title: Identification of amino acids in the N-terminal domain of corticotropin-releasing factor receptor 1 that are important determinants of high-affinity ligand binding.
abstract: The aim of the present study was to identify the N-terminal regions of human corticotropin-releasing factor (CRF) receptor type 1 (hCRF-R1) that are crucial for ligand binding. Mutant receptors were constructed by replacing specific residues in hCRF-R1 with amino acids from the corresponding position in the N-terminal region of the human vasoactive intestinal peptide receptor type 2 (hVIP-R2). In cyclic AMP stimulation and CRF binding assays, it was established that two regions within the N-terminal domain were crucial for the binding of CRF receptor agonists and antagonists: one region mapping to amino acids 43-50 and a second amino acid sequence extending from position 76 to 84 of hCRF-R1. Recently, it was found that the latter sequence plays a very important role in determining the high ligand selectivity of the Xenopus CRF-R1 (xCRF-R1). Replacement of amino acids 76-84 of hCRF-R1 with residues from the same segment of the hVIP-R2 N terminus markedly reduced the binding affinity of CRF ligands. Mutation of Arg76 or Asn81 but not Gly83 of hCRF-R1 to the corresponding amino acids of xCRF-R1 or hVIP-R2 resulted in 100-1,000-fold lower affinities for human/rat CRF, rat urocortin, and astressin. These data underline the importance of the N-terminal domain of CRF-R1 in high-affinity ligand binding.
authors.affiliation: Department of Molecular Neuroendocrinology, Max-Planck Institute for Experimental Medicine, Göttingen, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Wille
authors.forename: S
authors.initials: S
authors.lastname: Sydow
authors.forename: M R
authors.initials: MR
authors.lastname: Palchaudhuri
authors.forename: J
authors.initials: J
authors.lastname: Spiess
authors.forename: F M
authors.initials: FM
authors.lastname: Dautzenberg
mesh_terms: D000596:Amino Acids / Q000032:analysis*; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002478:Cells, Cultured; D003346:Corticotropin-Releasing Hormone / Q000378:metabolism / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D018076:DNA, Complementary; D006801:Humans; D007668:Kidney / Q000166:cytology; D008024:Ligands; D008969:Molecular Sequence Data; D016296:Mutagenesis / Q000502:physiology; D018696:Neuroprotective Agents / Q000378:metabolism / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D051381:Rats; D018019:Receptors, Corticotropin-Releasing Hormone / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D000596:Amino Acids; D018076:DNA, Complementary; D008024:Ligands; D018696:Neuroprotective Agents; D010446:Peptide Fragments; D018019:Receptors, Corticotropin-Releasing Hormone; C104416:astressin; D003346:Corticotropin-Releasing Hormone; D000242:Cyclic AMP