SUBUNIT-B OF THE MEMBRANE MOIETY (F-0) OF ATP SYNTHASE (F1F0) FROM ESCHERICHIA-COLI IS INDISPENSABLE FOR H+ TRANSLOCATION AND BINDING OF THE WATER-SOLUBLE F1 MOIETY

The ATP synthase complex, designated F1F0, of Escherichia coli is composed of a water-soluble portion (F1; membrane-associated ATPase, EC 3.6.1.3) with ATP-hydrolyzing activity and a membrane-integrated part (F0) with H+-translocating activity. F0 is built from 3 kinds of subunits (a, b and c). The F0 portion was isolated directly from membranes of an E. coli strain (KY 7485) that overproduces the enzyme several fold. Subunit b was extracted from purified F0 by 2 methods. One method included prolonged incubation of the F0 complex in the presence of trichloroacetate (2.5 M) and the separation of subunit b and an a-c complex by gel filtration. Alternatively, subunit b was extracted by deoxycholate and separated from the a-c complex by hydrophobic-interaction chromatography. Integrated into liposomes, the a-c complex exhibited neither H+ uptake nor binding of F1. However, a functional F0 compex was reconstituted by adding stoichiometric amounts of subunit b to the a-c complex.