REPAIR OF O6-METHYLGUANINE AND O4-METHYLTHYMINE BY THE HUMAN AND RAT O6-METHYLGUANINE-DNA METHYLTRANSFERASES

In order to compare the ability of the human and rat O6-methylguanine- DNA methyltransferases (transferases) to repair in vitro O6-methylguan ine (O6-MeGua) and O4-methylthymine (O4-MeThy) residues, which are two mutagenic DNA adducts formed by alkylating agents, we have purified b oth proteins to homogeneity. Gel electrophoresis of the proteins shows that the O4-MeThy repair is due to the transfer of the methyl group f rom the alkylated base to the transferase molecules. However, both pro teins repair with different efficiencies the O6-MeGua and O4-MeThy res idues present in alkylated DNA, poly[d(G.C)], poly(dG.dC), or in alkyl ated poly[d(A.T)] and poly(dA.dT), respectively. Reaction of both prot eins with either methylated residues follows a second-order kinetics. The rate constants are 1 x 10(9) M-1 min-1 for both proteins acting on O6-MeGua and 4.8 x 10(6) or 1.8 x 10(5) M-1 min-1 for the rat or huma n protein acting on 04-MeThy, respectively. The activity of the mammal ian transferases on O4-MeThy present in a poly(dA.dT) substrate is inh ibited by double-stranded DNA.