Strategies for the purification and on-column cleavage of glutathione-S-transferase fusion target proteins
Strategies for the purification and on-column cleavage of glutathione-S-transferase fusion target proteins
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DOI：
10.1016/s1570-0232(01)00637-7
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发表时间：
2002-03-25
影响因子：
3
通讯作者：
Birse, D
中科院分区：
文献类型：
--
作者：
Dian, C;Eshaghi, S;Birse, D
在本报告中，我们描述了一种灵活、高效和快速的蛋白质纯化策略，用于分离和裂解谷胱甘肽- s -转移酶（GST）融合蛋白。纯化和柱上切割策略被开发用于纯化困难蛋白和靶蛋白，其中有效的融合标签切割对于下游过程（如结构和功能研究）至关重要。为了测试和证明这种方法的灵活性，七个不同的不相关的靶蛋白进行了分析。描述了一种纯化技术，可以应用于广泛的可溶性和膜插入重组靶蛋白的不同功能，结构和化学性质。该策略在单个色谱步骤中执行，采用柱上切割方法，在200杯至40 mg/l的范围内产生“天然”蛋白质，纯度为95-98%。(C) 2002年Elsevier Science B.V.出版
In this report, we describe a flexible, efficient and rapid protein purification strategy for the isolation and cleavage of glutathione-S-transferase (GST) fusion proteins. The purification and on-column cleavage strategy was developed to work for the purification of difficult proteins and for target proteins where efficient fusion-tag cleavage is essential for downstream processes, such as structural and functional studies. To test and demonstrate the flexibility of this method, seven diverse unrelated target proteins were assayed. A purification technique is described that can be applied to a wide range of both soluble and membrane inserted recombinant target proteins of differing function, structure and chemical nature. This strategy is performed in a single chromatographic step applying an on-column cleavage method, yielding "native" proteins in the 200 mug to 40 mg/l scale of 95-98% purity. (C) 2002 Published by Elsevier Science B.V.