Trapping the ATP binding state leads to a detailed understanding of the F1-ATPase mechanism.

ABSTRACT

The rotary motor enzyme FoF1-ATP synthase uses the proton-motive force across a membrane to synthesize ATP from ADP and Pi (H2PO4(-)) under cellular conditions that favor the hydrolysis reaction by a factor of 2 × 10(5). This remarkable ability to drive a reaction away from equilibrium by harnessing an external force differentiates it from an ordinary enzyme, which increases the rate of reaction without shifting the equilibrium. Hydrolysis takes place in the neighborhood of one conformation of the catalytic moiety F1-ATPase, whose structure is known from crystallography. By use of molecular dynamics simulations we trap a second structure, which is rotated by 40° from the catalytic dwell conformation and represents the state associated with ATP binding, in accord with single-molecule experiments. Using the two structures, we show why Pi is not released immediately after ATP hydrolysis, but only after a subsequent 120° rotation, in agreement with experiment. A concerted conformational change of the α3ß3 crown is shown to induce the 40° rotation of the γ-subunit only when the ßE subunit is empty, whereas with Pi bound, ßE serves as a latch to prevent the rotation of γ. The present results provide a rationalization of how F1-ATPase achieves the coupling between the small changes in the active site of ßDP and the 40° rotation of γ.