A Study On Hypoglycemic Mechanism Of Mulberry Leaf Extract 1-Deoxynojirimycin

1-Deoxyrijimycin?DNJ?,which is widely distributed in microorganisms and plants,is a kind of polyhydroxy alkaloid extracted from plants.Pharmacological studies showed that DNJ exhibited the inhibition against diabetes related proteases,e.g.,protein tyrosine phosphatase 1B?PTP1B?,peroxisome proliferator-activated receptor??PPAR??,proliferator peroxisome-activated receptor?,?-Amylase,?-Glucosidase,etc.It has shown good potential application value in the treatment of obesity and diabetes.At present,the research on DNJ is mainly focused on the optimization of the separation and extraction process,biological activity detection of the extractives,and physicochemical property studies.However,the biosynthetic mechanism and hypoglycemic mechanism are seldom reported.This paper presents a theoretical study on DNJ,a hypoglycemic substance substantially reported in Domestic and foreign papers.Chpater 2 reports that 5 diabetes related proteases,i.e.,PTP1B,PPAR?,PPAR?,?-Amylase,and?-Glucosidase,and initial inference to DNJ's hypoglycemic targets is?-amylase and?-Glucosidase.In Chapter 3,we performed a 1 ns molecular dynamics?MD?simulation for each system,and calculated the binding free enery using MM/GBSA method.It is found that binding of DNJ to?-Glucosidase was predominat.Therefore,we speculate that?-Glucosidase is a potential target of DNJ.In order to investigate its binding mode and mechanism of action?MOA?,an extended 20 ns of MD simulation and MM/GBSA binding free energy calculation were performed in Chapter 3.The results showed that Asp202,Glu271,and Asp333 were main key residues,which played an important role in DNJ binding to?-Glucosidase through steady hydrogen bondings.Asn331 and His332 were minor key residues which worked by hydrophobic interaction and electrostatic attraction.To verify and supplement of research results,in 4 chapter we are applying the method of spectrum,ultraviolet fluorescence,CD to further explore the the interaction of DNJ and alpha glycosidase enzymes,DNJ-glycosidase enzymes system of quenching mechanism of combination mode,combining site,etc,and further look the influence of DNJ to alpha glycosidase enzymes'secondary structure and binding sites in approximate physiological conditions.The concentration of the partial inhibition of DNJ and?-glucosidase reactions was 0.297 ug/mL.The linewey-burk double-inverse method come to a conclusion that inhibition type of DNJ to?-glucosidase is competitive inhibition.The reaction is static quenching with generating ground-state complexes,which make alpha glycosidase endogenous fluorescence quenched.The process is an entropy-driven endothermic reaction.The main driving factor of the reaction is hydrophobic force.In different temperature?273K,298K,310K?,fluorescence quenching kinetic constant K_A is 1.48×10⁴ L·mol^-1,1.29×10⁴ L·mol^-1 and 1.12×10⁴ L·mol^-1.DNJ made conformation of alpha glycosidase changed,whose secondary structure rearranged.The changed conformation induces the enzyme active pocket to close,and substrate is not good for binding to the active site,this is the mechanisation that DNJ lower the activity of alpha glycosidase.This is consistent with our conclusion of molecular docking and molecular dynamics simulation.This study associate computer simulation and spectroscopy to clarify the mechanism of DNJ binding to?-Glucosidase in simulate human physiology condition,which may provide clues to design of hypoglycemic drugs and development of hypoglycemic mulberry leaf foods.