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- Title
Identification of Amino Acids in the N-Terminal Domain ofCorticotropin-Releasing Factor Receptor 1 that Are Important Determinants ofHigh-Affinity Ligand Binding.
- Authors
Wille, Sandra; Sydow, Sabine; Palchaudhuri, Monika R; Spiess, Joachim; Dautzenberg, Frank M.
- Abstract
Abstract: The aim of the present study was to identify theN-terminal regions of human corticotropin-releasing factor (CRF) receptor type1 (hCRF-R1) that are crucial for ligand binding. Mutant receptors wereconstructed by replacing specific residues in hCRF-R1 with amino acids fromthe corresponding position in the N-terminal region of the human vasoactiveintestinal peptide receptor type 2 (hVIP-R2). In cyclic AMP stimulation andCRF binding assays, it was established that two regions within the N-terminaldomain were crucial for the binding of CRF receptor agonists and antagonists:one region mapping to amino acids 43-50 and a second amino acid sequenceextending from position 76 to 84 of hCRF-R1. Recently, it was found that thelatter sequence plays a very important role in determining the high ligandselectivity of the Xenopus CRF-R1 (xCRF-R1). Replacement of aminoacids 76-84 of hCRF-R1 with residues from the same segment of the hVIP-R2 Nterminus markedly reduced the binding affinity of CRF ligands. Mutation ofArg[sup 76] or Asn[sup 81] but not Gly[sup 83] of hCRF-R1 tothe corresponding amino acids of xCRF-R1 or hVIP-R2 resulted in 100-1,000-foldlower affinities for human/rat CRF, rat urocortin, and astressin. These dataunderline the importance of the N-terminal domain of CRF-R1 in high-affinityligand binding.
- Subjects
CORTICOTROPIN releasing hormone; RADIOLIGAND assay
- Publication
Journal of Neurochemistry, 1999, Vol 72, Issue 1, p388
- ISSN
0022-3042
- Publication type
Article
- DOI
10.1046/j.1471-4159.1999.0720388.x