Isolation And Inhibition And Mechanism Of Peptides Against ?-glucosidase And DPP-? From Sheep Whey Protein

Diabetes mellitus(DMs),is a common metabolic disease with a genetic predisposition,of which type II diabetes is characterized by insulin resistance,hyperinsulinemia,and pancreatic ?-cell failure.?-glucosidase(AG),which plays an important role during the absorption of glucose,and DPP-?,which is very important in the course of glucose metabolism,these two enzyme inhibitors are widely used in treatment of type II diabetes.In this thesis,sheep whey protein was used as the raw material to obtain peptide solutions with AG inhibitory activity and DPP-? enzyme inhibitory activity using enzymatic digestion,and the obtained peptides were isolated,purified and identified;further screening was performed using molecular docking software,and the inhibition mechanism of the peptides on the target enzymes was investigated.After synthesizing the screened peptides,enzyme activity method and cell culture assay were used to validate their inhibitory activities in vitro.This research provides new ideas and options for the development of drugs for type II diabetes and related health food products.The study was conducted using whey protein powder as the substrate for hydrolysis,The inhibition activities of hydrolysates against AG and DPP-? enzymes were used as reference index to screen the suitable reaction conditions.The results showed that the hydrolysate obtained at 4.0 h of trypsin hydrolysis was 24.50±0.05%and had the highest AG inhibition of the three hydrolytic enzymes with a value of21.71±0.50%,and the hydrolysate of this enzyme also exhibited the highest DPP-? enzyme inhibition activity of 77.80±1.87%.DEAE-52 ion exchange chromatography was used to separate the hydrolysate of sheep whey protein treated with trypsin.The fraction with two enzyme inhibitory activities among the five fractions was H4,with the highest inhibitory activity40.19±2.88% against AG and 24.66±2.31% against DPP-?.The Sphadex G-15 dextran gel column was selected for further separation and two fractions were obtained.The H4-1 and H4-2 of the components had AG inhibitory activity,and H4-1's inhibition rate of AG was higher,16.45±1.96%.The DPP-? inhibition rate of H4-2fraction was 20.36±3.10%.H4-1 didn't exhibit DPP-? inhibitory activity.The H4-1,H4-2 fractions were identified by mass spectrometry,60 peptides were obtained from H4-1 fraction and 86 peptides were obtained from H4-2 fraction.Molecular docking was used for peptide screening.The AG inhibitory peptides were obtained as RYEEDMYWR(RY9)and RMMNAMER(RM8),and the DPP-? inhibitory peptides were RLYLHENK(RL8)and MQEHFTCCR(MQ9).RY9,RM8 were mainly bound to AG molecules through hydrogen bonding and hydrophobic interactions,and RL8 and MQ9 were mainly bound to DPP-? molecule at the active site.Caco-2 cells were used to verify the DPP-? inhibitory activity.This experiment provides ideas for the discovery of potential hypoglycemic foods and sources for the development of hypoglycemic drugs.