Subunit rotation in Escherichia coli F0F1-ATP synthase during oxidative phosphorylation

We report evidence for proton-driven subunit rotation in membrane-bound FoF1-ATP synthase during oxidative phosphorylation, A beta D380C/gamma C87 crosslinked hybrid F-1 having epitope-tagged beta D380C subunits (beta(flag)) exclusively in the two noncrosslinked positions was bound to F-o in F-1-depleted membranes, After reduction of the beta-gamma crosslink, a brief exposure to conditions for ATP synthesis followed by reoxidation resulted in a significant amount of beta(flag) appearing in the beta-gamma crosslinked product. Such a reorientation of gamma C87 relative to the three beta subunits can only occur through subunit rotation, Rotation was inhibited when proton transport through F-o was blocked or when ADP and P-i were omitted, These results establish FoF1 as the second example in nature where proton transport is coupled to subunit rotation.