The barbituate drug amytal inhibits complex I from passing electrons to Coenzyme Q. Circle your choice for each pair of options, if amytal is present. 9. a. Do you expect the mitochondria to contain NADH + H* -or- NAD ? FADH, -or- FAD? b. What is the expected redox state (oxidized "Ox" or reduced "Red") for complexes I-IV of ETC? i. Complex I Ox -or- Red iii. Complex IIl Ox -or- Red ii. Complex Il Ox -or- Red iv. Complex IV Ox -or- Red c. Will ATP be produced? Yes -or- No d. Is ETC/OxPhos uncoupled? Yes -or- No e. Will a proton gradient be present? Yes -or- No I
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- 1. Cyanide, oligomycin, and 2,4-dinitrophenol are all inhibitors of oxidative phosphorylation in mitochon- dria. Provide an explanation to the following conditions regarding these potent inhibitors. (a) Explain why adding cyanide to an active in vitro suspension of mitochondria blocks ATP synthesis. What happens to the rate of ATP synthesis when 2,4-dinitrophenol is added to this mitochon- drial suspension after it was treated with cyanide? (b) Explain why the rate of oxygen consumption decreases in an in vitro suspension of mito- chondria when oligomycin is added. What happens to the rate of oxygen consumption in this oligomycin- inhibited system after adding 2,4-dinitrophenol? Explain.9. The barbituate drug amytal inhibits complex I from passing electrons to Coenzyme Q. Circle your choice for each pair of options, if amytal is present. a. Do you expect the mitochondria to contain NADH + H* -or- NAD+ ? FADH, -or- FAD? b. What is the expected redox state (oxidized "Ox" or reduced "Red") for complexes I-IV of ETC? i. Complex I Ox -or- Red iii. Complex Il Ox -or- Red ii. Complex II Ox -or- Red iv. Complex IV Ox -or- Red C. Will ATP be produced? Yes -or- No d. Is ETC/QxPhos uncoupled? Yes -or- No Yes -or- No Will a proton gradient be present? e. 10 The inbibitor shown below is described as interacting with an enzvme by at least one T- 1 interaction and the D. Focus 目 States) w OE DELL (8)The ATP hydrolysis reaction (shown below) has a AG of -31 kJ mol-1. NH2 NH2 Do you predict that AS is positive or negative for this reaction. Explain in one sentence. а. .N H20 -OCH2 OCH2 Energy Phosphate ion ÓH ÓH Adenosine triphosphate (ATP) Adenosine diphosphate (ADP) b. How does the hydrophobic effect contribute to the AS you predicted in part a. Make sure to discuss the ordering of water molecules in your answer. Include a minimum of 3 sentances in your explaination.
- Compound A below is a key intermediate in the synthesis of keto-myo-inositol B. Suggest a synthetic route for the preparation of A from D-glucose C giving structures for all of the intermediates in your scheme. Ph3CO. Но, HO HO HO, HO 'HO. ОН ОН Ph CH3 А В(a) Consider the oxidation of malate to oxaloacetate by NAD*: malate + NAD+ → oxaloacetate + NADH + H+ In yeast mitochondria, where the pH = 8.1, this reaction is exergonic only at low oxaloacetate concentrations. Assuming a pH = 8.1, a temperature of 37 °C, and the steady-state concentrations given below, calculate the maximum concentration of oxaloacetate at which the reaction will still be exergonic. malate + NAD*→ oxaloacetate + NADH + H* lactate + NAD →→ pyruvate + NADH + H+ half reaction Pyruvate + 2H+ + 2e → lactate Pyruvate + CO₂ + H + 2e → malate Intracellular steady state concentrations: malate = 410 μM; NAD = 20.0 mM; pyruvate = 3.22 mM; NADH = 290 μM; AG=+29.7 kJ/mol AG¹ = +25.1 kJ/mol E° (V) - 0.190 - 0.330 lactate 1.1 mM CO₂ = 15.5 torr6. Malate dehydrogenase catalyzes the following reversible reaction: COO- HO-C-H CH₂ COO™ L-Malate NAD+ y malate NADH + H+ dehydrogenase COO- 0=C CH₂ COO™ Oxaloacetate AG'° = 29.7 kJ/mol Malate + NAD+→NADH + H+ + oxaloacetate Calculate AG" and the ratio or products and reactants for the malate dehydrogenase reaction to proceed from left to right as shown. (The Faraday constant. 3, is 96.48 kJ/V-mol; RT(37°C)= 2.58kJ/mol) Steps: 1. Explain how you determined which molecule is an electron donor Malate and which is an acceptor NAD*. -2- 2. Calculate AED (write equation, then show calculations, for standard reduction potentials (E_values) see table in the posted lecture) 3. Calculate AG (write equation, then show calculations) 4. Calculate the ratio of products and reactants needed to for Malate + NAD+→→NADH + H+ + oxaloacetate reaction to proceed forward (write equation, then show calculations)
- (b) the activity of the malate-aspartate shuttle (MAS) system of isolated rat brain mitochondria suspended in an isotonic me- dium buffered to pH 7.4. Diagram A illustrates NADH fluo- rescence emission upon addition of Glutamate in the ab- sence and presence of Aspartate. Diagram B illustrates sim- ilarly NADH fluorescence emission upon addition of Gluta- mate in the presence of Aspartate followed by additions of submicromolar concentrations of Ca2+. As is well estab- lished, the MAS in brain, skeletal muscle, and cardiac mus- Diagrams A and B on the right show changes in Glu A Glu В -No Asp + 0 +0.12 -0.48 + 16 0.81 +1.8 ++ Asp 10 min 2 min cle mitochondria is activated by cytosolic concentrations of Ca2* < 3 µM. To simulate the cytosolic part of the MAS, the following reagents were added to the medium: 4 units/ml glutamate-oxaloacetate transaminase, 6 units/ml malate dehydrogenase, 66 µM NADH, 5 mM aspartate, 5 mM malate, 0.5 mM ADP, 200 nM ruthenium red (to block the mitochondrial…In oxidative phosphorylation.... 1. Succinate contributes 2e- to Complex II and 2H+ to the mitochondrial proton gradient. 2. NADH in the matrix passes 2 e- to coenzyme Q via Complex I. 3. Complexes I, II, III, and IV each contribute to the matrix proton gradient. 4. O2 stabilizes the catalytically active conformation of Complex V. 5. Reversible protonation of c subunits leads to rotation of the Complex V gamma subunit. 6. Each β subunit can bind ATP tightly under the right conditions. 7. For every 3 protons that pass across the inner mitochondrial membrane, 1 ATP is produced. Choose all options that are true.1. Identify the oxidized coenzyme (letter abbreviation only) that participates in this reaction of the Kreb’s cycle. Succinate --> Fumarate 2. How many mol of NADH can be obtained upon the beta oxidation of stearic acid? 3. How many mol of ATP can be obtained upon the complete oxidation of 1 mol stearic acid? 4. How many steps in glycolysis in which ATO is converted to ADP?
- 2. Formation of OAA in mitochondria. In the last reaction of the TCA cycle, malate is dehydrogenated to regenerate the OAA necessary for the entry of acetyl-coA into the cycle: L-Malate + NAD+ → OAA + NADH + H+ AGo = 30.0 kJ/mol (a) Calculate the equilibrium ponstant for this reaction at 25 C. b) Because AGo assumes a standard pH of 7, the equilibrium constant calculated in (a) corresponds to K'eg _JOAA][NADH] [L-malate][NAD*] The measured concentration of L-malate in rat liver mitochondria is about 0.2 mM when the ratio [NAD*] / [NADH] is 10. Calculate the concentration of OAA at pH 7 in these mitochondria. (c) To appreciate the magnitude of the mitochondrial OAA concentration, calculate the number of OAA molecules in a single rat liver mitochondrion. Assume the mitochondrion is a sphere of diameter 2.0 µm. (From problem 10, chapter 16)23. An important step in glycolysis is the formation of ATP and pyruvate from phosphoenol-pyruvate (PEP) and ADP. PEP ADP pyruvate+ATP The equilibrium constant (Keq) for this reaction is approximately 2.5x10°. Calculate standard free energy change (AG°') for this reaction. Show your work. onege b. Is the reaction exergonic or endergonic at standard conditions? If, at equilibrium, the concentrations of ADP and ATP are 0.2 mM and 2.0 mM, respectively, what is the equilibrium concentration ratio of [pyruvate] to [PEP]? Show your work. с.2. Please, determine the answers of these multiple choices, they’re in a,b,c’s. A) The role of oxygen in oxidative phosphorylation (cellular breathing) is: -To be the last electron acceptor in the electron transfer chain -To hydrolyse carbohydrates - To add hydrogen ions to pyruvic acid at the end of glycolysis -To provide electrons for NADP reduction -To provide hydrogen ions B) Which molecule has the most potential energy? -glucose -phosphate -fructose 1-6 diphosphate -ethanol -I'ATP C) What is the product of fermentation in yeasts? -carbonic acid -oxygen -ATP -lactic acid -ethanol