Purification and biological activity of recombinant human bone morphogenetic protein-2 produced by E. coli expression system / 대한치주과학회지
The Journal of the Korean Academy of Periodontology
; : 41-50, 2008.
Article
in Ko
| WPRIM
| ID: wpr-170630
Responsible library:
WPRO
ABSTRACT
PURPOSE: Bone morphogenetic protein-2(BMP-2) has been shown to possess significant osteoinducitve potential. There have been attempts to overcome a limitation of mass production, and economical efficiency of BMP. The aim of this study was to produce recombinant human BMP-2(rhBMP-2) from E. coli in a large scale and evaluate its biological activity. MATERIALS AND METHODS: The E.coli strain BL21(DE3) was used as a host for rhBMP-2 production. Dimerized rhBMP-2 was purified by affinity chromatography using Heparin column. To determine the physicochemical properties of the rhBMP-2 expressed in E. coli, we examined the HPLC profile and performed Western blot analysis. The effect of the purified rhBMP-2 dimer on osteoblast differentiation was examined by alkaline phosphatase (ALP) activity and representing morphological change using C2C12 cell. RESULTS: E. coli was genetically engineered to produce rhBMP-2 in a non-active aggregated form. We have established a method which involves refolding and purifying a folded rhBMP-2 dimer from non-active aggregates. The purified rhBMP-2 homodimer was characterized by SDS-PAGE as molecular weight of about 28kDa and eluted at 34% acetonitrile, 13.27 min(retention time) in the HPLC profile and detected at Western blot. The purified rhBMP-2 dimer stimulated ALP activity and induced the transformation from myogenic differentiation to osteogenic differentiation. CONCLUSION: rhBMP-2 was produced in E. coli using genetic engineering. The purified rhBMP-2 dimer stimulated ALP activity and induced the osteogenic differentiation of C2C12 cells.
Key words
Full text:
1
Index:
WPRIM
Main subject:
Osteoblasts
/
Acetonitriles
/
Sprains and Strains
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Recombinant Proteins
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Heparin
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Genetic Engineering
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Blotting, Western
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Chromatography, Affinity
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Chromatography, High Pressure Liquid
/
Transforming Growth Factor beta
Limits:
Humans
Language:
Ko
Journal:
The Journal of the Korean Academy of Periodontology
Year:
2008
Type:
Article