Recombinant ubiquitin ligase PTB-U-box fusion gene and expression vector and application thereof

A ubiquitin ligase, ptb-u-box technology, applied in gene therapy, drug combination, genetic engineering and other directions, to achieve the effect of reducing cell proliferation and invasion ability

Active Publication Date: 2013-06-05
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is no report on the use of CHIP ubiquitin ligase activity to construct recombinant ubiquitin ligases for gene therapy research.

Method used

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  • Recombinant ubiquitin ligase PTB-U-box fusion gene and expression vector and application thereof
  • Recombinant ubiquitin ligase PTB-U-box fusion gene and expression vector and application thereof
  • Recombinant ubiquitin ligase PTB-U-box fusion gene and expression vector and application thereof

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Embodiment Construction

[0035] The present invention utilizes the PTB domain (Phosphotyrosine Binding domain) of linker molecule IRS-1 (Insulin Receptor Substrate 1) to be able to recognize and bind to the characteristics of insulin-like growth factor receptor (Insulin-like Growth Factor Receptor 1, IGF-1R) and CHIP To construct a recombinant ubiquitin ligase PTB-U-box that can target and degrade the oncogenic-related protein IGF-1R. The constructed fusion gene PTB-U-box is to clone the PTB domain gene of IRS-1 and the U-box domain gene of CHIP with ubiquitin ligase activity respectively, and use recombinant PCR to fuse; wherein, the PTB domain is responsible for For the targeted binding of IGF-1R, U-box is responsible for connecting ubiquitin molecules to substrates to promote ubiquitination and degradation of substrates. In the following, the present invention will be further described in detail in conjunction with the construction of the fusion gene PTB-U-box, eukaryotic expression vector and reco...

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Abstract

The invention discloses a recombinant ubiquitin ligase PTB-U-box fusion gene and an expression vector and application thereof. The recombinant ubiquitin ligase PTB-U-box fusion gene is formed by connecting a PTB domain gene of IRS-1 and a U-box domain gene of carboxy terminus of Hsc70 Interacting protein (CHIP), and the specific nucleotide sequence is shown as SEQ ID No. 1. The constructed PTB-U-box fusion gene can be cloned to different expression vectors so as to enter tumor cells through different paths to play a role, and the expression vectors comprise eukaryotic expression vectors or adenovirus expression vectors. After the fusion gene transfects the tumor cells, the carcinogenic related protein IGF-1R of the host tumor cells can be regulated down, the cell proliferation and intrusion capacities of the tumor cells are remarkably reduced, and the fusion gene shows proliferation inhibition and intrusion inhibition on the tumor cells so as to achieve the anti-tumor effect.

Description

technical field [0001] The invention belongs to the field of biotechnology, relates to gene recombination and its expression, in particular to a recombinant ubiquitin ligase PTB-U-box fusion gene and its expression vector and application. Background technique [0002] 1. Overview of CHIP [0003] CHIP (Carboxy terminus of Hsc70 Interacting Protein) is a kind of evolutionarily conserved and widely distributed cytoplasmic protein, which was originally considered as a co-chaperone. Since its molecule contains a U-box domain, it essentially belongs to a class of E3 ubiquitin ligases. CHIP plays an important role in protein quality control and stability regulation by promoting protein ubiquitination and degradation. In breast cancer tissue, the expression of CHIP is negatively correlated with tumor grade; experimental evidence also shows that CHIP can inhibit tumor proliferation and metastasis. These studies suggest that enhancing the role of CHIP may be used as a new tumor tr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/62C12N15/79C12N15/861A61K48/00A61P35/00
Inventor 药立波李霞王秦豪茹懿申亮亮张璟
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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