Photochemical and ligand-exchange properties of the cyanide complex of fully reduced cytochrome c oxidase.
Cytochrome oxidase, in its fully reduced state, forms a complex with CN having a Kd of 230 microM with a stoicheiometry of 1 CN molecule per cytochrome oxidase. We do not detect a second CN-binding site as seen by i.r. spectroscopy [Yoshikawa & Caughey (1990) J. Biol. Chem. 265, 7945-7958]. The ferrocytochrome a3-CN complex, like the analogous ferrocytochrome a3-CO complex, is photosensitive but with a 15-fold lower quantum yield for photolysis. Analysis of the recombination kinetics after CN photolysis establishes a simple bimolecular binding constant of 235 M-1.s-1, in agreement with the value obtained from stopped-flow studies [Antonini, Brunori, Greenwood, Malmström & Rotillo (1971) Eur. J. Biochem. 23, 396-400]. A rate of 0.07 s-1 for the first-order dissociation of CN from cytochrome a3 is found by the rate of exchange of CO with ferrocytochrome a3-CN, and is consistent with the value calculated from the equilibrium binding constant and the CN on rate. However, O2 is able to oxidize the fully reduced CN compound at a rate well in excess of the CN off rate. The product of this oxidation reaction is a partially reduced CN complex. This implies that O2 either promotes CN dissociation or is able to oxidize the CN-bound enzyme directly. These results are discussed in the context of the structure and dynamics of the ligand-binding site of cytochrome oxidase.
Hill
B C
BC
Department of Chemistry and Biochemistry, Concordia University, Montreal, Canada.
Marmor
S
S
D000818:Animals; D001665:Binding Sites; D002248:Carbon Monoxide / Q000378:metabolism; D002417:Cattle; D003576:Electron Transport Complex IV / Q000378:metabolism*; D007700:Kinetics; D008929:Mitochondria, Heart / Q000201:enzymology; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D010782:Photolysis; D011190:Potassium Cyanide / Q000378:metabolism*; D013053:Spectrophotometry
D002248:Carbon Monoxide; D003576:Electron Transport Complex IV; D011190:Potassium Cyanide; D010100:Oxygen