数字农科院2.0

Enhancing the endo-activity of the thermophilic chitinase to yield chitooligosaccharides with high degrees of polymerization

文献类型: 外文期刊

作者: Feifei Guan;Xiaoqian Tian;Ruohan Zhang;Yan Zhang;Ningfeng Wu;Jilu Sun;Honglian Zhang;Tao Tu;Huiying Luo;Bin Yao;Jian Tian;Huoqing Huang

作者机构:

关键词: Chitin;Chitinase;Endo-activity;Functional oligosaccharides;Mutants

期刊名称: Bioresources and Bioprocessing

ISSN: 2197-4365

年卷期: 2024 年 11 卷 1 期

页码:

收录情况: SCIE(2024版) ; ; EI(2024版)

摘要: Thermophilic endo-chitinases are essential for production of highly polymerized chitooligosaccharides, which are advantageous for plant immunity, animal nutrition and health. However, thermophilic endo-chitinases are scarce and the transformation from exo- to endo-activity of chitinases is still a challenging problem. In this study, to enhance the endo-activity of the thermophilic chitinase Chi304, we proposed two approaches for rational design based on comprehensive structural and evolutionary analyses. Four effective single-point mutants were identified among 28 designed mutations. The ratio of (GlcNAc)3 to (GlcNAc)2 quantity (DP3/2) in the hydrolysates of the four single-point mutants undertaking colloidal chitin degradation were 1.89, 1.65, 1.24, and 1.38 times that of Chi304, respectively. When combining to double-point mutants, the DP3/2 proportions produced by F79A/W140R, F79A/M264L, F79A/W272R, and M264L/W272R were 2.06, 1.67, 1.82, and 1.86 times that of Chi304 and all four double-point mutants exhibited enhanced endo-activity. When applied to produce chitooligosaccharides (DP ≥ 3), F79A/W140R accumulated the most (GlcNAc)4, while M264L/W272R was the best to produce (GlcNAc)3, which was 2.28 times that of Chi304. The two mutants had exposed shallower substrate-binding pockets and stronger binding abilities to shape the substrate. Overall, this research offers a practical approach to altering the cutting pattern of a chitinase to generate functional chitooligosaccharides. Graphical abstract: (Figure presented.)

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