Identification of a novel quinone-binding site in the cytochrome bo complex from Escherichia coli.
The cytochrome bo complex is a heme BO-type heme-copper quinol oxidase in the aerobic respiratory chain of Escherichia coli and functions as an electron transfer-linked proton pump. To study the protein-mediated electron transfer from substrates to metal centers, we carried out quantitative and qualitative analyses of a bound quinone in the purified oxidase and found that it has a novel high affinity ubiquinone-binding site distinct from the quinol oxidation site. Enzymatic and spectroscopic studies suggest that the quinone-binding site is located close to both the quinol oxidation site in subunit II and low-spin heme B in subunit I. The quinone-binding site of a bound ubiquinone-free oxidase was reconstituted with the potent quinol oxidation site inhibitor 2,6-dichloro-4-nitrophenol, which decreased the Vmax value of the ubiquinol-1 oxidase activity to one-fourth of the control activity. These results indicate that the quinone-binding site is essential for the catalytic functions of the cytochrome bo complex and mediates electron transfer from the quinol oxidation site to the low-spin heme.
Department of Plant Sciences, Graduate School of Science, University of Tokyo, Japan.
M
M
Sato-Watanabe
T
T
Mogi
T
T
Ogura
T
T
Kitagawa
H
H
Miyoshi
H
H
Iwamura
Y
Y
Anraku
D000388:Air; D003573:Cytochrome b Group; D003580:Cytochromes / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D007700:Kinetics; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism; D011809:Quinones / Q000378:metabolism*; D013379:Substrate Specificity
D003573:Cytochrome b Group; D003580:Cytochromes; D029968:Escherichia coli Proteins; D011809:Quinones; C017168:cytochrome bo, E coli; D010088:Oxidoreductases; C020094:duroquinol oxidase