The effect of energy state on the capacity of the H+-ATPase of inverted inner membrane vesicles of rat liver mitochondria to interact with a homogeneous inhibitor peptide from the same source [Cintron, N. M., & Pedersen, P. L. (1979) J. Biol. Chem. 254, 3439-3443] has been examined in some detail. The study has been conducted by using an assay procedure which allows both ATP synthetic and hydrolytic activities of the H+-ATPase to be monitored in the same assay system. When the purified inhibitor is incubated with inverted inner membrane vesicles in the presence of MgATP and then sedimented to remove excess inhibitor and MgATP, the ATPase activity of the H+-ATPase is markedly inhibited. When ATP synthesis is induced in the same assay system by initiation of respiration (addition of succinate), the synthetic rate proceeds with a brief lag phase, in the order of seconds, and then assumes a linear steady-state rate. Under these conditions, most of the peptide inhibitor remains associated with the inner membrane vesicles. It is released into the supernatant only when respiration is allowed to proceed several minutes. Inhibitor release is accompanied by a parallel rise in the capacity of the H+-ATPase to catalyze ATPase activity under nonenergized conditions (succinate absent). These results emphasize that binding of the peptide inhibitor to the H+-ATPase complex of rat liver and its release therefrom correlate well with the capacity of the enzyme to catalyze ATP hydrolysis rather than ATP synthesis. The lag phase in ATP synthesis when inhibitor is present is very brief and may reflect the time required for a respiration-induced electrochemical gradient to weaken the binding of the inhibitor to the enzyme surface. It would seem that if the peptide inhibitor is a regulatory molecule, one of its major roles in intact rat liver mitochondria may be to preserve newly synthesized ATP following a burst of “phosphorylating” respiration.
|Original language||English (US)|
|Number of pages||7|
|State||Published - Oct 1981|
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