Design, expression and functional characterization of a thermostable xylanase from Trichoderma reesei.


Xylanases isolated from microorganisms such as the Trichoderma reesei have attracted considerable research interest because of their potential in various industrial applications. However, naturally isolated xylanases cannot withstand harsh conditions such as high temperature and basic pH. In this study, we performed structural analysis of the major T. reesei xylanase (Xyn2), and novel flexible regions of the enzyme were identified based on B-factor, a molecular dynamics (MD) parameter. To improve thermostability of the Xyn2, disulfide bonds were introduced into the unstable flexible region by using site-directed mutagenesis and two recombinant xylanases, XM1 (Xyn2Cys12-52) and XM2 (Xyn2Cys59-149) were successfully expressed in Pichia pastoris. Secreted recombinant Xyn2 was estimated by SDS-PAGE to be 24 kDa. Interestingly, the half-lives of XM1 and XM2 at 60°C were 2.5- and 1.8- fold higher, respectively than those of native Xyn2. The XM1 also exhibited improved pH stability and maintained more than 60% activity over pH values ranging from 2.0 to 10.0. However, the specific activity and catalytic efficiency of XM1 was decreased as compared to those of XM2 and native Xyn2. Our results will assist not only in elucidating of the interactions between protein structure and function, but also in rational target selection for improving the thermostability of enzymes.

Submission Details

ID: fxAJ3y3r

Submitter: Shu-Ching Ou

Submission Date: March 7, 2019, 4:48 p.m.

Version: 1

Publication Details
He J;Tang F;Chen D;Yu B;Luo Y;Zheng P;Mao X;Yu J;Yu F,PLoS One (2019) Design, expression and functional characterization of a thermostable xylanase from Trichoderma reesei. PMID:30650138
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
99.5 Endo-1,4-beta-xylanase G0RUP7 XYN2_HYPJQ
99.5 Endo-1,4-beta-xylanase P36217 XYN2_HYPJR
94.2 Endo-1,4-beta-xylanase B5A7N4 XYN2_TRIHA
93.7 Endo-1,4-beta-xylanase P48793 XYN_TRIHA