Analysis of Arrestin Recruitment to Chemokine Receptors by Bioluminescence Resonance Energy Transfer.
Methods Enzymol
; 570: 131-53, 2016.
Article
em En
| MEDLINE
| ID: mdl-26921945
ABSTRACT
Chemokine receptors recruit the multifunctional scaffolding protein beta arrestin in response to binding of their chemokine ligands. Given that arrestin recruitment represents a signaling axis that is in part independent from G-protein signaling, it has become a hallmark of G protein-coupled receptor functional selectivity. Therefore, quantification of arrestin recruitment has become a requirement for the delineation of chemokine and drug candidate activity along different signaling axes. Bioluminescence resonance energy transfer (BRET) techniques provide methodology for such quantification that can reveal differences between nonredundant chemokines binding the same receptor, and that can be upscaled for high-throughput testing. We here provide protocols for the careful setup of BRET-based arrestin recruitment assays, and examples for the application of such systems in dose-response or time-course experiments. Suggestions are given for troubleshooting, optimizing test systems, and the interpretation of results obtained with BRET-based assays, which indeed yield an intricate blend of quantitative and qualitative information.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Arrestinas
/
Receptores de Quimiocinas
/
Técnicas de Transferência de Energia por Ressonância de Bioluminescência
Tipo de estudo:
Qualitative_research
Limite:
Humans
Idioma:
En
Revista:
Methods Enzymol
Ano de publicação:
2016
Tipo de documento:
Article
País de afiliação:
Canadá