Salidroside production through cascade biocatalysis with a thermostability-enhanced UDP-glycosyltransferase

Salidroside is a phenylpropanoid glycoside with wide applications in the food, pharmaceutical, and cosmetic industries; however, the plant genus Rhodiola, the natural source of salidroside, has slow growth and limited distribution. In this study, we designed a novel six-enzyme biocatalytic cascade for the efficient production of salidroside, utilizing cost-effective bio-based L-Tyrosine as the starting material. A preliminary analysis revealed that the poor thermostability of the Bacillus licheniformis UDP-glycosyltransferase (EC 2.4.1.384) BlYjiC M6 is a bottleneck in the cascade. Therefore, a combined computational strategy was used to engineer it and finally obtained a mutant TSM6 (T304V/G307A/N309W/F123W/T344V/D271G) with a 134-fold longer half-life at 40 °C and a 13 °C higher T_m^app compared to M6. The integration of TSM6 into the cascade improved salidroside productivity significantly, while reducing residual intermediates. After further optimization, the whole-cell biocatalytic cascade achieved a high salidroside titer of 12.8 g·L^-1 in a 5 L bioreactor, giving a productivity of 0.53 g·L^-1·h^-1. This study provides a green and efficient biosynthetic process for salidroside production and highlights the potential of enzyme engineering to enhance the biocatalytic cascade.