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KMID : 0848120170420020071
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International Journal of Oral Biology
2017 Volume.42 No. 2 p.71 ~ p.78
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Development of Refolding Process to Obtain Active Recombinant Human Bone Morphogenetic Protein-2 and its Osteogenic Efficacy on Oral Stem Cells
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Lee Ji-Hye
Jang Young-Joo
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Abstract
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BMP-2 is a well-known TGF-beta related growth factor, having a significant role in bone and cartilage formation. It has been employed to promote bone formation in some clinical trials, and to differentiate mesenchymal stem cells into osteoblasts. However, it is difficult to obtain this protein in its soluble and active form. hBMP-2 is expressed as an inclusion body in the bacterial system. To continuously supply hBMP-2 for research, we optimized the refolding of recombinant hBMP-2 expressed in E. coli, and established an efficient method by using detergent and alkali. Using a heparin column, the recombinant hBMP-2 was purified with the correct refolding. Although combinatorial refolding remarkably enhanced the solubility of the inclusion body, a higher yield of active dimer form of hBMP-2 was obtained from one-step refolding with detergent. The refolded recombinant hBMP-2 induced alkaline phosphatase activity in mouse myoblasts, at ED50 of 300-480ng/ml. Furthermore, the expressions of osteogenic markers were upregulated in hPDLSCs and hDPSCs. Therefore, using the process described in this study, the refolded hBMP-2 might be cost-effectively useful for various differentiation experiments in a laboratory.
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KEYWORD
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morphogenetic protein-2 (rhBMP-2) , Refolding , Heparin column purification , Osteogenic induction
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