Isolation and some properties of the restriction endonuclease BcnI from Bacillus centrosporus

A specific type-II restriction endonuclease BcnI from Bacillus centrosporus has been purified to electrophoretic homogeneity in three chromatographic steps. Around 15 micrograms of such a preparation can be isolated from 1 g of the cell paste. The yield of the enzyme is higher than that of any type-II restriction endonuclease so far reported. The molecular weight of the enzyme determined by gel filtration and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate equals 27 500 and 28 000 respectively. The activity of the restriction endonuclease is maximal at pH 9.2 and 40--45 degrees C. The optimal magnesium concentration was estimated to be 7.5 mM. The activity of BcnI may also be observed in the presence of Co2+, Mn2+, Ni2+ and Zn2+ but it is markedly less than in the presence of Mg2+.