The synthesis of recombinant membrane proteins in yeast for structural studies.
The synthesis of recombinant membrane proteins in yeast for structural studies.
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DOI：
10.1016/j.ymeth.2015.09.027
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发表时间：
2016-02
期刊：
影响因子：
4.8
通讯作者：
S. Routledge;Lina Mikaliunaite;Anjana Patel;Michelle Clare;Stephanie P. Cartwright;Z. Bawa;Martin D B Wilks;F. Low;D. Hardy;A. Rothnie;R. Bill
中科院分区：
文献类型：
--
作者：
S. Routledge;Lina Mikaliunaite;Anjana Patel;Michelle Clare;Stephanie P. Cartwright;Z. Bawa;Martin D B Wilks;F. Low;D. Hardy;A. Rothnie;R. Bill
从历史上看，重组膜蛋白的生产一直是一个主要的挑战，这意味着发表的膜蛋白结构比那些可溶性蛋白少得多。然而，最近，蛋白质数据库中储存的膜蛋白质结构的数量几乎呈指数级增加。这表明，现在可以使用经验性的方法来确保这些困难目标所需的蛋白质供应。酵母是重组真核膜蛋白的重要来源，本文综述了酵母生产蛋白质的方法。我们概述了优化表达载体、宿主细胞和培养条件的方法，以及用于结晶试验的功能蛋白的提取和纯化，为结构研究做准备。
Historically, recombinant membrane protein production has been a major challenge meaning that many fewer membrane protein structures have been published than those of soluble proteins. However, there has been a recent, almost exponential increase in the number of membrane protein structures being deposited in the Protein Data Bank. This suggests that empirical methods are now available that can ensure the required protein supply for these difficult targets. This review focuses on methods that are available for protein production in yeast, which is an important source of recombinant eukaryotic membrane proteins. We provide an overview of approaches to optimize the expression plasmid, host cell and culture conditions, as well as the extraction and purification of functional protein for crystallization trials in preparation for structural studies.