title: Characterization of a b2delta complex from Escherichia coli ATP synthase.
abstract: 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 F1-ATPase, was fully capable of reconstituting energy-dependent fluorescence quenching in membrane vesicles that had been depleted of F1. A complex of delta with the cytoplasmic domain of the b subunit of F0 was demonstrated and characterized by analytical ultracentrifugation using bST34-156, a form of the b domain lacking aromatic residues. Molecular weight determination by sedimentation equilibrium supported a b2delta 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 b2delta complex reaching from the membrane to the membrane-distal portion of the F1 sector, as required if it is to serve as a second stalk.
authors.affiliation: Department of Biochemistry, University of Western Ontario, London, Ontario N6A 5C1, Canada. sdunn@julian.uwo.ca
authors.forename: S D
authors.initials: SD
authors.lastname: Dunn
authors.forename: J
authors.initials: J
authors.lastname: Chandler
mesh_terms: D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D006180:Proton-Translocating ATPases / Q000737:chemistry* / Q000302:isolation & purification; D011796:Quinacrine; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D013050:Spectrometry, Fluorescence
chemical_list: D017931:DNA Primers; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D006180:Proton-Translocating ATPases; D011796:Quinacrine