A single residue (arg46) located within the N-terminus of the V1a vasopressin receptor is critical for binding vasopressin but not peptide or nonpeptide antagonists

Stuart R Hawtin, Victoria J Wesley, Rosemary A Parslow, John Simms, Alice Miles, Kim McEwan, Mark Wheatley

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

A fundamental issue in molecular endocrinology is to define how agonist:receptor interaction differs from antagonist:receptor interaction. The vasopressin V1a receptor (V1aR) is a member of a subfamily of related G protein-coupled receptors that are activated by the hormone AVP or related peptides. The N-terminus of the V1aR has recently been shown to be critical for binding agonists but not antagonists. Using a combination of N-terminally truncated constructs and alanine-scanning mutagenesis, individual residues that provide these agonist-specific binding epitopes have now been identified in this study. Our data establish that a single residue, Arg46, is critical for AVP binding to the V1aR. Systematic substitution revealed that Arg was required at this locus and could not be substituted by Lys, Glu, Leu, or Ala. In contrast, antagonist binding (cyclic or linear, peptide or nonpeptide) was unaffected. Disruption of Arg46 also resulted in defective intracellular signaling. Arginine is conserved at this locus in all members of the neurohypophysial peptide hormone receptor family cloned to date, indicative of a fundamental role in receptor function. In addition to Arg46, the residues Leu42, Gly43, Asp45 form a patch contributing to AVP binding. This study provides molecular insight into the role of the V1aR N-terminus and key differences between agonist and antagonist binding requirements.

Original languageEnglish
Pages (from-to)600-609
Number of pages10
JournalMolecular and Cellular Endocrinology
Volume16
Issue number3
DOIs
Publication statusPublished - Mar 2002

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Posterior Pituitary Hormones
Vasopressin Receptors
Vasopressins
Peptides
Peptide Receptors
Endocrinology
G-Protein-Coupled Receptors
Mutagenesis
Alanine
Arginine
Epitopes
Hormones

Keywords

  • Amino Acid Sequence
  • Animals
  • Arginine
  • Arginine Vasopressin
  • Binding Sites
  • Cell Line
  • Humans
  • Molecular Sequence Data
  • Mutagenesis
  • Radioligand Assay
  • Rats
  • Receptors, Vasopressin
  • Second Messenger Systems
  • Structure-Activity Relationship
  • Transfection
  • Tritium
  • Vasopressins
  • Journal Article
  • Research Support, Non-U.S. Gov't

Cite this

A single residue (arg46) located within the N-terminus of the V1a vasopressin receptor is critical for binding vasopressin but not peptide or nonpeptide antagonists. / Hawtin, Stuart R; Wesley, Victoria J; Parslow, Rosemary A; Simms, John; Miles, Alice; McEwan, Kim; Wheatley, Mark.

In: Molecular and Cellular Endocrinology, Vol. 16, No. 3, 03.2002, p. 600-609.

Research output: Contribution to journalArticle

Hawtin, Stuart R ; Wesley, Victoria J ; Parslow, Rosemary A ; Simms, John ; Miles, Alice ; McEwan, Kim ; Wheatley, Mark. / A single residue (arg46) located within the N-terminus of the V1a vasopressin receptor is critical for binding vasopressin but not peptide or nonpeptide antagonists. In: Molecular and Cellular Endocrinology. 2002 ; Vol. 16, No. 3. pp. 600-609.
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AU - Wesley, Victoria J

AU - Parslow, Rosemary A

AU - Simms, John

AU - Miles, Alice

AU - McEwan, Kim

AU - Wheatley, Mark

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N2 - A fundamental issue in molecular endocrinology is to define how agonist:receptor interaction differs from antagonist:receptor interaction. The vasopressin V1a receptor (V1aR) is a member of a subfamily of related G protein-coupled receptors that are activated by the hormone AVP or related peptides. The N-terminus of the V1aR has recently been shown to be critical for binding agonists but not antagonists. Using a combination of N-terminally truncated constructs and alanine-scanning mutagenesis, individual residues that provide these agonist-specific binding epitopes have now been identified in this study. Our data establish that a single residue, Arg46, is critical for AVP binding to the V1aR. Systematic substitution revealed that Arg was required at this locus and could not be substituted by Lys, Glu, Leu, or Ala. In contrast, antagonist binding (cyclic or linear, peptide or nonpeptide) was unaffected. Disruption of Arg46 also resulted in defective intracellular signaling. Arginine is conserved at this locus in all members of the neurohypophysial peptide hormone receptor family cloned to date, indicative of a fundamental role in receptor function. In addition to Arg46, the residues Leu42, Gly43, Asp45 form a patch contributing to AVP binding. This study provides molecular insight into the role of the V1aR N-terminus and key differences between agonist and antagonist binding requirements.

AB - A fundamental issue in molecular endocrinology is to define how agonist:receptor interaction differs from antagonist:receptor interaction. The vasopressin V1a receptor (V1aR) is a member of a subfamily of related G protein-coupled receptors that are activated by the hormone AVP or related peptides. The N-terminus of the V1aR has recently been shown to be critical for binding agonists but not antagonists. Using a combination of N-terminally truncated constructs and alanine-scanning mutagenesis, individual residues that provide these agonist-specific binding epitopes have now been identified in this study. Our data establish that a single residue, Arg46, is critical for AVP binding to the V1aR. Systematic substitution revealed that Arg was required at this locus and could not be substituted by Lys, Glu, Leu, or Ala. In contrast, antagonist binding (cyclic or linear, peptide or nonpeptide) was unaffected. Disruption of Arg46 also resulted in defective intracellular signaling. Arginine is conserved at this locus in all members of the neurohypophysial peptide hormone receptor family cloned to date, indicative of a fundamental role in receptor function. In addition to Arg46, the residues Leu42, Gly43, Asp45 form a patch contributing to AVP binding. This study provides molecular insight into the role of the V1aR N-terminus and key differences between agonist and antagonist binding requirements.

KW - Amino Acid Sequence

KW - Animals

KW - Arginine

KW - Arginine Vasopressin

KW - Binding Sites

KW - Cell Line

KW - Humans

KW - Molecular Sequence Data

KW - Mutagenesis

KW - Radioligand Assay

KW - Rats

KW - Receptors, Vasopressin

KW - Second Messenger Systems

KW - Structure-Activity Relationship

KW - Transfection

KW - Tritium

KW - Vasopressins

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

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DO - 10.1210/mend.16.3.0795

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JO - Molecular Endocrinology

JF - Molecular Endocrinology

SN - 0303-7207

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ER -