Cai Li-Nian, Xu Sheng-Nan, Lu Tao, Lin Dong-Qiang, Yao Shan-Jing
Key Laboratory of Biomass Chemical Engineering of Ministry of Education, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, China.
College of Environment, Zhejiang University of Technology, Hangzhou, 310014, China.
Bioresour Bioprocess. 2022 Apr 21;9(1):44. doi: 10.1186/s40643-022-00533-3.
The cellulase cocktail of marine Aspergillus niger exhibited salt-tolerant and thermostable properties, which is of great potential in industrial application. In order to excavate the single tolerant cellulase components from complex cellulase cocktail, constitutive homologous expression was employed for direct obtainment of the endoglucanase (AnEGL). Enzymatic property study revealed that AnEGL exhibited a property of salt tolerance and a strong thermostability in high salinity environment. Significantly, its activity increased to 129% and the half-life at 65 °C increased to 27.7-fold with the presence of 4.5 M NaCl. Molecular dynamics simulation revealed that Na and Cl could form salt bridges with charged residues, and then influenced the activity of loops and the stability of substrate binding pocket, which accounted for the salt tolerance and thermostability. Further, site-specific mutagenesis study proved that the residues Asp95 and Asp99 in the pocket were of great concern for the tolerant properties. The salt-tolerant and thermostable AnEGL was of great value in lignocellulosic utilization and the conjectural mechanisms were of referential significance for other tolerant enzymes.
海洋黑曲霉的纤维素酶混合物具有耐盐和热稳定特性，在工业应用中具有巨大潜力。为了从复杂的纤维素酶混合物中挖掘出单一的耐受性纤维素酶组分，采用组成型同源表达直接获得内切葡聚糖酶（AnEGL）。酶学性质研究表明，AnEGL在高盐环境中表现出耐盐性和较强的热稳定性。值得注意的是，在4.5 M NaCl存在的情况下，其活性提高到129%，在65℃下的半衰期增加到27.7倍。分子动力学模拟表明，Na和Cl可以与带电荷的残基形成盐桥，进而影响环的活性和底物结合口袋的稳定性，这解释了其耐盐性和热稳定性。此外，定点诱变研究证明，口袋中的Asp95和Asp99残基对耐受性特性至关重要。耐盐且热稳定的AnEGL在木质纤维素利用方面具有重要价值，推测的机制对其他耐受性酶具有参考意义。
