Three-dimensional imaging of in situ specimens with low-dose electron tomography to analyze protein conformation.
Assay Drug Dev Technol
; 2(5): 561-7, 2004 Oct.
Article
em En
| MEDLINE
| ID: mdl-15671654
ABSTRACT
We describe a novel three-dimensional (3-D) imaging tool for analysis of protein conformation of in situ samples. Sidec (Sidec Technologies AB, Stockholm, Sweden) electron tomography (SET) uses low-dose electron tomography and a refinement algorithm to reconstruct individual proteins and macromolecular complexes. The approach has successfully reconstructed therapeutic proteins in solution. In this study, we investigate the use of SET to visualize ion channels in cells and tissue samples. SET successfully resolved the volume and structural features of the target complex, showing that it was a tetrameric channel with a central pore. The technology could distinguish and provide 3-D images of the intra- and extracellular domains in the ion channel. In addition, SET was able to show that the channel associates in the form of a tetramer with the four subunits preorganized into dimers. While additional studies using smaller antibody markers are needed to resolve the subunit assembly further, this study demonstrates that SET is a valuable tool for visualization of in situ specimens and can provide important information on the subunit assembly of these macromolecular complexes, and thereby aid in the screening assay process in drug development.
Buscar no Google
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Tomografia
/
Imageamento Tridimensional
/
Elétrons
/
Canais Iônicos
Limite:
Animals
Idioma:
En
Revista:
Assay Drug Dev Technol
Assunto da revista:
FARMACOLOGIA
Ano de publicação:
2004
Tipo de documento:
Article
País de afiliação:
Suécia