Clarke C M, Hartley B S
Biochem J. 1979 Jan 1;177(1):49-62. doi: 10.1042/bj1770049.
The restriction endonuclease BstI was purified from 70kg of Bacillus stearothermophilus. The final product is at least 97% pure as judged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis; this major protein species co-migrates with the enzyme activity on native polyacrylamide-gel electrophoresis and isoelectric focusing. Pure restriction endonuclease BstI has a subunit mol.wt. of 26,000 and is probably a loosely associated dimer. The enzyme shows maximum activity at pH values between 7 and 9.5, and in the presence of 0.5-2mM-Mg2+. NaCl inhibits the restriction enzyme activity. Restriction endonuclease BstI cleaves DNA in a position identical with that cleaved by endonuclease BamHI (for Bacillus amyloliquefaciens), i.e.: (formula: see text). In the presence of high concentrations of enzyme, DNA cleavage occurs at secondary sites. This side-specificity is enhanced by the addition of glycerol. Preliminary studies indicate that these sites are of the type: (formula: see text).
限制性内切酶BstI是从70千克嗜热脂肪芽孢杆菌中纯化得到的。通过十二烷基硫酸钠/聚丙烯酰胺凝胶电泳判断，最终产物的纯度至少为97%；在天然聚丙烯酰胺凝胶电泳和等电聚焦中，这种主要蛋白质条带与酶活性迁移位置相同。纯的限制性内切酶BstI的亚基分子量为26,000，可能是一种松散结合的二聚体。该酶在pH值7至9.5之间以及存在0.5 - 2mM Mg2+的情况下表现出最大活性。NaCl会抑制限制性酶的活性。限制性内切酶BstI切割DNA的位置与淀粉酶芽孢杆菌的内切酶BamHI切割DNA的位置相同，即：（公式：见原文）。在高浓度酶存在的情况下，DNA在二级位点发生切割。添加甘油会增强这种位点特异性。初步研究表明这些位点的类型为：（公式：见原文）。
