Hyperuricemia is mainly caused by insufficient renal urate excretion. Urate transporter 1 (URAT1), an organic anion transporter, is the main protein responsible for urate reabsorption. In this study, we utilized artificial intelligence-based AlphaFold2 program to construct URAT1 structural model. After molecular docking and conformational evaluation, four e-pharmacophoric models were constructed based on the complex structures of probenecid-URAT1, benzbromarone-URAT1, lesinurad-URAT1, and verinurad-URAT1. Combining pharmacophore modeling, molecular docking, MM/GBSA calculation and ADME prediction, 25 flavonoids were selected from the natural products database containing 10,968 molecules. Then, a model of HEK-293T cells overexpressing URAT1 was constructed, and the inhibitory activity to URAT1 of 25 flavonoids was evaluated by measuring their effect on cellular uptake of 6-carboxyfluorescein (6-CFL). Fisetin, baicalein, and acacetin showed the best activity with IC50 values of 12.77, 26.71, and 57.30 µM, respectively. Finally, the structure-activity relationship of these three flavonoids was analyzed by molecular docking and molecular dynamics simulations. The results showed that the carbonyl group on C-4 and hydroxyl group on C-7, C-4', and C-5’ in flavonoids were conducive for URAT1 inhibitory effects. This study facilitates the application of flavonoids in the development of URAT1 inhibitors.Communicated by Ramaswamy H. Sarma
Pei He,Haiqi Xu,Can Yang,Dehong Yu,Yi Liu,Jiana Du,Yanfang Li
Xanthine oxidase (XO) is a key target for gout treatment. Great efforts have been made towards the discovery and development of new XO inhibitors. Aureusidin (AUR), a natural compound, emerges as the second reported XO inhibitor with an aurone skeleton with an IC50 value of 7.617 ± 0.401 μM in vitro. The inhibitory mechanism of AUR against XO was explored through enzyme kinetic studies, multi-spectroscopic methods, computer simulation techniques, and ADME prediction. The results showed that AUR acts as a rapid reversible and mixed-type XO inhibitor and its binding to XO was driven by hydrogen bonding and hydrophobic interaction. Moreover, AUR presented a strong fluorescence quenching effect through a static quenching process and induced a conformation change of XO. Its binding pattern with XO was revealed through molecular docking, and its affinity toward XO was enhanced through interactions with key amino acid residues in the active pocket of XO. Further, AUR demonstrated good stability and pharmacokinetic behavior properties in molecular dynamics simulation and ADME prediction. In short, the current work clarified in depth the inhibitory mechanism of AUR on XO firstly and then provided fresh insights into its further development as a natural potent XO inhibitor with aurone skeleton.
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