Identification and characterization of the multifunctional glycosyltransferase AdUGT86A1 in Angelica decursiva

Based on the limited substrate diversity of known coumarin glycosyltransferases, we identified and characterized AdUGT86A1 , a novel O-glycosyltransferase from Angelica decursiva . In vitro enzyme activity assay and substrate promiscuity analysis showed that AdUGT86A1 catalyzed the glycosylation of various coumarin derivatives (e.g., esculetin and fraxetin) and certain flavonoids (e.g., tectorigenin) to produce the corresponding glycosides. Enzymological characterization revealed that AdUGT86A1 exhibited optimal catalytic activity at a pH of 7.0 and 30°C, with Ca 2+ ions significantly enhancing the catalytic activity. Homology modeling and molecular docking showed that AdUGT86A1 had a GT-B fold architecture with substrate-binding patterns, with H20, Q99, Y163, V212, and Y297 serving as crucial residues for its catalytic function. Notably, alanine substitution at Y297 and Q99 increased the yields of esculin and fraxin by 2–3 fold, respectively. AdUGT86A1 represents not only an efficient biocatalytic tool for coumarin glycoside biosynthesis but also a molecular template for engineering multifunctional glycosyltransferases owing to its unique substrate promiscuity. This study offers a significant reference for enhancing natural product structural modification and drug development. • AdUGT86A1 , a coumarin O-glycosyltransferase, shows high substrate promiscuity and catalytic efficiency. • Ca 2+ boosts AdUGT86A1 activity, while Fe 2+ /Cu 2+ strongly inhibit it. • H20, Q99, Y163, V212, and Y297 form the substrate-binding pocket. • Y297A/Q99A mutants increase esculin/fraxin yield 2–3 × vs wild type.