title: Site-directed mutagenesis of glutamic acid 172 to glutamine completely inactivated human O6-alkylguanine-DNA-alkyltransferase.
abstract: DNA repair by O6-alkylguanine-DNA-alkyltransferase involves the stoichiometric transfer of the O6-alkyl group from the guanine lesion to the active-site cysteine residues of the protein. Site-directed mutagenesis of glutamic acid 172 of human O6-alkylguanine-DNA-alkyltransferase (EC 2.1.1.63) to glutamine totally abolished the alkyltransferase activity of the protein. This suggests that glutamic acid 172 is crucial to the alkyl transfer. It may act as a general acid (as CO2H) or base (as CO2-), or have a role as a component of a salt-link (-CO2-.....+N-), vital for the structural integrity of the active site. This is the first mutational inactivation of a protein in this family of DNA repair molecules by means of a residue change outside the highly conserved pentet (PCHRV) which includes the active-site cysteine.
authors.affiliation: CRC Department of Carcinogenesis, Paterson Institute for Cancer Research, Christie Hospital NHS Trust, Manchester, U.K.
authors.forename: J A
authors.initials: JA
authors.lastname: Rafferty
authors.forename: J
authors.initials: J
authors.lastname: Tumelty
authors.forename: M
authors.initials: M
authors.lastname: Skorvaga
authors.forename: R H
authors.initials: RH
authors.lastname: Elder
authors.forename: G P
authors.initials: GP
authors.lastname: Margison
authors.forename: K T
authors.initials: KT
authors.lastname: Douglas
mesh_terms: D000595:Amino Acid Sequence; D004260:DNA Repair; D005971:Glutamates / Q000737:chemistry; D005973:Glutamine / Q000737:chemistry; D006801:Humans; D008780:Methyltransferases / Q000737:chemistry*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019853:O(6)-Methylguanine-DNA Methyltransferase; D013329:Structure-Activity Relationship
chemical_list: D005971:Glutamates; D005973:Glutamine; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase