Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center Omaha, NE, USA.
Front Pharmacol. 2013 Aug 23;4:106. doi: 10.3389/fphar.2013.00106. eCollection 2013.
A major problem for structural characterization of membrane proteins, such as connexins, by nuclear magnetic resonance (NMR) occurs at the initial step of the process, the production of sufficient amounts of protein. This occurs because proteins must be expressed in minimal based media. Here, we describe an expression system for membrane proteins that significantly improves yield by addressing two common problems, cell toxicity caused by protein translation and codon bias between genomes. This work provides researchers with a cost-effective tool for NMR and other biophysical studies, to use when faced with little-to-no expression of eukaryotic membrane proteins in Escherichia coli expression systems.
对于通过核磁共振(NMR)对连接蛋白等膜蛋白进行结构特征描述来说,一个主要问题出现在初始步骤,即需要产生足够量的蛋白质。这是因为蛋白质必须在最小基础培养基中表达。在这里,我们描述了一种膜蛋白表达系统,该系统通过解决两个常见问题,即由蛋白质翻译引起的细胞毒性和基因组之间的密码子偏倚,显著提高了产量。这项工作为研究人员提供了一种经济高效的 NMR 和其他生物物理研究工具,当在大肠杆菌表达系统中遇到真核膜蛋白表达量很少或没有时,可以使用该工具。