Bayburt T H, Carlson J W, Sligar S G
Department of Biochemistry, University of Illinois, Urbana, Illinois, 61801, USA.
J Struct Biol. 1998 Sep;123(1):37-44. doi: 10.1006/jsbi.1998.4007.
A phospholipid bilayer of nanometer dimension has been used as a support for the study of reconstituted functional single-membrane proteins. This nanobilayer consists of an approximately 10-nm-diameter circular phospholipid domain stabilized by apolipoprotein A1. As a demonstration of this methodology, we formed the nanobilayers in the presence of hepatic microsomal NADPH-cytochrome P450 reductase. Incubation of a solution of enzyme-containing nanobilayers with a freshly cleaved mica substrate resulted in the spontaneous formation of a fully oriented supported monolayer of discoidal phospholipid domains. The P450-reductase in the oriented monolayer retains its catalytic activity. Characterization by scanning force microscopy revealed isolated single-membrane proteins that could be stably imaged over time. These results define a novel technique for the study of single-membrane proteins in a bilayer environment.
纳米尺寸的磷脂双层已被用作研究重组功能性单膜蛋白的载体。这种纳米双层由载脂蛋白A1稳定的直径约为10纳米的圆形磷脂结构域组成。作为该方法的一个实例，我们在肝微粒体NADPH-细胞色素P450还原酶存在的情况下形成了纳米双层。将含酶纳米双层溶液与新鲜劈开的云母底物孵育，导致盘状磷脂结构域完全取向的支持单层自发形成。取向单层中的P450还原酶保留其催化活性。通过扫描力显微镜表征发现了可以随时间稳定成像的孤立单膜蛋白。这些结果定义了一种在双层环境中研究单膜蛋白的新技术。
