ENERGY COUPLING, TURNOVER, AND STABILITY OF THE F0F1 ATP SYNTHASE AREDEPENDENT ON THE ENERGY OF INTERACTION BETWEEN GAMMA-SUBUNIT AND BETA-SUBUNIT

Replacement of the F0F1 ATP synthase gamma subunit Met-23 with Lys (ga mma M23K) perturbs coupling efficiency between transport and catalysis (Shin, K., Nakamoto, R. K., Maeda, M., and Futai, M. (1992) J. Biol C hem. 267, 20835-20839). We demonstrate here that the gamma M23K mutati on causes altered interactions between subunits. Binding of delta or e psilon subunits stabilizes the alpha(3) beta(3) gamma complex, which b ecomes destabilized by the mutation. Significantly, the inhibition of F-1 ATP hydrolysis by the epsilon subunit is no longer relieved when t he gamma M23K mutant F-1 is bound to F-0. Steady state Arrhenius analy sis reveals that the gamma M23K enzyme has increased activation energi es for the catalytic transition state. These results suggest that the mutation causes the formation of additional bonds within the enzyme th at must be broken in order to achieve the transition state. Based on t he x-ray crystallographic structure of Abrahams et al. (Abrahams, J.P. , Leslie, A. G. W., Lutter, R., and Walker, J.E. (1994) Nature 370, 62 1-628), the additional bond is likely due to gamma M23K forming an ion ized hydrogen bond with one of the beta Glu-381 residues. Two second s ite mutations, gamma Q269R and gamma R242C, suppress the effects of ga mma M23K and decrease activation energies for the gamma M23K enzyme. W e conclude that gamma M23K is an added function mutation that increase s the energy of interaction between gamma and beta subunits. The addit ional interaction perturbs transmission of conformational information such that epsilon inhibition of ATPase activity is not relieved and co upling efficiency is lowered.