A covalent complex between horse heart cytochrome c and yeast cytochrome c peroxidase: kinetic properties.
The kinetic properties of a 1:1 covalent complex between horse-heart cytochrome c and yeast cytochrome c peroxidase (ferrocytochrome-c:hydrogen-peroxide oxidoreductase, EC 1.11.1.5) have been investigated by transient-state and steady-state kinetic techniques. Evidence for heterogeneity in the complex is presented. About 50% of the complex reacts with hydrogen peroxide with a rate 20-40% faster than that of native enzyme; 20% of the complex exists in a conformation which does not react with hydrogen peroxide but converts to the reactive form at a rate of 20 +/- 5 s-1; 30% of the complex does not react with hydrogen peroxide to form the oxidized enzyme intermediate, cytochrome c peroxidase Compound I. Intramolecular electron transfer between covalently bound ferrocytochrome c and an oxidized site in cytochrome c peroxidase Compound I is too fast to measure, but a lower limit of 600 s-1 can be estimated at 5 degrees C in a 10 mM potassium phosphate buffer at pH 7.5. Free ferrocytochrome c reduces cytochrome c peroxidase Compound I covalently bound to ferricytochrome c at a rate 10(-4) to 10(-5)-times slower than for free Compound I. The transient-state ferrocytochrome c reduction rates of Compound I covalently linked to ferricytochrome c are about 70-times too slow to account for the steady-state catalytic properties of the 1:1 covalent complex. This indicates that hydrogen peroxide can interact with the 1:1 complex at sites other than the heme of cytochrome c peroxidase, generating additional species capable of oxidizing free ferrocytochrome c.
D001205:Ascorbic Acid; D002384:Catalysis; D003574:Cytochrome c Group / Q000378:metabolism*; D003578:Cytochrome-c Peroxidase / Q000378:metabolism*; D004579:Electron Transport; D006861:Hydrogen Peroxide / Q000378:metabolism; D007700:Kinetics; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000378:metabolism*; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D003574:Cytochrome c Group; D006861:Hydrogen Peroxide; D010544:Peroxidases; D003578:Cytochrome-c Peroxidase; D001205:Ascorbic Acid