S-ADENOSYL METHIONINE ALTERS THE DNA CONTACTS OF THE ECOKI METHYLTRANSFERASE

The EcoKI methyltransferase methylates two adenines on opposite strand s of its bipartite DNA recognition sequence AAC(N-6)GTGC. The enzyme h as a strong preference for hemimethylated DNA substrates, but the meth ylation state of the DNA does not influence its binding affinity. Meth ylation interference was used to compare the contacts made by the EcoK I methyltransferase with unmodified, hemimethylated or fully modified DNAs. Contacts were seen at or near the N7 position of guanine, in the major groove, for all of the guanines in the EcoKI recognition sequen ce, and at two guanines on the edge of the intervening spacer sequence . The presence of the cofactor and methyl donor S-adenosyl methionine had a striking effect on the interference pattern for unmodified DNA w hich could not be mimicked by the presence of the cofactor analogue S- adenosyl homocysteine. In contrast, S-adenosyl methionine had no effec t on the interference patterns for either kind of hemimethylated DNA, or for fully modified DNA. Differences between the interference patter ns for the unmodified DNA and any of the three forms of methylated DNA provide evidence that methylation of the target sequence influences t he conformation of the protein-DNA interface, and illustrate the impor tance of S-adenosyl methionine in the distinction between unmodified a nd methylated DNA by the methyltransferase.