title: Structural perspectives of an engineered β-1,4-xylanase with enhanced thermostability.
abstract: The glycoside hydrolase 10 (GH10) xylanase from Streptomyces sp. 9 (XynAS9) can operate in a broad range of pH and temperature, and thus is a potential candidate for commercial applications. Recently, we engineered XynAS9 via mutating several residues in accordance with the consensus sequences of GH10 thermophilic xylanases in an attempt to improve the enzyme thermostability and thermotolerance. The most promising effects were observed in the double mutant V81P/G82E. In order to investigate the molecular mechanism of the improved thermal profile of XynAS9, complex crystal structures of the wild type (WT) and mutant (MT) enzyme were solved at 1.88-2.05Å resolution. The structures reveal a classical GH10 (β/α)8 TIM-barrel fold. In MT XynAS9, E82 forms several interactions to its neighboring residues, which might aid in stabilizing the local structure. Furthermore, the MT structure showed lower B factors for individual residues compared to the WT structure, reflecting the increased MT protein rigidity. Analyses of the XynAS9 structures also delineate the detailed enzyme-substrate interaction network. More importantly, possible explanations for the enhanced thermal profiles of MT XynAS9 are proposed, which may be a useful strategy for enzyme engineering in the future.
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Chun-Chi
authors.initials: CC
authors.lastname: Chen
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China.
authors.forename: Huiying
authors.initials: H
authors.lastname: Luo
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Xu
authors.initials: X
authors.lastname: Han
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Pin
authors.initials: P
authors.lastname: Lv
authors.affiliation: Institute of Biological Chemistry, Academia Sinica, Taipei 11529, Taiwan.
authors.forename: Tzu-Ping
authors.initials: TP
authors.lastname: Ko
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China; College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Peng
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Chun-Hsiang
authors.initials: CH
authors.lastname: Huang
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Wang
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Jian
authors.initials: J
authors.lastname: Gao
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
authors.forename: Yingying
authors.initials: Y
authors.lastname: Zheng
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China; College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China.
authors.forename: Yunyun
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Department of Biochemistry, Norman Bethune Medicine School, Jilin University, Changchun 130021, China.
authors.forename: Jianyu
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China. Electronic address: yaobin@caas-bio.net.cn.
authors.forename: Bin
authors.initials: B
authors.lastname: Yao
authors.affiliation: Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China. Electronic address: guo_rt@tib.cas.cn.
authors.forename: Rey-Ting
authors.initials: RT
authors.lastname: Guo
mesh_terms: D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D055550:Protein Stability; D013696:Temperature
chemical_list: D043364:Endo-1,4-beta Xylanases
keywords: Crystal structure; Protein engineering; Protein rigidity; Thermozyme; β-1,4-Xylanase