Biochemistry Combination of Cofactors

Biochemistry MULTIPLE CHOICE (36 points; 3 points each) ) Choose I. the best answer as requested in each question and writing the appropriate letter in the Ьох at the left. Which comЬination of cofactors are among those involved in the conversion of pyruvate to acetyl-CoA? А. в. с. D. Е. NAD, ТРР, PLP, PLP, PLP, PLP, and ТРР lipoic acid, and NAD FAD, and lipoic acid FAD, and ТРР NAD, and FAD All of the following are located in the mitochondria except the enzymes: А. В. С. D. Е. of the citric acid cycle. of fatty acid oxidation. of glycolysis. and cofactors of electron transfer to oxygen. and cofactors of oxidative phosphorylation. Oxidation of one mole of acetyl-CoA via the citric-acid cycle results in net: А. в. с. D. Е. production of one mole of citrate. consumption of one mole of oxaloacetate. production of 7 moles of АТР. production of 2 moles of СО 2 • production of one mole of succinate. Which of the following is not true of the citric-acid cycle? А. В. С. D. Е. Oxaloacetate, citrate, and succinate function catalytically (not stoichiometrically) in the cycle. In the presence of malonate, one would expect succinate to accumulate. Succinate dehydrogenase channels electrons directly into the electron transfer chain. The citrate synthase is subject to allosteric regulation Ьу АТР and NADH. All enzymes of the cycle are located in the cytoplasm, except succinate dehydrogenase, which is bound to the inner mitochondrial memЬrane. Page 1 III.. MATCHING (14 points) Match the attribute of the mechanism in the each enzyme that we studied. These enzymes column, with the most appropriate attribute There is one best answer for each. Items on .than. once. 21. 0 . ·.o ·o 0 .□ □ 0 . . . . 0 0 0 reactions they catalyze with are listed in the left listed in the right col.umn. the right can be used more A. enolic,intermediate 1. Isocitrate dehydrogenase 2. Citrate synthase 3. Fumarase 4. Aconitase D. 4Fe-4S cluster for re-dox 5. Succinyl-CoA synthetase E. Q-cycle 6. a-ketoglutarate dehydrogenase (El) 7. cytochrome oxidase 8. E3 of the PDH complex 9. Malate dehydrogenase 10. 011. 12. 0 □ Page 4 013. 14. Acetoacetate decarboxylase NADH Dehydrogenase (Complex I) Succinate Dehydrogenase Pyruvate decarboxylase CoQ:Cytochrome-c Oxidoreductase (Complex III) B. �hosphci�Histidine intermedi,te c. 4Fe-4S cluster for dehydration/ rehydration F. Manganese. cofactcir to. polarize· bond at.beta .carbon G. Acid/base catalysis to generate a carbanion for nucleophic attack after induced fit H. Schiff-base to polarize bond at beta carbon I. Use of TPP carbanion for decarboxylation alpha to a carbonyl J. Copper re-dox cofactors K. Disulfide exchange with lipoic acid L. Stereo-specific hydride abstraction v. Page 6 SHORT ANSWER (15 points) 23. Individuals with a thiamine-deficient diet have beriberi, a disease caused by thiamine deficiency, have elevated levels of blood pyruvate and a-ketoglutarate, especially after consuming a meal rich in glucose. Explain how these effects are related to a deficiency of thiamine in biochemical terms? (1 pt) 24. Biochemists have often delineated the metabolic pathways of organic compounds by using a radioactively labeled substrate and following the fate of the label. (3 pt) How can you determine whether a suspension of isolated mitochondria metabolizes added glucose to carbon dioxide and water? Suppose you add a brief pulse of [3- 14 C] pyruvate (labeled at the methyl carbon) to the suspension of mitochondria. After one turn of the citric acid cycle, what is the location of the 14 C in the oxaloacetate? 25. How many turns of the cycle are required to release all the [3- 14 C] pyruvate as 14 C-C02? .__________.I .____I ___ □ Ce llular respir ation can be studied in isolated mitochondria by measuring oxygen (5 pt) consumption under different conditions using a Warburg apparatus. If 0.01 m sodium malonate is added to actively respiring mitochondria that are using pyruvate as fuel, respiration soon stops and a m etabolic interm ediate accumulates, as Hans Krebs discovered. What is the structure of this intermediate? [what is the lette red box for this �------------------------, structure from Problem #22] What is the structure of malonate? I Explain why it accumulates. .I . Explain why oxyge n consumption stops. Aside from removal of the malonate, what can overcome this inhibition of respiration? 26. When investigators add the antibiotic valinomycin to actively respiring mitochondr ia, several things happ en: the yield of ATP decrease s, the rate of oxygen consumption increases, heat is rele ased, and the pH gradient across the inner mitochondrial m embrane increases. (2 pt) Does valinomycin act as an uncoup ler or as an inhibitor of oxidative phosphorylation? L ------------- What part of the proton motive force does valinomycin directly affect, the pH gradient or the membrane potential? II -------------' --' L - _ 27.[This que stion comes from doing the Case Study on Achieve for Chapter 19] You return to the hospital, whe re you meet the physicians working on Dr. Tamm's case. He is still unconscious, but they have some critical lab results for you. Due to these results, and your previous sleuthing to fi1d out which toxins your suspected pharmaceutical scientists, Dr. Sooss, Dr. Strainj e, and Dr . Hoo, we re in possession of (antimycin A, Rotenone, and KCN, respective ly), who tried to kill Dr. Tamm? (4 pt) Page 1 No. on Test Correct Answer 1 2 3 4 5 6 7 8 9 10 11 12 B C D E B C D C B D D E 13 14 15 F 16 T 17 18 F F 19 T 20 T 21 T F 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. F G A C B I J K L H D A I E 23 Thiamine is required for the synthesis of TPP, a prosthetic group in the pyruvate dehydrogenase and a-ketoglutarate dehydrogenase complexes. A thiamine deficiency reduces the activity of these enzyme complexes and causes the observed accumulation of precursors. 24 Add uniformly labeled 14C-glucose, and check for the release of 14C-C02 Equally distributed in C-2 and C-3 of oxaloacetate; an infinite number of turns 25 Malonate is a competitive inhibitor of succinate dehydrogenase. Box 1. A block in the citric acid cycle stops NADH formation, which stops electron transfer, which stops respiration. A large excess of succinate (substrate) overcomes the competitive inhibition 26 Valinomycin acts as an uncoupler. It combines with K+ to form a complex that passes through the inner mitochondrial membrane, dissipating the membrane potential. ATP synthesis decreases, which causes the rate of electron transfer to increase. This results in an increase in the H+ gradient, 0 2 consumption, and amount of heat released 27 None of the three inhibitors could have caused these results, so Dr. Tamm's condition is not due to attempted homicide after all! Page 2 No. on Test Correct Answer 22 A<0tyl-CoA о tн, t-5 Сод Cltr•lt Ox•lo-Acot.ato l Nдо • M•lal• �00 но- �н, о-с-соо 1 оо 'Y с с оо но-с-соо сн,-<:оо Citric acid cycle н, ,. . н с соо соо " \1< " соо (()() О{) ,ю 1АDн, FAD I сн,- оо 1 тн, с f111-coo Succ;in■t• СОО н GTP jАП,) fС , S-CoA GOP �DP/ Р, ! Suc<.Jnyl•CoA -K•to9lutarate isocitrate kts sl.tde. k°'s e.ve.r��k�9 � u. '1\.е,е,с{ �о соммt� �о м