Enzyme and Microbial Biochemistry Laboratory, Department of Chemistry, Indian Institute of Technology, Delhi, New Delhi, India.
Bioresour Technol. 2013 Oct;145:357-61. doi: 10.1016/j.biortech.2012.11.024. Epub 2012 Nov 17.
A moderately halophilic protease producer, Bacillus sp. strain isolated from sea water is described. The protease is purified to homogeneity by ammonium sulphate precipitation and CM cellulose chromatography. The serine protease has a molecular mass of 29 kDa. Enzymatic characterization of protease revealed K(m) 2.22 mg mL(-1), Vmax 1111.11 U mL(-1), pH optimum 9.0, t1/2 190 min at 60°C and salt optima 1% (w/v) NaCl. The protease is remarkably stable in hydrophilic and hydrophobic solvents at high concentrations. The purified preparation is unstable at room temperature. Ca(2+) ions are required for preventing this loss of activity. Interestingly, the activity and stability are modulated differentially. Whereas, divalent cation Ca(2+) are involved in maintaining stability in solution at room temperature by preventing unfolding, monovalent Na(+) and K(+) ions participate in regulating the activity and assist in refolding of the enzyme. Application of the protease is shown in efficient removal of blood stain.
从海水中分离到一株中度嗜盐蛋白酶产生菌，命名为芽孢杆菌。通过硫酸铵沉淀和 CM 纤维素层析法将蛋白酶纯化为均一状态。该丝氨酸蛋白酶的分子量为 29 kDa。蛋白酶的酶学特性研究表明，其 K(m) 为 2.22 mg/mL，Vmax 为 1111.11 U/mL，最适 pH 值为 9.0，在 60°C 下半衰期为 190 分钟，最适盐度为 1%（w/v）NaCl。该蛋白酶在高浓度的亲水和疏水溶剂中具有显著的稳定性。在室温下，纯化的酶制剂不稳定。钙离子的存在对于防止这种活性丧失是必需的。有趣的是，活性和稳定性的调节方式不同。二价钙离子通过防止酶的展开来维持溶液在室温下的稳定性，而单价的钠离子和钾离子则参与调节酶的活性并协助酶的重折叠。该蛋白酶在有效去除血渍方面得到了应用。
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