Nozawa Akira, Nanamiya Hideaki, Tozawa Yuzuru
Cell-Free Science and Technology Research Center, Ehime University, Ehime, Japan.
Methods Mol Biol. 2010;607:213-8. doi: 10.1007/978-1-60327-331-2_17.
Membrane proteins play crucial roles in various processes. However, biochemical characterization of the membrane proteins remains challenging due to the difficulty in producing membrane proteins in a functional state. Here, we describe a novel method for the production of functional membrane proteins based on a wheat germ cell-free translation system. Using this method, functional membrane proteins are successfully synthesized in the presence of liposomes and a detergent. In addition, the synthesized membrane proteins are easily purified from the cell-free translation mixture as proteoliposomes by sucrose density gradient ultracentrifugation. These advantages over conventional approaches are very helpful for the clarification of the function of membrane proteins.
膜蛋白在各种过程中发挥着关键作用。然而，由于难以产生功能状态的膜蛋白，膜蛋白的生化特性表征仍然具有挑战性。在此，我们描述了一种基于小麦胚无细胞翻译系统生产功能膜蛋白的新方法。使用该方法，在脂质体和去污剂存在的情况下成功合成了功能膜蛋白。此外，通过蔗糖密度梯度超速离心，合成的膜蛋白作为蛋白脂质体很容易从无细胞翻译混合物中纯化出来。相对于传统方法的这些优势对于阐明膜蛋白的功能非常有帮助。
