DNA damage induced by ascorbate in the presence of Cu2+.
DNA damage induced by ascorbate in the presence of Cu2+ was investigated by use of bacteriophage phi X174 double-stranded supercoiled DNA and linear restriction fragments as substrates. Single-strand cleavage was induced when supercoiled DNA was incubated with 5 microM-10 mM ascorbate and 50 microM Cu2+ at 37 degrees C for 10 min. The induced DNA damage was analyzed by sequencing of fragments singly labeled at their 5'- or 3'-end. DNA was cleaved directly and almost uniformly at every nucleotide by ascorbate and Cu2+. Piperidine treatment after the reaction showed that ascorbate and Cu2+ induced another kind of DNA damage different from the direct cleavage. The damage proceeded to DNA cleavage by piperidine treatment and was sequence-specific rather than random. These results indicate that ascorbate induces two classes of DNA damage in the presence of Cu2+, one being direct strand cleavage, probably via damage to the DNA backbone, and the other being a base modification labile to alkali treatment. These two classes of DNA damage were inhibited by potassium iodide, catalase and metal chelaters, suggesting the involvement of radicals generated from ascorbate hydroperoxide.
D001205:Ascorbic Acid / Q000494:pharmacology*; D010584:Bacteriophage phi X 174 / Q000187:drug effects / Q000235:genetics; D001483:Base Sequence; D002118:Calcium / Q000494:pharmacology*; D002413:Cations, Divalent; D004249:DNA Damage; D004279:DNA, Viral / Q000187:drug effects* / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data
D002413:Cations, Divalent; D004279:DNA, Viral; D001205:Ascorbic Acid; D002118:Calcium