Freshly prepared mitochondria were incubated with b -hyroxybutyrate, oxidized cytochrome c, ADP, Pi, and cyanide. b -Hyroxybutyrate is oxidized by an NAD+ -dependent dehydrogenase.

The experimenter measured the rate of oxidation of β-hyroxybutyrate and the rate of formation of ATP.
(a) Indicate the probable flow of electrons in this system.
(b) How many moles of ATP would you expect to be formed per mole of β-hyroxybutyrate oxidized in this system?
(c) Why is β-hyroxybutyrate added rather than NADH?
(d) What is the function of the cyanide?
(e) Write a balanced equation for the overall reaction occurring in this system (electron transport and ATP synthesis).
(f) Calculate the net standard free energy change (ˆ†G°€²) in this system, using E€²0 values from Table 14.1 and a ˆ†G°€² value for ATP hydrolysis of - 32.2 kJ / mol.

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COC CoO CH H-C-OH CH CH NAD+ NADH I H CH3 TABLE 14.1 Standard reduction potentials of interest in biochemistry Oxidant Acetate CO2 2H 2e Succinate+ CO2+ 2H+ + 2e- Acetate 3H+2e Femedoxin (oxidized e 2H 2e α-Ketoglutarate+ CO2+ 2H+十2e Acetoacetate 2H2e Pyruvate CO2H 2e NAD + + H+ + 2e- NADPt H+ 2e Lipoate (oxidized)2H 2e 1,3-Bisphosphoglycerate 2H 2e Glutathione (oxidized)+ 2H+ + 2e- FAD (free coenzyme)2H 2e Acetaldehyde 2H +2e Pyruvate2H 26 oxaloacetate + 2H + + 2e- Reductant Pyruvate H20 a-Ketoglutarate H20 Acetaldehyde H2O Femedoxin (reduced) E" (V) -0.70 -0.67 -0.60 0.43 -0.42 -0.38 -0.35 -0.33 -0.32 0.32 -0.29 -0.29 -0.23 -0.22 -0.20 -0.19 lsocitrate B-Hydroxybutyrate Malate NADH NADPH Lipoate (reduced Glyceraldehyde-3-phosphate P 2 Glutathione (reduced) FADH2 Ethanol Lactate Malate 2 2 Oz (superoxide) Glutamate H2O FADH2 (enzyme-bound) -0.16 O2+ e a-Ketoglu FAD (enzyme-bound) 2H 2e Methylene blue (oxidized)+ 2H+ + 2e Fumarate2H+2e tarate NH4 2H 2e 2 -0.14 0 to -0.30 0.01 0.03 0.04 0.06 0.07 0.23 0.25 0.29 0.30 0.36 0.42 0.55 0.77 0.82 ethylene blue (reducea Succinate QH2 Ascorbate Cytochrome b (+2) Cytochrome c1(+2) Q2H2e Dehydroascorbate2H 2e Cytochrome b (+3) e Cytochrome ci(+3) e Cytochrome c (+3e Cytochrome a (+3)e 02 2H+ + 2e- Fericyanide2e NO3 (Nitrate) 2H2e Cytochrome a2(+3)e Fe (+3)e O22H2e 2 Cytochromec (+2) Cytochromea (+2) H202 Ferocyanide NO2 (Nitrite)+H2O Cytochrome a (+2) Fe(+2) H2O 2 2