Alkaline antibacterial peptide as well as targeting design and application thereof

An antibacterial peptide, alkaline technology, applied in the field of alkaline antibacterial peptide and its targeted design and application, can solve the problems of hemolytic toxicity, lack of targeted antibacterial and anticancer properties, antibiotic resistance, etc., and achieve strong anticancer active effect

Active Publication Date: 2015-05-06
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, what follows is a serious problem caused by the abuse of antibiotics: antibiotic resistance (Drug resistance)
Antimicrobial peptides are gen...

Method used

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  • Alkaline antibacterial peptide as well as targeting design and application thereof
  • Alkaline antibacterial peptide as well as targeting design and application thereof
  • Alkaline antibacterial peptide as well as targeting design and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment 1 Minimum Inhibitory Concentration (MIC) Determination

[0045] According to the polypeptide sequence in Table 1, various polypeptides were synthesized by fmoc solid-phase synthesis by Shanghai Botai Biotechnology Co., Ltd., and purified by RP-HPLC; the polypeptides include antimicrobial peptides and targeted antimicrobial peptides, wherein the targeted antimicrobial peptides are It is obtained by linking the antimicrobial peptide and the complementarity determining region of the antibody through several leucines.

[0046] The bacteria used were Escherichia coli (MG1655), Pseudomonas aeruginosa (1.2464, Beijing China General Microorganism Culture Collection Management Center), Staphylococcus aureus (ATCC6538), multi-drug resistant Staphylococcus aureus Y5 (Zhang Ying et al., together with food Molecular typing of Staphylococcus aureus in poisoning events, Chinese Journal of Preventive Medicine, 2008, 42(9):672-676; Ran He et al. A combinatorial yeast overlay ...

Embodiment 2

[0074] Embodiment 2 hemolytic experiment

[0075] 1. Blood Processing

[0076] (1) Collect 5 ml of fresh human blood and put it into a tube containing 0.5 ml of 3.8% sodium citrate, blow and beat the blood in the anticoagulant tube to make it fully mix with the anticoagulant.

[0077] (2) The above blood was centrifuged at 2000 rpm for 8 minutes, and the supernatant was removed. Rinse the blood with 10 mM PBS, pH=7.4, centrifuge at 2000 rpm for 5 minutes, discard the supernatant, and repeat the operation until the erythrocyte suspension is clear and free of impurities such as serum, then discard the supernatant. Then it was dissolved in PBS at 5% (v / v) to obtain erythrocyte suspension.

[0078] 2. Antimicrobial peptide treatment: the antimicrobial peptide was dissolved in PBS to obtain antimicrobial peptide solutions with final concentrations of 40 μM, 80 μM, 160 μM, and 320 μM.

[0079] 3. System mixing

[0080] Draw 50 microliters of the treated antimicrobial peptide sol...

Embodiment 3

[0086] Example 3 Escherichia coli membrane permeability experiment

[0087]β-galactosidase is a hydrolytic enzyme located in the cytoplasm of bacteria that can hydrolyze o-nitrophenyl β-D-galactopyranoside (ONPG) into galactose and o-nitrophenol (yellow). Add a certain amount of ONPG to the system, and measure the change of the absorbance value of the culture medium at 420 nanometers to determine the degree of hydrolysis of ONPG, so as to determine whether β-galactosidase hydrolyzes ONPG. Generally, ONPG cannot enter the cell because the enzyme is located inside the cell. But once the permeability of the cell membrane changes, ONPG enters the cell and is hydrolyzed, and the culture medium quickly turns yellow, A 420 The value rises rapidly in a short period of time. Therefore, this method can be used to detect the effect of antimicrobial peptides on cell membrane permeability. High membrane permeability indicates that antimicrobial peptides can form membrane channels.

[0...

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Abstract

The invention belongs to the field of biotechnology, and discloses an alkaline antibacterial peptide as well as targeting design and application thereof. The alkaline antibacterial peptide is composed of at least one alkaline amino acid and one or two hydrophobic amino acids; the alkaline antibacterial peptide includes 12-24 amino acids, wherein the proportion of the alkaline antibacterial peptide is more than 33.3%, and the alkaline antibacterial peptide has a broad-spectrum bacteriostatic action on escherichia coli, pseudomonas aeruginosa, staphylococcus aureus, multi-drug resistance staphylococcus aureus, saccharomyces cerevisiae and the like; the alkaline antibacterial peptide is prepared as a targeted antibacterial peptide, and the efficiency of the antibacterial peptide in killing lung cancer cells at 0.1mM reaches 100.3%; the antibacterial peptide or the targeted antibacterial peptide disclosed by the invention is applicable to bacterium inhabitation and cancer resistance, and can be widely applied to the fields of medicine, agriculture, food preservative and the like.

Description

technical field [0001] The present invention relates to the field of biotechnology, more specifically, to basic antimicrobial peptides and their targeted design and application. Background technique [0002] Antibiotics are a class of natural or synthetic compounds that can kill bacteria or inhibit the growth of bacteria. With the continuous development of science and technology, the definition of antibiotics has also been continuously expanded, and antimicrobial compounds, including antifungal compounds, have been included in the scope of antibiotics. [0003] Since the discovery of the first antibiotic penicillin in 1940 and its clinical application, a new era of antibiotic treatment has begun. Many infectious diseases that once seriously endangered human life and health have been effectively controlled due to the use of antibiotics, and the mortality rate of infants and the infection rate after surgery have been greatly reduced, and the average life expectancy of human b...

Claims

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Application Information

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IPC IPC(8): C07K7/08C07K14/00C07K19/00A61K38/10A61K38/16A61K47/48A61P31/04A61P31/10A61P35/00
CPCA61K38/10A61K38/16A61K47/50C07K7/08C07K14/00C07K19/00Y02A50/30
Inventor 刘秋云何建国周文良翁少萍邢梦付敏彭静立兰崇峰
Owner SUN YAT SEN UNIV
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