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Expression and purification of recombinant tenascin-R domains and their functions
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- Author:
- No author available
- Journal Title:
- JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
- Issue:
- 10
- DOI:
- 10.3321/j.issn:1000-2790.2000.10.004
- Key Word:
- 肌腱蛋白-R;细胞外基质;表达;纯化;功能
Abstract： 目的将肌腱蛋白-R不同功能片段在原核中表达、纯化,并研究其初步的生物学功能. 方法将编码肌腱蛋白-R不同功能片段的表达质粒pGEX-EGFL, pGEX-EGFS, pGEX-FN1-2, pGEX-FN6-8, pGEX-FG及空载pGEX-KG转化入E.Coli JM109中. 用IPTG诱导表达,以谷胱甘肽琼脂糖为填充料进行亲合层析,纯化所表达的融合蛋白. 以纯化的融合蛋白作为培养液成分, GST为对照,观察纯化的肌腱蛋白-R各功能片段对体外培养脊髓细胞活性及突起生长的影响. 结果 肌腱蛋白R的5个片段在大肠杆菌中都有表达,通过谷胱甘肽琼脂糖亲合层析,都得到了初步纯化. 通过对培养细胞进行MTT染色,NSE染色后图像分析,结果表明,EGFL,FN1-2促进神经元的存活,EGFL,EGFS和FN6-8对突起的生长有明显的促进作用. 结论肌腱蛋白-R的不同功能片段可以通过大肠杆菌表达, 经纯化获得蛋白,肌腱蛋白-R不同的片段对体外培养的脊髓细胞有不同的作用.
- [1]J L, Cleland,S E, Builder,J R, Swartz,etc.Polyethylene glycol enhanced protein refolding.[J].1992,10(9).
- [2]M, Schachner,J, Taylor,U, Bartsch,etc.The perplexing multifunctionality of janusin, a tenascin-related molecule.[J].1994,2(1).
- [3]肖华胜,陈镔复.肌腱蛋白R（TENASCIN-R）研究进展[J].1998,(1).
- [4]Xiao, Z.-C.,Taylor, J.,Montag, D.,etc.Distinct Effects of Recombinant Tenascin-R Domains in Neuronal Cell Functions and Identification of the Domain Interacting with the Neuronal Recognition Molecule F3/11[J].1996,8(4).
- [5]Penka Pesheva,Gianfranco Gennarini,Christo Goridis,etc.The F3/11 cell adhesion molecule mediates the repulsion of neurons by the extracellular matrix glycoprotein J1-160/180[J].1993,10(1).
- [6]肖华胜,程希平,张萍,等.快速高效质粒转化方法的研制[J].1998,(4).
- [7](美)J.萨姆布鲁克, E.F.弗里奇, T.曼尼阿蒂斯著 金冬雁, 黎孟枫.分子克隆实验指南[M].1992.
- [8]Eva Sabine Wintergerst,Babette Fuss,Udo Bartsch.Localization of Janusin mRNA in the Central Nervous System of the Developing and Adult Mouse[J].1993,5(4).
- [9]P, Pesheva,R, Probstmeier,A P, Skubitz,etc.Tenascin-R (J1 160/180 inhibits fibronectin-mediated cell adhesion--functional relatedness to tenascin-C.[J].1994,107 ( Pt 8).
- [10].The J1 glycoproteina novel nervous system cell adhesion molecule of the L2/HNK-1 family[J].1985,316(6024).