Enhancing the Catalytic Activity of Type II L-Asparaginase from Bacillus licheniformis through Semi-Rational Design.
Enhancing the Catalytic Activity of Type II L-Asparaginase from Bacillus licheniformis through Semi-Rational Design.
复制标题
通过半理性设计增强地衣芽孢杆菌II型L-天冬酰胺酶的催化活性
DOI：
10.3390/ijms23179663
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发表时间：
2022-08-26
影响因子：
5.6
通讯作者：
中科院分区：
文献类型：
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作者：
低催化活性是限制II型l -天冬酰胺酶（ASNase）在食品和制药工业中广泛应用的关键因素。本研究采用半理性设计构建智能文库，提高地衣芽孢杆菌ⅱ型ASNase的催化活性。通过饱和突变和组合突变筛选了催化效率显著提高的突变体。最终获得了一个五重突变体ILRAC，比活性为841.62 IU/mg， kcat/Km为537.15 min−1·mM−1，分别是野生型ASNase的4.24倍和6.32倍。地衣芽孢杆菌ⅱ型ASNase的比活性和kcat/Km均为首次报道。此外，突变体ILRAC的pH稳定性和热稳定性都得到了提高。同时，结构比对和分子动力学模拟表明，高结构稳定性和强底物结合有利于突变体ILRAC的热稳定性和酶活性的提高。这是首次采用半理性方法提高地衣芽孢杆菌II型ASNase的酶活性，为l -天冬酰胺酶的酶修饰和工业应用提供了新的见解。
Low catalytic activity is a key factor limiting the widespread application of type II L-asparaginase (ASNase) in the food and pharmaceutical industries. In this study, smart libraries were constructed by semi-rational design to improve the catalytic activity of type II ASNase from Bacillus licheniformis. Mutants with greatly enhanced catalytic efficiency were screened by saturation mutations and combinatorial mutations. A quintuple mutant ILRAC was ultimately obtained with specific activity of 841.62 IU/mg and kcat/Km of 537.15 min−1·mM−1, which were 4.24-fold and 6.32-fold more than those of wild-type ASNase. The highest specific activity and kcat/Km were firstly reported in type II ASNase from Bacillus licheniformis. Additionally, enhanced pH stability and superior thermostability were both achieved in mutant ILRAC. Meanwhile, structural alignment and molecular dynamic simulation demonstrated that high structure stability and strong substrate binding were beneficial for the improved thermal stability and enzymatic activity of mutant ILRAC. This is the first time that enzymatic activity of type II ASNase from Bacillus licheniformis has been enhanced by the semi-rational approach, and results provide new insights into enzymatic modification of L-asparaginase for industrial applications.
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影响因子：
4.8
作者：
Bansal, Saurabh;Srivastava, Ankit;Kundu, Bishwajit
通讯作者：
Kundu, Bishwajit
影响因子：
2.9
作者：
Aghaiypour, K;Wlodawer, A;Lubkowski, J
通讯作者：
Lubkowski, J
影响因子：
4
作者：
Karamitros, Christos S.;Konrad, Manfred
通讯作者：
Konrad, Manfred
影响因子：
6.3
作者：
Genheden S;Ryde U
通讯作者：
Ryde U
具有丙烯酰胺缓解潜力的戈登分枝杆菌新型 L-天冬酰胺酶的表征
DOI：
10.3390/foods10112819
发表时间：
2021-11-16
期刊：
Foods (Basel, Switzerland)
影响因子：
--
作者：
Chi H;Chen M;Jiao L;Lu Z;Bie X;Zhao H;Lu F
通讯作者：
Lu F