Kinetic, Fluorescence Spectroscopy and Molecular Docking Studies of Tyrosinase Inhibition by Ellagic Acid

Abstract

In this study, inhibition kinetics, fluorescence spectroscopy and molecular docking simulation were used to systematically investigate the inhibitory effect and mechanism of EA on mushroom tyrosinase. The in vitro study and kinetic results showed that EA significantly inhibited tyrosinase activity with a half maximal inhibitory concentration (IC50) of 0.05 mg/mL in a reversible mixed-type manner; the binding constant KI was smaller than KIS, indicating that EA bound more tightly to the free enzyme than to the enzyme-substrate complex. The fluorescence of tyrosinase was quenched statically by EA, and they combined to generate a complex through a spontaneous endothermal process, with hydrophobic interaction being the main force; there was only one binding site or class of binding sites. Simultaneous and three-dimensional fluorescence spectroscopy analysis showed that EA increased the polarity of the microenvironment of tyrosinase, decreased the hydrophobicity, and brought the Trp residues of tyrosinase closer to the binding site. Molecular docking simulation analysis further complemented and validated the above results by showing visually that EA was a mixed-type tyrosinase inhibitor, binding to the free enzyme or enzyme-substrate complex mainly through hydrophobic interactions and hydrogen bonding, ultimately leading to reduced enzyme activity. This study is of reference significance for the application of EA as a preservative in the food industry.

Keywords

• mushroom tyrosinase; ellagic acid; spectroscopic analysis; inhibition mechanism; molecular docking