Extra Copper(П) Enhances The DNA Cleavage Activity Supported With Wild-Type CuZn Superoxide Dismutase Via Multiple Pathways

It is well known that the primary function of wild type Cu,Zn superoxide dismutase (CuZnSOD) is to catalyze the conversion of the superoxide anion to hydrogen peroxide and dioxygen as an antioxidant enzyme. However, The aberrant copper-mediated oxidation in the enzyme (including its mutation forms) that damages nucleic acids, proteins including itself and cell membrane has attracted an extensive attention in the past decade.The present study examined the hydrogen peroxide-dependent DNA cleavage activity supported with the combinations between CuZnSOD and extra copper (CuZnSOD + nCu(II)) in details by a series of biochemical experiments.The UV-visible absorption titration of CuZnSOD with copper showed that some interactions may occur between the extra Cu(II) and CuZnSOD. The DNA cleavage experiments indicated that the presence of extra copper can significantly enhance the DNA cleavage activity supported with CuZnSOD. The relative activity and rates assay also showed that the DNA cleavage activity of CuZnSOD + nCu(II) was enhanced upon addition of extra Cu(II), and a cooperative effect between CuZnSOD and the extra Cu(II) occurs in DNA cleavage.The effect of free radical scavengers and inhibitors on the DNA cleavage activity was investigated, and the results provide an indirect support to the hypothesis that the reactive species causing DNA cleavage are more likely copper-bound hydroxyl radical, superoxide or peroxide species, as well as hydroxide anion. The favorable regions for DNA cleavage were observed to be pH 3.6-5.6 and pH 9.0-10, suggesting that the species responsible for DNA cleavage are different in different pH regionsOur results do not support a widely accepted conclusion that the copper-bound or free hydroxyl radicals provided by CuZnSOD with its end product hydrogen peroxide lead to release of copper from its host, the Fenton reaction between the free copper and the hydrogen peroxide being responsible for the aberrant copper-mediated oxidation chemistry in CuZnSOD. The results suggest that the cooperative effect between CuZnSOD and extra Cu(II) made the enhancement effect on DNA cleavage activity.