Insights Into The Binding Modes Of Flavones To Human Serum Albumin

Pharmaceutical interactions with human serum albumin (HSA) are of great interest, because HSA is a pharmacokinetic determinant and a good model for exploring the protein-ligand interactions. Due to the hydrophobic nature, naturally occurring flavones, which possess various pharmacological activities, bind to HSA in human plasma. Here, we have identified the binding modes of two representative flavones—baicalin (BLI) and its aglycone, baicalein (BLE)—to HSA using a combination of experimental and computational approaches. The association properties were measured applying spectroscopic methods and a higher affinity were found for BLE. As evidenced by displacement and chemical unfolding assays, both ligands bind at Sudlow site I. Furthermore, molecular docking was utilized to characterize the models of HSA-flavone complexes, and molecular dynamics (MD) simulations as well as free energy calculations were undertaken to examine the energy contributions and the roles of various amino acid residues of HSA in flavones binding; the mechanism whereby glycosylation affects the association was also discussed.By using a combination of experimental and computational methods, we presented here reasonable association models for elucidating the modes of action of flavones binding to HSA. Binding properties of two flavones, BLE and BLI, were determined using fluorescence spectroscopy, via measuring changes of the Trp214 fluorescence. Displacement binding and chemical unfolding assays were undertaken, and the result revealed that the binding sites of both flavones are located at Sudlow site I, adjacent to Trp214 and Tyr263. CD spectroscopy was used to investigate the secondary structures and interactions between HSA and flavones. Based on these findings, the binding models were mapped by molecular docking and MD simulations, thus providing residue-specific information about complexes formation. BLE and BLI were found to be located at Sudlow site I near Trp214 and Tyr263, overlapping rooms with ANS and warfarin. Additionally, the model showed the microenvironment of both flavones binding sites to be rich in nonpolar residues. To the end, we used post-MD free energy calculations following the MM-GBSA approach to determine the role of nonpolar, electrostatics, and entropic contributions in binding. The results suggested nonpolar interactions play an important role in the HSA-flavone binding. Glycosylation was found to decrease the ability of the flavone binding to HSA, due to more unfavorable electrostatic component of the solvation free energy and more unfavorable entropy penalty. Both experimental arid computational data fit each other well.For decades, pharmaceutical interactions with HSA have been of great importance in terms of drug discovery, because HSA is a key determinant for pharmacokinetic properties of many drugs. As a result, much effort has been exerted and many approaches have been utilized in this field. Our work present here suggests that MD-based computational methods, especially when combining with other experimental means, could be a valuable tool for understanding interactions between HSA and drug-like ligands.