Generation of a proteolytic signal: E3/E2-mediated polyubiquitination of IκBα.
Methods Mol Biol
; 1280: 339-54, 2015.
Article
em En
| MEDLINE
| ID: mdl-25736759
ABSTRACT
A key regulatory node in NF-κB signaling is the removal of the IκBα inhibitor, whose levels are tightly controlled by the ubiquitin-proteasome system. In response to signal activation and transmission, ubiquitin E1, E2, and E3 enzymes are employed to generate a lysine 48-linked ubiquitin chain that triggers degradation of IκBα by the proteasome. In this chapter we describe an in vitro biochemical approach to reconstitute the ubiquitination system. To do so, we detail methods for the preparation of the relevant enzymes and substrate, as well as for the execution of the reaction with high efficiency. This sensitive and highly reproducible readout can be applied to the study of proteins, small molecules, and other factors that modulate IκBα ubiquitination, thereby producing outcomes that impact NF-κB signaling to advance the course of improving human health.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Transdução de Sinais
/
Proteínas I-kappa B
/
Ubiquitina
/
Ubiquitinação
Limite:
Humans
Idioma:
En
Revista:
Methods Mol Biol
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2015
Tipo de documento:
Article
País de afiliação:
Estados Unidos