Improving the Thermal Stability of GH11 Xylanase XynASP through Cord Region Engineering

The thermostability and catalytic activity of GH11 xylanase XynASP from Aspergillus saccharolyticus JOP 1030-1 were improved by systematically engineering the cord region. Ultimately, mutant DSM4 was developed through iterative combinations of mutations. Compared to the wild-type XynASP, DSM4 showed a 130.9- and 9.3-fold increase in t_1/2^{50 °C} and catalytic efficiency, respectively. Reducing the flexibility of the cord region boosted the overall rigidity, resulting in improved thermal stability. The extensive catalytic cleft and prolonged contact between catalytic residues and the substrate were likely key factors in enhancing catalytic activity. Maintaining the thumb highly flexible can offset the negative impact on catalytic activity during the thermal stability modification of the cord region. This study indicates that the cord region is an effective target for enhancing the thermostability and catalytic activity of GH11 xylanase through engineered modifications.