1,N2-ethenoguanine, a mutagenic DNA adduct, is a primary substrate of Escherichia coli mismatch-specific uracil-DNA glycosylase and human alkylpurine-DNA-N-glycosylase

The promutagenic and genotoxic exocyclic DNA adduct 1 N-2-ethenoguanine (1,N-2-epsilonG) is a major product formed in DNA exposed to lipid peroxidation-derived aldehydes in vitro. Here, we report that two structurally unrelated proteins, the Escherichia coli mismatch-specific uracil-DNA glycosylase (MUG) and the human alkylpurine-DNA-N-glycosylase (ANPG), can release 1,N-2-epsilonG from defined oligonucleotides containing a single modified base. A comparison of the kinetic constants of the reaction indicates that the MUG protein removes the 1,N-2-epsilonG lesion more efficiently (k(cat)/K-m = 0.95 X 10(-3) min(-1) nM(-1)) than the ANPG protein (k(cat)/K-m = 0.1 X 10(-3) min(-1) nM(-1)). Additionally, while the nonconserved, N-terminal 73 amino acids of the ANPG protein are not required for activity on 1,N-6-ethenoadenine, hypoxanthine, or N-methylpurines, we show that they are essential for 1,N-2-epsilonG-DNA glycosylase activity. Both the MUG and ANPG proteins preferentially excise 1,N-2-epsilonG when it is opposite dC; however, unlike MUG, ANPG is unable to excise 1,N-2-EG when it is opposite dG. Using cell-free extracts from genetically modified E. coli and murine embryonic fibroblasts lacking MUG and mANPG activity, respectively, we show that the incision of the 1,N-2-epsilonG-containing duplex oligonucleotide has an absolute requirement for MUG or ANPG. Taken together these observations suggest a possible role for these proteins in counteracting the genotoxic effects of 1,N-2-epsilonG residues in vivo.