Antimicrobial peptide lf-th and its Escherichia coli recombinant preparation method

A technology of LF-TH and Escherichia coli, which is applied in biochemical equipment and methods, recombinant DNA technology, chemical instruments and methods, etc., can solve the problem of low antibacterial activity of antimicrobial peptides, achieve broad antibacterial spectrum, broad application potential, antibacterial The effect of activity improvement

Inactive Publication Date: 2011-12-14
NORTHEAST AGRICULTURAL UNIVERSITY
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Problems solved by technology

[0005] The purpose of the present invention is to solve the problem of low antibacterial activity of existing antimicrobial peptides, and provide antimicrobial peptide LF-TH and its Escherichia coli recombinant preparation method

Method used

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  • Antimicrobial peptide lf-th and its Escherichia coli recombinant preparation method
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  • Antimicrobial peptide lf-th and its Escherichia coli recombinant preparation method

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specific Embodiment approach 1

[0015] 具体实施方式一:本实施方式抗菌肽LF-TH的氨基酸序列包括33个氨基酸,其氨基酸序列如下:PheLysCysArgArgTrpGlnTrpArgTrpLysLysLeuGlyAlaLysProValProIleIleTyrCysAsnArgArgThrGlyLysCysGlnArgMet,其编码基因序列如下:TTCAAATGCCGTCGTTGGCAGTGGCGTTGGAAAAAACTGGGTGCGAAACCGGTTCCGATCATCTACTGCAACCGTCGTACCGGTAAATGCCAGCGTATG。

specific Embodiment approach 2

[0016] Specific embodiment two: The Escherichia coli recombinant preparation method of the antimicrobial peptide LF-TH in this embodiment is carried out according to the following steps:

[0017] 1. Select the 1st to 15th amino acid sequence Lfcin B (1 to 15) of LfcinB, wherein the tenth amino acid is replaced by Met with Trp, and select the 4th to 21st amino acid sequence Th (4 to 21) of Thanatin. Lfcin B (1-15) is connected with Th (4-21) to design the antimicrobial peptide LF-TH, and then design the coding gene of the antimicrobial peptide LF-TH according to the codon preference of Escherichia coli;

[0018] 2. Link the gene encoding the antimicrobial peptide LF-TH to the plasmid pET-32a(+), construct a recombinant Escherichia coli expression vector, transform the host cell E.coli BL21(DE3), induce expression with IPTG, collect the cells by centrifugation, and ultrasonicate Cleavage and centrifuge to collect the supernatant, use Ni-NTA chromatography column and protein puri...

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Abstract

Antimicrobial peptide LF-TH and its Escherichia coli recombinant preparation method. It solves the problem of low antibacterial activity of the existing antibacterial peptides. The amino acid sequence of the antimicrobial peptide LF-TH includes 33 amino acids, its amino acid sequence is shown in SEQ ID No.1, and its coding gene sequence is shown in SEQ ID No.2. Methods: 1. Design the antimicrobial peptide LF-TH and the coding gene of the antimicrobial peptide LF-TH; 2. Link the coding gene of the antimicrobial peptide LF-TH to the plasmid and construct a recombinant Escherichia coli expression vector, transform E.coliBL21(DE3), and The fusion protein is obtained after induced expression and purification, the fusion protein is cleaved by enterokinase to release the antimicrobial peptide LF-TH, and the antimicrobial peptide LF-TH is obtained after purification. The antibacterial peptide LF-TH of the present invention has high antibacterial activity, and has obvious inhibitory effect on Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Salmonella typhimurium.

Description

technical field [0001] The present invention relates to antibacterial peptide and its preparation method. Background technique [0002] A series of problems brought about by the long-term and extensive use of antibiotics, such as flora imbalance, the increase of drug-resistant strains, and the side effects of drugs, are becoming more and more serious. Especially, drug resistance of bacterial strains has become a problem that antibiotics cannot solve. Research and development of new Effective antimicrobial drugs have become a top priority. Antimicrobial peptides are a class of polypeptides with important immune functions in organisms, and are an important part of the body's non-specific immune function. Antimicrobial peptides have the characteristics of small molecular weight, thermal stability, good water solubility, and broad antibacterial spectrum. Antimicrobial peptides have a unique mechanism of action different from antibiotics, and strains are not easy to develop res...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/70C12R1/19
Inventor 冯兴军盛尊来马德星郭佳音李静
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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