Antivirus associated protein and application thereof

A protein, purpose technology, applied in the fields of biotechnology and immunology, which can solve problems such as lack of antiviral drugs

Inactive Publication Date: 2012-09-12
SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the past two years, the large-scale spread of AIDS, the worldwide spread of SARS-CoV, and the emergence of a new type of influenza virus (Avian-Flu) have sounded the alarm to people: there is still a lack of effective antiviral drugs

Method used

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  • Antivirus associated protein and application thereof
  • Antivirus associated protein and application thereof
  • Antivirus associated protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0148] Example 1. Discovering TRIM21 interacts with IRF3 by co-immunoprecipitation (co-IP)

[0149] It was previously known that there are a variety of protein molecules that have direct interactions with IRF3. The present invention focuses on protein molecules that directly interact with IRF3 after the IRF3 signaling pathway is activated by the virus, because such molecules may play an important role in the anti-virus process of host cells. In the present invention, the method of exogenously transfecting the pcDNA3.1 vector (purchased from Invitrogen) carrying the TBK1 (Tank Binding Kinase) gene (GenBank accession number NM_013254.2) was used to simulate the activation of the IRF3 signaling pathway of 293T cells by the virus. After the cells were collected, they were lysed, and the supernatant of the lysed cells was obtained by centrifugation, and then a co-IP test was performed using a monoclonal antibody to IRF3 (purchased from Santa Cruz). After identification by silver s...

Embodiment 2

[0150] Example 2. Cloning of the cDNA sequence of TRIM21

[0151] TRIM21 is a protein of about 52kDa, which is classified in the TRIM (Tripartitemotif-containing) protein family. This family of proteins has some common features: three zinc finger structures (a Ring-finger and two B-Box structures) at the N-terminus, a coiled-coil region, and a SPRY domain at the C-terminus. This protein is a component of the nuclear protein complex (RoSSA ribonucleoprotein), and it has been reported in the literature that it has the ability to bind DNA and RNA. The gene encoding human TRIM21 protein is located on chromosome 11 of the human genome and consists of 7 discontinuous exons. After the RNA of 293T cells was extracted using the RNA extraction kit, cDNA was obtained by reverse transcription using the universal primer Oligo dT, and then amplified using the two-terminal sequence primers. The 5' end has an EcoRI restriction endonuclease site, and the 3' end has an XhoI restriction endonu...

Embodiment 3

[0152] Example 3. Prokaryotic expression of TRIM21 and antibody preparation

[0153] After obtaining the DNA coding sequence of TRIM21, it was digested with EcoRI and XhoI, and then transformed into the corresponding sites of Escherichia coli expression vectors pET-28a and pGEX-6p-1. Both vectors are prokaryotic expression vectors with Lac operon, which are induced by IPTG in E. coli. Among them, the pET-28a vector has a histidine tag (His 6 -tag) sequence, the corresponding TRIM21 expression product is connected with His 6 -tag fusion protein can be affinity-purified using metal anhydride resins. The PGEX-6p-1 vector encodes a glutathione transferase (GST), so the corresponding TRIM21 expression product is a fusion protein linked with GST, which can be affinity purified by glutathione (GSH) resin. At the same time, there is a protease cutting point on the GST-TRIM21 fusion protein. Through proteolysis on the column, TRIM21 can be separated from the GST protein to avoid aff...

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PUM

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Abstract

The invention relates to antivirus associated protein and application thereof. The invention discloses TRIM21 protein and application of an accelerating agent or an inhibiting agent thereof to the preparation of compositions for regulating congenital immunity reaction signal transduction passages. The invention also discloses a method for screening potential substances for regulating the congenital immunity reaction signal transduction passages by using the TRIM21 protein as a target spot. The invention discloses the novel target spot for regulating the congenital immunity reaction signal transduction passages for the first time, and provides a novel path for the development of the antivirus medicine.

Description

technical field [0001] The invention belongs to the fields of biotechnology and immunology, and more specifically, the invention relates to an antiviral related protein and its application. Background technique [0002] Pathogen infection has always been a major disease that threatens human health. The World Health Organization (WHO) pointed out in a report that six major infectious diseases that threaten human beings are: acute respiratory disease, diarrhea and dysentery, acquired immune deficiency syndrome ( AIDS), tuberculosis, malaria and measles. A large part of it is caused by viral infection. In the past two years, the large-scale spread of AIDS, the worldwide spread of SARS virus (SARS-CoV), and the emergence of a new type of influenza virus (Avian-Flu) have all sounded the alarm to people: there is still a lack of effective antiviral drugs. [0003] Human research on viruses can be traced back thousands of years. As early as the Song Dynasty, ancient Chinese peopl...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/395A61P31/12C07K14/435C12N15/13C12Q1/68
Inventor 王琛阳凯石贺欣刘心义单玉飞
Owner SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
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