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Development of a human vasopressin V1a-receptor antagonist from an evolutionary-related insect neuropeptide.

ABSTRACT: Characterisation of G protein-coupled receptors (GPCR) relies on the availability of a toolbox of ligands that selectively modulate different functional states of the receptors. To uncover such molecules, we explored a unique strategy for ligand discovery that takes advantage of the evolutionary conservation of the 600-million-year-old oxytocin/vasopressin signalling system. We isolated the insect oxytocin/vasopressin orthologue inotocin from the black garden ant (Lasius niger), identified and cloned its cognate receptor and determined its pharmacological properties on the insect and human oxytocin/vasopressin receptors. Subsequently, we identified a functional dichotomy: inotocin activated the insect inotocin and the human vasopressin V1b receptors, but inhibited the human V1aR. Replacement of Arg8 of inotocin by D-Arg8 led to a potent, stable and competitive V1aR-antagonist ([D-Arg8]-inotocin) with a 3,000-fold binding selectivity for the human V1aR over the other three subtypes, OTR, V1bR and V2R. The Arg8/D-Arg8 ligand-pair was further investigated to gain novel insights into the oxytocin/vasopressin peptide-receptor interaction, which led to the identification of key residues of the receptors that are important for ligand functionality and selectivity. These observations could play an important role for development of oxytocin/vasopressin receptor modulators that would enable clear distinction of the physiological and pathological responses of the individual receptor subtypes.

SUBMITTER: Di Giglio MG

PROVIDER: S-EPMC5286520 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature

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Development of a human vasopressin V<sub>1a</sub>-receptor antagonist from an evolutionary-related insect neuropeptide.

Di Giglio Maria Giulia MG Muttenthaler Markus M Harpsøe Kasper K Liutkeviciute Zita Z Keov Peter P Eder Thomas T Rattei Thomas T Arrowsmith Sarah S Wray Susan S Marek Ales A Elbert Tomas T Alewood Paul F PF Gloriam David E DE Gruber Christian W CW

Scientific reports 20170201

Characterisation of G protein-coupled receptors (GPCR) relies on the availability of a toolbox of ligands that selectively modulate different functional states of the receptors. To uncover such molecules, we explored a unique strategy for ligand discovery that takes advantage of the evolutionary conservation of the 600-million-year-old oxytocin/vasopressin signalling system. We isolated the insect oxytocin/vasopressin orthologue inotocin from the black garden ant (Lasius niger), identified and c ...[more]

PMID: 28145450

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Project description:The neuropeptide vasopressin (VP) and its three G protein-coupled receptors (V1aR, V1bR and V2R) are of high interest in a wide array of drug discovery programs. V1aR is of particular importance due to its cardiovascular functions and diverse roles in the central nervous system. The structure-activity relationships underpinning ligand-receptor interactions remain however largely unclear, hindering rational drug design. This is not least due to the high structural flexibility of VP in its free as well as receptor-bound states. In this work, we developed a novel approach to reveal features of conformational selectivity upon VP-V1aR complex formation. We employed virtual screening strategies to probe VP's conformational space for transiently adopted structures that favor binding to V1aR. To this end, we dissected the VP conformational space into three sub-ensembles, each containing distinct structural sets for VP's three-residue C-terminal tail. We validated the computational results with experimental nuclear magnetic resonance (NMR) data and docked each sub-ensemble to V1aR. We observed that the conformation of VP's three-residue tail significantly modulated the complex dissociation constants. Solvent-exposed and proline trans-configured VP tail conformations bound to the receptor with three-fold enhanced affinities compared to compacted or cis-configured conformations. The solvent-exposed and more flexible structures facilitated unique interaction patterns between VP and V1aR transmembrane helices 3, 4, and 6 which led to high binding energies. The presented "virtual conformational space screening" approach, integrated with NMR spectroscopy, thus enabled identification and characterization of a conformational selection-type complex formation mechanism that confers novel perspectives on targeting the VP-V1aR interactions at the level of the encounter complex - an aspect that opens novel research avenues for understanding the functionality of the evolutionary selected conformational properties of VP, as well as guidance for ligand design strategies to provide more potent and selective VP analogues.

Project description:The identification of structurally related hypothalamic hormones that regulate blood pressure and diuresis (vasopressin, VP; CYFQNCPRG-NH2) or lactation and uterine contraction (oxytocin, OT; CYIQNCPLG-NH2) was a major advance in neuroendocrinology, recognized in the award of the Nobel Prize for Chemistry in 1955. Furthermore, the discovery of central actions of VP and OT as regulators of reproductive and social behavior in humans and other mammals has broadened interest in these neuropeptides beyond physiology into psychology. VP/OT-type neuropeptides and their G-protein coupled receptors originated in a common ancestor of the Bilateria (Urbilateria), with invertebrates typically having a single VP/OT-type neuropeptide and cognate receptor. Gene/genome duplications followed by gene loss gave rise to variety in the number of VP/OT-type neuropeptides and receptors in different vertebrate lineages. Recent advances in comparative transcriptomics/genomics have enabled discovery of VP/OT-type neuropeptides in an ever-growing diversity of invertebrate taxa, providing new opportunities to gain insights into the evolution of VP/OT-type neuropeptide function in the Bilateria. Here we review the comparative physiology of VP/OT-type neuropeptides in invertebrates, with roles in regulation of reproduction, feeding, and water/salt homeostasis emerging as common themes. For example, we highlight recent reports of roles in regulation of oocyte maturation in the sea-squirt Ciona intestinalis, extraoral feeding behavior in the starfish Asterias rubens and energy status and dessication resistance in ants. Thus, VP/OT-type neuropeptides are pleiotropic regulators of physiological processes, with evolutionarily conserved roles that can be traced back to Urbilateria. To gain a deeper understanding of the evolution of VP/OT-type neuropeptide function it may be necessary to not only determine the actions of the peptides but also to characterize the transcriptomic/proteomic/metabolomic profiles of cells expressing VP/OT-type precursors and/or VP/OT-type receptors within the framework of anatomically and functionally identified neuronal networks. Furthermore, investigation of VP/OT-type neuropeptide function in a wider range of invertebrate species is now needed if we are to determine how and when this ancient signaling system was recruited to regulate diverse physiological and behavioral processes in different branches of animal phylogeny and in contrasting environmental contexts.

Dataset Information

Development of a human vasopressin V1a-receptor antagonist from an evolutionary-related insect neuropeptide.

Publications

Development of a human vasopressin V<sub>1a</sub>-receptor antagonist from an evolutionary-related insect neuropeptide.

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