The mode of action of dinitrophenols on the different phosphorylating steps

Abstract

1. 1. Determinations of the concentrations (caq)opt of a dinitrophenol that induced maximal respiration in a medium deficient in phosphate and phosphate acceptor was carried out with different alkylnitrophenols and substrates (succinate, pyruvate (+ malate), glutamate, β-hydroxybutyrate) at various pH's. 2. 2. The pH optimum of maximally uncoupled succinate oxidation was 7.4, of maximally uncoupled pyruvate oxidation 6.8. 3. 3. The p(c1)opt, where c1 is the calculated concentration of uncoupling phenol in the lipid phase, is a linear function of pH, the slope of the line relating these quantities being o with succinate and — 0.5 for NAD-linked subtrates; an exceptional case is formed when the respiration of pyruvate (+ malate) is uncoupled by alkyldinitrophenols, in which case the slope is o. 4. 4. From a consideration of the reaction kinetics it appears that for systems in which dp(c1)opt/dpH is zero, uncoupling and inhibition by phenols (″) is best explained by the reactions. A∼I+o→A+I−o I−o⇋I+o where A is an oxidized member of the respiratory chain, and A∼I an intermediate of oxidative phosphorylation. When the slope is −0.5 the sequence A∼I+o→A+I−o I−o+H+⇋I+o appears to be the most likely. 5. 5. On basis of the difference in the relation between p(caq)opt and pH for different NAD-linked substrates it is concluded that mitochondrial NAD+ associated with the different substrates is present in different compartments, and that the whole phosphorylation system connected with a particular substrate must be localized in the same mitochondrial compartment. The compartments differ in their lipophilic character.