Observations of rotation within the FOFl-ATP synthase:: deciding between rotation of the FOc subunit ring and artifact

F0F1-ATP synthase mediates coupling of proton flow in F-0 and ATP synthesis/hydrolysis in F(0)c through rotation of central rotor subunits. A ring structure of F(0)c subunits is widely believed to be a part of the rotor. Using an attached actin filament as a probe, we have observed the rotation of the F(0)c subunit ring in detergent-solubilized F0F1-ATP synthase purified from Escherichia coli. Similar studies have been performed and reported recently [Sambongi ct al, (1999) Science 286, 1722-1724], However. in our hands this rotation has been observed only for the preparations which show poor sensitivity to dicyclohexylcarbodiimde, an F-0 inhibitor. We hare found that detergents which adequately disperse the enzyme for the rotation assay also tend to transform F0F1-ATP synthase into an F-0 inhibitor-insensitive state in which Fl can hydrolyze ATP regardless of the state of the F-0. Our results raise the important issue of whether rotation of the F(0)c ring in isolated F0F1-ATP synthase can be demonstrated unequivocally with the approach adopted here and also used by Sambongi et al. :(C) 2000 Federation of European Biochemical Societies.