A kind of ph-regulated self-assembled antimicrobial peptide and its preparation method and application

An antibacterial peptide, self-assembly technology, applied in the direction of antibacterial drugs, chemical instruments and methods, peptides, etc., can solve the problems of low biocompatibility and low stability of natural antibacterial peptides, which limit the application, and achieve low toxicity and good antibacterial properties. effect, good biocompatibility

Active Publication Date: 2021-07-30
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the low biocompatibility and low stability of natural antimicrobial peptides under physiological conditions limit its application in clinical and animal husbandry

Method used

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  • A kind of ph-regulated self-assembled antimicrobial peptide and its preparation method and application
  • A kind of ph-regulated self-assembled antimicrobial peptide and its preparation method and application
  • A kind of ph-regulated self-assembled antimicrobial peptide and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Embodiment 1: the design of antibacterial peptide

[0016] Design principle: In order to obtain antibacterial peptides that can effectively kill pathogenic bacteria and have good biocompatibility, a single peptide molecule is used to form a supramolecular nanostructure through non-covalent self-assembly, and its antibacterial activity and biocompatibility are tested. Through detection, a self-assembled antimicrobial peptide with high application value was obtained. Among the twenty kinds of amino acids that constitute natural proteins, except for Gly, the α-carbon atoms of other protein amino acids are all achiral carbon atoms. Due to the different side chains of amino acids, they can provide different interaction forces for stabilizing the three-dimensional structure of proteins, such as hydrophobic forces, electrostatic forces and hydrogen bonds, etc., to achieve the stability of protein stereostructure chemistry and physical chemistry, and to achieve the minimum free...

Embodiment 2

[0021] Example 2: Preparation of assembled peptides

[0022] 1. Polypeptides were synthesized by Fmoc solid-phase synthesis method and freeze-dried. The purity of the product was determined by electrospray mass spectrometry (ESI-MS) and reversed-phase high-performance liquid chromatography (RP-HPLC), and the purity of the antimicrobial peptide was greater than 95%.

[0023] 2. Completely dissolve the freeze-dried antimicrobial peptide powder that has passed the quality inspection in ultrapure water with pH = 6.0, sonicate for 30 minutes, and place it at room temperature for 6-7 days to allow the peptide molecules to self-assemble into nanoparticles.

Embodiment 3

[0025] 1. Morphological observation of assembled peptides

[0026] Apply 10 μl of peptide solution on a copper grid (400 square mesh), absorb for 5 minutes, then stain with 0.1% phosphotungstic acid for 30 seconds, dry for 15 minutes, and observe with a microscope. The result is as Figure 1 As shown, the self-assembling peptide SAP forms spherical nanostructures in an acidic environment.

[0027] 2. Determination of Antimicrobial Peptide Activity

[0028] A single colony was isolated from the agar plate and cultured in 5 ml of 3% tryptic soy broth (TSB) medium with shaking at 200 rpm overnight at 37°C. Add 50 μl of bacterial solution to 5 ml of fresh 3% TSB, shake at 200 rpm, and cultivate to logarithmic phase. OD 600 The bacterial solution of =0.4 was diluted 1000 times, treated with different concentrations of peptides, incubated at 37°C for 3 hours, then serially diluted the bacterial solution, inoculated on TSB agar, and incubated overnight at 37°C. Count the number ...

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Abstract

The invention discloses a pH-regulating self-assembled antibacterial peptide, its preparation method and application. The sequence of the antibacterial peptide SAP is shown in SEQ ID No.1. Design a histidine His-functionalized β-sheet peptide containing hydrophobic amino acids Trp and Val. His and Trp are respectively placed at the non-hydrogen bond sites adjacent to the β-sheet structure, distributed in a diagonal form, and other hydrogen bond sites distribute equal amounts of His and Val, respectively, D Pro-Gly acts as a β-turn to connect the two peptide chains of the β-sheet structure. The freeze-dried powder of the antimicrobial peptide is dissolved in ultrapure water with pH=6.0, and the monomers of the antimicrobial peptide fully self-assemble into nanoparticles in the aqueous solution. The antimicrobial peptide is used in the preparation of medicines for treating Escherichia coli infectious diseases in acidic environment. Antimicrobial peptides exert effective antibacterial effects in acidic environments, have low hemolytic toxicity, and have good salt ion stability, which overcomes the weakness of low stability of natural antimicrobial peptides.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a pH-regulating self-assembled antibacterial peptide, a preparation method and an application. Background technique [0002] The misuse and overuse of antibiotics has promoted the widespread spread of multi-drug resistant bacteria around the world, resulting in no or only a few drugs that can be used in clinical treatment of drug-resistant bacterial infections, and the world has entered a "post-antibiotic era", which has extensive social and economic impacts , threatening the achievement of the Sustainable Development Goals. Therefore, new antibacterial drugs should be developed while rationally using antibiotics, especially drugs with new antibacterial mechanisms to deal with the huge casualties and property losses caused by multidrug-resistant bacteria to human society. Antimicrobial peptides (AMPs), also known as host defense peptides, can destroy the integrity of bac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/00A61K38/16A61P31/04
CPCA61K38/00A61P31/04C07K14/001Y02A50/30
Inventor 单安山李丘轲李金泽丑淑丽
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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