Interactions between βD372 and γ subunit N-terminus residues γK9 and γS12 are important to catalytic activity catalyzed by Escherichia coli F1F0-ATP synthase

Substitution of Escherichia coli F1F0 ATP synthase residues beta D372 or gamma S12 with groups that are unable to form a hydrogen bond at this location decreased ATP synthase-dependent cell growth by 2 orders of magnitude, eliminated the ability of F1F0 to catalyze ATPase-dependent proton pumping in inverted E. coli membranes, caused a 15-20% decrease in the coupling efficiency of the membranes as measured by the extent of succinate-dependent acridine orange fluorescence quenching, but increased soluble F-1-ATPase activity by about 10%. Substitution of gamma K9 to eliminate the ability to form a salt bridge with beta D372 decreased soluble F-1-ATPase activity and ATPase-driven proton pumping by 2-fold but had no effect on the proton gradient induced by addition of succinate. Mutations to eliminate the potential to form intersubunit hydrogen bonds and salt bridges between other less highly conserved residues on the gamma subunit N-terminus and the beta subunits had little effect on ATPase or ATP synthase activities. These results suggest that the beta D372-gamma K9 salt bridge contributes significantly to the rate-limiting step in ATP hydrolysis of soluble F, while the beta D372-gamma S12 hydrogen bond may serve as a component of an escapement mechanism for ATP synthesis in which alpha beta gamma intersubunit interactions provide a means to make substrate binding a prerequisite of proton gradient-driven gamma subunit rotation.