The ATP-to-oxygen stoichiometries of oxidative phosphorylation by rat liver mitochondria. An analysis of ADP-induced oxygen jumps by linear nonequilibrium thermodynamics.

Uncertainty exists as to the proton stoichiometries of mitochondrial oxidative phosphorylation and consequently as to the ATP stoichiometries. In rat liver mitochondria, ADP/O ratios were determined from the total and extra oxygen consumed during ADP-stimulated respiration under conditions of quantitative conversion of ADP to ATP. For succinate, glutamate plus malate, 3-hydroxybutyrate, and 2-oxoglutarate, respectively, ADP/total O was 1.71, 2.71, 2.61, and 3.45. ADP/extra O was 2.03, 3.04, 3.23, and 4.15. The results were interpreted in terms of linear nonequilibrium thermodynamics. It was shown that ADP/extra O = Z/q where Z is the phenomenological stoichiometry and q is the degree of coupling. q was determined from the dependence of respiratory rate on delta Gp, the phosphorylation potential, and was about 0.98 for all substrates. The results were consistent with ideal ATP/O stoichiometries of 2 for succinate, 3 for glutamate plus malate, 3 or 3 1/4 for 3-hydroxybutyrate, and 4 for 2-oxoglutarate. Taking into account the oxidation-reduction free-energy changes measured across Sites 1 + 2 at static head (J.J. Lemasters, R. Grunwald, and R.K. Emaus J. Biol. Chem. 259, 3058-3063), an ideal ATP/O stoichiometry of 3 1/4 for 3-hydroxybutyrate is proposed. The lower ATP/O for glutamate plus malate is then accounted for by proton translocation linked to glutamate/aspartate exchange. The data suggest a new 13-proton scheme of chemiosmotic coupling in which proton stoichiometries are 3 for the F1Fo-ATPase, 1 for the exchange of ATP for ADP and Pi, and 5, 4, and 4 for Sites 1, 2, and 3.