Antibacterial peptide hp10 and its preparation method and application

A technology of antimicrobial peptide and synthesis method, which is applied in the field of antimicrobial peptide HP10 and its preparation, which can solve the problems of low antibacterial activity, toxic and side effects of animal cells, and restrictions on the popularization and application of antimicrobial peptides, and achieve a strong killing effect and is not easy to produce drug resistance , no residue to produce drug resistance effect

Active Publication Date: 2018-04-03
GUANGZHOU AONONG BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most of the antimicrobial peptides from natural sources have defects such as large molecular weight, immunogenicity, low antibacterial activity, toxic side effects on animal cells, or hemolysis, which to a certain extent limits the popularization and application of antimicrobial peptides in pig breeding

Method used

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  • Antibacterial peptide hp10 and its preparation method and application
  • Antibacterial peptide hp10 and its preparation method and application
  • Antibacterial peptide hp10 and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1: Preparation of antimicrobial peptide HP10:

[0021] The antibacterial peptide HP10 is intercepted and modified on the basis of the natural antibacterial peptide PMAP36 (GRFRRLRKKTRKRLKKIGKVLKWIPPIVGSIPLGCG). Since histidine is an essential amino acid for juvenile animals, and its antibacterial activity is stronger than that of arginine and lysine, so Replace arginine and lysine with histidine.

[0022] The chemical synthesis method of antimicrobial peptide HP10 comprises the following steps:

[0023] (1) According to the amino acid sequence HHHLHHIHKP of the antimicrobial peptide HP10, the antimicrobial peptide HP10 was activated and cross-linked one by one from the C-terminal to the N-terminal by the FMOC protection synthesis method, which was synthesized by Shanghai Gil (GL) Biochemical Co., Ltd. Synthesize the full sequence of antimicrobial peptides through a fully automatic peptide synthesizer (ABI433);

[0024] (2), the product of step (1) is desalte...

experiment example 1

[0029] Experimental example 1: Antibacterial peptide HP10 antibacterial test:

[0030] (1), Test strains: Escherichia coli, Salmonella, Staphylococcus aureus, Staphylococcus epidermidis, Bacillus subtilis and Bacillus megaterium.

[0031] (2), polypeptide mother solution (0.2% BSA+0.01% acetic acid): Weigh 0.2g bovine serum albumin (BSA) and add 80mLddH 2 Mix well in O water, then add 10 μL of acetic acid solution, then wash with ddHO 2 Dilute to 100 mL with O water, filter and sterilize with a filter membrane with a pore size of 0.22 μm, and store at 4°C for later use.

[0032] The antibacterial effect was reflected by measuring the minimum inhibitory concentration (MIC) of the antimicrobial peptide HP10, which was the lowest sample concentration at which no bacterial growth could be detected. Cultivate the test bacteria overnight at 37°C for 16 hours, then transfer 100 μL of the overnight cultured bacterial solution to fresh MHB broth medium, and cultivate for 2.5 to 3 h...

experiment example 2

[0036] Experimental example 2: Effects of antimicrobial peptide HP10 on growth performance, intestinal health and immune function of weaned piglets:

[0037] A total of 320 weaned piglets at the age of 21 days were selected, and were randomly divided into 4 treatment groups according to body weight and sex according to the experimental design scheme shown in Table 2. The negative control group was the basal diet, and the HP10- 50mg / kg antimicrobial peptide HP10 was added to the basal diet in the 50 group; 100mg / kg antimicrobial peptide HP10 was added to the basal diet in the HP10-100 group, and 100mg / kg aureomycin was added to the basal diet in the positive control group. 10 repetitions, 8 pigs per repetition, 14 days of test period.

[0038] Table 2 Experimental Design Scheme

[0039]

[0040] The effects of antimicrobial peptide HP10 on production performance, diarrhea rate and immunoglobulin of weaned piglets are shown in Table 3, figure 1 and Table 4. As can be see...

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Abstract

The antibacterial peptide HP10 of the present invention and its preparation method and application belong to the technical field of peptide synthesis containing 5-20 amino acids and the specific therapeutic activity technical field of compounds and pharmaceutical preparations. The amino acid sequence of the antimicrobial peptide HP10 is HHHLHHIHKP; its molecular weight is 1292.4Da, and its isoelectric point is 8.79. Its preparation comprises the following steps: (1), the antibacterial peptide HP10 that amino acid sequence is HHHLHHIHKP is activated and cross-linked one by one from C-terminus to N-terminus by FMOC protection synthetic method; (2), the product of step (1) is subjected to HPLC Antimicrobial peptide HP10 was obtained by desalting and purification by reverse phase column chromatography. The antimicrobial peptide HP10 in the present invention has the advantages of small molecular weight, convenient artificial synthesis, strong thermal stability, strong bactericidal effect, and no drug resistance; it also has the advantages of significantly improving the intestinal health of piglets and strengthening the body when applied in piglet feed. Advantages of immunity, improving piglet growth performance.

Description

technical field [0001] The invention belongs to the technical field of peptide synthesis containing 5-20 amino acids and the technical field of specific therapeutic activity of compounds and pharmaceutical preparations, and specifically relates to antibacterial peptide HP10 and its preparation method and application. Background technique [0002] Feed antibiotics have played a vital role in the development of the global livestock industry. In recent years, in order to pursue greater economic benefits, the abuse of antibiotics in animal production has become increasingly serious, which has not only caused the problem of veterinary drug residues in animal foods, but also led to the emergence of drug-resistant strains, which has threatened the sustainable development of animal husbandry. caused serious trouble. Therefore, developed countries such as the European Union and South Korea have completely banned the use of antibiotics in livestock and poultry feed, and finding alter...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/06A23K20/147A23K50/60C07K1/20C07K1/10C07K1/06
Inventor 姜文周盛昌苏斌朝施建成肖丽萍叶俊标
Owner GUANGZHOU AONONG BIOTECH
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