Abstract

Operational stability under high temperature is required for enzyme application in industrial processes. Error-prone PCR and B-factor analysis were employed to enhance the thermostability of a xylanase from GH family 11 in this study. Based on the top 10 mutants screened from the random mutation libraries, mutant Xyn371 was derived from the optimal mutant Xyn370 by integrating the beneficial residues identified in the other 9 screened mutants. Subsequently, a best-saturation mutant Xyn372 originated from Xyn371 was selected with a 60-min half-life at 70 °C (0.5-min half-life for the wild-type enzyme). According to the site-saturated mutagenesis of 10 residues with higher B-factors in Xyn372, mutants Xyn375 and Xyn376 were screened; their half-lives at 70 °C were 410 and 360 min, respectively. The substituted residues located in the “palm” region of the N-terminus and the newly generated hydrogen bonds in the mutants might contribute to improved thermostability. The significantly improved thermostability of mutants will pave the way for applications in different industrial areas.