Imaging of RNA-protein interactions in splicing complexes with dark-field STEM.
J Struct Biol
; 110(2): 111-21, 1993.
Article
em En
| MEDLINE
| ID: mdl-8338715
ABSTRACT
Splicing complexes were assembled in vitro on pre-messenger RNA. These complexes represent stages in the processing of pre-mRNA by snRNPs and accessory factors to remove the introns, thus forming messenger RNA. Complexes were purified using biotinylated oligonucleotide tags together with streptavidin-gold and simple centrifugation. The oligonucleotide tags with attached streptavidin-gold also serve as unambiguous labels of the splicing complexes for EM observation. The complexes were observed using dark-field scanning transmission electron microscopy (STEM). The high contrast and the high efficiency in detecting scattered electrons make it possible to visualize specimens at high resolution with low radiation dose. STEM visualization of the complexes allows a unique view of the events occurring in the splicing process. Three different classes of complex were identified. These complexes support the currently postulated steps in RNA splicing and demonstrate the dynamic nature of splicing complexes. This approach has broad application in high-resolution structural studies of nucleic acids and nucleic acid-protein interactions.
Buscar no Google
Base de dados:
MEDLINE
Assunto principal:
Precursores de RNA
/
Splicing de RNA
/
Spliceossomos
/
Ribonucleoproteínas Nucleares Pequenas
/
Microscopia Eletrônica de Transmissão e Varredura
Idioma:
En
Ano de publicação:
1993
Tipo de documento:
Article