title: Molecular basis of agonist docking in a human GPR103 homology model by site-directed mutagenesis and structure-activity relationship studies.
abstract: OBJECTIVE
  The neuropeptide 26RFa and its cognate receptor GPR103 are involved in the control of food intake and bone mineralization. Here, we have tested, experimentally, the predicted ligand-receptor interactions by site-directed mutagenesis of GPR103 and designed point-substituted 26RFa analogues.
  
  
  METHODS
  Using the X-ray structure of the β2 -adrenoceptor, a 3-D molecular model of GPR103 has been built. The bioactive C-terminal octapeptide 26RFa(19-26) , KGGFSFRF-NH2 , was docked in this GPR103 model and the ligand-receptor complex was submitted to energy minimization.
  
  
  RESULTS
  In the most stable complex, the Phe-Arg-Phe-NH2 part was oriented inside the receptor cavity, whereas the N-terminal Lys residue remained outside. A strong intermolecular interaction was predicted between the Arg(25) residue of 26RFa and the Gln(125) residue located in the third transmembrane helix of GPR103. To confirm this interaction experimentally, we tested the ability of 26RFa and Arg-modified 26RFa analogues to activate the wild-type and the Q125A mutant receptors transiently expressed in CHO cells. 26RFa (10(-6)  M) enhanced [Ca(2+) ]i in wild-type GPR103-transfected cells, but failed to increase [Ca(2+) ]i in Q125A mutant receptor-expressing cells. Moreover, asymmetric dimethylation of the side chain of arginine led to a 26RFa analogue, [ADMA(25) ]26RFa(20-26) , that was unable to activate the wild-type GPR103, but antagonized 26RFa-evoked [Ca(2+) ]i increase.
  
  
  CONCLUSIONS
  Altogether, these data provide strong evidence for a functional interaction between the Arg(25) residue of 26RFa and the Gln(125) residue of GPR103 upon ligand-receptor activation, which can be exploited for the rational design of potent GPR103 agonists and antagonists.
authors.affiliation: Inserm U982, Laboratory of Neuronal and Neuroendocrine Cell Differentiation and Communication, Neurotrophic Factors and Neuronal Differentiation Team, Institute for Research and Innovation in Biomedicine (IRIB); Cell Imaging Platform of Normandy (PRIMACEN), IRIB; Normandie Univ, France.
authors.forename: C
authors.initials: C
authors.lastname: Neveu
authors.forename: F
authors.initials: F
authors.lastname: Dulin
authors.forename: B
authors.initials: B
authors.lastname: Lefranc
authors.forename: L
authors.initials: L
authors.lastname: Galas
authors.forename: C
authors.initials: C
authors.lastname: Calbrix
authors.forename: R
authors.initials: R
authors.lastname: Bureau
authors.forename: S
authors.initials: S
authors.lastname: Rault
authors.forename: J
authors.initials: J
authors.lastname: Chuquet
authors.forename: J A
authors.initials: JA
authors.lastname: Boutin
authors.forename: L
authors.initials: L
authors.lastname: Guilhaudis
authors.forename: I
authors.initials: I
authors.lastname: Ségalas-Milazzo
authors.forename: D
authors.initials: D
authors.lastname: Vaudry
authors.forename: H
authors.initials: H
authors.lastname: Vaudry
authors.forename: J Sopkova-de Oliveira
authors.initials: JS
authors.lastname: Santos
authors.forename: J
authors.initials: J
authors.lastname: Leprince
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009479:Neuropeptides / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry; D043562:Receptors, G-Protein-Coupled / Q000819:agonists / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013329:Structure-Activity Relationship
chemical_list: D009479:Neuropeptides; D009842:Oligopeptides; C476582:QRFPR protein, human; D018343:Receptors, Adrenergic, beta-2; D043562:Receptors, G-Protein-Coupled