Summary 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

In Paraeoeeus denitrifieans four classes of redox proteins are involved in the electron transfer from methylamine to oxygen: methylamine dehydrogenase (MADH), amicyanin, cytochrome e and cytochrome e oxidase. MADH and its electron acceptor amicyanin are indispensable for growth on methylamine. At least three different cytochromes e and two types of cytochrome e oxidase, cytochromes aa3 and ebb 3 , have previously been proposed to participate in the electron transfer pathways from methylamine to oxygen. In this study, participation of both cytochrome e oxidases and of the quinol oxidase (cytochrome bb 3 ) has indeed been confirmed by analysis of a series of oxidase mutants. Interestingly, a P. denitrifieans cytochrome e oxidase mutant (l1aaiehh 3 ) retains the capacity to oxidise methylamine. It is demonstrated that the oxidation ofthe cytochrome e pool in this mutant does not proceed via an alternative cytochrome e oxidase, but rather via an 'uphill' electron transfer through the bel complex to ubiquinone, driven by the membrane potential. The subsequent oxidation of ubiquinol proceeds via the only remaining terminal oxidase, the hh3-type quinol oxidase.