Detection of β-Arrestin-Mediated G Protein-Coupled Receptor Ubiquitination Using BRET.

Journal Article

Ubiquitination of G protein-coupled receptors (GPCRs) is an important dynamic posttranslational modification that has been linked to the intracellular trafficking of internalized GPCRs to lysosomes. Ubiquitination of GPCRs is mediated by specific E3 ubiquitin ligases that are scaffolded by the adaptor proteins called β-arrestins. Traditionally, detection of GPCR ubiquitination is achieved by using ubiquitin antibodies to Western blot immunoprecipitates of detergent-solubilized GPCRs expressed in heterologous cells. However, studies have also shown that bioluminescence resonance energy transfer (BRET)-based techniques can reveal ubiquitination of GPCRs in intact cells and in real time. This chapter describes a step-by-step protocol to evaluate ubiquitination of GPCRs using the BRET methodology.

Full Text

Duke Authors

Cited Authors

  • Nagi, K; Shenoy, SK

Published Date

  • 2019

Published In

Volume / Issue

  • 1957 /

Start / End Page

  • 93 - 104

PubMed ID

  • 30919349

Pubmed Central ID

  • 30919349

Electronic International Standard Serial Number (EISSN)

  • 1940-6029

Digital Object Identifier (DOI)

  • 10.1007/978-1-4939-9158-7_6

Language

  • eng

Conference Location

  • United States