Study On The Structure Activity Relationship Of Casein-derived Bioactive Peptide And The Primary Exploration Of Directed Hydrolyzing Of Casein

This paper, studied on the structure-activity relationship of casein-derived angiotensin converting enzyme inhibitory peptide (ACEIP) and casein phosphopeptide (CPP), and primarily explored the directed hydrolyzing of casein.To investigate the relationship between the molecular structure of phosphopeptides and their calcium-binding property,6 different peptides with 0-3 successive or discontinuous phosphorylated serines based on the core structure of casein phosphopeptides were synthesized, and the capacity to inhibit the formation of calcium phosphate precipitate, the released ratio of the solubilized calcium through the dialysis membrane, and the apparent association constant of the synthesized peptides were tested, compared with each other and with that of CPP obtained by pancreatic hydrolysis of casein and subsequent purification. The results showed that Both the number of phosphorerine residue in the molecule of the peptides and the position of phosphorerine were important factors for calcium-binding activities. The phosphopeptide with 2 discontinuous phosphorylated serines had the best calcium-binding ability and the highest Ca permeation ratio through the dialysis membrane among all the phosphopeptides.Then the structure-activity relationship of ACE inhibitory peptide were studied with pharmacophore models. A data set consisting of 28 inhibitory peptides collected from the references was selected as the training set to generate the pharmacophore models. The best pharmacophore hypothesis consisted of five features (two hydrophobic functions, two hydrogen bond acceptors, and a negative ionizable function). The model was then used as query to search 3D databases of food-derived peptides. Three novel peptides with various fit values were synthesized and their ACE inhibitory activities were tested in vitro. The results showed that the in vitro activity of peptidesⅠ,ⅡandⅢwas consistent with their molecular modeling results.Finally casein was simulately hydrolyzed into small peptide by 39 protease using the software of peptidecutter on computer, and the most effective protease was chosen in terms of the amount of oligo-peptide produed by enzymatic hydrolysis, trypsin was approved to be the most suitable protease, and the rate of casein transformed to TCA (trichloroacetic acid) soluble product by trypsin is 90.32% under the condition: temperature 40℃and pH 7.5. The hydrolyzate was seperated by ultro-filtration, the ACE inhibitory rate of the MWCO<1K fraction increased from 15.19% to 92.31% after seperation at the tested concentration of 2.36 mg/ml, and the MWCO>1K fraction was rich of phosphopeptides, the N/P molar ratio of it was decreased to 23.36 to compare with that of 42.85 before seperation.