Isotope and thermal effects in chemiosmotic coupling to the membrane ATPase of Streptococcus.

We measured rates of ATP synthesis by the proton-translocating ATPase of the motile Streptococcus strain V4051. Starved cells were energized artificially by exposing their membranes to a variable electrical potential difference (internal medium negative) and a fixed pH difference (internal medium alkaline). The initial rates of ATP synthesis increased exponentially with protonmotive force. The results were the same in D2O and H2O; there was no solvent isotope effect. At a fixed protonmotive force, the rates were strongly dependent on temperature, as expected for a reaction with a large enthalpy of activation. At a different protonmotive force, the rates varied with temperature in an identical fashion; there was no change in the enthalpy of activation. We conclude that protonation-deprotonation steps are not rate limiting and that the protons that cross the membrane drive ATP synthesis by mass action. The transmembrane electric field acts by changing the concentrations of the reactants, not by changing the configuration of the enzyme-substrate complex.