Oxidative phosphorylation in yeast II. An oxidative phosphorylation-deficient mutant

Abstract

1. Mitochondria from the mutant Saccharomyces cerevisiae DH 1 oxidized members of the tricarboxylic acid cycle, d(−)- and l(+)-lactates, ethanol, NADH, and N, N, N′, N′-tetramethyl- p-phenylenediamine with P/O ratios found in manometric experiments to be as low as 0.1 to 0.4. The low phosphorylation efficiency was not improved by a large excess of ADP or glucose + hexokinase, by serum albumin, or by the supernatant from an homogenate of non-mutant yeast. The weak phosphorylation was suppressed by oligomycin. In short-term polarographic experiments, higher P/O ratios, determined by a radioactive phosphate incorporation technique, were consistently found. 2. The oxidation activity of the mutant mitochondria was the same in the presence and absence of hexokinase + glucose and was not raised by ADP. 3. 2,4-Dinitrophenol and carbonylcyanide p-trifluoromethoxyphenylhydrazone at uncoupling concentrations stimulated the oxidation of all substrates with the exception of l(+)-lactate. This indicates that the coupling mechanism was not abolished in the mutant mitochondria. In media of about 0.15 osmolarity, Mg 2+ was required for dinitrophenol to stimulate the oxidation. 4. ATPase (ATP phosphohydrolase, EC 3.6.1.3) activity of mutant mitochondria displayed two pH optima like that from non-mutated yeast. However, the oligomycinsensitive component with pH optimum at pH 9.5 was substantially reduced in the mutant mitochondria. 5. Cytochrome a content was found to be lower in the mutant mitochondria. 6. Possible changes in the oxidative phosphorylation system of the mutant mitochondria are discussed.