The activity of the ATP synthase from Escherichia coli is regulated by the transmembrane proton motive force

The ATP synthase from Escherichia coli was reconstituted into liposomes from phosphatidylcholine/phosphatidic acid. The proteoliposomes were energized by an acid-base transition and a K+/valinomycin diffusion potential, and one second after energization, the electrochemical proton gradient was dissipated by uncouplers, and the ATP hydrolysis measurement was started. In the presence of ADP and P-i, the initial rate of ATP hydrolysis was up to 9-fold higher with pre-energized proteoliposomes than with proteoliposomes that had not seen an electrochemical proton gradient. After dissipating the electrochemical proton gradient, the high rate of ATP hydrolysis decayed to the rate without preenergization within about 15 s. During this decay the enzyme carried out approximately 100 turnovers. In the absence of ADP and P-i, the rate of ATP hydrolysis was already high and could not be significantly increased by pre-energization. It is concluded that ATP hydrolysis is inhibited when ADP and P-i are bound to the enzyme and that a high Delta<(mu)over tilde>(H)+ is required to release ADP and P-i and to convert the enzyme into a high activity state. This high activity state is metastable and decays slowly when Delta<(mu)over tilde>(H)+ is abolished. Thus, the proton motive force does not only supply energy for ATP synthesis but also regulates the fraction of active enzymes.