Inhibitory Effects Of Pyrogallol And Tannic Acid On The Activity Of α-glucosidase

BackgroundThe incidence of type 2 diabetes is increasing year by year and there is a tendency to rejuvenate.The a-glucosidase inhibitor is closely related to the treatment of type 2 diabetes.How to effectively prevent the occurrence and development of chronic diseases has become a hot spot in the field of public health research.Some phenolic compounds in the environment may have application value in the prevention and treatment of chronic diseases.Pyrogallol and tannic acid are polyphenolic organic compounds widely present in the environment:pyrogallol can inhibit the in situ spread of breast ductal carcinoma and increase in cancer cells,anti-infection,induce apoptosis of lung cancer cells,etc.;tannic acid has anti-oxidation,anti-neuroinflammation,alleviation of Alzheimer’s disease,and delays of diabetic nephropathy.According to previous reports,they inhibit various enzyme activities,but the mechanism of inhibition of alpha-glucosidase(a-GC)activity is still unknown.In addition,their industrial production wastewater will also cause certain pollution to the environment.Therefore,it is very important to effectively use phenol-containing wastewater.In this experiment,the combination of enzyme inhibition kinetics and computer simulation was used to investigate the effects of pyrogallol and tannic acid on the inhibition of a-GC activity and its structure and function,which helped to understand the phenolic substances to a-GC.The mechanism of activity inhibition provides some new theoretical directions and basis for the development of new drug research for type 2 diabetes.MethodThe experimental methods mainly include:(1)measuring the effects of different concentrations of pyrogallol and tannic acid on the activity of a-GC;(2)evaluating whether resorcinol and tannic acid reversibly inhibit a-GC and the type of inhibition;(3)The effects of different concentrations of pyrogallol and tannic acid on the activity of the enz)yme were determined under different action times;(4)the conformation of the enzyme and the change of its surface hydrophobicity were analyzed by endogenous fluorescence method and ANS combined with fluorescence;(5)using molecular dynamics Learning and computer simulation docking to predict the three-dimensional structure of the enzyme and the site of the binding of pyrogallol and tannic acid to a-GC and their related residues.ResultsPyrogallol binds to a-GC in a reversible and relatively tight manner with typical mixed inhibition,and IC50=0.72±0.051 mM,Kl=0.37±0.018 mM,which mediated a-GC inactivation did not show significant time-kinetics.Endogenous fluorescence and ANS-combined with fluorescence analysis showed that pyrogallol can induce tertiary structure changes of a-glucosidase,which was associated with loss of enzyme activity(K=2.98±0.41 mM-1;n=1.01±0.09).Pyrogallol interacts with residues of q-GC activity sites(ASP68,MET69,TYR71,PHE157,PHE158,PHE177,GLN181,HIS348,ASP349,ASP406,VAL407,ASP408,ARG439 and ARG443).Tannic acid can rapidly and partially reversibly inhibit the activity of a-GC.It belongs to slope-parabolic linear type of mixed inhibition,with the IC50 is 1.21±0.03μM and Kl is 0.41±0.032μM.Tannic acid-mediated a-GC inactivation can occur rapidly without significant time dynamics.Tannic acid induced partial structural changes in α-GC rather than overall conformational changes and did not cause hydrophobic surface exposure(K=0.A2±0.44μM-1;n=1.89±0.11).The tannic acid interacts with residues near the a-glucosidase active site(MET69,TYR71,PHE177,ARG212,ASP214,GLU276,HIS348,ASP349,and ARG439).ConclusionBoth pyrogallol and tannic acid can inhibit the activity of α-GC,and the inhibition of tannic acid is stronger.Pyrogallol and tannic acid inhibit α-GC activity in a typical pure mixed type and the slope-parabolic mixed inhibition type,respectively.And pyrogallol induced the overall structural changes of α-GC;tannic acid induced partial regional structural changes of α-GC.Pyrogallol and tannic acid interact with residues near the active center of α-GC,and the phenolic hydroxyl groups in their structures play an important role in the mechanism of ligand binding and inhibition.