College of Biological and Pharmaceutical Engineering, Anhui Engineering Laboratory for Conservation and Sustainable Utilization of Traditional Chinese Medicine Resources, Anhui Province Key Laboratory for Quality Evaluation and Improvement of Traditional Chinese Medicine, West Anhui University, Lu'an city 237012, China.
College of Biological and Pharmaceutical Engineering, Anhui Engineering Laboratory for Conservation and Sustainable Utilization of Traditional Chinese Medicine Resources, Anhui Province Key Laboratory for Quality Evaluation and Improvement of Traditional Chinese Medicine, West Anhui University, Lu'an city 237012, China; School of Pharmacy, Anhui University of Traditional Chinese Medicine, HeFei 230012, China.
Food Chem. 2024 Dec 1;460(Pt 2):140698. doi: 10.1016/j.foodchem.2024.140698. Epub 2024 Jul 31.
Resveratrol is a promising functional ingredient applied in food products. However, low bioavailability and poor water solubility, which can be improved by glycosylation, hinder its application. A uridine diphosphate-dependent glycosyltransferase (UGT) from Bacillus subtilis 168 (named UGT) presents potential application for resveratrol glycosylation; nonetheless, imprecise regioselectivity renders the synthesis of resveratrol-3-O-β-D-glucoside (polydatin) difficult. Therefore, molecular evolution was applied to UGT. A triple mutant Y14I/I62G/M315W was developed for 3-OH glycosylation of resveratrol and polydatin accounted for 91% of the total product. Kinetic determination and molecular docking indicated that the enhancement of hydrogen bond interaction and altered conformation of the binding pocket increases the enzyme's affinity for the 3-OH group, stabilizing the enzyme-substrate intermediate and promoting polydatin formation. Furthermore, a fed-batch cascade reaction by periodic addition of resveratrol was conducted and nearly 20 mM polydatin was obtained. The mutant Y14I/I62G/M315W can be used for polydatin manufacture.
白藜芦醇是一种有应用前景的功能性食品配料。然而，其生物利用度低、水溶性差，可以通过糖基化来改善。枯草芽孢杆菌来源的尿苷二磷酸依赖性糖基转移酶（UGT）（命名为 UGT）具有将白藜芦醇进行糖基化的应用潜力；然而，其区域选择性不精确使得白藜芦醇-3-O-β-D-葡萄糖苷（虎杖苷）的合成变得困难。因此，对 UGT 进行了分子进化。开发了三重突变体 Y14I/I62G/M315W 用于白藜芦醇的 3-OH 糖基化，虎杖苷占总产物的 91%。动力学测定和分子对接表明，氢键相互作用的增强和结合口袋构象的改变增加了酶对 3-OH 基团的亲和力，稳定了酶-底物中间物，促进了虎杖苷的形成。此外，通过周期性添加白藜芦醇进行了分批补料级联反应，得到了近 20 mM 的虎杖苷。突变体 Y14I/I62G/M315W 可用于虎杖苷的生产。
