Some spectral and steady-state kinetic properties of Pseudomonas cytochrome oxidase.
Some spectra of Pseudomonas cytochrome oxidase are reported, both for comparison with those of other workers and to illustrate the differences between the ascorbate- and dithionite-reduced forms of the enzyme. A spectrum of the reduced enzyme-CO complex, prepared in the absence of added reductants by incubation under CO, is also included. Ultracentrifugation studies yielded a value for the sedimentation coefficient (s20,w) of 7.5S, and an isoelectric point of pH6.9 was determined by isoelectric focusing. Steady-state kinetic constants of the electron donors, quinol, sodium ascorbate, reduced Pseudomonas azurin and Pseudomonas ferrocytochrome c551 were investigated giving Km values of 30mM, 4mM, 49muM and 5.6muM respectively. The two protein substrates were observed to be subject to product inhibition and the Ki for oxidized Pseudomonas azurin was evaluated at 4.9muM. Steady-state kinetics were also used to investigate the effects of the oxidation products of dithionite on the oxidase and nitrite reductase activities of Pseudomonas cytochrome oxidase. These experiments showed that whereas the oxidase activity was inhibited, the nitrite reductase activity was slightly enhanced.
D001205:Ascorbic Acid; D001400:Azurin; D002248:Carbon Monoxide; D003574:Cytochrome c Group; D004227:Dithionite; D003576:Electron Transport Complex IV / Q000032:analysis*; D006873:Hydroquinones; D007700:Kinetics; D009572:Nitrite Reductases; D010084:Oxidation-Reduction; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D013057:Spectrum Analysis
D003574:Cytochrome c Group; D006873:Hydroquinones; D001400:Azurin; D004227:Dithionite; D002248:Carbon Monoxide; D009572:Nitrite Reductases; D003576:Electron Transport Complex IV; D001205:Ascorbic Acid