Spectroscopic and quantitative analysis of the oxygenated and peroxy states of the purified cytochrome d complex of Escherichia coli.
Oxygenated and peroxy states of the cytochrome d complex of Escherichia coli have been proposed as intermediates in the reaction mechanism of this ubiquinol oxidase. In this report, several stable states of the purified enzyme were examined spectroscopically at room temperature. As purified, the cytochrome d complex exists in an oxygenated state characterized by an absorbance band at 650 nm. Removal of oxygen results in loss of absorbance at this wavelength, which is restored upon the return of oxygen. The presence of one oxygen molecule in the oxygenated state was quantified by measuring oxygen released when excess hydrogen peroxide was added to the oxygenated state by passage of argon generates a "partially reduced" state with an absorbance peak at 628 nm, apparently due to reduced cytochrome d. Addition of equimolar hydrogen peroxide to the fully oxidized state produces the peroxy state. This peroxy state is also formed upon addition of excess hydrogen peroxide to the oxygenated state via a stable intermediate termed "peroxy intermediate." It is likely that 1) the oxygenated state consists of one molecule of oxygen bound to reduced heme d, and 2) there are at least two stable states that have bound peroxide at room temperature, the peroxy state and a newly discovered peroxy intermediate.
Lorence
R M
RM
Department of Chemistry, University of Illinois, Urbana 61801.
Gennis
R B
RB
D002374:Catalase / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology; D017937:Cytochrome d Group; D003580:Cytochromes / Q000378:metabolism* / Q000648:ultrastructure; D004926:Escherichia coli / Q000201:enzymology*; D005290:Ferric Compounds; D005292:Ferricyanides; D005296:Ferrous Compounds; D006861:Hydrogen Peroxide; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism; D013057:Spectrum Analysis
D003580:Cytochromes; D005290:Ferric Compounds; D005292:Ferricyanides; D005296:Ferrous Compounds; D017937:Cytochrome d Group; D006861:Hydrogen Peroxide; D010544:Peroxidases; D002374:Catalase; D010100:Oxygen