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

Publication ,  Journal Article
Wu, S; Birnbaumer, M; Guan, Z
Published in: Anal Chem
August 1, 2008

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.

Duke Scholars

Published In

Anal Chem

DOI

EISSN

1520-6882

Publication Date

August 1, 2008

Volume

80

Issue

15

Start / End Page

6034 / 6037

Location

United States

Related Subject Headings

  • Tandem Mass Spectrometry
  • Serine
  • Receptors, Vasopressin
  • Receptors, G-Protein-Coupled
  • Phosphorylation
  • Phosphoproteins
  • Humans
  • Cyclic AMP-Dependent Protein Kinases
  • Chromatography
  • Binding Sites
 

Citation

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Wu, S., Birnbaumer, M., & Guan, Z. (2008). Phosphorylation analysis of G protein-coupled receptor by mass spectrometry: identification of a phosphorylation site in V2 vasopressin receptor. Anal Chem, 80(15), 6034–6037. https://doi.org/10.1021/ac8008548
Wu, Shilan, Mariel Birnbaumer, and Ziqiang Guan. “Phosphorylation analysis of G protein-coupled receptor by mass spectrometry: identification of a phosphorylation site in V2 vasopressin receptor.Anal Chem 80, no. 15 (August 1, 2008): 6034–37. https://doi.org/10.1021/ac8008548.
Wu, Shilan, et al. “Phosphorylation analysis of G protein-coupled receptor by mass spectrometry: identification of a phosphorylation site in V2 vasopressin receptor.Anal Chem, vol. 80, no. 15, Aug. 2008, pp. 6034–37. Pubmed, doi:10.1021/ac8008548.
Journal cover image

Published In

Anal Chem

DOI

EISSN

1520-6882

Publication Date

August 1, 2008

Volume

80

Issue

15

Start / End Page

6034 / 6037

Location

United States

Related Subject Headings

  • Tandem Mass Spectrometry
  • Serine
  • Receptors, Vasopressin
  • Receptors, G-Protein-Coupled
  • Phosphorylation
  • Phosphoproteins
  • Humans
  • Cyclic AMP-Dependent Protein Kinases
  • Chromatography
  • Binding Sites