title: Unique interaction pattern for a functionally biased ghrelin receptor agonist.
abstract: Based on the conformationally constrained D-Trp-Phe-D-Trp (wFw) core of the prototype inverse agonist [D-Arg(1),D-Phe(5),D-Trp(7,9),Leu(11)]substance P, a series of novel, small, peptide-mimetic agonists for the ghrelin receptor were generated. By using various simple, ring-constrained spacers connecting the D-Trp-Phe-D-Trp motif with the important C-terminal carboxyamide group, 40 nm agonism potency was obtained and also in one case (wFw-Isn-NH(2), where Isn is isonipecotic acid) ~80% efficacy. However, in contrast to all previously reported ghrelin receptor agonists, the piperidine-constrained wFw-Isn-NH(2) was found to be a functionally biased agonist. Thus, wFw-Isn-NH(2) mediated potent and efficacious signaling through the Gα(q) and ERK1/2 signaling pathways, but in contrast to all previous ghrelin receptor agonists it did not signal through the serum response element, conceivably the Gα(12/13) pathway. The recognition pattern of wFw-Isn-NH(2) with the ghrelin receptor also differed significantly from that of all previously characterized unbiased agonists. Most importantly, wFw-Isn-NH(2) was not dependent on GluIII:09 (Glu3.33), which otherwise is an obligatory TM III anchor point residue for ghrelin agonists. Molecular modeling and docking experiments indicated that wFw-Isn-NH(2) binds in the classical agonist binding site between the extracellular segments of TMs III, VI, and VII, interacting closely with the aromatic cluster between TMs VI and VII, but that it does so in an opposite orientation as compared with, for example, the wFw peptide agonists. It is concluded that the novel peptide-mimetic ligand wFw-Isn-NH(2) is a biased ghrelin receptor agonist and that the selective signaling pattern presumably is due to its unique receptor recognition pattern lacking interaction with key residues especially in TM III.
authors.affiliation: Laboratory for Molecular Pharmacology, Department of Neuroscience and Pharmacology, University of Copenhagen, Blegdamsvej 3b, DK-2200, Copenhagen, Denmark.
authors.forename: Bjørn
authors.initials: B
authors.lastname: Sivertsen
authors.forename: Manja
authors.initials: M
authors.lastname: Lang
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Frimurer
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: Holliday
authors.forename: Anders
authors.initials: A
authors.lastname: Bach
authors.forename: Sylvia
authors.initials: S
authors.lastname: Els
authors.forename: Maja S
authors.initials: MS
authors.lastname: Engelstoft
authors.forename: Pia S
authors.initials: PS
authors.lastname: Petersen
authors.forename: Andreas N
authors.initials: AN
authors.lastname: Madsen
authors.forename: Thue W
authors.initials: TW
authors.lastname: Schwartz
authors.forename: Annette G
authors.initials: AG
authors.lastname: Beck-Sickinger
authors.forename: Birgitte
authors.initials: B
authors.lastname: Holst
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000235:genetics / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D057786:Peptidomimetics / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D054440:Receptors, Ghrelin / Q000819:agonists* / Q000235:genetics / Q000378:metabolism*; D013373:Substance P
chemical_list: D057786:Peptidomimetics; D054440:Receptors, Ghrelin; D013373:Substance P; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D044365:GTP-Binding Protein alpha Subunits, G12-G13; D043802:GTP-Binding Protein alpha Subunits, Gq-G11