[Purification of recombinant DNA methyltransferase M2.BstSE from nickase-modification system NM.BstSEI and study of the enzyme properties].

ABSTRACT

The operon of nickase-modification system from Bacillus stearothermophilus SE-589 (recognition site 5'-GAGTC-3') includes two DNA methyltransferase genes bstSEIM1 and bstSEIM2. Gene encoding DNA methyltransferase M2.BstSEI was cloned in pJW vector and expressed in E. coli cells. The enzyme M2.BstSEI has been isolated by chromatographic purification. M2.BstSEI displays maximum activity at 55 degrees C and pH 7.5. The enzyme modifies adenine in DNA sequence 5'-GAGTC-3' and has substrate specificity 5'-GASTC-3'. The kinetic parameters of methylation reaction have been determined. The catalytic constant--2.2 min(-1), the Michaelis constant on T7 DNA--9.8 nM and on SAM--5.8 microM.