Structural and molecular insights into the binding mechanisms of β-glucosidase-ascorbic acid-rutin/isoquercetin ternary systems.

Elucidating the interaction within the β-glucosidase (BGL)-ascorbic acid (AA)-quercetin glycoside ternary system is essential for advancing the integrated utilisation of Rosa roxburghii Tratt (RRT) resources and BGL in the pharmaceutical and food industries. This study is the first to demonstrate the regulatory role of AA in a ternary system and its influence on BGL conformational changes during substrate binding. Our results show significant molecular aggregation in the BGL-AA-rutin/isoquercetin (Rut/Iso) systems, wherein Rut/Iso induced conformational alterations in BGL. Ternary system formation was primarily driven by hydrogen bonding, electrostatic interactions, and hydrophobic interactions, with Arg56, Asn115, Met258, Gly293 and Glu249 identified as key residues mediating these interactions. AA enhanced the binding affinity between Rut/Iso and BGL, and a positive synergy in the co-binding of AA and Rut with BGL. These results provide a theoretical foundation for the potential application of BGL as a carrier for drugs or functional foods and offer new insights into natural enzyme activators and the valorisation of RRT resources.