DNA strand breaks by metal-induced oxygen radicals in purified Salmonella typhimurium DNA.

Purified Salmonella typhimurium DNA was incubated for 1h at 37 degrees C with various concentrations (10-100 muM) of transition metal ions (Fe(2+), Fe(3+), Cu(2+), Ni(2+), Cd(2+)), with various concentrations (0.1-100 mM) of H(2)O(2), and with various concentrations of each transition metal ion in the presence of various concentrations of H(2)O(2). Damage to DNA was assessed by electrophoresis of the reaction mixtures in 1% agarose gel. Breakage of the DNA strands would produce a series of DNA fragments resulting in a smear in the gel, while intact DNA produced a single band. Results showed that no damage to the DNA was detectable after incubation with either H(2)O(2) alone or either of the metal ions alone. However, all of the metal ions investigated triggered DNA breakage in the presence of H(2)O(2). The extent of breakage depended on the metal ion and on its concentration, as well as on the H(2)O(2) concentration. Addition of either EDTA or catalase to the reaction mixture completely inhibited the DNA degradation, confirming the involvement of both the metal ion and the H(2)O(2) in the breakage of DNA strands. Production of the hydroxyl radical when H(2)O(2) and a metal ion were both present in the reaction mixture was evidenced by the thiobarbituric acid method. The most extensive damage was caused by Cu(2+) followed, in decreasing order, by Fe(2+), Fe(3+), Ni(2+), and Cd(2+).