Suicide inactivation of the E. coli O6-methylguanine-DNA methyltransferase.

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Abstract
The O6‐methylguanine‐DNA methyltransferase of Escherichia coli acts rapidly and stoichiometrically to convert a mutagenic O6‐methylguanine residue in DNA to unsubstituted guanine. Even at low protein concentrations and in the absence of any cofactors, the transfer of a methyl group to one of the protein's own cysteine residues occurs in less than 2 s at 37 degrees C. The entire kinetic process can be followed experimentally at 5 degrees C. Formation of S‐methylcysteine in the protein is accompanied by loss of activity and accounts for the exceptional suicide kinetics of this enzyme as well as for the sharp saturation of O6‐methylguanine repair observed in vivo. The enzyme can remove greater than 98% of the methyl groups from O6‐methylguanine present in alkylated DNA, but leaves N‐alkylated purines untouched. Single‐stranded DNA containing O6‐methylguanine is a poor substrate, with the methyl transfer occurring at approximately 0.1% of the rate for duplex DNA. This latter observation may explain the high frequency of mutations induced by alkylating agents at DNA replication forks.