Development of Refolding Process to Obtain Active Recombinant Human Bone Morphogenetic Protein-2 and its Osteogenic Efficacy on Oral Stem Cells.
10.11620/IJOB.2017.42.2.071
- Author:
Ji Hye LEE
1
;
Young Joo JANG
Author Information
1. Department of Nanobiomedical Science and BK21 PLUS Global Research Center for Regenerative Medicine, Dankook University, 29 Anseo-Dong, Cheonan 330-714, South Korea. yjjang@dankook.ac.kr
- Publication Type:Original Article
- Keywords:
Recombinant human bone morphogenetic protein-2 (rhBMP-2);
Refolding;
Heparin column purification;
Osteogenic induction
- MeSH:
Alkalies;
Alkaline Phosphatase;
Animals;
Cartilage;
Detergents;
Heparin;
Humans*;
Inclusion Bodies;
Mesenchymal Stromal Cells;
Methods;
Mice;
Myoblasts;
Osteoblasts;
Osteogenesis;
Solubility;
Stem Cells*;
Transforming Growth Factor beta
- From:International Journal of Oral Biology
2017;42(2):71-78
- CountryRepublic of Korea
- Language:English
-
Abstract:
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.
