Site-directed mutagenesis of glutamic acid 172 to glutamine completely inactivated human O6-alkylguanine-DNA-alkyltransferase.
Rafferty, Joseph A Tumelty, J Skorvaga, Milan Elder, Rhoderick H Margison, Geoffrey P Douglas, K T
Rafferty, Joseph A
Tumelty, J
Skorvaga, Milan
Elder, Rhoderick H
Margison, Geoffrey P
Douglas, K T
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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.
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1994-02-28
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Site-directed mutagenesis of glutamic acid 172 to glutamine completely inactivated human O6-alkylguanine-DNA-alkyltransferase. 1994, 199 (1):285-91 Biochem. Biophys. Res. Commun.