Heterodimer protein of recombinant human bone morphogenetic protein and efficient expression and renaturation method of heterodimer protein

A technology of morphogenic proteins and heterodimers, applied in chemical instruments and methods, recombinant DNA technology, using vectors to introduce foreign genetic material, etc., can solve the problems of easy pollution, limited wide application, and high price of BMP proteins, and achieve The effect of improving alkaline phosphatase activity, saving protein purification steps, and simple culture conditions

Inactive Publication Date: 2015-01-14
TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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Problems solved by technology

[0005] At present, BMP protein is generally obtained by extracting from bovine bone, which is costly, low in purity and easy to contaminate
And higher animals are immune to heterologous BMP, which will weaken the osteoinductive activity of BMP
The use of insect cells o

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  • Heterodimer protein of recombinant human bone morphogenetic protein and efficient expression and renaturation method of heterodimer protein
  • Heterodimer protein of recombinant human bone morphogenetic protein and efficient expression and renaturation method of heterodimer protein
  • Heterodimer protein of recombinant human bone morphogenetic protein and efficient expression and renaturation method of heterodimer protein

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Embodiment Construction

[0035] Below in conjunction with embodiment, the present invention is further described, and following embodiment is illustrative, not limiting, can not limit protection scope of the present invention with following embodiment.

[0036]The invention provides a recombinant human bone morphogenetic protein heterodimer protein, specifically BMP7 / BMP2 heterodimer protein, the sequence of the protein from the N-terminal to the C-terminal is BMP7-connecting peptide-BMP2, and the connecting peptide is composed of Composed of 3 repeated amino acid sequences of GlyGlyGlyGlySer. BMP2 of the present invention, BMP7 mature peptide nucleic acid sequence, BMP2 contains 104 amino acids, BMP7 contains 139 amino acids;

[0037] The method for expressing a large amount of BMP7 / BMP2 heterodimer protein in Escherichia coli and the main method for renaturation to produce active recombinant BMP protein according to the present invention, the steps are as follows:

[0038] (1) Construction of recom...

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Abstract

The invention relates to a heterodimer protein of a recombinant human bone morphogenetic protein and an efficient expression and renaturation method of the heterodimer protein. The method comprises the following steps: linking a BMP (Bone Morphogenetic Protein) 7 gene with a BMP2 gene through DNA fragments encoding (GlyGlyGlyGlySer)3 by virtue of a fusion PCR technology, inserting fusion gene into a pHis-NusA plasmid so as to construct a recombinant plasmid, introducing the recombinant plasmid into escherichia coli BL21 (DE3), culturing at 37 DEG C, and inducing the expression by virtue of 0.5mM IPTG. The results show that the BMP7/BMP2heterodimer protein can be efficiently expressed by virtue of the method, the expressed protein is present in a form of an inclusion body and a large number of renaturated proteins can be obtained by virtue of simple purification and renaturation of the inclusion body. According to the method, the operation for obtaining the BMP active protein is simple, the preparation cost is greatly reduced and the method is suitable for large-scale fermentation production and provides the possibility of clinical wide application of BMP in the treatment of bone damage.

Description

technical field [0001] The present invention relates to high-efficiency expression of BMP7 / BMP2 heterodimer protein by Escherichia coli prokaryotic expression system, and preparation of biologically active recombinant human bone morphogenetic protein BMP7 / BMP2 heterodimer protein through denaturation and dialysis renaturation new method. Background technique [0002] Bone morphogenetic protein (BMP) is a protein that was first discovered in 1965 and can induce normal and ectopic osteogenesis. Ability to induce new bone formation. At present, more than 20 BMPs have been identified and cloned, and they belong to the members of the transforming growth factor β (transforming growth factor, TGF-β) supergene family. BMP2 is one of the BMPs with the most extensive research and the strongest osteogenic activity . BMP7 is mainly expressed in bone and kidney. BMP7 has strong osteoinductive activity, can stimulate the proliferation of undifferentiated mesenchymal cells and different...

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Application Information

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IPC IPC(8): C07K19/00C12N1/21C12N15/70C12N15/62
Inventor 刁爱坡党建利李玉银时威威罗深恒
Owner TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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