Cyanidin‐3‐O‐Glucoside Covalent Inhibition of Insulin‐Degrading Enzyme Reveals the Potential Molecular Mechanism for Insulin Resistance Improvement

ABSTRACT Cyanidin‐3‐O‐glucoside (C3G), widely reported for ameliorative effects on insulin resistance and blood glucose levels, is often attributed to modulating insulin signaling pathways. However, previous studies based on the target fishing method of chemical proteomics have identified C3G's ability to covalently bind target proteins, including insulin‐degrading enzyme (IDE), an Alzheimer's disease pathway protein closely related to metabolism. The specific interactions of anthocyanins with their target proteins and their correlation with activity remain unverified. Therefore, elucidating the relationship between C3G's bioactivity and target proteins is crucial. In this study, we employed various methods to clarify the binding and activity effects of C3G on IDE. CETSA‐WB revealed that C3G binds to IDE and significantly enhances its thermal stability. Insulin and fluorescent peptide substrate measurements identified C3G's enzymatic inhibitory activity with an IC 50 of 109.6 μM. The preparation of recombinant human IDE ( rh IDE) allowed us to directly confirm the covalent binding of C3G to IDE. Furthermore, the silencing of IDE in cells negated the significant improvement in insulin resistance by C3G. These findings unveil a potential novel molecular mechanism by which C3G may regulate metabolic pathways, offering guidance for understanding how anthocyanins exert their functions through interactions with target proteins.