| Autor: |
Xia B; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China., Ma D; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China., Ye Z; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China., Yang J; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China., Zhang H; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China., Hu X; School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, Anhui, China. |
| Abstrakt: |
Dextranase is an enzyme that specifically hydrolyzes the α-1, 6 glucoside bond. In order to improve the activity of dextranase from Arthrobacter oxidans KQ11, site-directed mutagenesis was used to modify the amino acids involved in the "tunnel-like binding site". A saturating mutation at position 507 was carried out on this basis. The mutant enzymes A356G, S357W, W507Y, and W507F with improved enzyme activities and catalytic efficiency were successfully obtained. Compared with wild type (WT), W507Y showed the specific activity increasing by 3.00 times, the k cat value increasing by 3.62 times, the K m value decreasing by 54%, and the catalytic efficiency ( k cat / K m ) increasing by 8.98 times. The three-dimensional structure analysis showed that the increase of the number of hydrogen bonds and the distance between "tunnel-like binding sites" were important factors affecting enzyme activity. Compared with WT, W507Y had a shortened distance from the residues on the other side of the "tunnel-like binding site", which made it easier to generate hydrogen binding forces. Accordingly, the substrate hydrolysis and product efflux were accelerated, which dramatically increased the enzyme activity and catalytic efficiency. |
