Engineering of Glycosyltransferase for Efficient Biosynthesisof Salidroside

Salidroside has been widely used in the cosmetic and medicinal industries. Previously, potential glycosyltransferase UGT_BS was obtained for salidroside synthesis. However, the catalytic efficiency for salidroside was undesirable. In this study, a semirational design was applied to engineer UGT_BS. The quadruple mutant M4 (I62N/S129T/F168W/Y316S) showed significantly enhanced salidroside synthesis. A conversion rate of 94.7% was obtained using mutant M4, which was 2.2-fold higher than that of the wild-type. The regioselectivity of mutant M4 toward tyrosol hydroxyl was also improved, resulting in 97.9% salidroside in the total product, which was 1.2-fold higher than that of the wild type. Kinetic constants and molecular simulations indicated that increased affinity and altered conformation of the binding pocket accounted for the enhanced salidroside synthesis. Furthermore, a fed-batch cascade reaction strategy was used, and over 183 mM salidroside was obtained. The engineered mutant M4 demonstrated precise catalysis for salidroside formation.