title: Dominant sensitization variants of human O(6)-methylguanine-DNA-methyltransferase obtained by a mutational screen of surface residues.
abstract: A scanning mutagenesis experiment was performed on human O(6)-methylguanine methyltransferase (hMGMT), directed largely at non-conserved surface residues that have not previously been studied. Variants typically contained two or more substitutions. Two of the 16 variants characterized in detail are inactive for methyltransfer, but increase the cytotoxicity and mutagenic effects of methylating agents. This phenotype is reminiscent of a variant (C145A) that has a mutation in the methyl-accepting cysteine. C145A is inactive, but reportedly binds methylated DNA and confers sensitivity to methylating agents. The sensitization phenotype of the two new variants is more striking in strains that are wild-type for DNA repair than in strains that are deficient for repair, suggesting that these proteins inhibit functional DNA repair proteins by competitively binding to methylated DNA. Both variants have multiple substitutions in the last helix of the protein. These results suggest that the C-terminal helix is necessary for methyltransfer activity, but not for methylguanine-specific binding.
authors.affiliation: Department of Biophysics and Biophysical Chemistry, The Johns Hopkins University School of Medicine, WBSB 708, 725 N. Wolfe Street, Baltimore, MD 21205-2185, USA.
authors.forename: L R
authors.initials: LR
authors.lastname: Brown
authors.forename: J
authors.initials: J
authors.lastname: Deng
authors.forename: N D
authors.initials: ND
authors.lastname: Clarke
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics; D006801:Humans; D008769:Methylnitronitrosoguanidine / Q000633:toxicity; D008969:Molecular Sequence Data; D009152:Mutagenicity Tests; D009154:Mutation; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics*; D017434:Protein Structure, Tertiary
chemical_list: D008769:Methylnitronitrosoguanidine; D019853:O(6)-Methylguanine-DNA Methyltransferase