Phosphorylation analysis of G protein-coupled receptor by mass spectrometry: identification of a phosphorylation site in V2 vasopressin receptor.

Published

Journal Article

Phosphorylation plays vital roles in the regulation and function of the V2 vasopressin receptor (V2R), a G protein-coupled receptor (GPCR) that is responsible for maintaining water homeostasis in the kidney. Through a combination of immunoaffinity purification, immobilized metal affinity chromatography, and nanoflow liquid chromatography tandem mass spectrometry, we identified a novel phosphorylation site (Ser(255)) in the third intracellular loop of human V2R. We showed that the third intracellular loop could be phosphorylated in vitro by protein kinase A, but not by Akt kinase, although sequence motif analysis predicated otherwise. The analytical procedures and methodologies described in this study should be generally applicable for identifying the endogenous phosphorylation sites in other GPCRs, overcoming the limitations of conventional approaches such as sequence motif analysis and site-directed mutagenesis.

Full Text

Duke Authors

Cited Authors

  • Wu, S; Birnbaumer, M; Guan, Z

Published Date

  • August 1, 2008

Published In

Volume / Issue

  • 80 / 15

Start / End Page

  • 6034 - 6037

PubMed ID

  • 18578504

Pubmed Central ID

  • 18578504

Electronic International Standard Serial Number (EISSN)

  • 1520-6882

Digital Object Identifier (DOI)

  • 10.1021/ac8008548

Language

  • eng

Conference Location

  • United States