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Antibacterial peptides with high antibacterial activity and application thereof

A technology of antibacterial activity and antimicrobial peptides, applied in the field of biomedical engineering, can solve problems such as public health, food safety and food safety threats, and achieve the effect of significant antibacterial effect, mature preparation process, and convenient access channels

Active Publication Date: 2022-01-11
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

If antibiotics continue to be abused as antibacterial drugs, it will pose a great threat to global public health, food safety, food security, etc.

Method used

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  • Antibacterial peptides with high antibacterial activity and application thereof
  • Antibacterial peptides with high antibacterial activity and application thereof
  • Antibacterial peptides with high antibacterial activity and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Determination of growth curve of experimental strains

[0055] The growth curve of experimental strains was measured by turbidimetric method, and the specific steps were as follows:

[0056] (1) Strain activation: Take 20 L of bacterial solution stored in a glycerol tube and add it to 100 mL of MHB medium for 18 hours at 37°C.

[0057] (2) Numbering: Take 11 large test tubes filled with meat extract peptone liquid medium, and mark the culture time with a marker pen, that is, 0, 1.5, 3, 4, 6, 8, 10, 12, 14, 16 hours.

[0058] (3) Inoculation: Accurately draw 200L of E. coli culture solution with a pipette gun, and inoculate them into 11 numbered large test tubes of meat extract peptone liquid medium, shake after inoculation, and mix the cells evenly.

[0059] (4) Cultivation: the inoculated 11 test tubes were placed on a shaker, and cultured with shaking at 37°C. Take out the test tubes numbered corresponding to the time at 0, 1.5, 3, 4, 6, 8, 10, 12, 14, and 16 hours ...

Embodiment 2

[0063] 1. Antimicrobial peptide inhibition zone experiment

[0064] The disc agar diffusion method was used to measure the diameter of the antibacterial peptide inhibition zone, and the synthetic Cecropin-B antimicrobial peptide and ampicillin were used as positive controls. The method refers to the American Clinical and Laboratory Standards Institute (CLSI). The specific experimental steps are as follows:

[0065] (1) Make antimicrobial peptide drug-sensitive paper sheets with a drug content of 50 g.

[0066] (2) Separation and purification of test strains;

[0067] (3) Preparation of inoculum solution: Pick 3 colonies with a diameter of about 1 mm from the purified test bacteria plate and inoculate them in 3 mL of MH broth medium, and incubate at 35°C for 5-6 hours to make the test bacteria reach the target level. number of growing seasons. Subsequently, the concentration of the bacterial liquid in the logarithmic growth phase was corrected to 0.5 McFarland standard with p...

Embodiment 3

[0085] PI staining test

[0086] The fluorescent dye PI (propidium iodide) is a nuclear staining reagent that can stain DNA and is commonly used in the detection of cell apoptosis. It is an ethidium bromide analog that emits red fluorescence upon intercalation into double-stranded DNA. PI cannot pass through the living cell membrane, but it can pass through the damaged cell membrane and stain the nucleus. Based on this principle, we used Cecropin-B (H.cecropia) as a positive control, and used PI staining to observe the damage of E.coli K-12 cell membrane after the action of antimicrobial peptides, so as to explore its possible inhibitory effect. bacteria mechanism. The specific operation method is as follows:

[0087] (1) Strain purification.

[0088](2) Pick 3 colonies with a diameter of about 1 mm from the purified test bacteria plate and inoculate them in 3 mL of MH broth medium, and incubate at 35°C for 5-6 hours to allow the test bacteria to reach the logarithmic grow...

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Abstract

The invention discloses four antibacterial peptides with high antibacterial activity and application thereof. The amino acid sequence of the polypeptide is selected from one of the amino acid sequences shown as SEQ ID NO.1-SEQ ID NO.4. The four antibacterial peptides have bacteriostatic activity, AMP1, AMP2 and AMP3 show strong bacteriostatic activity, and preliminary toxicity analysis shows that the four antibacterial peptides have no hemolytic activity under the action concentration of 50g / mL, and have no significant influence on proliferation and cellular morphology of 293T cells.

Description

technical field [0001] The invention belongs to the technical field of biomedical engineering, and specifically relates to four antibacterial peptides with high antibacterial activity and applications thereof. Background technique [0002] Penicillin was first discovered in 1928, and together with sulfonamides as a therapeutic drug, the library of drugs used to treat microbial infections has gradually been enriched. The period from 1940 to 1960 was the golden age of antimicrobial drug development, and the vast majority of antibiotics in use today were discovered during this period. Antibiotics have been by far the most commonly prescribed drugs and have contributed greatly to reducing mortality and morbidity caused by microbial infections. However, due to the irregular use of antibiotics, bacteria gradually develop resistance to it, many bacteria such as Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanii, Pseudomonas aeruginosa Pseudomonas aeruginosa and ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/00A61K38/16A61P31/04A61K8/64A61Q17/00A61L15/32A61L15/46A23K20/147
CPCC07K14/001A61P31/04A61K8/64A61Q17/005A61L15/32A61L15/46A23K20/147A61K38/00A61L2300/404A61L2300/252A61K49/14A01N47/44A01P1/00Y02A50/30
Inventor 顾月清韩智豪尚乾许昊然耿轩
Owner CHINA PHARM UNIV
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