Mechanistic insights into α-amylase inhibition, binding affinity and structural changes upon interaction with gallic acid

Abstract

The inhibitory mechanism of gallic acid (GA) on α-amylase was investigated through enzymatic inhibition assays, multi-spectroscopy, Job plot analysis and in-silico molecular docking. The results reveal that GA competitively binds to the active site of α-amylase in a 1:1 stoichiometry, dominantly via four hydrogen bonds and one hydrophobic interaction. The α-amylase-GA complex showed an increased thermal stabilisation, accompanied by a rise in α-helical and β-sheet components, resulting in a partial folding of the protein structure. The compact structure prevents substrate binding and inhibits enzyme activity. The self-quenching ability of GA was considered in the intrinsic fluorescence analysis, revealing a distinct loss in fluorescence intensity with a blueshift in the α-amylase-GA complex spectra. This observation suggests a direct interaction between GA and tryptophan amino acid residues, as the residues move to a more hydrophobic environment. Findings in this study provide insights into the interaction mechanism between GA and α-amylase, which will aid in understanding the inhibitory mechanism of their complex forms found in nature.