DNA recognition by the EcoP15I and EcoPI modification methyltransferases

Abstract

The DNA-binding properties of the EcoP15I DNA methyltransferase (M·EcoP15I; MTase) were studied using electro-phoretic mobility shift assays. We show by molecular size-exclusion chromatography and dimethyl suberimidate cross-linking that M·EcoP15I is a dimer in solution. While M·EcoP15I binds approx, threefold more tightly to its recognition sequence, 5'-CAGCAG-3', than to non-specific sequences in the presence of AdoMet or its analogs, the discrimination between specific and non-specific sequences significantly increases in presence of ATP. These results suggest for the first time a role for ATP in DNA recognition by type-III restriction-modification enzymes. Furthermore, we show that although c2, EcoPI mutant MTases are defective in AdoMet binding, they are still able to bind DNA in a sequence-specific manner.