Improvement of Trichoderma reesei xylanase II thermal stability by serine to threonine surface mutations.

Three simple mutants, S80T, S146T, and S149T, and a double mutant, S80T–S149T, were constructed and expressed in Escherichia coli to replace Serine on the surface of the Trichoderma reesei xylanase protein with Threonine residues. While the Wild-type (WT) xylanase showed a half-life time (t1/2) of 20min at 55°C, the double mutant was more thermostable exhibiting a t1/2 value of 37min, followed by the S80T and S149T mutants whose t1/2 values were 25 and 23min, respectively. At 55°C, the S146T mutant showed a decrease in thermostability with a t1/2 value of 3min. While the WT enzyme retained only 32% of residual activity after incubation for 5min at 60°C, the S80T, S149T, and the S80T–S149T mutant enzymes retained 45%, 41%, and 60%, respectively. Molecular modeling attributed the increase in the thermostability of the S80T and S149T mutants to a new hydrogen bond formation and a packing effect, respectively.