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Method for efficiently expressing recombinant liraglutide

A technology for high-efficiency expression of liraglutide, applied in the field of biomedicine, can solve the problems of high technical difficulty, unfavorable large-scale production of liraglutide, and high production cost

Inactive Publication Date: 2015-07-01
杭州北斗生物技术有限公司
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  • Abstract
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AI Technical Summary

Problems solved by technology

Novo Nordisk's liraglutide is prepared by biological methods such as genetic engineering, which is technically difficult and costly, which is not conducive to the large-scale production of liraglutide

Method used

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  • Method for efficiently expressing recombinant liraglutide
  • Method for efficiently expressing recombinant liraglutide
  • Method for efficiently expressing recombinant liraglutide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0027] A. Cloning of the gene encoding liraglutide

[0028] Primers were designed according to the liraglutide gene sequence, and codons unfavorably expressed in Escherichia coli were replaced to obtain an optimized gene sequence. Two gene sequences were added to the 5' end of the gene sequence, which were enterokinase cleavage site sequences EK and HIS tag sequence HIS6, the expressed gene fragment HIS6-EK-liraglutide was obtained, the sequence is shown in SEQ ID NO:1.

[0029] B. Construction of expression vector

[0030] Then clone the full-length gene sequence of the molecular chaperone troponin C, the sequence is shown in SEQ ID NO: 2, and then connect it to the gene fragment HIS 6 -The 5' end of EK-liraglutide is ligated with the plasmid and gene fragments that have undergone the same enzyme digestion by T4DNA ligase, and the ligated product is transformed into E. coli BL21 by electroporation or 42°C heat shock method, that is, recombinant into Vector pET-22b-troponin ...

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Abstract

The invention relates to the biomedical field, and in particular relates to a method for efficiently expressing recombinant liraglutide. The method for efficiently expressing recombinant liraglutide is used for performing gene cloning and transformed expression. The recombinant liraglutide is constructed, and liraglutide protein is prepared through an escherichia coli expression technology. Compared with the two methods, the method disclosed by the invention is simple and convenient to operate, high in yield and low in cost.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a method for efficiently expressing recombinant liraglutide. Background technique [0002] Liraglutide is a human glucagon-like peptide-1 (GLP-1) analogue used in the treatment of diabetes. [0003] Liraglutide is an analogue of human glucagon-like peptide-1 (GLP-1), which has an amino acid difference compared with the molecular structure of natural GLP-1, and adds a 16-carbon palmitoyl fatty acid side chain, but it is different from the natural GLP-1 molecular structure. Human GLP-1 has 95% homology and retains the efficacy of natural GLP-1. It can bind and activate GLP-1 receptors, promote the secretion of insulin in the glucose concentration-dependent manner of pancreatic β cells, and at the same time increase the glucose concentration-dependent Mode to reduce excessive glucagon secretion. The liraglutide injection developed by Novo Nordisk has been proven to have a good effect in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/16C12N15/70C12N1/21C07K14/605C07K1/22C07K1/18
Inventor 张加慧封小燕刘沐荣刘翔
Owner 杭州北斗生物技术有限公司
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