Regulation of ATP hydrolysis in hepatoma 22a mitochondria.

A decrease in the rate of ATP hydrolysis was observed after preincubation of intact mitochondria from hepatoma 22a with an uncoupler. This effect is due both to a decrease in the rate of ATP transport and to an inactivation of the F0F1-ATPase. The former effect is shown to result from an uncoupler-induced ADP efflux. In de-energized mitochondria from hepatoma (but not from mice liver), the concentration of adenine nucleotides in the matrix equilibrates with the medium concentration via a carboxyatractyloside (CATR)-insensitive transport system. CATR-insensitive accumulation of medium ADP and stoichiometric exchange of added ATP are observed in energized hepatoma mitochondria. The dependence of the uncoupler-induced inactivation of ATPase activity on delta mu H+, pH, and ATP is consistent with the effect being caused by the natural protein inhibitor (IF1) of F0F1. ATP- and pH-dependent inactivation of the enzyme is also observed after disruption of mitochondria with the detergent Lubrol-WX. Almost all F0F1 in hepatoma mitochondria have IF1 bound in a noninhibitory manner. In the presence of uncoupler, this complex converts, via a reversible pH-dependent and an irreversible ATP-dependent process, to an inhibitory complex. The pH-dependent step can be blocked by Zn2+ and Cd2+ ions which probably bind to negatively charged residues on IF1, thereby preventing their protonation and conversion of the protein to an inhibitory conformation.