a-Amylase inhibition, anti-glycation property and characterization of the binding interaction of citric acid with a-amylase using multiple spectroscopic, kinetics and molecular docking approaches
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Date
2022-05
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Elsevier
Abstract
The quest to suppress complications associated with diabetes mellitus is ever increasing, while food addi- tives and preservatives are currently being considered to play additional roles besides their uses in food enhancement and preservation. In the present study, the protective prowess of a common food preserva- tive (citric acid, CA) against advanced glycation end-products (AGEs) formation and its binding interac- tion mechanism with a-amylase (AMY), an enzyme linked with hyperglycemia management, were examined. Enzyme inhibition kinetics, intrinsic fluorescence, synchronous and 3D fluorescence spectro- scopies, ultraviolet–visible (UV–Vis) absorption spectroscopy, Fourier transform-infrared (FT-IR) spec- troscopy, thermodynamics, and molecular docking analyses were employed. Results obtained showed that citric acid decreased a-amylase activity via mixed inhibition (IC50 = 5.01 ± 0.87 mM, Kic = 2.42 mM, Kiu = 160.34 mM) and suppressed AGEs formation (IC50 = 0.795 ± 0.001 mM). The intrinsic fluorescence of free a-amylase was quenched via static mechanism with high bimolecular quenching constant (Kq) and binding constant (Ka) values. Analysis of thermodynamic properties revealed that AMY-CA complex was spontaneously formed (DG < 0), entropy driven (TDS > DH), with involvement of electrostatic forces. UV–Vis, FT-IR and 3D fluorescence spectroscopies affirmed alterations in a- amylase native conformation due to CA binding interaction. CA interacted with His-101, Asp-197, His- 299, and Glu-233 within AMY active site. Our findings indicated that CA could impair formation of AGEs and interact with a-amylase to slow down starch hydrolysis; vital properties in management of type 2 diabetes complications.
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Keywords
a-Amylase inhibition, Citric acid, Anti-glycation, Hyperglycemia, Spectroscopy, Molecular docking