[Isolation of SAU 96 I restrictase, its physical and catalytic properties]

Abstract

Restrictase Sau 96 I was isolated from Staphylococcus aureus PS 96 and purified by chromatography on DEAE-cellulose, phosphocellulose and hydroxylapatite. The preparation was studied by gel filtration on Toyopearl HW-55 and polyacrylamide gel electrophoresis under denaturing conditions. The active form of the enzyme is a dimer with a molecular weight of 54,000 +/- 5000 composed of two identical subunits. Catalytic properties of the restrictase were determine; the pH optimum is 8.5-9.0, the optimal concentration of NaCl and Mg2+ is 15-100 mM and 10 mM, respectively. Mn2+ ions at a concentration of 2 mM can replace Mg2+, while Zn2+, Ca2+, Cu2+ ions cannot replace Mg2+. The optimal temperature is 30-43 degrees. Ethanol and glycerol at concentrations more than 10% inhibit the enzyme without changing its specificity; p-chloromercuribenzoate inhibits the enzyme at a concentration of 0.05 mM.