Structural insights into xylanase mutant 254RL1 for improved activity and lower pH optimum

• Published in: Enzyme and microbial technology 2021-06, Vol.147, p.109786-109786, Article 109786
• Main Authors: Xiang, La, Wang, Meixing, Wu, Lian, Lu, Zhenghui, Tang, Jingya, Zhou, Jiahai, Huang, Weixue, Zhang, Guimin
• Format: Article
• Language: English
• Subjects: Crystal structure; Glycoside hydrolase; Mutation; pKa; Xylanase
• ISSN: 0141-0229; 1879-0909
• DOI: 10.1016/j.enzmictec.2021.109786

•The crystal structure of 254RL1 was determined at 2.21 Å resolution.•Better substrates access and products release leaded to improved activity of 254RL1.•Decreased the pKa value of acid-base catalyst E159 reduced the pH optimum. Xylanases degrade xylan to valuable end products. In our previous study, the alkaline xylanase S7-xyl from Bacillus halodurans S7 was engineered by rational design and the best mutant xylanase 254RL1 exhibited 3.4-fold improvements in specific activity at pH 9.0. Further research found that the enzyme activity at pH 6.0 was almost 2-fold than that at pH 9.0. To elucidate the reason of enhanced performance of 254RL1 at decreased pH optimum, we determined the X-ray crystal structure of 254RL1 at 2.21 Å resolution. The structural analysis revealed that the mutations enlarged the opening of the access tunnel and shortened the tunnel. Moreover, the mutations changed the hydrogen bond network around the catalytic residue and decreased the pKa value of acid-base catalyst E159 which reduced the pH optimum of the xylanase. The result provided the basis for the acid-alkaline engineering of the glycoside hydrolases.