Exploration of a N-terminal disulfide bridge to improve the thermostability of a GH11 xylanase from Aspergillus niger

The Journal of General and Applied Microbiology

Xin-Shu Zhu²  Chen-Yan Zhou²  Jing Kang²  Tong-Biao Li²  Yong-Tao Wang¹ 

[1] The First Affiliated Hospital, Xinxiang Medical University;School of Life Science and Technology, Xinxiang Medical University

关键词: disulfide bridge;    site-directed mutagenesis;    thermostability;    xylanase;   

DOI  :  10.2323/jgam.62.83

学科分类：微生物学和免疫学

来源: Applied Microbiology, Molecular and Cellulrar Biosciences Research Foundation

【 摘 要 】

To improve the thermostability of xylanase XynZF-2 from Aspergillus niger XZ-3S, a disulfide bridge was introduced in the N-terminal domains by site-directed mutagenesis (V1C and E27C). Simultaneously, the active sites of XynZF-2 were predicted by bioinformatics software and verified by site-directed mutagenesis (E103D and E194D). The mutated active sites xynED- and the mutated disulfide bridge xynDC-encoding genes were constructed and expressed in Escherichia coli BL21 (DE3). Compared to the native xylanase, it was found that the residual activity of the mutated XynED was 0.17%. The optimum temperature of the variant XynDC was increased to 45°C from 40°C of XynZF-2. After treatment at 40°C for 60 min, the variant XynDC retained 66.77% of their original activity, while the XynZF-2 retained about 44.36% residual activity. t1/245°C of the variant XynDC also increased from 7 min to 14 min. The results of the mutated xylanases indicated that the active center of XynZF-2 mainly consisted of two catalytic residues (Glu103 and Glu194), and the introduction of a disulfide bridge in the N-terminal domains can improve the thermostability of XynZF-2.

【 授权许可】

Unknown