The FoF1-ATP synthases of energy transducing membranes couple condensation of ADP with Pi to proton electrochemical gradients generated by electron transport processes. Fo is an integral membrane protein complex which mediates transmembrane proton complex which mediates transmembrane proton conduction. F1 is a peripheral complex containing the catalytic sites. When removed from the membrane, F1 is an ATPase. Five different subunits comprise F1 in the stoichiometry, alpha3beta3gammadeltaepsilon. The catalytic sites, which are on beta subunits, appear to be distant from the membrane in the intact synthase. The catalytic alpha3beta3 complex is connected to Fo by a narrow stalk comprised of other F1 subunits and parts of Fo subunits that protrude from the membrane. The working hypothesis generally considered for coupling ATP synthesis to the proton electrochemical gradient suggests that proton flow through Fo generates conformational changes which are propagated through the stalk to the major subunits of F1 where affinities of catalytic sites for substrates and product are ultimately affected. In addition to catalytic sites, F1 contains three other binding sites for adenine nucleotides. These are located at interfaces of alpha and beta subunits. Since a specific regulatory role has yet to be defined for these sites, they are called noncatalytic sites. The work proposed is focused on the function and cooperative interactions of three sites in F1 that are involved in catalysis or control of catalysis. These are catalytic sites, noncatalytic sites, and an anionic segment of beta subunits which binds inhibitory, amphiphilic cations. The anionic site is postulated to be an interaction site in beta for one or more subunits that comprise the stalk. Methods proposed to probe these sites include characterization of chemically modified or mutant forms of the enzyme which have impaired activity resulting from an alteration in one of the functional sites and selective crosslinking to establish points of interaction of those sites which are at interfaces of subunits. Elucidation of the mechanism and control of the ATP synthase will greatly enhance understanding of processes fundamental to the bioenergetics of mammalian cells.
