Molecular Insights Into β-Glucuronidase Inhibition by Alhagi Graecorum Flavonoids: A Computational and Experimental Approach.

Abstract

In this study, we aimed to investigate the inhibitory mechanisms of β-glucuronidase by flavonoids derived from Alhagi graecorum through both experimental and computational approaches. The activity of β-glucuronidase was assessed using an in vitro enzyme inhibition assay, where myricetin and chrysoeriol were identified as potent inhibitors based on their low IC₅₀ values. Kinetic studies were conducted to determine the inhibition type, revealing that both compounds exhibit noncompetitive inhibition of β-glucuronidase-catalyzed hydrolysis of PNPG. Molecular docking was employed to explore the binding affinities of the flavonoids, showing that myricetin formed the highest number of polar interactions with the enzyme. Additionally, molecular dynamics (MD) simulations were performed to evaluate the stability of the enzyme-inhibitor complexes, demonstrating consistent trajectory behavior for both compounds, with significant energy stabilization. Interaction energy analyses highlighted the dominant role of electrostatic forces in myricetin's inhibition mechanism, while Van der Waals forces were more prominent for chrysoeriol. The MM/PBSA method was used to calculate the binding free energies, with myricetin and chrysoeriol exhibiting the lowest values. Potential energy landscape analysis further revealed that β-glucuronidase adopts a more closed conformation when bound to these inhibitors, limiting substrate access. These findings suggest that flavonoids from Alhagi graecorum hold promise for clinical applications, particularly in managing drug-induced enteropathy.