Role of a bound ubiquinone on reactions of the Escherichia coli cytochrome bo with ubiquinol and dioxygen.
To probe the functional role of a bound ubiquinone-8 in cytochrome bo-type ubiquinol oxidase from Escherichia coli, we examined reactions with ubiquinol-1 and dioxygen. Stopped-flow studies showed that anaerobic reduction of the wild-type and the bound ubiquinone-free (delta UbiA) enzymes with ubiquinol-1 immediately takes place with four kinetic phases. Replacement of the bound ubiquinone with 2,6-dibromo-4-cyanophenol (PC32) suppressed the anaerobic reduction of the hemes with ubiquinol-1 by eliminating the fast phase. Flow-flash studies in the reaction of the fully reduced enzyme with dioxygen showed that the heme b to heme o electron transfer occurs with a rate constant of approximately 10(4) s-1 in all three preparations. These results support our previous proposal that the bound ubiquinone is involved in facile oxidation of substrates in subunit II and subsequent intramolecular electron transfer to low-spin heme b in subunit I.
Mogi
T
T
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan. mogi@biol.s.u-tokyo.ac.jp
Sato-Watanabe
M
M
Miyoshi
H
H
Orii
Y
Y
D002384:Catalysis; D003573:Cytochrome b Group; D003580:Cytochromes / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006418:Heme / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D013059:Spectrum Analysis, Raman; D013997:Time Factors; D014451:Ubiquinone / Q000031:analogs & derivatives* / Q000378:metabolism / Q000502:physiology*
D003573:Cytochrome b Group; D003580:Cytochromes; D029968:Escherichia coli Proteins; D014451:Ubiquinone; D006418:Heme; C017168:cytochrome bo, E coli; C030778:ubiquinone 8; D010088:Oxidoreductases; C020094:duroquinol oxidase; C003741:ubiquinol; D010100:Oxygen