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Non-specific receptor binding type fungus targeting antibacterial peptide, and preparation method and application thereof

An antimicrobial peptide and binding technology, which is applied in the field of non-specific receptor binding fungal targeting antimicrobial peptides and its preparation, can solve the problems of non-serious, negative effects of antifungal drugs that cannot be ignored, and difficult treatment methods, and achieve precise target The effects of specificity, high cell viability, and high application potential

Active Publication Date: 2020-08-14
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Candida albicans, as an opportunistic pathogen, will seriously affect the economic benefits of livestock and poultry production. However, compared with prokaryotic bacterial diseases, there are still a large number of drugs such as antibiotics. Medicinal products such as the ones that successfully destroy bacterial cells without harming normal human cells, fungal diseases make it more difficult for treatments to kill fungal cells without harming normal human cells because fungal cell structures are similar to human eukaryotic cells. Most chemical treatments are toxic to fungi and host cells and cannot be applied
There are currently four types of treatment available commercially for antifungal infections, however, the negative effects of antifungal drugs cannot be ignored, although most of these side effects are not serious, such as watery diarrhea, diarrhoea, stomach pain or nausea and a few allergic reaction
In addition, the use of antifungal drugs can disrupt the normal microbial balance in the gut and vagina, allowing harmful microorganisms to multiply and cause lesions

Method used

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  • Non-specific receptor binding type fungus targeting antibacterial peptide, and preparation method and application thereof
  • Non-specific receptor binding type fungus targeting antibacterial peptide, and preparation method and application thereof
  • Non-specific receptor binding type fungus targeting antibacterial peptide, and preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Use R language to statistically analyze the amino acid sequence characteristics of fungal antimicrobial peptides and broad-spectrum antimicrobial peptides in the natural antimicrobial peptide library, compare the parameters of the two types of antimicrobial peptides, especially the amino acid composition, and screen to obtain the best amino acid composition, namely Ala, Cys ,Ser,Leu,Ile; on this basis, the symmetrical structure of amino acid centered on tryptophan (++hyh W hyh++), (h, hydrophobic amino acid; +, positively charged amino acid; y, hydrophilic amino acid ); To obtain a short-chain targeting antimicrobial peptide sequence with low toxicity, high efficiency and less likely to cause immune regulation disorder.

[0015] Amino acid sequence of table 1 antimicrobial peptide

[0016]

Embodiment 2

[0018] Synthesis of pheromone-labeled antimicrobial peptides by solid-phase chemical synthesis

[0019] 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;

[0020] 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 evap...

Embodiment 3

[0024] Determination of Antimicrobial Peptide Activity

[0025] 1. Determination of antibacterial activity: the antimicrobial peptides are configured into a certain 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 (bacteria) and RPMI1640 (fungus) added with MOPS as diluents, 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 individual / 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 (for fungi, cultivate at 28°C for 48 hours), and the minimum inhibi...

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Abstract

The invention discloses a non-specific receptor binding type fungus targeting antibacterial peptide, and a preparation method and application thereof. The sequence of the antibacterial peptide P19 isshown as SEQ ID No. 1. The preparation method comprises the following steps: statistically analyzing related structure function parameters of the antibacterial peptide with activity to bacteria or fungi in a natural antibacterial peptide library by using R language; comparing the related parameters; screening by using the high-frequency amino acid composition, the high-frequency charge number, thehydrophobicity and the structure characteristics of the antifungal antibacterial peptide to obtain the optimal amino acid composition; and comparing the antibacterial activity and strain selectivityof each peptide chain to obtain the antibacterial peptide P19 controlled by hydrophobicity and amphipathicity of the adjusted amino acid sequence. The antibacterial peptide is applied to the preparation of drugs for treating fungal infectious diseases. The antibacterial peptide only has a relatively strong killing effect on fungal strains, especially candida albicans, has no influence on other bacterial strains, and shows relatively accurate targeting specificity.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a non-specific receptor-binding fungal targeting antimicrobial peptide and its preparation method and application. Background technique [0002] Candida albicans, as an opportunistic pathogen, will seriously affect the economic benefits of livestock and poultry production. However, compared with prokaryotic bacterial diseases, there are still a large number of drugs such as antibiotics. Medicinal products such as the ones that successfully destroy bacterial cells without harming normal human cells, fungal diseases make it more difficult for treatments to kill fungal cells without harming normal human cells because fungal cell structures are similar to human eukaryotic cells. Most chemical treatments are toxic to fungi and host cells and cannot be applied. There are currently four types of treatment available commercially for antifungal infections, however, the negative e...

Claims

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Application Information

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IPC IPC(8): C07K7/06C07K1/04C07K1/06A61K38/08A61P31/10
CPCC07K7/06A61P31/10A61K38/00
Inventor 单安山丑淑丽李丘轲李金泽
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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