THE ALPHA BETA SUBUNIT INTERACTION IN H+-ATPASE (ATP SYNTHASE) - AN ESCHERICHIA-COLI ALPHA-SUBUNIT MUTATION (ARG-ALPHA-296 -] CYS) RESTORESCOUPLING EFFICIENCY TO THE DELETERIOUS BETA-SUBUNIT MUTANT (SER-BETA-174 -] PHE)/

The Ser-beta 174 residue of the Escherichia coli H+-ATPase beta subuni t has been shown to be near the catalytic site together with Gly-beta 149, Gly-beta 172, Glu-beta 192, and Val-beta 198 (Iwamoto, A., Park, M.-Y., Maeda, M., and Futai, M. (1993) J. Biol. Chem. 268, 3156-3160). In this study, we introduced various residues at position 174 and fou nd that the larger the side chain volume of the residue introduced, th e lower the enzyme activity became. The Phe-beta 174 mutant was defect ive in energy coupling between catalysis and transport, whereas the Le u-beta 174 mutant could couple efficiently, although both mutants had essentially the same ATPase activities (similar to 10% of the wild typ e). The defective energy coupling of the Phe-beta 174 mutant was suppr essed by the second mutation (Arg-alpha 296 --> Cys) in the alpha subu nit. The Cys-alpha 296/Phe-beta 174 mutant had essentially the same me mbrane ATPase activity as the Phe-beta 174 single mutant when assayed under the conditions that stabilize the double mutant enzyme. These re sults indicate the importance of the alpha/beta interaction, especiall y that between the regions near Arg-alpha 296 and Ser-beta 174, for en ergy coupling in the H+-ATPase. The 2 residues (Ser-beta 174 and Arg-a lpha 296) may be located nearby at the interface of the two subunits. About 1 mol of N-[C-14]ethylmaleimide could bind to 1 mol of the alpha subunit of Cys-alpha 296/Phe-beta 174 or Cys-alpha 296 mutant ATPase, but could not inhibit the enzyme activity. This is the first intersub unit mutation/suppression approach to ATPase catalysis and its energy coupling.