The symbiotically essential cbb(3)-type oxidase of Bradyrhizobium japonicum is a proton pump

Purified cbb(3)-type oxidase of Bradyvhizobirrmjapo-nicum was reconstituted into phospholipid vesicles. Tight vesicles were obtained as shown by the d isturbance of Delta pH with CCCP and the membrane potential with valinomyci n, which led to a sixfold increase in cytochrome c oxidase activity. The ve sicles were thus suitable for proton translocation experiments. In the pres ence of valinomycin, a pulse with reduced cytochrome c caused an acidificat ion with a subsequent alkalinization, whereas the same pulse caused only an alkalinization in the presence of valinomycin plus CCCP. We conclude that the cbb(3)-type oxidase of B. japonicum is a proton pump. (C) 2000 Federati on of European Biochemical Societies.