Fast ADP release in F1-ATPase.

Abstract

Recently, we proposed a theory-basaed method to analyze fast (10-100 µs timestep) single-molecule imaging trajectories in F1-ATPase. A key quantity in this method is the angular velocity vs. rotation angle extracted from both experimental data and computer simulations. When applying the method on Thermophilic Bacillus F1-ATPase rotation data, we detected a short-lived substep previously not detectable in the angular histograms. The comparison between the experimental and theory reveals that an 80O substep of the “concerted” ATP binding and ADP release involves an intermediate state reminiscent of a 3-occupancy structure. Its lifetime (∼10 µs) is about six orders of magnitude smaller than the lifetime for “spontaneous” ADP release from a singly occupied state. The theory-based method was also applied to single-molecule imaging data from Paracoccus Denitrificans F1-ATPase and it yielded a similar hidden state in the transitions between subsequent long dwells. The ∼20 µs lifetime is several times shorter then the experimental imaging frame time (100 µs), so by detecting this short-lived state the method was used to achieve temporal "super-resolution". Our recent findings indicate a common mechanism for the acceleration of ADP release in the F1-ATPase motor of the two species.