The structure and function of the V1a vasopressin receptor
The neurohypohysial hormone family includes the nonapeptides vasopressin (A VP) and oxytocin
(OT). There are four distinct Family A (rhodopsin-like) G-protein coupled receptors (GPCRs) for
these agonists which are members of the neurohypophysial receptor sub-family of GPCRs. the
VJa receptor, the VJb receptor, the V2 receptor and the oxytocin receptor (OTR). V,"R. V'hR and
OTR are coupled to Gq/ ll and stimulate inostiol trisphosphate production. The V 2R couples to Ci,
and signals via adenylyl cyclase.
This study utilizes a combination of mutagenesis and molecular pharmacology to elucidate
important molecular determinants of structure and function for the rat V JaR. Initially, an available
homology model of V laR was analysed for the likely boundaries of the second extracellular loop
region (ECL2). "Alanine-scanning" mutagenesis of this ECL2 domain identification of
functionally important residues. The mutant receptors were compared with wild-type in terms of
their A VP-affinity and the interactions with three available peptide and non-peptide antagonists.
These data allowed the identification of a series of aromatic amino acid residues (Phe189
) and a putative disulphide-forming cysteine (Cys205) required for wild-type ligand
binding, activation and cell-surface localization.
The precise molecular requirements at these functionally important loci within EeL2 were
subsequently analysed by further site-directed mutagenesis. Interestingly. the roles of these amino
acids were found to have different biophysical constraints including chain length. hydrophobicity
and aromaticity. These analyses have increased overall understanding of the ECL2 domain of
Mutagenesis was further employed to determine the importance of two conserved motifs (NPxxY
and NSCC) of the V laR in the distal transmembrane region (TM 7) and the putative intracellular
eighth helix (Helix 8). This study identifies a crucial role for the arginine residue of the
functionally conserved NPxxY region (Asn3SO
) for agonist-mediated signalling and agonist
affinity but not trafficking/cell surface stability.
Overall, these results provide significant new information toward understanding the structure and
function of the neurohypophysial hormone receptors. Furthermore, these data may contribute to
the development of new therapeutic treatments and our understanding of GPCR-activation as a