Role of a bound ubiquinone on reactions of the Escherichia coli cytochromebo with ubiquinol and dioxygen

To probe the functional role of a bound ubiquinone-8 in cytochrome be-type ubiquinol oxidase from Escherichia coli we examined reactions with ubiquino l-l and dioxygen. Stopped-flow studies showed that anaerobic reduction of t he mild-type and the bound ubiquinone-free (Delta UbiA) enzymes with ubiqui nol-l immediately takes place with four kinetic phases, Replacement of the bound ubiquinone with 2,6-dibromo-4-cyanophenol (PC32) suppressed the anaer obic reduction of the hemes with ubiquinol-l by eliminating the fast phase. Flow-flash studies in the reaction of the fully reduced enzyme with dioxyg en showed that the heme b to heme o electron transfer occurs with a rate co nstant of similar to 10(4) s(-1) in all three preparations. These results s upport our previous proposal that the bound ubiquinone is involved in facil e oxidation of substrates in subunit II and subsequent intramolecular elect ron transfer to low-spin heme 6 in subunit I. (C) 1999 Federation of Europe an Biochemical Societies.