Precisely engineering transglycosidases as robust biocatalysts toward glycoside reformation for innovative food processing.

Transglycosidases with multiple activities represent a distinct subgroup within the glycoside hydrolase family. They can not only catalyze hydrolysis reactions using water molecules as substrate acceptors but also mediate transglycosylation reactions with other carbohydrates toward glycoside reformation. It makes them become crucial enzymes for the artificial synthesis of glycosidic bonds. However, natural transglycosidases exhibit inherent limitations, such as narrow substrate specificity, low transglycosylation efficiency, and poor tolerance, which restrict their applications. Therefore, modifying existing transglycosidases via enzyme engineering approaches (e.g., non-rational, semi-rational, and rational design) to significantly enhance their relevant properties has become a research hotspot in the field of transglycosidases. Nevertheless, traditional methods present practical drawbacks, including long development cycles, heavy workloads, and high dependence on existing information. Notably, the emergence of artificial intelligence (AI)-assisted enzyme engineering is expected to address these issues. This review summarizes the research progress on traditional ways for transglycosidase tailoring, as well as studies related to AI-assisted technology. It also covers the applications of transglycosidases in innovative food processing, for instances, oligosaccharide production, the alcoholic beverage industry, baking industry, dairy industry, cyclodextrin synthesis, and food additive development. Precisely engineered transglycosidases exert their potential as powerful biocatalysts in the field of innovative food processing.