Molecular Insights into Covalent Binding between Anthocyanin C3G and α-Glucosidase: Interaction of Flavylium with Cysteine Thiol

Abstract Dietary anthocyanins have attracted considerable attention for their ability to modulate postprandial glycemia, yet their molecular mechanisms remain unclear. Here, we show that cyanidin-3-O-glucoside (C3G), a major anthocyanin, inhibits α-glucosidase through a cysteine-directed covalent interaction. C3G inhibited yeast α-glucosidase and a mouse intestinal α-glucosidase preparation with IC50 values of 169.7 and 719.1 μM, while time-dependent kinetic analysis yielded apparent kinact/KI values of 1.46 × 10–5 and 3.02 × 10–6 μM–1 min–1. Activity-based protein profiling (ABPP) and cysteine-blocking experiments supported cysteine-dependent probe engagement, with direct covalent probe labeling and reduced labeling after cysteine blockade. Mechanistically, the intrinsic electrophilic flavylium (FAV) scaffold undergoes nucleophilic addition with free thiols to form stable covalent adducts. Covalent docking suggested Cys9 as a candidate residue in mammalian α-glucosidase. Cysteine-directed reactivity provides a chemical basis for the sustained inhibitory effects of C3G. This mechanism reveals a previously underappreciated covalent component of anthocyanin–α-glucosidase interactions.