Electrical current generation and proton pumping catalyzed by the ba3‐type cytochrome coxidase from Thermus thermophilus

Several amino acid residues that have been shown to be essential for proton transfer in most cytochrome coxidases are not conserved in the ba3‐type cytochrome coxidase from the thermophilic eubacterium Thermus thermophilus. So far, it has been unclear whether the Th. thermophilusba3‐type cytochrome coxidase can nevertheless function as an electrogenic proton pump. In this study, we have combined charge translocation measurements on a lipid bilayer with two independent methods of proton pumping measurements to show that enzymatic turnover of the Th. thermophiluscytochrome coxidase is indeed coupled to the generation of an electrocurrent and proton pumping across the membrane. In addition to a ‘vectorial’ consumption of 1.0 H+/e−for water formation, proton pumping with a stoichiometry of 0.4–0.5 H+/e−was observed. The implications of these findings for the mechanism of redox‐coupled proton transfer in this unusual cytochrome coxidase are discussed.