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Cell-penetrating peptide and human Beta-defensin 3 fusion protein and preparation method and application thereof

A β-defensin and fusion protein technology, applied in the fields of peptide/protein components, biochemical equipment and methods, chemical instruments and methods, etc., can solve the problem that foreign proteins are not easy to penetrate the skin barrier

Inactive Publication Date: 2017-04-26
SHAANXI HUIKANG BIO TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] A technical problem to be solved by the present invention is to overcome the disadvantage that exogenous proteins are not easy to pass through the skin barrier, and provide a membrane-penetrating peptide-human β-defensin 3 fusion protein that can be transdermally transported

Method used

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  • Cell-penetrating peptide and human Beta-defensin 3 fusion protein and preparation method and application thereof
  • Cell-penetrating peptide and human Beta-defensin 3 fusion protein and preparation method and application thereof
  • Cell-penetrating peptide and human Beta-defensin 3 fusion protein and preparation method and application thereof

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Experimental program
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Effect test

Embodiment 1

[0043] Penetrating peptide-human β-defensin 3 fusion protein, the full length is 59 amino acids, the nitrogen terminal is the penetrating peptide sequence, the carbon terminal is the human β-defensin 3 sequence, and the two peptides are connected by two glycines and one serine , with a molecular weight of 6.7KD; the amino acid sequence of the penetrating peptide-human β-defensin 3 fusion protein is as follows:

[0044] 1 YARVRRRGPR RGGSGIINTL QKYYCRVRGG RCAVLSCLPK EEQIGKCSTR GRKCCRRKK

[0045] Wherein the amino acid sequence of the penetrating peptide is:

[0046] 1 YARVRRRGPR R

[0047] The amino acid sequence of human beta defensin 3 is:

[0048] 1 GIINTLQKYY CRVRGGRCAV LSCLPKEEQI GKCSTRGRKC CRRKK

[0049] The nucleotide sequence of the penetrating peptide-human beta defensin 3 fusion protein is:

[0050]

[0051] The preparation method of the above-mentioned penetrating peptide-human beta defensin 3 fusion protein is as follows:

[0052] 1. Gene acquisition

[0053...

Embodiment 2

[0109] Step 4 of the Pichia pastoris fermentation of the membrane-penetrating peptide-human β-defensin 3 fusion protein in this example is: inoculate the screened transformants in 400 ml of Pichia pastoris-induced expression medium, and shake culture at 30°C for 24 hours, as The first-level seeds were transferred to a 5L fermenter with 4L of basic salt medium, the temperature was set at 30°C, the pH value was 5.0, and they were cultivated for 16-20 hours. As the second-level seeds, they were transferred to 80L of basic salt culture medium. Base (basic salt medium consists of 40g glycerol, 18.2gK 2 SO 4 , 26.7mlH 3 PO 4 , 0.93g CaSO 4 2H 2 O, 14.9g MgSO 4 , 4.13gKOH, dissolved in 1000ml deionized water and mixed;) 150L large tank fermentation. During the fermentation process, the growth temperature is 30°C, the induction temperature is 26°C, the pH value is 4.5, and the dissolved oxygen is controlled at 30% to 40%. PTM1 trace element glycerol solution, the volume ratio o...

Embodiment 3

[0112] Step 4 of the Pichia pastoris fermentation of the membrane-penetrating peptide-human β-defensin 3 fusion protein in this example is: inoculate the screened transformants in 400 ml of Pichia pastoris-induced expression medium, and shake culture at 30°C for 24 hours, as The first-level seeds were transferred to a 5L fermenter with 4L of basic salt medium, the temperature was set at 30°C, the pH value was 5.0, and they were cultivated for 16-20 hours. As the second-level seeds, they were transferred to 80L of basic salt culture medium. Base (basic salt medium consists of 40g glycerol, 18.2gK 2 SO 4 , 26.7mlH 3 PO 4 , 0.93g CaSO 4 2H 2 O, 14.9g MgSO 4, 4.13gKOH, dissolved in 1000ml deionized water and mixed;) 150L large tank fermentation. During the fermentation process, the growth temperature is 30°C, the induction temperature is 28°C, the pH value is 5.5, and the dissolved oxygen is controlled at 30% to 40%. PTM1 trace element glycerol solution, the volume ratio of...

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Abstract

Cell-penetrating peptide and human Beta-defensin 3 fusion protein has 59 amino acids in full length, has a cell-penetrating peptide sequence at its nitrogen end and Beta-defensin 3 mature peptide sequence at its carbon end, and the two peptide segment are connected via double glycine and single serine and have molecular weight of 6.7 KD. The preparation method comprises the steps of acquiring a gene, constructing Pichia pastoris expression vector, constructing cell-penetrating peptide and human Beta-defensin 3 fusion protein Pichia pastoris genetically-engineered bacterium, fermenting cell-penetrating peptide and human Beta-defensin 3 fusion protein with Pichia pastoris, and purifying the cell-penetrating peptide and human Beta-defensin 3 fusion protein. Application of cell-penetrating peptide and human Beta-defensin 3 fusion protein in the preparation of bactericidal gels, application in the preparation of film cosmetics and application in the preparation of toner cosmetics are also provided.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a membrane-penetrating peptide-human beta defensin 3 fusion protein. Background technique [0002] Antibacterial peptides, also known as peptide antibiotics or antimicrobial peptides, are a class of peptide substances produced by the biological defense system to defend against exogenous pathogens. Antimicrobial peptides are widely distributed in plants, animals, insects and human bodies, have strong activity against microbial invasion, and are an important part of the body's immune system. Human defensin belongs to the family of antimicrobial peptides. It is a new class of arginine-rich cationic antibacterial peptides discovered in recent years. It can be divided into two categories according to its source and the connection mode of disulfide bonds in the molecule. : α-defensins, β-defensins, wherein β-defensins mainly exist in epithelial tissue cells such as human mucos...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/62C12N15/81C12N15/66C07K1/36C07K1/34C07K1/18A61K38/17A61K9/06A61P31/04A61K8/64A61Q17/00C12R1/84
Inventor 李敏侯增淼李晓颖赵金礼杨小琳
Owner SHAANXI HUIKANG BIO TECH CO LTD
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