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Intramolecular adjuvant-containing recombinant gene for preventing and treating helicobacter pylori, protein and biological product

A technology of Helicobacter pylori and amino acids, applied in the direction of medical preparations containing active ingredients, gene therapy, bacterial antigen components, etc., can solve the problems of restriction and toxicity application, and achieve the effect of preventing damage and protecting biological activity

Inactive Publication Date: 2014-11-19
SICHUAN VACCINE TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Vibrio cholerae enterotoxin (CT) has been shown to be a potent adjuvant for mucosal immunity, but its application is limited due to its toxicity

Method used

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  • Intramolecular adjuvant-containing recombinant gene for preventing and treating helicobacter pylori, protein and biological product
  • Intramolecular adjuvant-containing recombinant gene for preventing and treating helicobacter pylori, protein and biological product
  • Intramolecular adjuvant-containing recombinant gene for preventing and treating helicobacter pylori, protein and biological product

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Example 1: Design and construction of multi-target fusion polypeptide

[0065] Such as figure 1 Shown is a schematic diagram of the construction of a multi-target fusion polypeptide BIB vaccine: BIB described in the present invention is to add a section of intramolecular adjuvant CTB that removes the signal peptide to the N segment of the multi-target fusion polypeptide IB, forming such as SEQ ID NO: The amino acid sequence shown in 2. The DNA sequence corresponding to the above amino acid sequence is the nucleotide sequence shown in SEQ ID NO:1. The protein sequence length of this BIB is 289aa, and protein molecular weight is 32.82 kDa, and protein isoelectric point is 9.05 pI, and its amino acid sequence is as shown in SEQ ID NO: 2, and the protein sequence length of CTB is 100aa, and protein molecular weight is 11.32 kDa, and protein The isoelectric point is 7.97 pI.

Embodiment 2

[0066] Embodiment 2: the preparation of rBIB engineering bacteria

[0067] The length of the BIB gene sequence is 876bp, which was synthesized by Beijing Jinweizhi; then the fusion gene BIB was constructed on the pET28a (+) vector, and the restriction endonuclease site was Bam H I and Hind III; after that, transform into Escherichia coli BL21 (DE3), screen positive clones, and construct vaccine engineering bacteria BIB.

Embodiment 3

[0068] Embodiment 3: Preparation of rBIB protein

[0069] 3.1 Fermentation of BIB engineering bacteria

[0070] The improved TB medium was used as the fermentation medium, after high temperature and high pressure sterilization, inoculated with 10% inoculum, cultured at 37°C for 8-10 h, after that, added lactose with a final concentration of 5 mM for 10 h, and controlled the pH at 7.2±0.3, adjust the speed to control the dissolved oxygen above 30%. Finally, the yield of BIB cells was between 30-45 g / L; the protein expression was between 25%-35%.

[0071] 3.2 Preparation of BIB inclusion bodies

[0072] Take 1 kg / time of BIB engineering bacteria collected after fermentation and centrifugation, resuspend in ultrasonic lysis buffer at a ratio of 1:10 (W / V), and stir with a magnetic stirrer to mix evenly. Place the beaker of the uniformly mixed bacterial solution in ice cubes, pre-cool to 4°C, and use a sonicator at 250 W, on for 2 s, off for 2 s, and for 40 min. Take a small a...

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Abstract

The invention discloses an intramolecular adjuvant-containing recombinant gene for preventing and treating helicobacter pylori, protein and a biological product. the multi-target recombinant gene comprises a multi-target fusion polypeptide having an amino acid sequence shown in a SEQ ID NO:2, a multi-target recombinant gene for coding the multi-target fusion polypeptide having the nucleotide sequence shown in a SEQ ID NO:2, and invention also has an application of a specific antibody of the above multi-target recombinant gene or the above multi-target fusion polypeptide or the multi-target fusion polypeptide as the biological product for preventing and treating helicobacter pylori infection. According to the invention, an intramolecular adjuvant CTB for removing signal peptide is added at N segment of a recombinant helicobacter pylori multi-target fusion polypeptide (rIB), so that prevention and treatment effect for the helicobacter pylori infection can be increased.

Description

technical field [0001] The invention relates to the biotechnical field of vaccine and antibody pharmacy, in particular to a recombinant gene, protein and biological product containing an intramolecular adjuvant for preventing and treating Helicobacter pylori. Background technique [0002] Helicobacter pylori (Hp) is an important pathogenic bacterium discovered in 1982. Current studies have shown that this bacterium is closely related to MALT lymphoma and gastric cancer in addition to causing gastritis and gastric ulcer. Bacterial pathogens associated with human tumorigenesis published by WHO. Recent studies have also found that Hp is also highly correlated with cardiovascular diseases such as coronary heart disease. Hp can infect the elderly, children and young adults, but the infection rate varies greatly in different countries and regions due to different economic levels and living habits. The infection rate of the general population in my country is about 50-80%, and it...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/62A61K48/00A61K39/02A61K39/39A61K39/40A61P31/04
Inventor 潘兴祝洁周永君高礼珍
Owner SICHUAN VACCINE TECH
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