Preparation of ribosomes loaded with truncated nascent proteins to study ribosome binding to mammalian mitochondria.
Mitochondrion
; 6(2): 64-70, 2006 Apr.
Article
em En
| MEDLINE
| ID: mdl-16513430
ABSTRACT
Supporting a co-translational model of protein import into mitochondria, we have previously shown that ribosome-nascent chain complexes (RNCs) specifically bind to mitochondria. When producing RNCs using the rabbit reticulocyte lysate in vitro translation system, it was necessary to maximize ribosome loading with truncated nascent proteins because it had a direct impact on RNC binding. We describe here the optimal conditions for preparing RNCs. We show that translation temperature and reaction time are two critical factors, with 30 degrees Celsius and 15min being optimal, respectively. We also show that transcription reactions can be used directly in the translation reaction to create RNCs.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Ribossomos
/
Mitocôndrias
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
Mitochondrion
Ano de publicação:
2006
Tipo de documento:
Article
País de afiliação:
Estados Unidos