南方医科大学学报

南方医科大学学报 ›› 2006, Vol. 26 ›› Issue (08): 1083-1086.

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大肠杆菌表达的人血管内皮细胞生长因子的复性与纯化研究(英文)

胡志明； 马骊； 周明乾； 高基民； 王小宁；   

  1. 南方医科大学分子免疫学研究所； 南方医科大学分子免疫学研究所 广东广州510515； 广东广州510515； 广东广州510515华南理工大学生物科学与工程学院； 广东广州510641；
  • 出版日期:2006-08-20 发布日期:2006-08-20
  • 基金资助:
    Supported by National "973"Development Project for High-TechResearch (2001CB510008), Sci-tech Development Program ofGuangzhou (2003Z2-E0171) and Key Sci-Tech Research Project ofGuangdong Province (2004B31201010)

Refolding and purification of recombinant human VEGF-121 expressed as inclusion bodies in Escherichia coli

HU Zhi-ming1, MA Li1, ZHOU Ming-qian1, GAO Ji-min1, WANG Xiao-ning1, 2 1Institute of Molecular Immunology, Southern Medical University, Guangzhou 510515, China; 2School of Biosciences & Bioengineering, South China University of Technology, Guangzhou 510641, China   

  1. 南方医科大学分子免疫学研究所； 南方医科大学分子免疫学研究所 广东广州510515； 广东广州510515； 广东广州510515华南理工大学生物科学与工程学院； 广东广州510641；
  • Online:2006-08-20 Published:2006-08-20

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摘要： 以包涵体的形式在大肠杆菌中表达人血管内皮细胞生长因子(VEGF121) , 采用发酵罐进行工程菌的高密度发酵,得到菌干重 46 g/L, VEGF121 包涵体 4.5 g/L。包涵体经洗涤、溶解、超滤法复性, 得率为 81%。复性后的目标蛋白经离子交换色谱及 Sepharcry S-100 凝胶过滤色谱纯化, 目标蛋白的纯度达到 95%, 目标蛋白的总得率 31%。采用该工艺制备的 VEGF121 能刺激人脐静脉血管内皮细胞增殖, 具有天然 VEGF 生物学活性。 

Abstract: Vascular endothelial growth factor 121 (VEGF121) was expressed as inclusion bodies by recombinant Escherichia coli. High concentrations of both biomass (46 g dry cell/L) and VEGF121 inclusion bodies (4.5 g/L) were obtained by applying a high-cell-density culture. After the inclusion bodies were washed and dissolved, VEGF121 was refolded at 0.2 mg/ml by ultrafiltration in refolding buffer with a yield of 81%. Renatured VEGF121 was purified by anion chromatography and Sephacry S-100 chromatography with purity higher than 95% and final purification yield of 31%. The purified VEGF121 could stimulate the proliferation of human umbilical vein endothelial cells as demonstrated by a biological activity assay. 

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