The delta subunit of Escherichia coli ATP synthase has been expressed and purified, both as the intact polypeptide and as delta', a proteolytic fragment composed of residues 1-134. The solution structure of delta' as a five-helix bundle has been previously reported (Wilkens, S., Dunn, S. D., Chandler, J., Dahlquist, F. W., and Capaldi, R. A. (1997) Nat. Struct. Biol. 4, 198-201). The delta subunit, in conjunction with delta-depleted F-1-ATPase, was fully capable of reconstituting energy-dependent fluorescence quenching in membrane vesicles that had been depleted of F-1. A complex of delta with the cytoplasmic domain of the b subunit of F-0 was demonstrated and characterized by analytical ultracentrifugation using b(ST34-156), a form of the b domain lacking aromatic residues. Molecular weight determination by sedimentation equilibrium supported a b(2) delta subunit stoichiometry. The sedimentation coefficient of the complex, 2.1 S, indicated a frictional ratio of approximately 2, suggesting that delta and the b dimer are arranged in an end-to-end rather than side-by-side manner. These results indicate the feasibility of the b(2) delta complex reaching from the membrane to the membrane-distal portion of the F-1 sector, as required if it is to serve as a second stalk.
