When pure reduced cytochrome c is added to carefully prepared mitochondria along with ADP, Pi , antimycin A, and oxygen, the cytochrome c becomes oxidized, and ATP is formed, with a P/O ratio approaching 1.0.
(a) Indicate the probable flow of electrons in this system.
(b) Why was antimycin A added?
(c) What does this experiment tell you about the location of coupling sites for oxidative phosphorylation?
(d) Write a balanced equation for the overall reaction (including cyt c oxidation and ATP synthesis).

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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