TY - JOUR
T1 - Overview of electron crystallography of membrane proteins
T2 - Crystallization and screening strategies using negative stain electron microscopy
AU - Nannenga, Brent L.
AU - Iadanza, Matthew G.
AU - Vollmar, Breanna S.
AU - Gonen, Tamir
PY - 2013
Y1 - 2013
N2 - Electron cryomicroscopy, or cryoEM, is an emerging technique for studying the three-dimensional structures of proteins and largemacromolecular machines. Electron crystallography is a branch of cryoEM inwhich structures of proteins can be studied at resolutions that rival those achieved by Xray crystallography. Electron crystallography employs two-dimensional crystals of a membrane protein embedded within a lipid bilayer. The key to a successful electron crystallographic experiment is the crystallization, or reconstitution, of the protein of interest. This unit describes ways in which protein can be expressed, purified, and reconstituted into well-ordered two-dimensional crystals. A protocol is also provided for negative stain electron microscopy as a tool for screening crystallization trials. When large and well-ordered crystals are obtained, the structures of both protein and its surrounding membrane can be determined to atomic resolution.
AB - Electron cryomicroscopy, or cryoEM, is an emerging technique for studying the three-dimensional structures of proteins and largemacromolecular machines. Electron crystallography is a branch of cryoEM inwhich structures of proteins can be studied at resolutions that rival those achieved by Xray crystallography. Electron crystallography employs two-dimensional crystals of a membrane protein embedded within a lipid bilayer. The key to a successful electron crystallographic experiment is the crystallization, or reconstitution, of the protein of interest. This unit describes ways in which protein can be expressed, purified, and reconstituted into well-ordered two-dimensional crystals. A protocol is also provided for negative stain electron microscopy as a tool for screening crystallization trials. When large and well-ordered crystals are obtained, the structures of both protein and its surrounding membrane can be determined to atomic resolution.
KW - Electron crystallography
KW - Membrane proteins
KW - Negative stain electron microscopy
KW - Protein purification
KW - Protein solubilization
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U2 - 10.1002/0471140864.ps1715s72
DO - 10.1002/0471140864.ps1715s72
M3 - Article
C2 - 23546618
AN - SCOPUS:84879775726
SN - 1934-3655
JO - Current protocols in protein science
JF - Current protocols in protein science
IS - SUPPL.72
M1 - 17.15
ER -