Question

Mitochondria produce most of the cell's ATP by oxidative phosphorylation. Electrons from NADH and FADH2 formed in the TCA cycle are transported through the electron transport chain (ETC) which consists of four complexes (I, II, III and IV). Hereby, a proton gradient is formed across the inner membrane of the mitochondria which is used by the ATP synthase to form ATP. In isolated mitochondria, oxygen consumption rate (OCR) and rate of ATP synthesis can be determined, respectively. Seahorse and NOVOstar assays. It is desired to determine the consumption of oxygen and the rate of ATP synthesis in isolated mitochondria with succinate as a substrate instead of pyruvate. The experiment is carried out in the presence of two substances: 

Rotenone which inhibits ETC complex I 
Antimycin A which inhibits ETC complex III

QUESTION: Indicate how the consumption of oxygen and the rate of ATP synthesis will be affected by respectively. Rotenone and Antimycin A

Ps. Please only write a short and precise answer of how the Oxygen consumption and ATP production will be affected by BOTH Rotenone and Antimycin A.

Answer 1

1. Rotenone inhibit the mitochondrial respiratory chain. It inhibit the electron transfer from the iron-sulfur centers which cause the blockage in oxidative phosphorylation with decrease in ATP synthesis.

Effect on oxygen consumption :

1. On Rotenone concentration 0–1 nm:

• oxygen consumption rates remained between 100 and 97%.

2. On Rotenone increased above 1 nm:  

• oxygen consumption decrease rapidly due to decrease in activity of ATP synthesis in complex 1 of mitochondrial respiratory chain.

2. Antimycin A inhibit specifically oxidative phosphorylation result in cessation of ATP synthesis

• Antimycin A binding to the cytochrome c reductase which inhibit the oxidation of ubiquinone.

Effect on oxygen consumption :

Oxygen consumption stops due to cessation of oxidative phosphorylation and cessation of ATP synthesis.