INVOLVEMENT OF CYTOCHROME-C-OXIDASE SUBUNIT-III IN ENERGY COUPLING

The role of the conserved acidic residues of subunit III of cytochrome c oxidase (COII) in energy transduction was investigated. Using a COI I deletion mutant of Paracoccus denitrificans, complemented with a pla smid expressing either the wild type (wt) COII gene or site-directed m utants of the COII gene, we measured cytochrome c oxidase-dependent AT P synthesis, respiration, and membrane potential. Cytochrome c oxidase -dependent ATP synthesis was attenuated in nonacidic mutants of either Glu(98) (E98A and E98Q), or Asp(259) (D259a) but not in the acidic mu tant E98D. The rates of respiration in the energy conversion-defective mutants were as high as or higher than that in the wt. The cytochrome c oxidase-induced increment of membrane potential in the nonacidic mu tants was similar to or higher than that in the wt. In contrast, when succinate-driven ATP synthesis was mediated solely by ubiquinol oxidas e (e.g., in the presence of myxothiazol), the rates of ATP synthesis i n the nonacidic mutants were higher than that in the wt. Moreover, myx othiazol, which inhibited succinate respiration as well as ATP synthes is in wt and E98D, stimulated ATP synthesis, while inhibiting succinat e respiration, in the nonacidic mutants. These results indicate that t he attenuation of energy conversion in these mutants is limited to cyt ochrome c oxidase and thus suggest that subunit III plays a role in en ergy conversion by cytochrome c oxidase.