枯草芽孢杆菌HS18碱性蛋白酶分离纯化及其性质研究

Separation and purification of alkaline protease and its properties from Bacillus subtilis HS18

摘要: 从合浦珠母贝肠道中分离鉴定到酶菌株枯草芽孢杆菌（Bacillus subtilis）HS18,将枯草芽孢杆菌HS18的发酵液,经过硫酸铵盐析、DEAE-Sephadex A-50柱层析、Sephadex G-100柱层析3步纯化后得到了单一的酶蛋白,回收率为11.4%;经SDS-聚丙烯酰胺凝胶电泳测得酶蛋白分子量约为31ku;该蛋白酶最适反应温度为50℃,最适pH10.0;K+及Na+对酶有明显激活作用;丝氨酸蛋白酶特异性抑制剂（PMSF）能强烈抑制酶活性。表明所纯化到的蛋白酶为丝氨酸碱性蛋白酶。

Abstract: The protease produced by the fermentation of strain HS18, which isolated from the intestinal of Pinctada martensi and identified as Bacillus subtilis.A protease with high purity from the fermentation of Bacillus subtilis HS18 was purified by precipitation with ammonium sulfate, DEAE-Sephadex A-50column chromatography and Sephadex G-100 gel filtration chromatography and its recovery rate was 11.4%.The protease was determined by SDS-PAGE to have a molecular weight of 31ku.The optimal pH and temperature of the protease were 10.0 and 50℃, respectively.In addition, this protease could be activated by divalent cations such as K+ and Na+ and inhibited by serine-protease inhibitors (PMSF) .All results showed that this alkaline protease was a serine-protease.