Abstract
Purpose: The study on the interaction between various compounds and macromolecules such as enzymes has been very important for monitoring the alteration of structural and functional properties of them. Resveratrol (3, 5, 4-trihydroxy-stilbene; RES) is a biologically active phytoallexin found in grapes and other food products. This article shows an interaction of native bovine liver catalase (BLC) with natural antioxidant product, trans resveratrol (tRES) using multispectroscopic methods.Methods: The interaction between BLC and tRES is performed using UV-vis absorption, fluorescence and circular dichroism (CD) spectroscopy and molecular docking study.Results: In vitro kinetic studies indicated that tRES can decrease BLC activity through uncompetitive inhibition. The results of spectroscopic methods represented that the binding of tRES with BLC can change the micro-region around aromatic amino acids (tryptophan (Trp) and tyrosine (Tyr)) and quench intrinsic fluorescence of BLC by a static mechanism. According to fluorescence quenching data analysis, it was revealed that tRES has one binding site on BLC. The thermodynamic parameters were obtained, which demonstrated that tRES can bind to BLC by van der Waals forces and hydrogen bonds. Molecular docking results indicated that tRES binds to BLC away from heme group and near to the Tyr 324 and Phe 265. These results are in agreement with the experimental results.Conclusion: The inhibitory effect of tRES on BLC demonstrated that excessive consumption of the antioxidants could be resulted in hazardous effects.
Highlights
Catalase (H2O2:H2O2 oxidoreductase, EC 1.11.1.6), one of the main components of the antioxidative defense system, is an active and ubiquitous enzyme that exists in almost all living organisms
Decomposition of H2O2 by bovine liver catalase (BLC) leads to decrease in H2O2 UV absorption and this reduction was considered as BLC activity
This reduction is due to suicide inactivation of BLC by H2O2.20 Michaelis-Menten and Lineweaver-Burk graphs were plotted and kinetic parameters were calculated according to Lineweaver-Burk graph and equation (4): 1 km . 1 1 (4)
Summary
Catalase (H2O2:H2O2 oxidoreductase, EC 1.11.1.6), one of the main components of the antioxidative defense system, is an active and ubiquitous enzyme that exists in almost all living organisms. It protects human and animal tissues against toxic effects of hydrogen peroxide (H2O2) that acts as a free radical.[1,2] Catalase deficiency leads to many disorders such as aging, mutagenesis, acatalasemia, urinary tract diseases, diabetes, Alzheimer’s disease, vitiligo and tumors.[3] Bovine liver catalase (BLC) is a homotetramer enzyme with four subunits. NADPH is not necessary for the activity of catalase and prevents inactivating of enzyme at low concentration of H2O2.4 There are three channels in catalase structure, which one of them reaches the active site Through this channel substrate (H2O2) can enter and products can exist. Equation (2), compound I is reduced by another H2O2 molecule, result in formation resting form of enzyme and generation of O2 and second H2O molecule.[7,8,9]
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