Responses to the Major Acrolein-derived Deoxyguanosine Adduct inEscherichia coli

Acrolein, a reactive α,β-unsaturated aldehyde found ubiquitously in the environment and formed endogenously in mammalian cells, reacts with DNA to form an exocyclic DNA adduct, 3H-8-hydroxy-3-(β-d-2′-deoxyribofuranosyl)-5,6,7,8-tetrahydropyrido[3,2-a]purine-9-one (γ-OH-PdG). The cellular processing and mutagenic potential of γ-OH-PdG have been examined, using a site-specific approach in which a single adduct is embedded in double-strand plasmid DNA. Analysis of progeny plasmid reveals that this adduct is excised by nucleotide excision repair. The apparent level of inhibition of DNA synthesis is ∼70% in Escherichia coli ΔrecA, uvrA. The block to DNA synthesis can be overcome partially byrecA-dependent recombination repair. Targeted G → T transversions were observed at a frequency of 7 × 10−4/translesion synthesis. Inactivation ofpolB, dinB, and umuD,C genes coding for “SOS” DNA polymerases did not affect significantly the efficiency or fidelity of translesion synthesis. In vitroprimer extension experiments revealed that the Klenow fragment of polymerase I catalyzes error-prone synthesis, preferentially incorporating dAMP and dGMP opposite γ-OH-PdG. We conclude from this study that DNA polymerase III catalyzes translesion synthesis across γ-OH-PdG in an error-free manner. Nucleotide excision repair, recombination repair, and highly accurate translesion synthesis combine to protect E. coli from the potential genotoxicity of this DNA adduct.