Oxidative damage of DNA induced by the cytochrome C and hydrogen peroxide system.

To elaborate the peroxidase activity of cytochrome c in the generation of free radicals from H2O2, the mechanism of DNA cleavage mediated by the cytochrome c/H2O2 system was investigated. When plasmid DNA was incubated with cytochrome c and H2O2, the cleavage of DNA was proportional to the cytochrome c and H2O2 concentrations. Radical scavengers, such as azide, mannitol, and ethanol, significantly inhibited the cytochrome c/H2O2 system-mediated DNA cleavage. These results indicated that free radicals might participate in the DNA cleavage by the cytochrome c and H2O2 system. Incubation of cytochrome c with H2O2 resulted in a time-dependent release of iron ions from the cytochrome c molecule. During the incubation of deoxyribose with cytochrome c and H2O2, the damage to deoxyribose increased in a time-dependent manner, suggesting that the released iron ions may participate in a Fenton-like reaction to produce dOH radicals that may cause the DNA cleavage. Evidence that the iron-specific chelator, desferoxamine (DFX), prevented the DNA cleavage induced by the cytochrome c/H2O2 system supports this mechanism. Thus we suggest that DNA cleavage is mediated via the generation of dOH by a combination of the peroxidase reaction of cytochrome c and the Fenton-like reaction of free iron ions released from oxidatively damaged cytochrome c in the cytochrome c/H2O2 system.