Cost effective characterization process and molecular dynamic simulation of detergent compatible alkaline protease from Bacillus pumilus strain MP27
Cost effective characterization process and molecular dynamic simulation of detergent compatible alkaline protease from Bacillus pumilus strain MP27
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DOI：
10.1016/j.procbio.2017.04.024
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发表时间：
2017-07-01
影响因子：
4.4
通讯作者：
Shukla, Pratyoosh
中科院分区：
文献类型：
--
作者：
Baweja, Mehak;Singh, Puneet Kumar;Shukla, Pratyoosh
从南大洋水样中分离到一种从短小芽孢杆菌MP27菌株中分离到的耐冷碱性蛋白酶，其分子量约为53 kDa。经一步TPP部分纯化，纯度倍数为16.65。该酶在一定的温度和pH范围内具有广泛的稳定性，在50℃和pH 12时分别保持了52.25%和92%的活性。该酶与所测试的洗涤剂一起显示出良好的活性，在10 mg/ml的十二烷基硫酸钠(10 mg/ml)和-99%的潮汐洗涤剂(7 mg/ml)下表现出98%的稳定性。在此基础上，成功地将1152bp的碱性蛋白酶基因克隆到pGEM-T Easy载体中，并在大肠杆菌DH5α中表达。对基因序列进行进一步翻译、建模，并进行分子动力学模拟。模拟的蛋白质在最初的5 ns内高度不稳定，因此在1 ns的模拟后不能与配体形成键。
A psychrothermotolerant alkaline protease isolated from Bacillus pumilus MP27 with a molecular mass similar to 53 kDa was isolated from Southern ocean water samples. It was partially purified by single step TPP with purity fold of 16.65. The enzyme was found to be widely stable within a range of temperature and pH, maintaining 52.25% of its activity at 50 degrees C and 92% at pH 12. The enzyme exhibited an exceptional activity along with tested detergents, showing 98% stability with SDS (10 mg/ml) and- 99% stability with Tide detergent (7 mg/ml). Further, the alkaline protease gene of 1152 bp was successfully cloned in pGEM-T Easy vector in E. coli DH5 alpha. The gene sequence was further translated, modeled and molecular dynamic simulation was performed. The modeled protein was highly unstable during the first 5 ns and therefore could not able to form bonds with the ligand after 1 ns of simulation.