DNA single strand breakage by copper(II) complexes and hydrogen peroxide at physiological conditions

The conversion of the closed circular double stranded supercoiled DNA (pBR322) to the nicked circular form and linear form was used to investigate DNA nicking induced by the reactions of Cu(II) complexes with hydrogen peroxide (H2O2). As a result, the dose-response curve for the CuII(en) (en:ethylenediamine) mediated H2O2 dependent DNA nicking was observed. For a fixed concentration of CuII(en)2 (1 mM), the concentration of H2O2 producing a maximum extent of DNA nicking was 100 mM. On the other hand, for a fixed concentration of H2O2 (100 mM), the concentration of CuII(en)2 was 1 mM. That is, when the concentration ratio of H2O2 to CuII(en)2 was 100, a maximum extent of DNA nicking was observed. This result is a good accordance with that observed by thiobarbituric acid (TBA) and ESR-spin trapping methods. This result gives first experimental evidence that Cu(II) complex did cause the DNA damage in the presence of H2O2. On the other hand, CuII(edta) (edta: ethylenediaminetetraacetic acid) did not cause the DNA strand breaks with the reaction of H2O2, but it was activated by addition of the biological reductant such as ascorbic acid in the presence of H2O2 to cause the DNA strand breaks.