Intramolecular electron transfer processes in Cu(B)-deficient cytochrome bo studied by pulse radiolysis.
The Escherichia coli cytochrome bo is a heme-copper terminal ubiquinol oxidase, and functions as a redox-driven proton pump. We applied pulse radiolysis technique for studying the one-electron reduction processes in the Cu(B)-deficient mutant, His333Ala. We found that the Cu(B) deficiency suppressed the heme b-to-heme o electron transfer two order of the magnitude (4.0 x 10(2) s(-1)), as found for the wild-type enzyme in the presence of 1 mM KCN (3.0 x 10(2) s(-1)). Potentiometric analysis of the His333Ala mutant revealed the 40 mV decrease in the E(m) value for low-spin heme b and the 160 mV increase in the E(m) value of high-spin heme o. Our results indicate that Cu(B) not only serves as one-electron donor to the bound dioxygen upon the O-O bond cleavage, but also facilitates dioxygen reduction at the heme-copper binuclear centre by modulating the E(m) value of heme o through magnetic interactions. And the absence of a putative OH(-) bound to Cu(B) seems not to affect the uptake of the first chemical proton via K-channel in the His333Ala mutant.
Kobayashi
Kazuo
K
Institute of Scientific and Industrial Research, Osaka University, Mihogaoka, Ibaraki, Osaka, Japan.
Tagawa
Seiichi
S
Mogi
Tatsushi
T
D000409:Alanine / Q000235:genetics; D019943:Amino Acid Substitution / Q000187:drug effects; D003300:Copper / Q000378:metabolism*; D003573:Cytochrome b Group / Q000378:metabolism*; D004579:Electron Transport / Q000187:drug effects; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006418:Heme / Q000378:metabolism; D006639:Histidine / Q000235:genetics; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007700:Kinetics; D009154:Mutation / Q000235:genetics; D010084:Oxidation-Reduction / Q000187:drug effects; D011190:Potassium Cyanide / Q000494:pharmacology; D011199:Potentiometry; D011675:Pulse Radiolysis
D003573:Cytochrome b Group; D029968:Escherichia coli Proteins; D006418:Heme; D006639:Histidine; D003300:Copper; C017168:cytochrome bo, E coli; D011190:Potassium Cyanide; D000409:Alanine