CN-116514914-B - Small molecule active peptide and preparation method and application thereof

CN116514914BCN 116514914 BCN116514914 BCN 116514914BCN-116514914-B

Abstract

The invention relates to a small molecule active peptide, the amino acid sequence of which is LPAYPMP. The preparation method comprises the steps of carrying out enzymolysis on abalone viscera by using alkaline protease, inactivating enzyme after enzymolysis is finished to obtain enzymolysis liquid, carrying out centrifugation, rough filtration and ultrafiltration treatment on the enzymolysis liquid, collecting a plurality of polypeptides with molecular weight smaller than 1kDa, respectively freeze-drying the polypeptides to obtain powder, carrying out sequence identification on polypeptide freeze-dried powder to obtain a plurality of polypeptide sequences, deleting repeated sequences, screening to obtain a plurality of non-repeated polypeptides, carrying out virtual screening on the obtained polypeptides by using protein HMGR as a receptor through docking software, screening out polypeptides with the docking score of which the ranking is top, and carrying out solid-phase synthesis to obtain the small molecular active peptide. The small molecule active peptide can effectively inhibit the accumulation of total cholesterol and total triglyceride in cells, has high inhibition rate on the solubility of cholesterol micelles, has the efficacy of reducing blood fat, can be applied to preparing auxiliary blood fat reducing medicines, and has the advantages of high specificity and small toxic and side effects compared with the traditional blood fat reducing medicines.

Inventors

QIAO KUN
LIU ZHIYU
LIU LINA
CHEN BEI
XU MIN
SU YONGCHANG
LIU SHUJI

Assignees

福建省水产研究所（福建水产病害防治中心）

Dates

Publication Date: 20260512
Application Date: 20230608

Claims (2)

1. A small molecule active peptide is characterized in that the amino acid sequence is LPAYPMP.
2. The use of the small molecule active peptide of claim 1 in the preparation of a medicament for assisting in reducing blood lipid.

Description

Small molecule active peptide and preparation method and application thereof Technical Field The invention relates to the technical field of polypeptides, in particular to a small molecule active peptide, a preparation method and application thereof. Background At present, most of blood lipid reducing medicines used in clinic are statin medicines, and according to reliable researches, the use of the statin medicines has two main side effects, namely, the increase of transaminase and the adverse reaction risk of muscle pain. Therefore, the way of controlling blood fat through diet therapy is gradually favored by people. However, at present, the processing and utilization of abalone are mainly concentrated on cans, freezing, fresh marketing and the like, the intensive processing and utilization of abalone is low, and the waste of leftovers is serious. The existing research on abalone activity is mainly focused on the antioxidant activity of the leftovers such as gonads, viscera, shells, gills and the like. At present, research on development of natural hypolipidemic peptide is getting more and more popular, but abalone viscera-derived hypolipidemic peptide is freshly reported. Disclosure of Invention The invention aims to provide a small molecular active peptide, a preparation method and application thereof, which take abalone offal as a raw material to obtain polypeptide with the blood lipid reducing effect, provide a new idea for the high-value processing and utilization of the abalone offal, and expand resources for developing natural blood lipid reducing products. In order to achieve the aim, the invention discloses a small molecule active peptide, the amino acid sequence of which is LPAYPMP. The preparation method of the small molecule active peptide comprises the following steps: S1, preparation of raw materials Selecting viscera of abalone as raw materials; S2, enzymolysis The viscera of the abalone are subjected to enzymolysis by using alkaline protease, and after the enzymolysis is finished, the enzyme is deactivated to obtain an enzymolysis liquid; S3, filtering Centrifuging, rough filtering and ultra-filtering the enzymolysis liquid, collecting a plurality of polypeptides with molecular weight less than 1kDa, and respectively freeze-drying the polypeptides into powder; s4 sequence identification Performing sequence identification on polypeptide freeze-dried powder to obtain a plurality of polypeptide sequences, deleting repeated sequences, and screening to obtain a plurality of non-repeated polypeptides; S5, virtual screening And taking the protein HMGR as a receptor, virtually screening the obtained polypeptides through docking software, screening out the polypeptides with the top ranking of the docking scores, and performing solid-phase synthesis to obtain the small molecule active peptide. Preferably, in the step S2, the addition amount of alkaline protease is 8000-12000U/g of substrate, the pH value is 9.5-10.5, the ratio of the substrate to the ultrapure water is 1:40-60, the enzymolysis temperature is 40-60 ℃, and the enzymolysis time is 3-6 hours. Preferably, in the step S2, the alkaline protease is added in an amount of 10000U/g of substrate, the pH value is 10.0, the ratio of the substrate to the ultrapure water is 1:50, the enzymolysis temperature is 55 ℃, and the enzymolysis time is 4 hours. Preferably, in step S3, the enzymatic hydrolysate is centrifuged by a plate centrifuge, and is strained by a ceramic membrane, and the obtained filtrate is ultrafiltered by a membrane of < 1kDa, and the polypeptides with molecular weight less than 1kDa are collected. Preferably, in step S4, after desalting the polypeptide lyophilized powder by a desalting column, performing mass spectrometry by using a mass spectrometer of a nano-spray ion source to obtain a polypeptide sequence. Preferably, in step S5, the protein HMGR molecule is hydrogenated and dehydrated by using a docking software Discovery Studio, an energy minimization treatment is performed, the polypeptide molecule is mapped with Discovery Studio Client small molecules as ligands, finally the active pocket of the protein HMGR is determined, and virtual screening is performed under the conditions of pocket range 8 and box edge 4. The invention also discloses a compound of the protein and the polypeptide, wherein the polypeptide and the protein are combined with each other in the compound, the amino acid sequence of the polypeptide is LPAYPMP, and the protein is HMGR protein. Preferably, the HMGR protein has a crystal structure of 1HWK. In addition, the invention also discloses application of the small molecule active peptide and the small molecule active protein in preparing auxiliary hypolipidemic drugs. The invention has the following beneficial effects: 1. The small molecule active peptide can effectively inhibit the accumulation of total cholesterol and total triglyceride in cells, has high inhibition rate on the solubility of cholestero