Biochemistry Exam 2 Answer Key 26 Questions

Answer Key No. on Test Correct Answer 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 A C B C C A C C E A B E D D B 16 1. 2. 3. 4. 5. 17 1. H,D 2. J 3. C,G 4. K 5. K,E,I 6. F 7. A 8. A,B Page 1 D A C B E 18 no proton motive force/membrane to drive ATP synthesis Like all enzymes, the F1 subunit of the ATP synthase catalyzes a reaction in both directions: ADP + Pi ATP + H2O The standard free-energy change (∆G °') for ATP hydrolysis is -30.5 kJ/ mol. With no proton motive force to drive the reaction toward ATP synthesis, the hydrolysis (ATPase activity) occurs spontaneously. 19 (a) 2 -> 1 -> 3 -> 5 -> 4 -> 6 (b) No; Fe-S has a larger E o', so will probably be the terminal acceptor (c) Yes; NADH can only give up 2e– at a time and they must be give one at a time to cyto c. (d) 10; First calculate ∆E o' = E o'(oxidant) - E o'(reductant) = +1.08 (-0.42) = +1.5 V ∆G °'¸ = -n ¡∆E o'= (-2)(100 kJ/V•mol)(1.5 V)= -300 kJ/2e∆G °' for ATP synthesis is given as +30 kJ/mol, so theoretically the flow of 2 electrons from FPb to H2O2 could drive the synthesis of (300 kJ)/(30.5 kJ/mol) = 10 mol ATP. Page 2 Answer Key No. on Test Correct Answer No addition No addition Drug Drug +DNP and/or CN - Time 15 50 O2 consumed ATP synthesized 20 +DNP +CN - Time 21 F 22 T 23 F 24 F 25 (a) False. Mitochondrial ATP synthesis will cease, but to compensate, cells will accelerate the production of ATP by glycolysis, preventing ATP levels from dropping to zero. (b) False. The acceleration of glycolysis noted above will actually increase the rate of glucose consumption. (c) False. Because electron transfer through the respiratory chain is tightly coupled to ATP synthesis, blocking ATP synthase blocks electron flow and oxygen consumption. (d) False. The citric acid cycle is an oxidative pathway, producing NADH. When electron flow from NADH to O2 is blocked, NADH accumulates, NAD+ is depleted, and the citric acid cycle slows for lack of an electron acceptor (NAD+). 26 Answer Key No. on Test Correct Answer 26 Page 3 Biochemistry II I. MULTIPLE CHOICE (45 points; 3 points each) Choose the BEST answer to the question and write the letter in the appropriate box. 1. Entry of acetyl-CoA into the citric acid cycle is decreased when: A. B. C. D. 2. Oxaloacetate uniformly labeled with 14C (i.e., with equal amounts of 14C in each of its carbon atoms) is condensed with unlabeled acetyl-CoA. After a single pass through the citric acid cycle back to oxaloacetate, what fraction of the original radioactivity will be found in the oxaloacetate? A. 3. all B. 3/4 C. 1/2 D. 1/3 E. 1/4 Which of the following statements about the metabolic fate of pyruvate is incorrect ? A. B. C. D. 4. the ratio of [ATP]/[ADP] is high. [AMP] is high. the ratio of [NAD+]/[NADH] is high. NADH is rapidly oxidized through the respiratory chain. Under aerobic conditions, pyruvate is oxidatively decarboxylated and enters the citric acid cycle. In yeast growing anaerobically, pyruvate is oxidized to 3phosphoglycerate to use up excess NAD+ generated by the early steps of glycolysis. In anaerobic muscle, pyruvate is converted to lactate. The pyruvate dehydrogenase complex converts pyruvate to acetyl-CoA. Which of the following statements about the oxidation of pyruvate in aerobic conditions in animal cells is correct? A. B. C. D. E. The methyl (-CH3) group is eliminated as CO2. The process occurs in the cytosolic compartment of the cell. One of the products of the reactions of the pyruvate dehydrogenase complex is acetyl-CoA. The pyruvate dehydrogenase complex uses all of these as cofactors: NAD+, lipoic acid, pyridoxal phosphate (PLP), and FAD. The reaction is so important to energy production that pyruvate dehydrogenase operates at full speed under all conditions. Page 2 5. Which of the following statements is not true of the reaction catalyzed by the pyruvate dehydrogenase complex? A. B. C. D. 6. A new compound is isolated from mitochondria and a claim is made that it represents a previously unrecognized carrier in the electron transfer chain, and it is given the name coenzyme Z. Which line of evidence do you feel is the least conclusive in assigning this compound a position in the electron transfer chain? A. B. C. D. 7. E. ATP synthesis will continue, but the P/O ratio will drop to 1. electron transfer from succinate to O2 will continue unabated. all ATP synthesis from the mitochondria will stop. electron transfer from NADH will cease, but O2 uptake will continue. energy diverted from the cytochromes will be used to make ATP, and the P/ O ratio will rise. Which of the following enzymatic activities would you expect to be decreased by thiamine deficiency? A. B. C. D. E. 9. When added to a mitochondrial suspension, coenzyme Z very rapidly and specifically is taken up by the mitochondria. Removal of coenzyme Z from the mitochondria results in a decreased rate of oxygen consumption. Alternate oxidation and reduction of the mitochondrion-bound coenzyme Z can be readily demonstrated. The rate of oxidation and reduction of mitochondrion-bound coenzyme is of the same order of magnitude as the overall rate of electron transfer in mitochondria as measured by oxygen consumption. If electron transfer in tightly coupled mitochondria is blocked with antimycin A, then: A. B. C. D. 8. The reaction occurs in the mitochondrial matrix. Both NAD+ and a flavin nucleotide act as electron carriers. Biotin participates in the decarboxylation. Two different cofactors/co-substrates containing -SH groups participate. citrate synthase isocitrate dehydrogenase a-ketoglutarate dehydrogenase complex fumarase malate dehydrogenase The citric acid cycle is frequently described as the major pathway of aerobic catabolism, which means that it is an oxygen-dependent degradative process. However, none of the reactions of the cycle directly involves oxygen as a reactant. Why is the pathway oxygen-dependent? A. B. C. D. E. Oxygen is part of all degradation as it's needed for oxidation reactions. Oxygen is required for mitochondria to breath, and if they can't the CAC does not operate In the mitochondria, the CAC is regulated by an oxygen-dependent factor that binds allosterically to citrate synthase such that it cannot undergo it's conformational change needed to protect the carbanion nucleophile. The CAC produces carbons dioxide, which was mistaken for oxygen by the early scientists using the Warburg apparatus. In the mitochondria, the CAC is regulated by energy charge and the production of ATP is oxygen dependent Page 3 10. If the mitochondrial electron carriers are artificially fully oxidized and NADH is then added to the system, the last carrier to become reduced is: A. B. C. D. E. 11. In eukaryotic cells, the enzymes of glycolysis are in the __________, and the enzymes of the citric acid cycle are in the _____________. A. B. C. D. E. 12. B. C. D. E. D. E. Succinate is a signal molecule that releases CO 2 from lysoomes Succinate is a signal molecule that releases CO 2 from mitochondria Succinate can be used to make cellulose, which when burned produces many moles of CO 2 . Succinate acts catalytically and is regenerated by the cycle to produce more CO 2 . Succinate is actually polymer with many carbon atoms Assume that 10 protons are pumped from the matrix per 1/2 O 2 reduced by electrons from NADH. Even so, the P/O ratio for this NADH differ in species with ATP Synthases that contain 9 c-subunits in the F 0 complex versus those with 12 c-subunits in the F 0 complex, which is _______, respectively. A. B. C. D. E. 15. The Eo' value must be more negative than +0.05 to make electron flow to them favorably Flavins are not part of the Q-cycle Flavins can only accept one electron Complex III does not bind to flavins The Eo' value must be more positive than +0.05 to make electron flow to them favorably In 1935, Szent-Gyorgi showed that the production of CO 2 in an extract of muscle increased when succinate was added. For every mole of succinate added, many extra moles (>>4) of CO 2 were produced. Why? A. B. C. 14. inner mitochondrial membrane; mitochondrial matrix cytosol; mitochondrial matrix mitochondrial matrix; cytosol cytosol; inner mitochondrial membrane inner mitochondrial membrane; cytosol Electrons from reduced ubiquinone (coenzyme Q or CoQH2), which has a standard reduction potential (Eo') of +0.05 V, do not flow through a flavin or flavoprotein (Eo' for flavin is (–0.20 to –0.04 V) because ______. A. 13. cytochrome a 3. coenzyme Q. cytochrome b . flavoprotein. ubiquinone. 3 versus 4 9 versus 12 4 versus 3 2.5 versus 2 2 versus 2.5 Which of following is a membrane-bound enzyme of the Kreb's Cycle that forms one of the complexes in Electron Transport Chain? A. NADH dehydrogenase B. Succinate dehydrogenase C. Cytochrome dehydrogenase D. Cytochrome reductase E. Cytochrome oxidase II. Page 4 MATCHING (23 points) 16. Match the enzyme in the Kreb's cycle, listed on the left, with the enzyme in glycolysis or fermentation that use the same mechanistic strategy from the list on the right. Answers used only once and all answers are used. (10 pts) 1. malate dehydrogenase 2. isocitrate dehydrogenase 3. succinate dehydrogenase A. B. C. D. E. class II aldolase GAPDH enolase LDH pyruvate decarboxylase 4. succinyl-CoA synthetase 5. E1 of α-ketoglutarate dehydrogenase (KGDH) complex 17. Match the attribute or strategy used in enzyme-catalyzed reaction mechanisms listed on the right with the enzymes listed on the left. List all that apply and place the letter in the boxes provided. These attributes are only for what the enzyme does in the direction of the cycle, not the reverse. All choices on the right are used and some used more than once. (13 pts) 1. E1 of pyruvate dehydrogenase (PDH) complex 2. E3 of KGDH complex 3. Succinyl-CoA synthetase 4. Malate dehydrogenase 5. Isocitrate dehydrogenase 6. Citrate synthase 7. Fumarase 8. Aconitase A. rehydration of double bond B. dehydration C. mixed anhydride with phosphate D. TPP-catalyzed decarboxylation E. manganese cofactor F. only acid/base catalysis (no cofactors) G. phospho-His intermediate H. decarboxylation alpha to carbonyl I. decarboxylation beta to carbonyl J. disulfide exchange K. Hydride transfer from alcohol carbon III. SHORT ANSWER & PROBLEMS. 18. (14 points) Page 5 When the F1 portion of the ATP synthetase complex is removed from the mitochondrial membrane and studied in solution, it functions as an ATPase. Why does it not function as an ATP synthetase? (2 pt) (8 words or less for credit) 19. A recently discovered bacterium grows in a peroxide-contaning environment and carries out ATP synthesis coupled to the flow of electrons through a chain of carriers to some electron acceptor. The components of its electron transfer chain differ from those found in mitochondria; they are listed below with their standard reduction potentials. Electron carriers in the newly discovered bacterium: (8 pt) Number 1 2 3 4 5 Oxidant NAD+ flavoprotein b (FPb ) (oxidized) cytochrome c (Fe3+) Fe-S protein (oxidized) flavoprotein a (FPa ) (oxidized) (reduced) cytochrome c (Fe2+) Fe-S protein (reduced) flavoprotein a 1 +0.22 2 +0.88 2 +0.77 2 +1.08 (reduced) H2O H2O2 6 Reductant NADH flavoprotein b E o' (V) -0.32 -0.42 Electrons transferred 2 2 (a) Place the numbers for each electron carrier in the order in which they are most likely to act. (b) Is it likely that O2 (for which Eo' = 0.82 V) is the final electron acceptor in this organism? (b) (c) Is it likely that any of these carriers would be involved in a "Qcycle"? (c) (d) (d) Calculate the maximum number of ATP molecules that could theoretically be synthesized, under standard conditions, per pair of electrons transfered through this chain of carriers? (The Faraday constant, ¡, is ~100 kJ/V•mol.) Assume the ∆G°' is –30 kJ/mol for ATP hydrolysis. No addition No addition Drug Drug Time O2 consumed ATP synthesized 20. One drug, Cyanide ion (CN-) and another drug, 2,4-Dinitrophenol (DNP) affect mitochondrial oxidative phosphorylation. Add labeled curves to the graphs below to show the effects of adding each of these compounds separately to a suspension of mitochondria supplied with O2, succinate, ADP and Pi. (Arrow indicates time of addition of drug)(you need to draw and label 4 curves) (4 pts) Time III. TRUE/FALSE. (8 points) Page 6 21. The condensation to make ATP from ADP + Pi in the active site on ATP Synthase has a DG°' of +7.3 kcal/mole. 22. The mechanism of ATP Synthase is termed the Binding-Change Model. 23. The catabolism of fructose in the liver begins with the phosphorylation to Fru 6-P. 24. Succinyl-CoA is a feedback inhibitor of citrate synthase. 25. Consider a liver cell carrying out the oxidation of glucose under aerobic conditions. Suppose that we added a very potent and specific inhibitor of the mitochondrial ATP synthase, completely inhibiting this enzyme. Tell whether each of the following statements about the effect of this inhibitor is true or false; if false, explain in a sentence why it is false. (a) ATP production in the cell will quickly drop to zero. (b) The rate of glucose consumption by this cell will decrease sharply. (c) The rate of oxygen consumption will increase. (d) The citric acid cycle will speed up to compensate. Page 7 V. PATHWAYS. (10 pts) 26. Write out the three steps in the TCA cycle from isocitrate to succinate. For each step give the name of the enzyme that catalyzes the reaction (no abbreviations) to the left, any cofactors required, if any (abbreviations OK), write in labeled boxes, any cosubstrates/products, write in boxes connected by curved arrows (if more than one, use semicolon to separate), the name of each metabolic intermediate (at the right) and their structures in the labeled box (including isocitrate and succinate; you can abbreviate phosphate and/or phosphoryl by a circled "P").