Title: Enhanced in vivo repair of O(4)-methylthymine by a mutant human DNA alkyltransferase.
Authors: Encell, L P; Loeb, L A
Published In Carcinogenesis, (2000 Jul)
Abstract: The repair of O(6)-methylguanine (m(6)G) by human O(6)-alkylguanine-DNA alkyltransferase (hAGT) is approximately 5000-fold greater than that for O(4)-methylthymine (m(4)T). To evaluate each adduct's contribution to mutagenesis, we previously created a mutant hAGT with increased specificity for m(4)T in vitro. The mutant and wild-type (WT) hAGT have now been expressed in bacterial strains that allow for the specific detection of A:T-->G:C and G:C-->A:T mutations induced by m(4)T and m(6)G, respectively. After exposure to the mutagenic methylating agent, N-methyl-N'-nitro-N-nitrosoguanidine, A:T-->G:C substitutions were reduced >4-fold in cells expressing the mutant hAGT compared with 1. 1-fold for WT hAGT. G:C-->A:T substitutions were decreased >2.5-fold in cells expressing the mutant hAGT, whereas WT hAGT totally prevented G:C-->A:T mutations. These results demonstrate that the altered substrate specificity of hAGT observed in vitro also occurs in vivo, and that it is responsible for the observed differences in mutations.
PubMed ID: 10874019
MeSH Terms: DNA Damage; DNA Repair/physiology*; Enzyme Inhibitors/pharmacology; Escherichia coli/enzymology; Escherichia coli/genetics; Guanine/analogs & derivatives*; Guanine/metabolism; Guanine/pharmacology; Humans; Lac Operon; Methylnitronitrosoguanidine/toxicity; O(6)-Methylguanine-DNA Methyltransferase/antagonists & inhibitors; O(6)-Methylguanine-DNA Methyltransferase/genetics*; O(6)-Methylguanine-DNA Methyltransferase/metabolism*; Phenotype; Point Mutation; Substrate Specificity; Thymine/analogs & derivatives; Thymine/metabolism