Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Establishment of a Stable Cell Line Expressing Human BMP2/7-PTD for Efficient Osteogenic Induction

효과적인 뼈 세포분화 유도를 위한 유전자 재조합 PTD 융합 인간 뼈 형성촉진인자2/7(BMP2/7-PTD)를 발현하는 세포주 개발

  • Park, Seung-Won (Department of Agricultural Biology, National Academy of Agricultural Science, Rural Development Administration) ;
  • Kang, Seok-Woo (Department of Agricultural Biology, National Academy of Agricultural Science, Rural Development Administration) ;
  • Goo, Tae-Won (Department of Agricultural Biology, National Academy of Agricultural Science, Rural Development Administration) ;
  • Kim, Seong-Ryul (Department of Agricultural Biology, National Academy of Agricultural Science, Rural Development Administration) ;
  • Paik, Soon-Young (Department of Microbiology, College of Medicine, The Catholic University of Korea)
  • 박승원 (농촌진흥청 국립농업과학원 농업생물부 잠사양봉소재과) ;
  • 강석우 (농촌진흥청 국립농업과학원 농업생물부 잠사양봉소재과) ;
  • 구태원 (농촌진흥청 국립농업과학원 농업생물부 잠사양봉소재과) ;
  • 김성렬 (농촌진흥청 국립농업과학원 농업생물부 잠사양봉소재과) ;
  • 백순영 (가톨릭대학교 의과대학 미생물학교실)
  • Received : 2012.01.06
  • Accepted : 2012.03.07
  • Published : 2012.04.30
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Abstract

Heterodimeric recombinant human bone morphogenetic proteins (rhBMPs) are powerful tools for bone tissue engineering. However, BMPs have several important limitations in their application to bone regeneration. BMPs have a short half-life and must be used in high concentrations, which may be cost-inefficient. To overcome these problems, we established a stable cell line that expressed the fusion protein comprised of recombinant human BMP2/7 heterodimer protein and PTD (rhBMP2/7-PTD). This stable cell line enabled high process yields by continuously expressing rhBMP2/7-PTD products at high levels throughout cultivation. This synthesized BMP7 was fused to a BMP2 protein with four glycine residues (to allow free bond rotation of the domains) and PTD. To demonstrate that the rhBMP2/7-PTD protein that was secreted from an rhBMP2/7-PTD-expressing stable cell line exhibited biological activity consistent with its role as an osteogenic differentiation induction growth factor, we evaluated BMP-induced ALP activity. Our results suggest that this cell line may be a powerful and efficient tool for applications such as bone tissue regeneration.

유전자 재조합 이형이합체 인간 뼈 형성촉진인자(rhBMP)들은 뼈 재생을 위한 조직공학연구에 중요한 요소들이다. 그러나 실제 뼈 재생에 관한 연구를 수행함에 이들을 이용하는 것은 뼈 형성 촉진인자들이 짧은 반감기를 가지며 비교적 고농도의 단백질을 사용해야 하기 때문에 그만큼 많은 비용이 소요된다는 문제점을 가지고 있다. 이러한 한계를 뛰어넘기 위하여, 본 연구에서는 단백질 전달 서열(PTD)이 융합된 인간 뼈 형성촉진인자 2와 7이 이형이합체 유전자 재조합 단백질(rhBMP2/7-PTD)을 발현하는 세포주를 확립하였다. 이형이합체의 형성은 BMP 2와 BMP 7 단백질 및 PTD 영역의 사이에 각각 4개의 glycine 아미노산 염기서열이 첨가될 수 있도록 하여 각 단백질의 folding이 자유롭도록 디자인하였다. 이렇게 개발된 세포주는 고농도의 rhBMP2/7-PTD을 지속적으로 발현하여 배양액 내로 분비함으로 조직공학용 연구 및 개발에 효율적으로 이용 할 수 있다. 이상의 세포주에서 발현된 rhBMP2/7-PTD 단백질은 뼈 세포분화 유도를 확인 할 수 있는 ALP 활성을 나타냄으로써 뼈 성장촉진 단백질로서 생물학적인 활성을 가지고 있음을 보였다. 본 연구의 결과로 개발된 rhBMP2/7-PTD 형질전환 세포주는 향후 뼈 조직 재생과 같은 연구에 중요하고 효과적인 도구로 이용될 수 있을 것으로 사료된다.

Keywords

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