Structural and functional analysis of aa3‐type and cbb3‐type cytochrome c oxidases of Paracoccus denitrificans reveals significant differences in proton‐pump design

Abstract

In Paracoccusdenitrificans the aa₃‐type cytochrome c oxidase and the bb₃‐type quinol oxidase have previously been characterized in detail, both biochemically and genetically. Here we report on the isolation of a genomic locus that harbours the gene cluster ccoNOQP, and demonstrate that it encodes an alternative cbb₃‐type cytochrome c oxidase. This oxidase has previously been shown to be specifically induced at low oxygen tensions, suggesting that its expression is controlled by an oxygen‐sensing mechanism. This view is corroborated by the observation that the ccoNOQP gene cluster is preceded by a gene that encodes an FNR homologue and that its promoter region contains an FNR‐binding motif. Biochemical and physiological analyses of a set of oxidase mutants revealed that, at least under the conditions tested, cytochromes aa₃, bb₃. and cbb₃ make up the complete set of terminal oxidases in P. denitrificans. Proton‐translocation measurements of these oxidase mutants indicate that all three oxidase types have the capacity to pump protons. Previously, however, we have reported decreased H⁺/e coupling efficiencies of the cbb₃‐type