초록

＊ 인간의 BMP-2 아미노산 서열에 근거하고 대장균 유전자에서 선호하는 codon을 사용하여 BMP-2의 Synthetic gene을 합성함.
＊ 다양한 벡터와 host strain을 적용하여 발현 단백질의 가용화율을 높이고 발현 조건을 최적화 함.
＊ 정제한 단백질의 활성을 증명하기 위하여 in vitro assay system을 확립하고 in vivo 실험을 통하여 임상적으로 응용할 수 있는 기반을 마련함.

Abstract ▼

Bone morphogenetic protein-2 (BMP-2) is a low molecular weight glycoprotein belonging to a group of bone matrix proteins. The osteoinductive capacity of BMP-2 has been demonstrated in preclinical models and evaluated in clinical trials. BMPs can be isolated directly from bones; however, the yield is

Bone morphogenetic protein-2 (BMP-2) is a low molecular weight glycoprotein belonging to a group of bone matrix proteins. The osteoinductive capacity of BMP-2 has been demonstrated in preclinical models and evaluated in clinical trials. BMPs can be isolated directly from bones; however, the yield is lowand the purification scheme is complex. Recombinant BMPS have been obtained from mammalian cell cultures, but incomplete monomer processing and low yields are usual in these processes. Alternatively, the production of biologically active recombinant humans BMPs (rhBMPs) through in vitro refolding of Escherichia coli produced inclusion bodies has been reported. However, the refolding procedure is complicated and the overall yield is low. As well, the refolding buffer contains expensive reagents.
Our goal was to develop an effective method to improve the high production of the soluble form of an active BMP-2 without any refolding or solublization steps. For this purpose, A 345-bp gene that encodes human bone morphogenetic protein-2 (hBMP-2) has been synthesized. The codon usage of the resulting gene was modified to include those triplets that are utilized in highly expressed Escherichia coli genes. The hBMP-2 gene was efficiently expressed in E. coli as a soluble and active protein. Since the recombinant hBMP-2 was readily solublized, no further solublization steps were required throughout purification. No additional tagging residues were introduced into the synthetic hBMP-2 gene product. Afterwards, a high yielding method for a soluble expression of hBMP-2 with a biological activity was established.
The developed synthetic gene is a promising approach for scaling-up the soluble expression of hBMP-2.

목차 Contents

• 표지...1
• 제출문...2
• 보고서 요약서...3
• 요약문...4
• SUMMARY...5
• 연구 성과 실적 및 향후 계획...6
• 참여연구원 현황표...10
• II. 총괄연구과제 연구결과...11
• 1. 연구개발과제의 배경 및 필요성...12
• 2. 국내외 기술개발 현황...13
• 3. 연구개발과제의 추진체계...14
• 4. 연구개발수행 내용 및 결과...14
• 5. 목표달성도 및 관련분야 기여도...22
• 5.1. 총괄연구개발과제의 목표달성도...22
• 5.2. 관련분야 기여도...23
• 6. 향후 연구성과 추진 계획...24
• 7. 연구개발결과의 파급효과...24
• 7.1. 기술적 파급효과...24
• 7.2. 경제.산업적 파급효과...24
• 7.3. 사회적 파급효과...25
• 8. 연구개발결과의 활용계획...25
• 8.1. 연구개발결과의 활용계획 및 추가연구의 필요성...25
• 8.2. 사업화(기업화) 목표 달성도 및 계획...26
• 8.3. 사업화 수준(계획 및 완료 포함)...26
• 9. 연구개발과정에서 수집한 해외과학기술정보...26
• 10. 참고문헌...26
• IV. 첨부서류...28
• 1. 자체평가의견서...30
• 2. 실적 증빙자료...34