Hydroxylation of deoxyguanosine at 5′ site of GG and GGG sequences in double-stranded DNA induced by carbamoyl radicals

Free radicals generated by chemicals can cause sequence-specific DNA damage and play important roles in mutagenesis and carcinogenesis. Carbamoyl group (CONH2) and its derived groups (CONR2) occur as natural products and synthetic chemical compounds. We have investigated the DNA damage by carbamoyl radicals (CONH2), one of carbon-centered radicals. Electron spin resonance (ESR) spectroscopic study has demonstrated that carbamoyl radicals were generated from formamide by treatment with H2O2 Plus Cu(II), and from azodicarbonamide by treatment with Cu(II). We have investigated sequence specificity of DNA damage induced by carbamoyl radicals using 12 P-labeled DNA fragments obtained from the human c-Ha-ras-1 and p53 genes. Treatment of double-stranded DNA with carbamoyl radicals induced an alteration of guanine residues, and subsequent treatment with piperidine or Fpg protein led to chain cleavages at 5'-G of GG and GGG sequences. Carbamoyl radicals enhanced Cu(II)/H2O2-mediated formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in double-stranded DNA more efficiently than that in single-stranded DNA. These results shows that carbamoyl radicals specifically induce hydroxylation of deoxyguanosine at 5' site of GG and GGG sequences in double-stranded DNA.