Mitochondrial ATPase : biochemical and molecular genetic analysis

S.cerevisiae mutants were isolated showing nuclear-coded resistance to the antibiotic venturicidin, a known F0ATPase binding antibiotic. Two types of mutant were identified, one of which had cross-resistance to a variety of antibiotics and appeared linked to the leu1 locus on chromosome VII, and one which was cross-resistant to chloramphenicol only and not linked to leu1. The level of resistance to venturicidin was not increased in isolated mitochondria, therefore resistance shown in both groups is believed to be due to a decrease in plasma membrane permeability to these antibiotics. The fluorescence properties of several organotin compounds, derivatives of the substituted flavones 3-hydroxyflavone (hof) and penta-hydroxyflavone (morin), were investigated on incubation with rat liver mitochondria. The compound Bu2SnBr(of) was found to show fluorescence enhancement when added to mitochondrial preparations, which could be lowered by addition of the non-fluorescent compound Bu3SnAc. Addition of Bu2SnBr(of) did not affect mitochondrial membrane potential, and conversely the energetic state of the mitochondrial inner membrane had no effect on Bu2SnBr(of) fluorescence. This compound was shown to be an inhibitor of mitochondrial ATPase, and is thought to have its binding site on the F0 moiety of that enzyme complex. The nuclear gene causing respiratory deficiency in the complementation group G57 was cloned and sequenced. This gene (PET57) encoded a protein of 36 Kdal which did not show significant homology with any known protein. The mutant strain was deficient in mitochondrial ATPase activity, but the major F1ATPase subunits were detected in mutant mitochondria, although in reduced amounts. Mutant F1 showed abnormal membrane binding and could not be isolated by standard methods. The protein encoded by the gene PET57 is transported into mitochondria and is thought to contribute to processing or assembly of one or more of the cytoplasmic subunits of the F1F0ATPase complex.