The N-terminal juxtamembrane segment of the V1a vasopressin receptor provides two independent epitopes required for high-affinity agonist binding and signaling

Stuart R Hawtin, Victoria J Wesley, John Simms, Cymone C H Argent, Khalid Latif, Mark Wheatley

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

It is fundamentally important to define how agonist-receptor interaction differs from antagonist-receptor interaction. The V1a vasopressin receptor (V1aR) is a member of the neurohypophysial hormone subfamily of G protein-coupled receptors. Using alanine-scanning mutagenesis of the N-terminal juxtamembrane segment of the V1aR, we now establish that Glu54 (1.35) is critical for arginine vasopressin binding. The mutant [E54A]V1aR exhibited decreased arginine vasopressin affinity (1700-fold) and disrupted signaling, but antagonist binding was unaffected. Mutation of Glu54 had an almost identical pharmacological effect as mutation of Arg46, raising the possibility that agonist binding required a mutual interaction between Glu54 and Arg46. The role of these two charged residues was investigated by 1) substituting Glu54; 2) inserting additional Glu/Arg in transmembrane helix (TM) 1; 3) repositioning the Glu/Arg in TM1; and 4) characterizing the reciprocal mutant [R46E/E54R]V1aR. We conclude that 1) the positive/negative charges need to be precisely positioned in this N terminus/TM1 segment; and 2) Glu54 and Arg46 function independently, providing two discrete epitopes required for high-affinity agonist binding and signaling. This study explains why Glu and Arg, part of an -R(X3)L/V(X3)E(X3)L- motif, are conserved at these loci throughout this G protein-coupled receptor subfamily and provides molecular insight into key differences between agonist and antagonist binding requirements.

Original languageEnglish
Pages (from-to)2871-2871
Number of pages11
JournalMolecular and Cellular Endocrinology
Volume19
Issue number11
DOIs
Publication statusPublished - Nov 2005

Keywords

  • Amino Acid Motifs
  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Arginine
  • Arginine Vasopressin
  • Cells, Cultured
  • Glutamic Acid
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis
  • Protein Structure, Secondary
  • Receptors, Vasopressin
  • Signal Transduction
  • Journal Article
  • Research Support, Non-U.S. Gov't

Fingerprint Dive into the research topics of 'The N-terminal juxtamembrane segment of the V1a vasopressin receptor provides two independent epitopes required for high-affinity agonist binding and signaling'. Together they form a unique fingerprint.

  • Cite this