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Antibacterial and antiviral hybrid peptide as well as preparation method and application thereof

An antiviral and hybrid technology, applied in the field of genetic engineering and biopharmaceuticals, can solve the problems of less research on active peptides, achieve the effects of less drug resistance, improved antibacterial effect, and great application value

Inactive Publication Date: 2016-09-28
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Relieving the increasingly serious threat of drug-resistant bacteria to human health has become an urgent problem to be solved. Therefore, the discovery and development of a new class of antimicrobial substances has become a research hotspot in the world, but there are few active peptides that are both antibacterial and antiviral.

Method used

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  • Antibacterial and antiviral hybrid peptide as well as preparation method and application thereof
  • Antibacterial and antiviral hybrid peptide as well as preparation method and application thereof
  • Antibacterial and antiviral hybrid peptide as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1 Preparation of Antibacterial and Antiviral Hybrid Polypeptide C-L

[0029] Inoculate a single colony of Pichia pastoris GS115 in 3-5ml YPD (2% tryptone, 2% glucose, 1% yeast powder, pH 7.0) liquid medium, shake and culture at 30°C for about 12 hours, press 1:100- Inoculate in 100ml YPD liquid medium at a volume ratio of 1:50, culture with shaking at 30°C until OD 600 =1.3-1.5, stop culturing. The culture solution was transferred into a centrifuge tube, placed on ice for 10 minutes, and then centrifuged at 1500g for 5 minutes at 4°C to collect the bacteria. Add pre-cooled sterile water to gently suspend the cells, place on ice for 15-30 minutes, and then centrifuge at 1500g at 4°C for 5 minutes. Discard the supernatant, add 10ml of pre-cooled 1M sterile sorbitol solution, and resuspend the bacteria to make a competent cell suspension. Aliquot 200 μl per tube into spare Pichia competent cells, use immediately or store at -70°C.

[0030]According to the po...

Embodiment 2

[0036] The thermostability test of embodiment 2 polypeptide C-L

[0037] Take 200 μl of the recombinant protein purified in Example 1 and treat it for 10, 20, and 30 min respectively under the conditions of 40°C, 60°C, 80°C, and 100°C in a water bath, use untreated recombinant protein as a control, and standard Staphylococcus aureus As an indicator bacterium, the antibacterial effect of the samples before and after treatment was measured by the method of inhibition zone diameter. The experiment was repeated 3 times, and the average value was taken as the measurement result (Table 1).

[0038] The antibacterial zone diameter of recombinant target protein C-L to Staphylococcus aureus after table 1 heat treatment

[0039]

[0040] The results showed that the effects of different heating temperatures and times on the antibacterial effect of the recombinant target protein were different. When the heating temperature did not exceed 60°C, the diameter of the inhibition zone basic...

Embodiment 3

[0041] Example 3 The inhibitory effect of polypeptide C-L on common bacteria

[0042] Put the filter paper (diameter 6mm) that has been sterilized by dry heat (160°C, 2h) into the recombinant target protein solution (50μg / ml) and fully soak it for 24h. Apply it on the surface of the culture medium to make a bacteria-containing plate, take out the fully soaked filter paper and stick it on the bacteria-containing plate, stick one piece for each petri dish, place it in a constant temperature incubator at 37°C for 24 hours, observe and measure The diameter of the zone of inhibition (Table 2).

[0043] Table 2 The diameter of the inhibition zone of the hybrid peptide C-L to each common bacterium

[0044]

[0045]

[0046] Note: CVCC refers to the China Veterinary Microbiological Culture Collection Management Center; this laboratory refers to the Feed Biotechnology Laboratory of China Agricultural University.

[0047] It can be seen from Table 2 that the antibacterial hybrid...

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Abstract

The invention relates to an antibacterial and antiviral hybrid peptide as well as a preparation method and application thereof. An amino acid sequence and a nucleotide sequence of the antibacterial and antiviral hybrid peptide are as shown in SEQ ID No. 1 and 2. A gene for coding the antibacterial and antiviral hybrid peptide is cloned to an expression vector pPICZ[alpha]A to obtain a recombinant vector, then Pichia pastoris is converted, methyl alcohol is used for inducing expression, and an expression product is purified to obtain the target antibacterial antiviral hybrid peptide. The antibacterial and antiviral hybrid peptide can prevent and kill various bacteria and viruses, has low probability of resistance, and therefore, can serve as an ideal antibiotic substituent, a people and livestock biological antibacterial and antiviral medicine, a feed additive, a preservative, a sanitizer or the like.

Description

technical field [0001] The invention relates to the fields of genetic engineering and biopharmaceuticals, in particular to a novel antibacterial and antiviral hybrid polypeptide and its preparation method and application. Background technique [0002] Antibiotics play an important role in the control, prevention and treatment of various infectious diseases. Due to the use of antibiotics, by the 1980s, humans could almost conquer most infectious diseases. However, with the passage of time and the abuse of antibiotics, there are more and more drug-resistant strains. Surveillance has found that among the most representative drug-resistant strains, drug-resistant staphylococcus has risen to 40%; drug-resistant coagulase-negative staphylococcus exceeds 77%. Now, drug-resistant bacteria that can resist all antibiotics have appeared, and the problem of antibiotic resistance has seriously threatened the safety and health of humans and animals. Relieving the increasingly serious th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/81A61K38/17A61P31/04A61P31/12A61P31/16A01N47/44A01P1/00A01P3/00A23K20/195
CPCC07K14/4723A01N47/44A61K38/00C07K14/43563C07K2319/00C12N15/815C12N2800/102
Inventor 张日俊卫旭彪黄燕张璐璐武如娟郑召君斯大勇廖秀冬
Owner CHINA AGRI UNIV
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