DNA synthesis blocking lesions induced by singlet oxygen are targeted to deoxyguanosines

In vitro DNA synthesis on single stranded templates damaged by singlet oxygen was investigated in the supF tRNA gene sequence, using several DNA polymerases. Singlet oxygen was generated by the thermal decomposition of the water soluble with the endoperoxide of disodium 3,3'-(1,4-naphthylidene) dipropionate (NDPO2). The data demonstrated that damage at deoxyguanosine residues interrupts DNA polymerization. Modified T7 phage and Thermus aquaticus DNA polymerases were found to synthesize DNA fragments which terminated opposite deoxyguanosine, while T4 phage DNA polymerase and avian myeloblast virus reverse transcriptase were blocked one nucleotide 3' to deoxyguanosine positions on the template. DNA polymerase I (Klenow fragment) from Escherichia coli was inhibited at both positions, before and at the putative damaged sites. The blocking lesions, induced by 5 mM NDPO2, were estimated to be approximately 1.5 per 260 nucleotides, corresponding to 2% of deoxyguanosines. The distribution of lesions in the supF gene did not reveal any specific sequence context which showed distinct susceptibility to the attack of singlet oxygen.