Control of the Rate of Reverse Electron Transport in Submitochondrial Particles by the Free Energy

The effects of the substrates and products on the rate and direction of ATP-dependent reverse electron transport from succinate to nicotinamide adenine dinucleotide (NAD) in bovine heart submitochondrial particles (ETPH) were investigated. Except at very low and very high concentrations of the reactants (relative to their apparent K_ms), the rate of reverse electron transport depends linearly on log [ATP]/[ADP] [phosphate] at constant redox potential and on log [reduced NAD] [fumarate]/[NAD] [succinate] at constant phosphate potential. The process can be converted from reverse to forward electron transport by poising the redox potential at appropriate values and lowering the phosphate potential. Similarly, poising the phosphate potential and increasing the redox potential convert reverse electron transport into forward electron transport. In the state with no net electron transport, the free-energy change associated with ATP hydrolysis is lower than the change associated with the redox reaction's free energy by 3 to 4 kcal/mol. This indicates that the stoichiometry ATP/2 e^- of the process is 1.3 or greater. These findings are discussed in relation to the chemiosmotic mechanism of coupling. It is suggested that the observed stoichiometry might be the result of an H⁺/ATP ratio of 3 and an H+/2 e^- ratio of 4 for the span succinate → NAD. The effect of uncouplers on adenosine triphosphatase (ATPase) and reverse electron transport and the effect of NADH on the rate of the ATPase reaction suggest that the intrinsic coupling in ETPH particles is quite tight, at least in the first site, despite their low P/O ratio and lack of respiratory control.