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Title: Structure and function of V1b vasopressin receptor
Author: Goto, Yukie
ISNI:       0000 0004 2697 7514
Awarding Body: University of Birmingham
Current Institution: University of Birmingham
Date of Award: 2010
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The V1b vasopressin receptor (V1bR) is a receptor for a neurohypophysial hormone [arginine8] vasopressin (AVP). V1bR is a G-protein coupled receptor (GPCR) belonging to the Family A GPCR superfamily. The structures of seven α-helical transmembrane domains of this family members can be predicted based on the crystal structure of bovine rhodopsin (bRho) and human β2 adrenergic receptor (β2AR) obtained by X-ray crystallography. This study aimed to identify amino acid residues which participate in ligand binding of the V1bR by site-directed mutagenesis with the aid of molecular models of vasopressin receptors based on the crystal structure of bRho. The V1bR is a potential drug target in treating stress-related conditions such as depression, anxiety and post-traumatic stress disorders. Since it is the latest subtype identified among the mammalian neurohypophysial hormone receptors, it remains as the least studied subtype. A closely related subtype V1a receptor (V1aR) has been studied in far more detail for its potential of being a drug target in treating cardiac conditions and epilepsy. Hence, effective means of studying the V1bR can be accomplished by exploring the information already available on the V1aR and thereby defining the differences and similarities existing between the two. Detailed subtype comparisons are also fundamental for designing subtype selective drugs for effective therapy with fewer side-effects. This project was designed also to elucidate amino acid residues which determine selectivity of ligands for the V1bR over the V1aR. This study demonstrated that the charged residues Glu1.35 and Arg3.26, polar residues Gln2.61 and Tyr5.38, and cyclic residue Pro2.60 are crucial in AVP binding to the V1bR. These residues are all located at the extracellular facing surface of transmembrane domains (TMs). Also at the vicinity, charged residues Arg1.27 and Asp2.65, and aromatic Trp2.64 are shown to be important components of the AVP binding cavity. The study demonstrated that two residues Trp6.48 and Phe6.51 located at the TM6 are required for high affinity binding of non-peptide antagonists to the V1bR. The reciprocal mutagenesis between V1bR and V1aR residues revealed that Phe7.35 located at the exofacial surface of TM7 is a key residue required for a high affinity binding of a non-peptide antagonist with selectivity for the V1bR over V1aR. Also on the TM7, Met7.39 was shown to be involved in a high affinity binding of an antagonist with selectivity for the V1bR over V1aR. Two residues on the top of TM5, Leu5.39 and Thr5.42 were also shown to participate in V1bR-selective binding of non-peptide antagonists. From the perspectives of drug development, the variants of V1bR were studied in two streams: firstly to characterise pharmacological properties of single nucleotide polymorphism (SNP) variants of human V1bR; and secondly to determine differences in TM architectures between rat and human V1bRs. The study showed that a SNP variant of the V1bR with a residue substitution of Gly to Arg at position 191 is more readily expressed on the cell-surface in comparison to the wild-type. The variant was also found to generate InsP-InsP3 accumulation effectively in response to AVP stimulation. The mutagenesis involving introduction of specific rat V1bR residues into human V1bR revealed that there is a slight difference in the local environment of TM4 between the V1bRs found in the two species.
Supervisor: Not available Sponsor: Biotechnology and Biological Sciences Research Council ; Schering-Plough
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QR Microbiology