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Dynamique structurale de complexes RCPG - neuropeptides

Abstract : G protein-coupled receptors (GPCR) recognize an extracellular ligand in order to transmit the corresponding information inside the cell. They are major players in physiology and predominant pharmacological targets. Here, we studied two GPCR for which the endogenous ligands are peptides. a) The ĸ opioid receptor (KOP), which recognizes dynorphin, and which is involved in the regulation of pain and reward mechanism (and thus in addiction). The discovery, pharmacology and structure - activity relationship of dynorphin are described in this manuscript as a review published in "Vitamins and Hormones" in 2019. GPCR activity requires a complex conformational dynamics and nuclear magnetic resonance (NMR) is an excellent technique allowing such characterization. However, it requires isotope labeling of the proteins of interest and most of GPCR structural studies used receptors expressed in insect cells. We produced KOP using an E. coli expression strategy, which is the tool of choice for isotope labeling. The recombinant receptor was successfully refolded in detergent micelles and was shown to be functional in terms of ligand binding. It offers new possibilities to study its structure and dynamics by NMR. b) The growth hormone secretagogue receptor (GHSR) is naturally activated by ghrelin, a digestive peptide hormone of 28 amino-acids residues. This hormone - receptor system is involved in a multitude of physiological process such as the regulation of food intake, growth hormone secretion and glucose homeostasis. However, structural information about peptide - GPCR interaction is sparse because of the difficulty in crystallizing such complexes. Using a perdeuterated receptor expressed in E. coli and refolded in lipid nanodiscs in the group of J.L. Banères (IBMM, Montpellier), we used several liquid state NMR techniques (15N relaxation, transferred NOE, saturation transfer difference) to characterize the 3D structure and dynamics of ghrelin bound to GHSR. Our work particularly shed the light on the role of a post-translational modification (acylation on Ser 3) on the receptor specific recognition mechanism.
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Guillaume Ferré. Dynamique structurale de complexes RCPG - neuropeptides. Pharmacologie. Université Paul Sabatier - Toulouse III, 2019. Français. ⟨NNT : 2019TOU30141⟩. ⟨tel-02651269⟩

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