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A role for K268 in V2R folding.

Christian Le Gouill1, Thomas Darden, Michael T Madziva

  • 1Department of Anesthesiology, UCLA School of Medicine, Los Angeles, CA 90095, USA.

FEBS Letters
|August 24, 2005
PubMed
Summary

The V2 vasopressin receptor (V2R) requires specific amino acids in its third intracellular loop for Gs activation. Mutations at positions 231 and 268 diminish signaling, suggesting an interaction crucial for receptor function.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Pharmacology

Background:

  • The V2 vasopressin receptor (V2R), a G protein-coupled receptor (GPCR), regulates water reabsorption in the kidneys.
  • V2R activation of Gs is critical for its physiological function.

Purpose of the Study:

  • To investigate the role of the third intracellular loop of V2R in Gs activation.
  • To identify specific amino acid residues essential for V2R signal transduction.

Main Methods:

  • Utilized a chimeric receptor approach by inserting V2R segments into the Gq-coupled V1aR.
  • Employed site-directed mutagenesis to confirm findings in the native V2R.
  • Assessed receptor expression, ligand binding, internalization, phosphorylation, and signaling activity.

Main Results:

  • Identified glutamate 231 and glutamine 225 at the amino terminus of loop 3 as necessary for V2R signal transduction.
  • A double-mutant (Lys231/Glu268) showed enhanced expression and normal binding/internalization but significantly reduced signaling.
  • Positively charged amino acid at codon 268 is essential for V2R expression.

Conclusions:

  • The identified amino acids (231 and 268) in the V2R third intracellular loop are crucial for Gs protein activation.
  • These residues likely interact and play a role in promoting GDP/GTP exchange, a key step in G protein signaling.
  • Mutations affecting these residues impair V2R signaling despite maintaining receptor expression and ligand binding.

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