Comparative Analysis of Biochemical Properties of Site Specific Methyl-directed DNA Endonucleases BisI, BlsI and EcoBLI Recognizing 5’-G(5mC)NGC-3’/3’-CGN(5mC)G-5’

VA Chernukhin, DA Gonchar, MA Abdurashitov, EV Kileva, OA Belichenko, VS Dedkov, NA Mikhnenkova, SKh Degtyarev
RJPBCS, vol 7(2), 2016, p.p. 388-394

A comparative analysis of biochemical properties of three similar site specific methyl-directed DNA endonucleases BisI, BlsI and EcoBLI recognizing 5’-G(5mC)NGC-3’/3’-CGN(5mC)G-5’ has been done. Optimal concentrations of MgCl2, NaCl or KCl, pH of reaction buffers as well as the optimal reaction temperatures for these enzymes were determined. It was shown that BisI has the maximal activity in the buffer 10 mM Tris-HCl (pH 9.0 at 250 C), 15 mM MgCl2, 150 mM KCl, 1 mM DTT. An optimal buffer for MD-endonuclease BlsI is 10 mM Tris-HCl (pH 8.0 at 250 C), 15 mM MgCl2, 100 mM NaCl, 1 mM DTT. The enzyme EcoBLI displays a maximal activity in the buffer 10 mM Tris-HCl (pH 8.5 at 250 C), 10 mM MgCl2, 25 mM NaCl, 1 mM DTT. Optimal temperature for all enzymes is 370 C

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