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Helicobacter pylori urease B antigen epitope polypeptide and application thereof

A technology of Helicobacter pylori and antigenic epitopes, applied in the direction of application, virus antigen components, peptides, etc., can solve the problems of not being able to inhibit the activity of urease well, and cannot completely prevent the colonization of Helicobacter pylori, so as to prevent and control infection , enhanced inhibitory and clearance effects, good immunogenicity effects

Active Publication Date: 2010-10-20
INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have found that monoclonal antibodies against different epitopes of UreB can inhibit the activity of urease, thereby blocking the infection of Helicobacter pylori, but immunizing animals with intact urease can not inhibit the activity of urease well, so it cannot completely prevent the infection of Helicobacter pylori. Helicobacter colonization in the stomach

Method used

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  • Helicobacter pylori urease B antigen epitope polypeptide and application thereof
  • Helicobacter pylori urease B antigen epitope polypeptide and application thereof
  • Helicobacter pylori urease B antigen epitope polypeptide and application thereof

Examples

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Effect test

Embodiment 1

[0049] Example 1. Identification of Antigen Recognition Epitopes of Anti-Helicobacter Pylori Urease B Subunit Monoclonal Antibodies A1H10, B3D9, and A3C10

[0050] Antigen epitopes were screened by expressing urease B fragments by multiple truncations, and the positional relationship of each truncated fragment in the Helicobacter pylori urease B subunit is as follows figure 1 shown.

[0051] 1. The first segmented expression of UreB truncated protein and the preliminary identification of antigenic epitope

[0052] 1. Recombinant expression of truncated Helicobacter pylori UreB segmented antigens UP1 (sequence 5), UP2 (sequence 6), UP3 (sequence 7), UP4 (sequence 8):

[0053] 1) Culture of Helicobacter pylori

[0054] Helicobacter pylori (Helicobacter pylori) 26695 (ATCC No.700392) adopts solid culture, culture medium is: Campylobacter jejuni agar base (China Diarrheal Disease Control Shanghai Reagent Supply Research Center) 43g / L, defibrinated sheep blood (purchased by the m...

Embodiment 2

[0151] Example 2. Synthesis of epitope polypeptides to further confirm the epitopes targeted by monoclonal antibodies A1H10, B3D9, and A3C10

[0152] 1. Synthesis of epitope peptides

[0153] For the monoclonal antibody recognition epitope determined in Example 1, epitope polypeptides were synthesized by chemical synthesis (sent to Beijing Zhongke Yaguang Biological Co., Ltd. for synthesis), and named respectively as UP32 (the amino acid sequence is sequence 31 in the sequence listing), The purity of UP35 (the amino acid sequence is sequence 34 in the sequence listing) and UP38 (the amino acid sequence is sequence 37 in the sequence listing) is above 90%, and the synthesis amount is 4 mg.

[0154] 2. Determine the epitopes targeted by monoclonal antibodies A1H10, B3D9, and A3C10 by ELISA

[0155] The three synthetic epitope polypeptides UP32, UP35, and UP38 (10 μg / ml) were respectively coated on ELISA plates (100 ul / well) to detect the epitopes targeted by the A1H10, B3D9, an...

Embodiment 3

[0157] Embodiment 3, the recognition experiment of fusion epitope peptide and synthetic epitope peptide and patient's serum

[0158] In order to detect whether specific antibodies against the antigenic epitopes identified in the present invention are produced in the serum of clinically infected H. pylori patients, we prepared fusion epitope peptides (3 μg / ml) GST-UP32, GST-UP35, Epitope peptide (10 μ g / ml) UP32, UP35, UP38 synthesized by GST-UP38 and embodiment 2 are respectively coated with ELISA plates (100ul / hole), and are collected with this laboratory and use S001 Helicobacter pylori urease detection kit (colloid Gold method, Beijing Kangmei Tianhong Biotechnology Co., Ltd.) identified as UreB-positive 3 parts of patient serum and 2 parts of normal human serum as the primary antibody (1:100) for ELISA analysis, the specific operation steps are the same as step 4 of Example 1.

[0159] The result of the reaction is as Figure 7 As shown, the fusion epitope protein and the...

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Abstract

The invention discloses a helicobacter pylori urease B antigen epitope polypeptide and application thereof. The amino acid sequence of the antigen epitope polypeptide is the sequence 31 in a sequence table, and the coding gene sequence of the antigen epitope polypeptide is the sequence 25 in the sequence table. The antigen epitope polypeptide provided by the invention can stimulate an organism togenerate helicobacter pylori resistance protective immunity reaction, thereby enhancing the effect of inhibiting helicobacter pylori infection, improving germ eliminating capacity of the organism, and playing an improving role in corresponding researches on development of vaccine and pathogenic mechanism and clinical diagnosis.

Description

technical field [0001] The invention relates to a Helicobacter pylori (Helicobacter pylori) urease B subunit (UreB) protein B cell antigen epitope polypeptide and application thereof. Background technique [0002] In 1982, Australian scholars Warren and Marshall first isolated and cultured Helicobacter pylori (H. pylori) from the gastric mucosa of patients with chronic active gastritis. The discovery of this bacteria opened a new era of gastric microbe research. Helicobacter pylori is widely distributed, and the infection rate of the population worldwide is as high as about 50%. It has been confirmed that Helicobacter pylori infection is the main cause of chronic gastritis, gastric ulcer and duodenal ulcer, and the persistent infection of Helicobacter pylori is also closely related to the occurrence of gastric adenocarcinoma and gastric lymphoma. relevant. In 1994, the World Health Organization classified it as a Type I carcinogen. [0003] The current treatment for Helic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08C12N15/31A61K38/10A61K39/12A61K39/395A61K48/00A61P31/04C12Q1/68C12Q1/04G01N33/569G01N33/577
Inventor 刘纯杰王艳春邱炎陶好霞
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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