Francis Dana M, Page Rebecca
Brown University, Providence, Rhode Island.
Curr Protoc Protein Sci. 2010 Aug;Chapter 5(1):5.24.1-5.24.29. doi: 10.1002/0471140864.ps0524s61.
Recombinant protein expression in Escherichia coli (E. coli) is simple, fast, inexpensive, and robust, with the expressed protein comprising up to 50 percent of the total cellular protein. However, it also has disadvantages. For example, the rapidity of bacterial protein expression often results in unfolded/misfolded proteins, especially for heterologous proteins that require longer times and/or molecular chaperones to fold correctly. In addition, the highly reductive environment of the bacterial cytosol and the inability of E. coli to perform several eukaryotic post-translational modifications results in the insoluble expression of proteins that require these modifications for folding and activity. Fortunately, multiple, novel reagents and techniques have been developed that allow for the efficient, soluble production of a diverse range of heterologous proteins in E. coli. This overview describes variables at each stage of a protein expression experiment that can influence solubility and offers a summary of strategies used to optimize soluble expression in E. coli.
在大肠杆菌中进行重组蛋白表达简单、快速、成本低且稳定，表达的蛋白可占细胞总蛋白的50%。然而，它也有缺点。例如，细菌蛋白表达的快速性常常导致蛋白未折叠/错误折叠，特别是对于那些需要更长时间和/或分子伴侣才能正确折叠的异源蛋白。此外，细菌细胞质高度还原的环境以及大肠杆菌无法进行几种真核生物的翻译后修饰，导致那些需要这些修饰来折叠和发挥活性的蛋白以不溶性形式表达。幸运的是，已经开发出多种新型试剂和技术，可在大肠杆菌中高效、可溶性地生产多种异源蛋白。本综述描述了蛋白表达实验每个阶段可能影响溶解性的变量，并总结了用于优化大肠杆菌中可溶性表达的策略。
