Biochemical and Biophysical Research Communications, Vol.481, No.1-2, 139-145, 2016
The disruption of two salt bridges of the cold-active xylanase XynGR40 results in an increase in activity, but a decrease in thermostability
Cold-active xylanases are of great interest due to their large potential for application in the food industry. In this study, salt bridges of the eight glycoside hydrolase (GH) family 10 cold-active xylanases reported to date were predicted and the salt bridges specific to the cold-active xylanase XynGR40 were identified. Seven mutants were constructed to disrupt salt bridges specific to XynGR40. The results suggested that five mutants lost their xylanase activity, while the other two mutants, D3ON and D83N, displayed different properties when compared with the wild-type XynGR40. First, both mutations showed an obvious decrease in thermostability, with the Tip of D3ON and D83N at 50 degrees C being about one half and one sixth of the wild-type, respectively. Second, both D3ON and D83N had a higher specific activity than the wild-type, with activities about 13 and 163% higher, respectively. Third, both D3ON and D83N had high kat and K. values, which resulted in a higher catalytic efficiency of the mutant D83N, but a lower catalytic efficiency of the mutant D3ON compared to the wild-type. Our results suggested that salt bridges play important roles in both the activity and thermostability of the cold-active xylanase XynGR40. The mutant D83N had a higher k(cat) and higher relative activity at low temperatures than the wild-type, and is a good candidate for application in the food industry. (C) 2016 Elsevier Inc. All rights reserved.