Symmetrical-terminals antibacterial peptide PP and preparation method and application thereof

An antibacterial peptide, symmetrical technology, applied in the field of terminal symmetric antibacterial peptide PP and its preparation and application, can solve the problems of low cell selectivity, high hemolytic activity and toxicity

Inactive Publication Date: 2018-03-02
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Porcine antimicrobial peptide Protegrin-1 (PG-1) contains 18 amino acid β-sheet antibacterial peptides, which ha

Method used

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  • Symmetrical-terminals antibacterial peptide PP and preparation method and application thereof
  • Symmetrical-terminals antibacterial peptide PP and preparation method and application thereof
  • Symmetrical-terminals antibacterial peptide PP and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Design of Antimicrobial Peptides

[0018] Symmetric-terminal antimicrobial peptides were designed by embedding β-turns within two symmetrical α-helical units. First, amino acids of different structural types are selected, including hydrophobic amino acids Ile, Phe and Trp; positively charged amino acids His, Lys and Arg, and an α-helix combined six-membered residue (IHKFWR) with high hydrophobicity and perfect amphipathic structure is designed, Then, the amino acid sequence CRRRFC of the β-turn position of the porcine antimicrobial peptide Protegrin-1 (PG-1) was embedded in two α-helical monomers to form a terminally symmetrical antimicrobial peptide.

[0019] Amino acid sequence of table 1 terminal symmetrical antimicrobial peptide

[0020]

[0021] The charge number of PP is +7, and the hydrophobic value is 0.49 respectively. The porcine antimicrobial peptides Protegrin-1 (PG-1) and PP were amidated at the carboxyl terminals to increase a positive charge and incr...

Embodiment 2

[0023] PP was synthesized by solid-phase chemical synthesis.

[0024] 1. The preparation of antimicrobial peptides is carried out one by one from the C-terminal to the N-terminal, and is completed by a peptide synthesizer. First, Fmoc-X (X is the first amino acid at the C-terminal of each antimicrobial peptide) is inserted into Wang resin, and then the Fmoc group is removed to obtain X-Wang resin; then Fmoc-Y-Trt-OH (9 -Fmoxy-trimethyl-Y, Y is the second amino acid at the C-terminus of each antimicrobial peptide); according to this procedure, it is synthesized from the C-terminus to the N-terminus until the synthesis is completed, and the side of the Fmoc group is removed chain protection resin;

[0025] 2. Add a cleavage reagent to the peptide resin obtained above, react for 2 hours at 20°C in the dark, filter; wash the precipitate with TFA (trifluoroacetic acid), mix the washing liquid with the above filtrate, concentrate with a rotary evaporator, and then add 10 times Abo...

Embodiment 3

[0029] Determination of antimicrobial activity of antimicrobial peptides

[0030] 1. Determination of antibacterial activity: Prepare the peptide as a storage solution for use. The minimum inhibitory concentrations of several antimicrobial peptides were determined by the broth microdilution method. Using 0.01% acetic acid (containing 0.2% BSA) as the diluent, a series of gradient antimicrobial peptide solutions were sequentially prepared using the double dilution method. Take 100 μl of the above solution and place it in a 96-well cell culture plate, then add an equal volume of the bacteria solution to be tested (~10 5 pc / ml) in each well. Positive controls (containing bacterial fluid but not antimicrobial peptides) and negative controls (neither bacterial fluid nor peptides) were set up. Incubate at a constant temperature of 37°C for 20 hours, and the minimum inhibitory concentration is the one where no turbidity is seen at the bottom of the well with the naked eye. The te...

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Abstract

The invention provides symmetrical-terminals antibacterial peptide PP and a preparation method and an application thereof. A sequence of antibacterial peptide PP is shown as a sequence table SEQ ID No.1. The preparation method comprises the following steps: employing two symmetrical alpha spiral units, a beta-corner part amino acid sequence CRRRFC of PG-1 is embedded in the symmetrical alpha spiral units to form the symmetrical-terminals antibacterial peptide, and the symmetrical-terminals antibacterial peptide PP is designed. The invention also provides an application of the symmetrical-terminals antibacterial peptide PP in preparation of a medicine for treating gram-positive bacteria or gram-negative bacteria infectious diseases, the antibacterial peptide PP has high bacteriostatic activity and low toxicity, a therapeutic index is 80 times of PG-1, and the antibacterial peptide PP has high cell selectivity. The antibacterial peptide PP can purposely kill pathogen in organism, and increases the development potential and biological value of the antibacterial peptide.

Description

technical field [0001] The invention mainly relates to a terminal symmetrical antimicrobial peptide PP and its preparation method and application. Background technique [0002] For a long time, antibiotics have been widely used in animal husbandry. As a feed additive, they can promote the growth of livestock and poultry and improve the production efficiency of animal husbandry. However, the long-term abuse of antibiotics leads to the multiplication of drug-resistant strains and drug residues, which not only limits the development of animal husbandry but also damages human health. Screening new antimicrobial agents without drug resistance and residue is an urgent problem to be solved. Antimicrobial peptide is a small molecular polypeptide with broad-spectrum antibacterial activity, antifungal activity, antiviral activity, antiprotozoal activity and is not easy to produce drug resistance. Different from antibiotics, antimicrobial peptides mainly act on the plasma membrane of...

Claims

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Application Information

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IPC IPC(8): C07K7/08C07K1/04C07K1/20A61K38/10A61P31/04
CPCC07K7/08A61K38/00
Inventor 董娜李仲玉徐欣瑶单安山
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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