KINETIC-STUDIES OF MVAI DNA METHYLTRANSFERASE INTERACTION WITH MODIFIED OLIGONUCLEOTIDE DUPLEXES

We have measured steady-state kinetics of a N4-cytosine methylase, M . MvaI, using as substrates modified non-selfcomplementary tettadecanuc leotide duplexes containing the CCWGG target sequence. The inner or ou ter localisation of the dI residue in the MvaI recognition site seems to be of little importance since the specificity constants k(cat)/K-M are only 2 to 7 fold smaller than that of the canonical substrate. Rep lacement of dG residues by dI in both strands resulted in a 25 to 60-f old decrease of the specificity constant. Modifications of the phospha te backbone or opening of the sugar ring of one of the dG residues had only little influence on the action of M . MvaI. The enzyme appears t o be rather tolerant to different kinds of modifications in its substr ate in the mainor groove.