Molecular mechanism and in vitro digestion of β-galactosidase binding to three small molecules in milk tea

Abstract

In this study, folic acid (FA) inhibited β-galactosidase (β-Gal) activity, whereas EGCG and lutein activated β-galactosidase hydrolytic activity and the binding of small molecules made β-Gal more resistant to pepsin-induced protein hydrolysis. Various spectroscopic experiments indicated that FA, EGCG, and lutein can change their ratios of α-helix, β-folding, and irregular coiling, which could affect the microenvironment around the tryptophan, and vibrate chemical bonds. Thermodynamic calculations suggested that these three small molecules interact with proteins mainly through van der Waals forces and hydrogen bonding, which is consistent with the results of molecular docking simulation experiments. The possible binding sites for β-galactosidase and the three small molecules were investigated. β-galactosidase showed the strongest binding capacity for lutein, while EGCG showed a relatively weak binding capacity. The results of density-functional theory (DFT) analysis also confirmed these findings and described that small molecules are more elastic when bound to β-galactosidase than free small molecules, which also demonstrated their stronger interaction ability. Molecular dynamics (MD) experiments also confirmed the structural flexibility of the small molecules when β-Gal was bound to the three small molecules mentioned above, in the order of β-Gal- EGCG, β-Gal-FA, and β-Gal-lutein. The present study provides new insights into the interactions of the three small molecules with β-Gal, which may facilitate the application of functional β-Gal in the food industry.