Membrane bioenergetic parameters in uncoupler-resistant mutants of Bacillus megaterium

Mutants of Bacillus megaterium displaying malate-driven ATP synthesis resistant to uncouplers of oxidative posphorylation are further characterized. Both the pH gradient and electrical potential generated across the membrane by malate respiration are equally sensitive to uncouplers in the wild type and uncoupler-resistant mutants. The mutants possess 0 to 10% of the wild type ATPase activity which is not activated by pretreatment with heat or trypsin. Despite this inability to measure ATPase activity, the mutants demonstrate acid-pulse-driven ATPase synthesis which is sensitive to uncouplers as well as malate-driven ATP synthesis which becomes uncoupler sensitive at pH 5.5. N,N' -Dicyclohexylcarbodiimide and valinomycin plus potassium inhibition of ATP synthesis is reversed by uncouplers in the mutants but not in the wild type. The data support the existence of a specific site on the ATPase complex for uncoupler binding which, if altered by mutation, affects uncoupler binding to the complex. The retention of malate-driven ATP synthesis in the absence of a significant pH gradient or electrical potential suggests that an alternative intermediate is involved in coupling oxidation to phosphorylation.