PURIFICATION AND CHARACTERIZATION OF THE METHYLTRANSFERASE FROM THE TYPE-1 RESTRICTION AND MODIFICATION SYSTEM OF ESCHERICHIA-COLI K12

The DNA methyltransferase component of the type I restriction and modi fication enzyme of Escherichia coli K12 has been purified. The active component, a trimer of molecular mass 170 kDa consisting of one DNA re cognition subunit (S) and two modification subunits (M), showed the ex pected preference for modifying a hemimethylated substrate rather than an unmethylated one. Small amounts of the dimers M2 and M1S1 were als o isolated. Subunit rearrangements of the three protein species occurr ed on ion exchange and heparin-agarose chromatography. Denaturation of the trimer gave folding intermediates, and these and the dimer forms isolated during purification may reflect the assembly of the protein i n vivo. Enzyme activity was recovered on refolding the denatured prote in by dilution of the denaturant. A comparison of the predicted isoele ctric points of all known S subunits of type I restriction and modific ation enzymes revealed values that correlated with the arrangement of type I systems in several families. Electrostatic interactions may exp lain the different subunit stoichiometries observed during purificatio n of type I enzymes and the differing preferences for hemimethylated D NA displayed by the three type I families.