Discovery and engineering of UDP-glycosyltransferase and its applications in natural sweetener glycosylation.

With the growing health concern on sugar consumption, the high sweetness and low calorie natural sweeteners, such as steviol glycosides, mogrosides, glycyrrhizin, and trilobatin, offer a viable alternative. Glycosylation is an effective means to enhance the functional properties of natural sweeteners. UDP-glycosyltransferases (UGTs), key enzymes that catalyze transglycosylation reactions, play a crucial role in the glycosylation of natural sweeteners. However, natural UGTs often face limitations, such as low activity, narrow substrate specificity, and undesirable regioselectivity, which hinder their applications in industrial-scale production of glycosylated natural sweeteners. This review summarizes the advancements in omics-based approaches, as well as machine learning tools for mining and discovering novel UGTs. Subsequent protein engineering methods have successfully optimized UGTs performance, resulting in enhanced activity, higher stability, improved regioselectivity, and broader substrate promiscuity. Finally, the applications of engineered UGTs in the synthesis of high-value natural sweeteners are also discussed. From our perspective, the discovery and engineering of UGTs present a promising strategy for developing new natural sweeteners.