LIPOSOMES ON A STREPTAVIDIN CRYSTAL: A SYSTEM TO STUDY MEMBRANE PROTEINS BY CRYO-EM

被引：22

作者：

Wang, Liguo ^{[1
]}

Sigworth, Fred J. ^{[1
]}

机构：

[1] Yale Univ, Dept Cellular & Mol Physiol, New Haven, CT 06520 USA

来源：

METHODS IN ENZYMOLOGY, VOL 481: CRYO-EM, PART A - SAMPLE PREPARATION AND DATA COLLECTION | 2010年 / 481卷

关键词：

BIOTINYLATED MACROMOLECULES; 2-DIMENSIONAL CRYSTALS; ELECTRON-MICROSCOPY; CORE STREPTAVIDIN; POTASSIUM CHANNEL; LIPID-MEMBRANE; RESOLUTION; BINDING; RECONSTITUTION; ARRAYS;

D O I：

10.1016/S0076-6879(10)81007-9

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

In this chapter, we describe the preparation of cryo-EM specimens for random spherically constrained (RSC) single-particle reconstruction of membrane proteins. The specimen consists of liposomes into which the purified membrane protein is reconstituted at low density. The substrate is a 2D streptavidin crystal, which serves as an affinity surface that tethers the liposomes, which are doped with biotinylated lipids; the crystal can also serve as an image-quality and image-calibration reference. After subtraction of the crystal and lipid membrane contributions to the image, the remaining particle images can be used for 3D reconstruction.

引用

页码：147 / 164

页数：18