Ritchie T K, Grinkova Y V, Bayburt T H, Denisov I G, Zolnerciks J K, Atkins W M, Sligar S G
Department of Medicinal Chemistry, University of Washington, Seattle, Washington, USA.
Methods Enzymol. 2009;464:211-31. doi: 10.1016/S0076-6879(09)64011-8.
Self-assembled phospholipid bilayer Nanodiscs have become an important and versatile tool among model membrane systems to functionally reconstitute membrane proteins. Nanodiscs consist of lipid domains encased within an engineered derivative of apolipoprotein A-1 scaffold proteins, which can be tailored to yield homogeneous preparations of disks with different diameters, and with epitope tags for exploitation in various purification strategies. A critical aspect of the self-assembly of target membranes into Nanodiscs lies in the optimization of the lipid:protein ratio. Here we describe strategies for performing this optimization and provide examples for reconstituting bacteriorhodopsin as a trimer, rhodopsin, and functionally active P-glycoprotein. Together, these demonstrate the versatility of Nanodisc technology for preparing monodisperse samples of membrane proteins of wide-ranging structure.
自组装磷脂双分子层纳米盘已成为模型膜系统中用于功能性重组膜蛋白的一种重要且多功能的工具。纳米盘由包裹在载脂蛋白A-1支架蛋白工程衍生物内的脂质结构域组成，可进行定制以产生具有不同直径的均匀盘状制剂，并带有表位标签，便于在各种纯化策略中使用。将目标膜自组装成纳米盘的一个关键方面在于脂质与蛋白质比例的优化。在此，我们描述了进行这种优化的策略，并提供了将细菌视紫红质重组为三聚体、视紫红质和功能活性P-糖蛋白的示例。这些共同证明了纳米盘技术在制备具有广泛结构的膜蛋白单分散样品方面的多功能性。
