Revealing binding mechanism of β-casein to chrysin, apigenin, and luteolin and locating its binding pockets by molecular docking and molecular dynamics.

Revealing the interaction mechanism of proteins with bioactive molecules and the location of their binding pockets is crucial for predicting the structure-function relationship of proteins in drug discovery and design. Despite some published papers on the interaction of β-casein with small bioactive molecules, the ambiguity of the location and constituent amino acids of β-casein binding pockets prompted us to identify them by in silico simulation of its interaction with three polyphenols, chrysin, apigenin, and luteolin. Molecular docking revealed that the primary β-casein binding pocket for chrysin consists of five nonpolar amino acids (Leu73, Phe77, Pro80, Ile89, and Pro196), three polar neutral amino acids (Ser137, Gln138, and Gln197), and two polar charged amino acids (Glu136, and Arg198). For β-casein/apigenin and β-casein/luteolin complexes, Asn83 also contributes to forming the pocket. Molecular dynamics provided more details, such as the relative contribution of determinative amino acids and the role of various forces. For example, we found that Glu210, Glu132, and Glu35 are the most destructive residues in the binding of chrysin, apigenin, and luteolin to β-casein, respectively. Also, we observed that hydrophobic forces mainly stabilize β-casein/chrysin and β-casein/apigenin, and polar solvation (including hydrogen bonds) stabilizes β-casein/luteolin, all by spontaneous processes.