A single residue (Arg⁴⁶) located within the N-terminus of the V_1a vasopressin receptor is critical for binding vasopressin but not peptide or nonpeptide antagonists

A fundamental issue in molecular endocrinology is to define how agonist:receptor interaction differs from antagonist:receptor interaction. The vasopressin V_1a receptor (V_1aR) is a member of a sub-family of related G protein-coupled receptors that are activated by the hormone AVP or related peptides. The N-terminus of the V_1aR 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 V_1aR. 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 V_1aR N-terminus and key differences between agonist and antagonist binding requirements.