Journal of Periodontal and Implant Science

Korean Academy of Periodontology (대한치주과학회)
권호 표지

Purification and biological activity of recombinant human bone morphogenetic protein-2 produced by E. coli expression system

E. coli 발현 시스템에 의해 생산된 recombinant human bone morphogenetic protein-2의 정제와 생물학적 활성

  • Choi, Kyung-Hee (Research Development Institute, Cowellmedi Co. LTD.) ;
  • Moon, Keumok (Research Development Institute, Cowellmedi Co. LTD.) ;
  • Kim, Soo-Hong (Research Development Institute, Cowellmedi Co. LTD.) ;
  • Yun, Jeong-Ho (Department of Dentistry, College of Medicine, Kwandong University, Myongji Hospital) ;
  • Jang, Kyung-Lib (Depatment of Biological Science, College of Natural science, Pusan National University) ;
  • Cho, Kyoo-Sung (Department of Periodontology, Research Institute for Periodontal Regeneration, College of Dentistry, Yonsei University)
  • 최경희 ((주) 코웰메디, 기술연구소) ;
  • 문금옥 ((주) 코웰메디, 기술연구소) ;
  • 김수홍 ((주) 코웰메디, 기술연구소) ;
  • 윤정호 (관동대학교 의과대학 명지병원 치과) ;
  • 장경립 (부산대학교 생명과학부) ;
  • 조규성 (연세대학교 치과대학 치주과학교실, 치주조직재생연구소)
  • Published : 2008.03.31
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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.

Keywords

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