightly bound inorganic ions described in terms of two half-reactions. each (iron or molybdenum, for example) capable of partici with a characteristic standard reduction pating in electron transfers Certain flavoproteins act in a quite different role as a When two electrochemical half-cells. each light receptors. Cryptochromes are a family of flavo- containing the components of a half-reaction, proteins, widely distributed in the eukaryotic phyla, that connected. electrons tend to flow to the mediate the effects of blue light on plant development half-cell with the higher reduction potential and the effects of light on mammalian circadian rhythms (oscillations in physiology and biochemistry, with a The strength of this tendency is proportional to the difference between the two reduction 24-hour period). The cryptochromes are homologs of potentials(4E and is a function of the another family of flavoproteins, the photolyases. Found concentrations of oxidized and reduced in both prokaryotes and eukaryotes, photolyases use the energy of absorbed light to repair chemical defects in dna a The standard free-energy change for an We examine the function of flavoproteins as elec- xidation-reduction reaction is directly tron carriers in Chapter 19, when we consider their roles oportional to the difference in standard in oxidative phosphorylation (in mitochondria) and pho- reduction potentials of the two half-cells tophosphorylation (in chloroplasts), and we describe nF△E° the photolyase reactions in Chapter 25 a Many biological oxidation reactions are dehydrogenations in which one or two SUMMARY 13. 3 Biological Oxidation-Reduction hydrogen atoms(H +e)are transferred Reactions from a substrate to a hydrogen acceptor. Oxidation-reduction reactions in living cells a In many organisms, a central energy-conserving involve specialized electron carriers. process is the stepwise oxidation of glucose to a NAD and NADP are the freely diffusible COz, in which some of the energy of oxidation is coenzymes of many dehydrogenases. Both conserved in aTP as electrons are passed to O NAD and NADP accept two electrons and

OH N H H OH R H NH HCOH N O O N HCOH HCOH P O O O P O O H N N N O NH N N N O R NH N N N O H O H H FAD FMN •  O  O Flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) CH2 NH2 CH2 CH2 CH3 CH3 isoalloxazine ring H e  H e CH  3 CH3 N O FADH• (FMNH•) (semiquinone) CH3 CH3 FADH2 (FMNH2) (fully reduced) Flavoproteins are often very complex; some have, in ad￾dition to a flavin nucleotide, tightly bound inorganic ions (iron or molybdenum, for example) capable of partici￾pating in electron transfers. Certain flavoproteins act in a quite different role as light receptors. Cryptochromes are a family of flavo￾proteins, widely distributed in the eukaryotic phyla, that mediate the effects of blue light on plant development and the effects of light on mammalian circadian rhythms (oscillations in physiology and biochemistry, with a 24-hour period). The cryptochromes are homologs of another family of flavoproteins, the photolyases. Found in both prokaryotes and eukaryotes, photolyases use the energy of absorbed light to repair chemical defects in DNA. We examine the function of flavoproteins as elec￾tron carriers in Chapter 19, when we consider their roles in oxidative phosphorylation (in mitochondria) and pho￾tophosphorylation (in chloroplasts), and we describe the photolyase reactions in Chapter 25. SUMMARY 13.3 Biological Oxidation-Reduction Reactions ■ In many organisms, a central energy-conserving process is the stepwise oxidation of glucose to CO2, in which some of the energy of oxidation is conserved in ATP as electrons are passed to O2. ■ Biological oxidation-reduction reactions can be described in terms of two half-reactions, each with a characteristic standard reduction potential, E . ■ When two electrochemical half-cells, each containing the components of a half-reaction, are connected, electrons tend to flow to the half-cell with the higher reduction potential. The strength of this tendency is proportional to the difference between the two reduction potentials (E) and is a function of the concentrations of oxidized and reduced species. ■ The standard free-energy change for an oxidation-reduction reaction is directly proportional to the difference in standard reduction potentials of the two half-cells: G  n E . ■ Many biological oxidation reactions are dehydrogenations in which one or two hydrogen atoms (H e) are transferred from a substrate to a hydrogen acceptor. Oxidation-reduction reactions in living cells involve specialized electron carriers. ■ NAD and NADP are the freely diffusible coenzymes of many dehydrogenases. Both NAD and NADP accept two electrons and 516 Chapter 13 Principles of Bioenergetics FIGURE 13–18 Structures of oxidized and reduced FAD and FMN. FMN consists of the structure above the dashed line on the FAD (ox￾idized form). The flavin nucleotides accept two hydrogen atoms (two electrons and two protons), both of which appear in the flavin ring system. When FAD or FMN accepts only one hydrogen atom, the semi￾quinone, a stable free radical, forms.

Chapter 13 Further Reading 517 one proton. NAD and NADP are bound to flavoproteins. They can accept either one or dehydrogenases in a widely conserved two electrons. Flavoproteins also serve as light structural motif called the rossmann fold receptors in cryptochromes and photolyases. FAD and fmn. the flavin nucleotides. serve as tightly bound prosthetic groups of Key terms Terms in bold are defined in the glossary. autotroph XXX thioester XXX dehydrogenation XXX metabolism XXX inorganic pyrophosphatase XXX reducing equivalent XXX nucleoside diphosphate standard reduction potential metabolite xXX kinase xXX Er) XXX intermediary metabolism XXX adenylate kinase XXX pyridine nucleotide XXX anabolism xXX phosphagens XXX ein XXX standard transformed constants XXX polyphosphate kinase-1, -2 XXX flavin nucleotides XXX phosphorylation potential conjugate redox pair Xx photolyase XXX Further Reading Bioenergetics and Thermodynamics Morowitz, H..(1978) Foundations of Bioenergetics, Academic Atkins, PW(1984)The Second Law. Scientific American Books. Inc. New York. Clear, rigorous description of thermodynamics in biology. A well-illustrated and elementary discussion of the second lay Nicholls, D.G.& Ferguson, SJ.(2002)Bioenergetics 3. Academic Press, Inc. New York. Becker, W.M.( 1977) Energy and the Living Cell: An introduc Clear well-illustrated intermediate-level discussion of the tion to Bioenergetics, J. B. Lippincott Company, Philadelphia. theory of bioenergetics and the mechanisms of energy A clear introductory account of cellular metabolism, in terms of transductions Tinoco, I, Jr, Sauer, K,& Wang, J C. (1996)Physical Chem- Bergethon, P.R.(1998)The Physical Basis of Biochemistry Springer Verlag, New York. edn, Prentice-Hall, Inc, Upper Saddle River, Chapters 11 through 13 of this book, and the books by Tinoco Chapters 2 through 5 cover thermodynamics et al. and van Holde et al.(below), are excellent general refer- van Holde, K.E., Johnson, W.C.,& Ho, PS. (1998)Principles ences for physical biochemistry, with good discussions of the of Physical Biochemistry. Prentice-Hall, Inc, Upper Saddle River applications of thermodynamics to biochemistry. Edsall, J.T.& Gutfreund, H.( 1983) Biothermodynamics: The Chapters 2 and 3 are especially relevant. tudy of Biochemical Processes at Equilibrium, John Wiley Phosphoryl Group Transfers and ATP Harold, FM. (1986)The Vital Force: A Study of Bioenergetics, Alberty, R.A. (1994) Biochemical thermodynamics. Biochim W. H. Freeman and Company. New York. iophys. Acta 1207. 1-11 a beautifully clear discussion of thermodynamics in biological Explains the distinction between biochemical and chemical equations, and the calculation and meaning of transformed Harris, D.A.(1995)Bioenergetics at a Glance. Blackwel thermodynamic properties for ATP and other phosphorylat Science. Oxford. A short, clearly written account of cellular energetics, including W.A.& Henderson, J F.(1983)Cell ATP John Wiley introductory chapters on thermodynamics. Loewenstein, W.R. ( 1999) The Touchstone of Life: Molecular chemistry of ATP, its role in metabolic regulation, and its Information, Cell Communication, and the Foundations of tabolic and anabolic roles Life, Oxford University Press, New York. Frey, P.A.& Arabshahi, A.(1995)Standard free-energy change Beautifully written discussion of the relationship between for the hydrolysis of the a-B-phosphoanhydride bridge in ATP. and information Biochemistry 34, 11,, 310

one proton. NAD and NADP are bound to dehydrogenases in a widely conserved structural motif called the Rossmann fold. ■ FAD and FMN, the flavin nucleotides, serve as tightly bound prosthetic groups of flavoproteins. They can accept either one or two electrons. Flavoproteins also serve as light receptors in cryptochromes and photolyases. Chapter 13 Further Reading 517 Key Terms autotroph XXX heterotroph XXX metabolism XXX metabolic pathways XXX metabolite XXX intermediary metabolism XXX catabolism XXX anabolism XXX standard transformed constants XXX phosphorylation potential (Gp) XXX thioester XXX adenylylation XXX inorganic pyrophosphatase XXX nucleoside diphosphate kinase XXX adenylate kinase XXX creatine kinase XXX phosphagens XXX polyphosphate kinase-1, -2 XXX electromotive force (emf) XXX conjugate redox pair XXX dehydrogenation XXX dehydrogenases XXX reducing equivalent XXX standard reduction potential (E
) XXX pyridine nucleotide XXX oxidoreductase XXX flavoprotein XXX flavin nucleotides XXX cryptochrome XXX photolyase XXX Terms in bold are defined in the glossary. Further Reading Bioenergetics and Thermodynamics Atkins, P.W. (1984) The Second Law, Scientific American Books, Inc., New York. A well-illustrated and elementary discussion of the second law and its implications. Becker, W.M. (1977) Energy and the Living Cell: An Introduc￾tion to Bioenergetics, J. B. Lippincott Company, Philadelphia. A clear introductory account of cellular metabolism, in terms of energetics. Bergethon, P.R. (1998) The Physical Basis of Biochemistry, Springer Verlag, New York. Chapters 11 through 13 of this book, and the books by Tinoco et al. and van Holde et al. (below), are excellent general refer￾ences for physical biochemistry, with good discussions of the applications of thermodynamics to biochemistry. Edsall, J.T. & Gutfreund, H. (1983) Biothermodynamics: The Study of Biochemical Processes at Equilibrium, John Wiley & Sons, Inc., New York. Harold, F.M. (1986) The Vital Force: A Study of Bioenergetics, W. H. Freeman and Company, New York. A beautifully clear discussion of thermodynamics in biological processes. Harris, D.A. (1995) Bioenergetics at a Glance, Blackwell Science, Oxford. A short, clearly written account of cellular energetics, including introductory chapters on thermodynamics. Loewenstein, W.R. (1999) The Touchstone of Life: Molecular Information, Cell Communication, and the Foundations of Life, Oxford University Press, New York. Beautifully written discussion of the relationship between entropy and information. Morowitz, H.J. (1978) Foundations of Bioenergetics, Academic Press, Inc., New York. [Out of print.] Clear, rigorous description of thermodynamics in biology. Nicholls, D.G. & Ferguson, S.J. (2002) Bioenergetics 3, Academic Press, Inc., New York. Clear, well-illustrated intermediate-level discussion of the theory of bioenergetics and the mechanisms of energy transductions. Tinoco, I., Jr., Sauer, K., & Wang, J.C. (1996) Physical Chem￾istry: Principles and Applications in Biological Sciences, 3rd edn, Prentice-Hall, Inc., Upper Saddle River, NJ. Chapters 2 through 5 cover thermodynamics. van Holde, K.E., Johnson, W.C., & Ho, P.S. (1998) Principles of Physical Biochemistry, Prentice-Hall, Inc., Upper Saddle River, NJ. Chapters 2 and 3 are especially relevant. Phosphoryl Group Transfers and ATP Alberty, R.A. (1994) Biochemical thermodynamics. Biochim. Biophys. Acta 1207, 1–11. Explains the distinction between biochemical and chemical equations, and the calculation and meaning of transformed thermodynamic properties for ATP and other phosphorylated compounds. Bridger, W.A. & Henderson, J.F. (1983) Cell ATP, John Wiley & Sons, Inc., New York. The chemistry of ATP, its role in metabolic regulation, and its catabolic and anabolic roles. Frey, P.A. & Arabshahi, A. (1995) Standard free-energy change for the hydrolysis of the –-phosphoanhydride bridge in ATP. Biochemistry 34, 11,307–11,310.

518 Chapter 13 Principles of Bioenergetics Hanson, R W. (1989) The role of ATP in metabolism. Biochem. Biological Oxidation-Reduction Reactions Educ.17,86-92 Cashmore, AR, Jarillo, J.A, Wu, YJ Liu D(1999) Excellent summary of the chemistry and biology of ATP. Cryptochromes: blue light receptors for plants and animals Kornberg, A.(1999)Inorganic polyphosphate: a molecule of Science284,760-765 many functions. Annu. Rev. Biochem. 68, 89-125. Dolphin, D, Avramovic, 0, Poulson, R. (eds)(1987) Lipmann, F.(1941)Metabolic generation and utilization of Pyridine Nucleotide Coenzymes: Chemical, Biochemical, and Medical Aspects, John Wiley Sons, Inc, New York. The classic description of the role of high-energy phosphate An excellent two-volume collection of authoritative reviews. Among the most useful are the chapters by Kaplan, Pullman, B. Pullman, A (1960) ure of Westheimer. Veech, and Ohno and Ushio energy-rich phosphates. Radiat. Res, Suppl. 2, 160-181 fraaije, M.w.& Mattevi, A.(2000)Flavoenzymes: diverse cata- An advanced discussion of the chemistry of ATP and other lysts with recurrent features. Trends Biochem Sci. 25, 126-132. energy-rich"compounds. Massey, V(1994) Activation of molecular oxygen by flavins and Veech, R L, Lawson, J.W.R., Cornell, N W,& Krebs, H.A. flavoproteins. J. Bio/ Chem 269, 22,459-22,462 (1979)Cytosolic phosphorylation potential. J. Biol. Chem. 254, A short review of the chemistry of flavin-oxygen interactions in 6538-6547 Experimental determination of ATP, ADP, and P, concentrations Rees, D.C(2002)Great metalloclusters in enzymology. Annu. in brain, muscle, and liver, and a discussion of the problems in Rev. Biochem. 71, 221-246 determining the real free-energy change for ATP synthesis in Advanced review of the types of metal ion clusters found in enzymes and their modes of action Westheimer, F.H. (1987)Why nature chose phosphates. Science Williams, R.E.& Bruce, N C(2002) New uses for an old 235,1173-117 enzyme-the old yellow enzyme family of flavoenzymes A chemists description of the unique suitability of phosphate Microbiology 148, 1607-1614 esters and anhydrides for metabolic transformations Problems 1. Entropy Changes during Egg Development Con- lowing reactions at pH 7.0 and 25C, using the AG values sider a system consisting of an egg in an incubator. The white in Table 13-4 and yolk of the egg contain proteins, carbohydrates, and ipds. If fertilized, the egg is transformed from a single cell to a complex organism. Discuss this irreversible process in (a)Glucose 6-phosphate H2O glucose+ Pi and universe. Be sure that you first clearly define the system and surroundings. (b)Lactose H2O glucose galactose 2. Calculation of AG from an Equilibrium Constant Calculate the standard free-energy changes of the following metabolically important enzyme-catalyzed reactions at 25oC (c) Malate fumarate +Ho and pH 7.0, using the equilibrium constants giv 4. Experimental Determination of Keg and AG If 0.1 aspartate M solution of glucose 1-phosphate is incubated with a catalytic (a) Glutamate oxaloacetate= amount of phosphoglucomutase, the glucose 1-phosphate is transformed to glucose 6-phosphate. At equilibrium, the con- aspartate a-ketoglutarate Keq=6.8 centrations of the reaction components are triose phosphate Glucose 1-phosphate e glucose 6-phosphate 4.5×10-3M 9.6×10 (b)Dihydroxyacetone phosphate glyceraldehyde 3-phosphate Ke=0.0475 Calculate Keg and AGo for this reaction at 25oC phasphofructokin 5. Experimental Determination of AG for ATP Hy- (c) Fructose 6-phosphate ATP drolysis A direct measurement of the standard free-energy fructose 1, 6-bisphosphate ADP Ke=254 change associated with the hydrolysis of ATP is technically demanding because the minute amount of ATP remaining at 3. Calculation of the Equilibrium Constant fromAG equilibrium is difficult to measure accurately. The value of Calculate the equilibrium constants Keg for each of the fol- AG can be calculated indirectly. however, from the equilib-

518 Chapter 13 Principles of Bioenergetics Hanson, R.W. (1989) The role of ATP in metabolism. Biochem. Educ. 17, 86–92. Excellent summary of the chemistry and biology of ATP. Kornberg, A. (1999) Inorganic polyphosphate: a molecule of many functions. Annu. Rev. Biochem. 68, 89–125. Lipmann, F. (1941) Metabolic generation and utilization of phosphate bond energy. Adv. Enzymol. 11, 99–162. The classic description of the role of high-energy phosphate compounds in biology. Pullman, B. & Pullman, A. (1960) Electronic structure of energy-rich phosphates. Radiat. Res., Suppl. 2, 160–181. An advanced discussion of the chemistry of ATP and other “energy-rich” compounds. Veech, R.L., Lawson, J.W.R., Cornell, N.W., & Krebs, H.A. (1979) Cytosolic phosphorylation potential. J. Biol. Chem. 254, 6538–6547. Experimental determination of ATP, ADP, and Pi concentrations in brain, muscle, and liver, and a discussion of the problems in determining the real free-energy change for ATP synthesis in cells. Westheimer, F.H. (1987) Why nature chose phosphates. Science 235, 1173–1178. A chemist’s description of the unique suitability of phosphate esters and anhydrides for metabolic transformations. Biological Oxidation-Reduction Reactions Cashmore, A.R., Jarillo, J.A., Wu, Y.J., & Liu D. (1999) Cryptochromes: blue light receptors for plants and animals. Science 284, 760–765. Dolphin, D., Avramovic, O., & Poulson, R. (eds) (1987) Pyridine Nucleotide Coenzymes: Chemical, Biochemical, and Medical Aspects, John Wiley & Sons, Inc., New York. An excellent two-volume collection of authoritative reviews. Among the most useful are the chapters by Kaplan, Westheimer, Veech, and Ohno and Ushio. Fraaije, M.W. & Mattevi, A. (2000) Flavoenzymes: diverse cata￾lysts with recurrent features. Trends Biochem. Sci. 25, 126–132. Massey, V. (1994) Activation of molecular oxygen by flavins and flavoproteins. J. Biol. Chem. 269, 22,459–22,462. A short review of the chemistry of flavin–oxygen interactions in flavoproteins. Rees, D.C. (2002) Great metalloclusters in enzymology. Annu. Rev. Biochem. 71, 221–246. Advanced review of the types of metal ion clusters found in enzymes and their modes of action. Williams, R.E. & Bruce, N.C. (2002) New uses for an old enzyme—the old yellow enzyme family of flavoenzymes. Microbiology 148, 1607–1614. 1. Entropy Changes during Egg Development Con￾sider a system consisting of an egg in an incubator. The white and yolk of the egg contain proteins, carbohydrates, and lipids. If fertilized, the egg is transformed from a single cell to a complex organism. Discuss this irreversible process in terms of the entropy changes in the system, surroundings, and universe. Be sure that you first clearly define the system and surroundings. 2. Calculation of G
from an Equilibrium Constant Calculate the standard free-energy changes of the following metabolically important enzyme-catalyzed reactions at 25 C and pH 7.0, using the equilibrium constants given. aspartate aminotransferase (a) Glutamate oxaloacetate 3::::::::::::4 aspartate -ketoglutarate K eq  6.8 triose phosphate isomerase (b) Dihydroxyacetone phosphate 3:::::::::::4 glyceraldehyde 3-phosphate K eq  0.0475 phosphofructokinase (c) Fructose 6-phosphate ATP 3:::::::::::::::4 fructose 1,6-bisphosphate ADP Keq  254 3. Calculation of the Equilibrium Constant from G Calculate the equilibrium constants K eq for each of the fol￾lowing reactions at pH 7.0 and 25 C, using the G values in Table 13–4. (a) Glucose 6-phosphate H2O glucose Pi (b) Lactose H2O glucose galactose (c) Malate fumarate H2O 4. Experimental Determination of K eq and G If a 0.1 M solution of glucose 1-phosphate is incubated with a catalytic amount of phosphoglucomutase, the glucose 1-phosphate is transformed to glucose 6-phosphate. At equilibrium, the con￾centrations of the reaction components are Glucose 1-phosphate 34 glucose 6-phosphate 4.5 103 M 9.6 102 M Calculate K eq and G for this reaction at 25 C. 5. Experimental Determination of G for ATP Hy￾drolysis A direct measurement of the standard free-energy change associated with the hydrolysis of ATP is technically demanding because the minute amount of ATP remaining at equilibrium is difficult to measure accurately. The value of G can be calculated indirectly, however, from the equilib￾fumarase 3::::::4 b-galactosidase 3::::::::::4 glucose 6-phosphatase 3::::::::::4 Problems

Chapter 13 Problems 519 rium constants of two other enzymatic reactions having less (c) The phosphorylation of glucose in the cell is coupled favorable equilibrium constants to the hydrolysis of ATP; that is, part of the free energy of Glucose 6-phosphate +H20-glucose Pi Keq=270 TP hydrolysis is used to phosphorylate glue ATP+ glucose→→ADP+ glucose6 phosph (1) Glucose+P- glucose 6-phosphate H2O △G°=13.8 kJ/mol 2)ATP+H20→ADP+P1 Using this information, calculate the standard free energy of △G°=-30.5 k/mol hydrolysis of ATP at 25C. Sum: Glucose ATP- glucose 6-phosphate ADP 6. Difference between△G"°and△ g Consider the fol- lowing interconversion, which occurs in glycolysis( Chapter Calculate Ke for the overall reaction. For the ATP-dependent hosphorylation of glucose, what concentration of Fructose 6-phosphate e glucose 6-phosphate needed to achieve a 250 uM intracellular concentration of glu Ke=1.97 6-phosphate when the concentrations of ATP and ADP are 3.38 mM and 1. 32 mM, respectively? Does this coupling (a) What is△G° for the reaction(at25°0)? process provide a feasible route, at least in principle. for the (b)If the concentration of fructose 6-phosphate is ad phosphorylation of glucose in the cell? Explain justed to 1.5 M and that of glucose 6-phosphate is adjusted (d) Although coupling ATP hydrolysis to glucose phos- to0.50M, what is△G? phorylation makes thermodynamic sense, we have not yet (c) Why are△a°and△ G different? specified how this coupling is to take place. Given that cou- pling requires a common intermediate, one conceivable route 7. Dependence of AG on pH The free d is to use ATP hydrolysis to raise the intracellular concentra- by the hydrolysis of ATP under standard oH tion of P, and thus drive the unfavorable phosphorylation of 7.0 is-305 kJ/mol. If ATP is hydrolyzed un n- glucose by Pr. Is this a reasonable route?(Think about the ditions but at pH 5.0, is more or less free energy released? solubility products of metabolic intermediates. Explain (e) The ATP-coupled phosphorylation of glucose is cat- fucokinase. this 8. The AG for Coupled Reactions Glucose l-pho zyme binds ATP and glucose to form a glucose-ATP-enzyme phate is converted into fructose 6-phosphate in two succes- complex, and the phosphoryl group is transferred directly sive reactions: from AtP to glucose. Explain the advantages of this route Glucose 1; phosphate→→ glucose6 phosphate Glucose6 phosphate→→ fructose6pho 10. Calculations of AG for ATP-Coupled Reactions From data in Table 13-6 calculate the ago value for the Using the AG values in Table 13-4, calculate the equilibrium reactions onstant. K for the sum of the two reactions at 25C. (a)Phosphocreatine ADP- creatine ATP Glucose 1-phosphate→→ fructose6 phosphate (b)ATP fructose- ADP fructose 6-phosphate 11. Coupling ATP Cleavage to an Unfavorable Reaction 9.Strategy for Overcoming an Unfavorable Reaction: To explore the consequences of coupling ATP hydrolysis under ATP-Dependent Chemical Coupling The phosphoryla- physiological conditions to a thermodynamically unfavorable tion of glucose to glucose 6-phosphate is the initial step biochemical reaction, consider the hypothetical transformation @ Catabolism of giucose. The direct phosphorylation of glu- x→ Y for which△C°=2khnl cose by Pi is described by the equation (a) What is the ratio YyX] at equilibrium? Glucose+P→→ glucose6 phosphate+H20 Suppose X and Y participate in a sequence of reac AGo=13.8 kJmol tions during which ATP is hydrolyzed to ADP and Pr. The overall reaction is (a)Calculate the equilibrium constant for the above re- action. In the rat hepatocyte the physiological concentrations X+ATP+H2O→Y+ADP+P1 of glucose and P are maintained at approximately 4.8 mM. Calculate(YVIX) for this reaction at equilibrium. Assume tha What is the equilibrium concentration of glucose 6-phosphate the equilibrium concentrations of ATP. ADP and p, are I ained by the direct phosphorylation of (c)We know that [ATPL (ADPl and Pil are not I M un this reaction represent a reasonable metabolic step for the der physiological conditions. Calculate [YyX] for the AtP- catabolism of glucose? Explain. oupled reaction when the values of ATP [ADPl, and [Pll (b) In principle, at least, one way to increase the con- are those found in rat myocytes (Table 13-5) centration of glucose 6-phosphate is to drive the equilibrium I trations of glucose and p, doe sing the intracellular concen- 12. Calculations of AG at Physiological Concentrations reaction to the right by inci ing a fixed concentration of Calculate the physiological AG(not AG)for the reaction P at 4.8 mM, how high would the intracellular concentration of glucose have to be to give an equilibrium concentration of Phosphocreatine ADP-creatine ATP glucose 6-phosphate of 250 uM(the normal physiological con- at 25C, as it occurs in the cytosol of neurons, with phos- centration)? Would this route be physiologically reasonable, phocreatine at 4.7 mM, creatine at 1.0 mM, ADP at 0.73 mM, given that the maximum solubility of glucose is less than 1 M? and ATP at 2.6 mM

Chapter 13 Problems 519 rium constants of two other enzymatic reactions having less favorable equilibrium constants: Glucose 6-phosphate H2O 8n glucose Pi K eq  270 ATP glucose 8n ADP glucose 6-phosphate K eq  890 Using this information, calculate the standard free energy of hydrolysis of ATP at 25 C. 6. Difference between G and G Consider the fol￾lowing interconversion, which occurs in glycolysis (Chapter 14): Fructose 6-phosphate 34 glucose 6-phosphate K eq  1.97 (a) What is G for the reaction (at 25 C)? (b) If the concentration of fructose 6-phosphate is ad￾justed to 1.5 M and that of glucose 6-phosphate is adjusted to 0.50 M, what is G? (c) Why are G and G different? 7. Dependence of G on pH The free energy released by the hydrolysis of ATP under standard conditions at pH 7.0 is 30.5 kJ/mol. If ATP is hydrolyzed under standard con￾ditions but at pH 5.0, is more or less free energy released? Explain. 8. The G for Coupled Reactions Glucose 1-phos￾phate is converted into fructose 6-phosphate in two succes￾sive reactions: Glucose 1-phosphate 88n glucose 6-phosphate Glucose 6-phosphate 88n fructose 6-phosphate Using the G values in Table 13–4, calculate the equilibrium constant, K eq, for the sum of the two reactions at 25 C: Glucose 1-phosphate 88n fructose 6-phosphate 9. Strategy for Overcoming an Unfavorable Reaction: ATP-Dependent Chemical Coupling The phosphoryla￾tion of glucose to glucose 6-phosphate is the initial step in the catabolism of glucose. The direct phosphorylation of glu￾cose by Pi is described by the equation Glucose Pi 88n glucose 6-phosphate H2O G  13.8 kJ/mol (a) Calculate the equilibrium constant for the above re￾action. In the rat hepatocyte the physiological concentrations of glucose and Pi are maintained at approximately 4.8 mM. What is the equilibrium concentration of glucose 6-phosphate obtained by the direct phosphorylation of glucose by Pi ? Does this reaction represent a reasonable metabolic step for the catabolism of glucose? Explain. (b) In principle, at least, one way to increase the con￾centration of glucose 6-phosphate is to drive the equilibrium reaction to the right by increasing the intracellular concen￾trations of glucose and Pi . Assuming a fixed concentration of Pi at 4.8 mM, how high would the intracellular concentration of glucose have to be to give an equilibrium concentration of glucose 6-phosphate of 250 M (the normal physiological con￾centration)? Would this route be physiologically reasonable, given that the maximum solubility of glucose is less than 1 M? (c) The phosphorylation of glucose in the cell is coupled to the hydrolysis of ATP; that is, part of the free energy of ATP hydrolysis is used to phosphorylate glucose: (1) Glucose Pi 8n glucose 6-phosphate H2O G  13.8 kJ/mol (2) ATP H2O 8n ADP Pi G  30.5 kJ/mol Sum: Glucose ATP 8n glucose 6-phosphate ADP Calculate K eq for the overall reaction. For the ATP-dependent phosphorylation of glucose, what concentration of glucose is needed to achieve a 250 M intracellular concentration of glu￾cose 6-phosphate when the concentrations of ATP and ADP are 3.38 mM and 1.32 mM, respectively? Does this coupling process provide a feasible route, at least in principle, for the phosphorylation of glucose in the cell? Explain. (d) Although coupling ATP hydrolysis to glucose phos￾phorylation makes thermodynamic sense, we have not yet specified how this coupling is to take place. Given that cou￾pling requires a common intermediate, one conceivable route is to use ATP hydrolysis to raise the intracellular concentra￾tion of Pi and thus drive the unfavorable phosphorylation of glucose by Pi . Is this a reasonable route? (Think about the solubility products of metabolic intermediates.) (e) The ATP-coupled phosphorylation of glucose is cat￾alyzed in hepatocytes by the enzyme glucokinase. This en￾zyme binds ATP and glucose to form a glucose-ATP-enzyme complex, and the phosphoryl group is transferred directly from ATP to glucose. Explain the advantages of this route. 10. Calculations of G for ATP-Coupled Reactions From data in Table 13–6 calculate the G value for the reactions (a) Phosphocreatine ADP 8n creatine ATP (b) ATP fructose 8n ADP fructose 6-phosphate 11. Coupling ATP Cleavage to an Unfavorable Reaction To explore the consequences of coupling ATP hydrolysis under physiological conditions to a thermodynamically unfavorable biochemical reaction, consider the hypothetical transformation X n Y, for which G  20 kJ/mol. (a) What is the ratio [Y]/[X] at equilibrium? (b) Suppose X and Y participate in a sequence of reac￾tions during which ATP is hydrolyzed to ADP and Pi . The overall reaction is X ATP H2O 8n Y ADP Pi Calculate [Y]/[X] for this reaction at equilibrium. Assume that the equilibrium concentrations of ATP, ADP, and Pi are 1 M. (c) We know that [ATP], [ADP], and [Pi ] are not 1 M un￾der physiological conditions. Calculate [Y]/[X] for the ATP￾coupled reaction when the values of [ATP], [ADP], and [Pi ] are those found in rat myocytes (Table 13–5). 12. Calculations of G at Physiological Concentrations Calculate the physiological G (not G ) for the reaction Phosphocreatine ADP 8n creatine ATP at 25 C, as it occurs in the cytosol of neurons, with phos￾phocreatine at 4.7 mM, creatine at 1.0 mM, ADP at 0.73 mM, and ATP at 2.6 mM.

520 Chapter 13 Principles of Bioenergetics 13. Free Energy Required for ATP Synthesis under 18. Standard Reduction Potentials The standard re- Physiological Conditions In the cytosol of rat hepato- duction potential, E, of any redox pair is defined for the cytes, the mass-action ratio, Q. is half-cell reaction ADP|PT=53×X102M Oxidizing agent electrons- reducing agent The E values for the NAD" NADH and pyruvate/lactate con- Calculate the free energy required to synthesize ATP in a rat jugate redox pairs are -0.32 V and -0 19 V, respectively (a) Which conjugate pair has the greater tendency to lose electrons? E 14. Daily ATP Utilization by Human Adult (a)A total of 30.5 kJ/mol of free energy is needed to Which is the stronger g agent? Explain. synthesize ATP from ADP and P, when the reactants and (c) Beginning with 1 M co ions of each reactant products are at 1 M concentrations(standard state). Because and product at pH 7, in which will the following re- the actual physiological concentrations of ATP, ADP, and P action proceed? are not I M, the free energy required to synthesize ATP un- Pyruvate+ NADH+H'=lactate+ NAD der physiological conditions is different from AG. Calculate the free energy required to synthesize ATP in the human he- (d) What is the standard free-energy change (AG )at patocyte when the physiological concentrations of ATP. ADP. 25"C for the conversion of pyruvate to lactate? and Pi are 3.5, 1.50, and 5.0 mM, respective (e) What is the equilibrium constant )A68 kg(150 lb)adult requires a caloric intake of reaction? 2,000 kcal(8, 360 k)of food per day(24 h). The food is me- 19. Energy Span of the Respiratory Chain Electron tabolized and the free energy is used to synthesize ATP, whic then provides energy for the body's daily chemical and me- resented by the net reaction equation chanical work. Assuming that the efficiency of converting food energy into ATP is 50%, calculate the weight of ATP used by a human adult in 24 h What percentage of the body NADH +H+202= H20+ NAD weight does this represent? (c)Although adults synthesize large amounts of ATP mitochondrial electron transfer. Use Ero values from table ily, their body weight, structure, and composition do not change significantly during this period. Explain this apparent 13-7 contradiction. b) Calculate△G° for this reaction (c) How many ATP molecules can theoretically be gen- 15. Rates of Turnover of y and B Phosphates of ATP erated by this reaction if the free energy of ATP synthesis un- If a small amount of ATP labeled with radioactive phospho- der cellular conditions is 52 k/mol? rus in the terminal position, Iy PIATP, is added to a yeast 20. Dependence of Electromotive Force on Conce few minutes. but the concentration of atp remains un hanged. Explain If the same experiment is carried out us- tered by an electrode immersed in a solution containing the ing ATP labeled with"P in the central position, (B-PJATP following mixtures of NAD and NADH at pH 7.0 and 25C, the p does not appear in P within such a short time. Why? with reference to a half-cell of E000V (a)1.0 mM NAD* and 10 mM NADH 16. Cleavage of ATP to AMP and PPi during Metab (b) 1.0 mM NAD and 1.0 mM NADH lism The synthesis of the activated form of acetate(acetyl (c) 10 mM NAD and 1.0 mM NADH CoA) is carried out in an ATP-dependent proces 21. Electron Affinity of Compounds List the following Acetate CoA ATP- acetyl-CoA AMP +PP substances in order of increasing tendency to accept ele )The△C° for the hydrolysis of acetyl-CoA to acetatealoacetate()Oa(Wm、ate):()ax trons:(a)a-ketoglutarate +CO and CoA is-322 k/mol and that for hydrolysis of ATP to AMP and PP is.5 kJ/mol Calculate Ago for the atP. 22. Direction of Oxidation- Reduction Reactions Which dependent synthesis of acetyl-CoA of the following reactions would you expect to proceed in the rophosphatase, which catalyzes the hydrolysis of PP, to P. the appropriate enzymes are present to catalyze thema that (b) Almost all cells contain the enzyme inorganic py- direction shown, under standard conditions, assuming that What effect does the presence of this enzyme have on the (a) Malate+ NAD+- oxaloacetate NADH +H+ (b) Acetoacetate NADH +h- 17. Energy for H* Pumping The parietal cells of the (c)Pyruvate+NADH +H+- lactate +NAD+AD* B-hydroxybutyrate +NA stomach lining contain membranepumps"that transport hy (d) Pyruvate+ B-hydroxybutyrate→→ drogen ions from the cytosol of these cells(pH 7.0)into the lactate acetoacetate stomach, contributing to the acidity of gastric juice (pH 1.0) Calculate the free energy required to transport 1 mol of hy (e) Malate+ pyruvate→→ oxaloacetate+ lactate drogen ions through these pumps. (Hint: See Chapter 11.) ) Acetaldehyde+ succinate→→ ethanol+ fumarate Assume a temperature of 25C

520 Chapter 13 Principles of Bioenergetics 13. Free Energy Required for ATP Synthesis under Physiological Conditions In the cytosol of rat hepato￾cytes, the mass-action ratio, Q, is [A [ D A P T ] P [P ] i ]  5.33 102 M1 Calculate the free energy required to synthesize ATP in a rat hepatocyte. 14. Daily ATP Utilization by Human Adults (a) A total of 30.5 kJ/mol of free energy is needed to synthesize ATP from ADP and Pi when the reactants and products are at 1 M concentrations (standard state). Because the actual physiological concentrations of ATP, ADP, and Pi are not 1 M, the free energy required to synthesize ATP un￾der physiological conditions is different from G . Calculate the free energy required to synthesize ATP in the human he￾patocyte when the physiological concentrations of ATP, ADP, and Pi are 3.5, 1.50, and 5.0 mM, respectively. (b) A 68 kg (150 lb) adult requires a caloric intake of 2,000 kcal (8,360 kJ) of food per day (24 h). The food is me￾tabolized and the free energy is used to synthesize ATP, which then provides energy for the body’s daily chemical and me￾chanical work. Assuming that the efficiency of converting food energy into ATP is 50%, calculate the weight of ATP used by a human adult in 24 h. What percentage of the body weight does this represent? (c) Although adults synthesize large amounts of ATP daily, their body weight, structure, and composition do not change significantly during this period. Explain this apparent contradiction. 15. Rates of Turnover of and  Phosphates of ATP If a small amount of ATP labeled with radioactive phospho￾rus in the terminal position, [- 32P]ATP, is added to a yeast extract, about half of the 32P activity is found in Pi within a few minutes, but the concentration of ATP remains un￾changed. Explain. If the same experiment is carried out us￾ing ATP labeled with 32P in the central position, [- 32P]ATP, the 32P does not appear in Pi within such a short time. Why? 16. Cleavage of ATP to AMP and PPi during Metabo￾lism The synthesis of the activated form of acetate (acetyl￾CoA) is carried out in an ATP-dependent process: Acetate CoA ATP 8n acetyl-CoA AMP PPi (a) The G for the hydrolysis of acetyl-CoA to acetate and CoA is 32.2 kJ/mol and that for hydrolysis of ATP to AMP and PPi is 30.5 kJ/mol. Calculate G for the ATP￾dependent synthesis of acetyl-CoA. (b) Almost all cells contain the enzyme inorganic py￾rophosphatase, which catalyzes the hydrolysis of PPi to Pi . What effect does the presence of this enzyme have on the synthesis of acetyl-CoA? Explain. 17. Energy for H Pumping The parietal cells of the stomach lining contain membrane “pumps” that transport hy￾drogen ions from the cytosol of these cells (pH 7.0) into the stomach, contributing to the acidity of gastric juice (pH 1.0). Calculate the free energy required to transport 1 mol of hy￾drogen ions through these pumps. (Hint: See Chapter 11.) Assume a temperature of 25 C. 18. Standard Reduction Potentials The standard re￾duction potential, E , of any redox pair is defined for the half-cell reaction: Oxidizing agent n electrons 8n reducing agent The E values for the NAD/NADH and pyruvate/lactate con￾jugate redox pairs are 0.32 V and 0.19 V, respectively. (a) Which conjugate pair has the greater tendency to lose electrons? Explain. (b) Which is the stronger oxidizing agent? Explain. (c) Beginning with 1 M concentrations of each reactant and product at pH 7, in which direction will the following re￾action proceed? Pyruvate NADH H 34 lactate NAD (d) What is the standard free-energy change (G ) at 25 C for the conversion of pyruvate to lactate? (e) What is the equilibrium constant (K eq) for this reaction? 19. Energy Span of the Respiratory Chain Electron transfer in the mitochondrial respiratory chain may be rep￾resented by the net reaction equation NADH H  1 2  O2 34 H2O NAD (a) Calculate the value of E for the net reaction of mitochondrial electron transfer. Use E values from Table 13–7. (b) Calculate G for this reaction. (c) How many ATP molecules can theoretically be gen￾erated by this reaction if the free energy of ATP synthesis un￾der cellular conditions is 52 kJ/mol? 20. Dependence of Electromotive Force on Concen￾trations Calculate the electromotive force (in volts) regis￾tered by an electrode immersed in a solution containing the following mixtures of NAD and NADH at pH 7.0 and 25 C, with reference to a half-cell of E 0.00 V. (a) 1.0 mM NAD and 10 mM NADH (b) 1.0 mM NAD and 1.0 mM NADH (c) 10 mM NAD and 1.0 mM NADH 21. Electron Affinity of Compounds List the following substances in order of increasing tendency to accept elec￾trons: (a) -ketoglutarate CO2 (yielding isocitrate); (b) ox￾aloacetate; (c) O2; (d) NADP. 22. Direction of Oxidation-Reduction Reactions Which of the following reactions would you expect to proceed in the direction shown, under standard conditions, assuming that the appropriate enzymes are present to catalyze them? (a) Malate NAD 8n oxaloacetate NADH H (b) Acetoacetate NADH H 8n -hydroxybutyrate NAD (c) Pyruvate NADH H 8n lactate NAD (d) Pyruvate -hydroxybutyrate 8n lactate acetoacetate (e) Malate pyruvate 8n oxaloacetate lactate (f) Acetaldehyde succinate On ethanol fumarate