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| Expression of human acidic fibroblast growth factor in Pichia pastoris | |
| 作者姓名： | 余瑛 |
| 作者单位： | YU Ying1，2，3，CAI Shaoxi1，2，Harald G. WERIRICH 4，XIA Yuxian1，3，# 1College of Bioengineering，Chongqing University，Chongqing 400044，P.R. China 2Key Laboratory of Biomechanics and Tissue Engineering Under the State Ministry of Education，Chongqing University，Chongqing 400044，P.R. China 3Genetic Engineering Center，Chongqing University，Chongqing 400044，P.R. China 4Institute for Medical Chemistry and Biochemistry，University of Innsbruck，Austria Received 5 March 2003; revised 15 April 2003 |
| 基金项目： | the National Natural Science Foundation of China (No.19732003) |
| 摘 要： | 1. Introduction Acidic fibroblast growth factor (aFGF), a member of a family of structurally related polypeptides, is a kind of multifunctional protein that can stimulate cellular proliferation, migration and differentiation. In vivo, aFGF displays angiogenic activity promoting vascular endothelial cell mitogenesis as well as chemotaxis and induction of proteases involved in tissue regeneration 1]. Due to this wide range of biological activities, many potential therapeutic usages of aFGF … |
Expression of human acidic fibroblast growth factor in Pichia pastoris |
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| YU Ying,CAI Shaoxi,Harald G. WERIRICH,XIA Yuxian,# College of Bioengineering,Chongqing University,Chongqing,P.R. China Key Laboratory of Biomechanics and Tissue Engineering Under the State Ministry of Education,Chongqing University,C.Expression of human acidic fibroblast growth factor in Pichia pastoris[J].Journal of Chongqing University,2003,2(2). | |
| Authors: | YU Ying CAI Shaoxi Harald G WERIRICH XIA Yuxian # College of Bioengineering Chongqing University Chongqing PR China Key Laboratory of Biomechanics and Tissue Engineering Under the State Ministry of Education Chongqing University C |
| Institution: | 1. College of Bioengineering, Chongqing University, Chongqing 400044, P.R.China;Key Laboratory of Biomechanics and Tissue Engineering Under the State Ministry of Education, Chongqing University, Chongqino 400044.P.R.China;Genetic Engineering Center, Chonggqi 2. College of Bioengineering, Chongqing University, Chongqing 400044, P.R.China;Key Laboratory of Biomechanics and Tissue Engineering Under the State Ministry of Education, Chongqing University, Chongqino 400044.P.R.China 3. Institute for Medical Chemistry and Biochemistry, University of Innsbruck, Austria 4. College of Bioengineering, Chongqing University, Chongqing 400044, P.R.China;Genetic Engineering Center, Chongqing University, Chongqing 400044, P.R.China |
| Abstract: | Pichia pastoris expression system is similar to that of the mammal cell in modification of expressed protein, including refolding and glycosylation. A human aFGF gene was cloned into the intracellular expression vector pPIC9K. The Pichia pastoris KM71 strain was transformed with the recombined expression plasmid. Transgenic expression was observed after screening the transformants with G418. The expression and secretion of recombinant human aFGF (rhaFGF) into the culture medium were testified by ELISA assay. The yield peaked after two days of induction and was approximately 10 mgL-1 in shake-flask fermentation medium. The recombinant proteins were purified by the combination of heparin-Sepharose affinity chromatography and gel filtration chromatography. Two proteins with relative molecular masses (Mr) of 17 000 and 35 000 were purified as a single band in SDS-PAGE, whose biological activities were determined by MTT assay. It is found that the protein with Mr of 17 000 is nonglycosylated haFGF, and that with Mr of 35 000 is glycosylated haFGF; and the latter has a lower biological activity than the former. |
| Keywords: | aFGF Pichia pastoris expression |
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