Characterization and rational engineering of a novel thermostable xylanase from Streptomyces lomondensis S015 for...

Enzymatic hydrolysis of hemicellulose from the plant biomass to sustainably produce value-added compounds represents a promising approach for carbon neutrality, which requires efficient and robust xylanases. In this study, we report the biochemical characterization of a new GH11 family xylanase SlXyn11A from Streptomyces lomondensis S015, and the protein engineering of SlXyn11A for enhanced thermostability and operational properties. SlXyn11A exhibited optimal activity at 70 °C and pH 7.2, liberating xylotriose and xylotetraose as the primary products from beechwood xylan. A total of 3.96 ± 0.05 g/L xylooligosaccharides were produced from 7.5 g/L beechwood xylan by SlXyn11A within four hours. We then substituted the C- and N-terminal sequences of SlXyn11A with those from thermophilic xylanase, generating NF-SlXyn11A-NX with significantly improved thermostability. Further molecular dynamics simulations revealed that terminal substitution reduced the flexibility of several regions, specifically Tyr122-Thr132, and decreased the unfolding motions for the increased rigidity, which ameliorated conformation stability. This study not only identified a new xylanase with industrial application potential but also elevated the understanding of terminal substitution to the xylanase thermostability.