Arenicola cristata antibacterial peptide NZ17074 derived peptide N6 and application thereof

A technology for derivatizing peptides and antibacterial peptides, applied in application, antibacterial drugs, peptides, etc., can solve problems such as constraints, and achieve the effects of small molecular weight, good bactericidal effect, and easy artificial synthesis

Active Publication Date: 2017-09-22
FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, its further clinical application is

Method used

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  • Arenicola cristata antibacterial peptide NZ17074 derived peptide N6 and application thereof
  • Arenicola cristata antibacterial peptide NZ17074 derived peptide N6 and application thereof
  • Arenicola cristata antibacterial peptide NZ17074 derived peptide N6 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Design and synthesis of sea earthworm antimicrobial peptide NZ17074-derived peptide N6

[0023] 1. Based on the NZ17074 parent peptide sequence, by replacing and adding or subtracting amino acid residues, by appropriately increasing the antimicrobial peptide hydrophobicity and changing the number of disulfide bond pairs, a mutant antimicrobial peptide N6 was finally determined, trying to improve The bacteriostatic activity of the antimicrobial peptide on Gram-negative bacteria, the nucleotide sequence of the antimicrobial peptide N6 gene is shown in SEQ ID NO:2, and the amino acid sequence is shown in SEQ ID NO:1.

[0024] 2. Antimicrobial peptides are synthesized using a solid-phase synthesis method through a twelve-channel semi-automatic peptide synthesizer. The purity of the synthesized peptides (>90%) was determined by reverse-phase high-performance liquid chromatography C18 column, and the molecular weights of the derived peptides N6 and N6 were confirmed...

Embodiment 2

[0025] Example 2 Bacteriostatic experiment of sea earthworm antimicrobial peptide NZ17074 derived peptide N6

[0026] The pathogenic bacteria in this embodiment come from China Veterinary Microbial Culture Collection Management Center (CVCC), China Microbiological Culture Collection Management Committee General Microbiology Center (CGMCC) and China Industrial Microbiology Culture Collection Management Center (CICC), and the specific bacterial strains are as shown in the table 1.

[0027] The minimum inhibitory concentration (MIC, minimum inhibitory concentration) of the derivative peptide was determined with reference to the method established by the Clinical and Laboratory Standards Institute (CLSI, Clinical and Laboratory Standards Institute) (WIEGAND et al., Agar and broth dilution methods to determine the minimalinhibitory concentration (MIC )of antimicrobial substances.Nature protocols,2008,3(2):163-175), slightly modified according to the specific situation, the operatio...

Embodiment 3

[0035] Example 3 Cytotoxicity study of sea earthworm antimicrobial peptide NZ17074-derived peptide N6

[0036] 1. Determination of hemolysis

[0037] Hemolyticity is an important indicator to measure whether antimicrobial peptides are suitable for intravenous injection therapy. The specific operations are as follows:

[0038] 6-week-old ICR female mice with SPF grade were taken, blood was collected from eyeballs, and collected using heparin sodium anticoagulant tubes. The collected blood was centrifuged at 1500 rpm for 10 min at 4° C., and the erythrocytes were repeatedly washed three times with 10 mM PBS (pH 7.3) until the supernatant was colorless and transparent, and an 8% erythrocyte suspension was prepared.

[0039] The sample peptide was dissolved in sterile 0.9% physiological saline, prepared into a stock solution with a concentration of 512 μg / mL, and diluted 2-fold to a final concentration of 1 μg / mL. 100 μL of erythrocyte suspension and antimicrobial peptide soluti...

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Abstract

The invention provides an arenicola cristata antibacterial peptide NZ17074 derived peptide N6 and application thereof. An antibacterial peptide NZ17074 molecular structure is optimally designed to obtain a novel derived peptide N6, and the amino acid sequence of the novel derived peptide N6 is shown as SEQ ID NO: 1. The antibacterial peptide N6 provided by the invention has a very good sterilization effect on Gram-negative bacteria, and is used for remarkably reducing the haemolytic activity on mouse erythrocytes and the cytotoxicity on mouse original macrophages. A mouse toxemia model test result shows that the antibacterial peptide N6 also has an endoxemia resisting effect, and can be used for remarkably improving the survival rate of mice, and the effect of the antibacterial peptide N6 is better than that of an antibiotic polymyxin. The antibacterial peptide N6 has small molecular weight and is easily artificially synthesized; the antibacterial peptide N6 is a low-molecular-weight polypeptide with extremely good application value, can be used for preparing novel antibacterial and anti-infection medicines and the like, and has a good application prospect.

Description

technical field [0001] The present invention relates to antibacterial peptides and applications thereof, in particular to sea earthworm antibacterial peptide NZ17074-derived peptide N6 and applications thereof. Background technique [0002] Antimicrobial peptides are biologically active small molecule polypeptides formed by protease degradation of precursors produced by biological organisms, and mainly constitute the first line of defense of the body (ZASLOFF, Antimicrobial peptides of multicellular organisms. Nature, 2002, 415: 389- 395). The mechanism of action of antibacterial peptides on bacteria is mainly based on the membrane as the target site, through electrostatic force and receptor-mediated cell membrane force, causing changes in membrane permeability, reducing the stability of the cell membrane, or inserting into the cell membrane to form transmembrane holes , eventually leading to the leakage of bacterial contents and death. Due to the special mechanism of acti...

Claims

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Application Information

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IPC IPC(8): C07K14/435C12N15/12A61K38/17A61P31/04
CPCA61K38/00C07K14/43504
Inventor 王建华杨娜王秀敏滕达毛若雨郝娅
Owner FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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