The role of the fourth extracellular domain of the rat corticotropin-releasing factor receptor type 1 in ligand binding
1999 | journal article; research paper. A publication with affiliation to the University of Göttingen.
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The role of the fourth extracellular domain of the rat corticotropin-releasing factor receptor type 1 in ligand binding
Sydow, S.; Flaccus, A.; Fischer, A. & Spiess, J. (1999)
European Journal of Biochemistry, 259(1-2) pp. 55-62. DOI: https://doi.org/10.1046/j.1432-1327.1999.00007.x
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- Authors
- Sydow, S.; Flaccus, A; Fischer, Andre ; Spiess, Joachim
- Abstract
- The role of the fourth extracellular loop (e4) of rat corticotropin-releasing factor (CRF) receptor, type 1, in ligand binding was investigated using chimeric receptor molecules. e4 of CRF receptor, type I, was replaced by the corresponding domains of two other G protein-coupled receptors, the rat glucagon receptor or the human pituitary adenylate cyclase activating polypeptide (PACAP) receptor. Both chimeras were transported properly to the cell membranes of transfected chinese hamster ovary cells as indicated by immunocytochemical analysis. Ovine CRF (oCRF) was bound specifically, but with low affinity (K-d = 2-5 mu M). Cyclic AMP was not accumulated intracellularly in response to increasing concentrations of oCRF. Based on these data, it is concluded that e4 of rat CRF receptor, type 1, is involved in ligand binding. To confirm the importance of e4 in binding CRF, three negatively charged amino acids of e4, Glu336, Asp337 and Glu338, were replaced by Gin, Asn and Gin, respectively. No effect on ligand binding and cyclic AMP accumulation was observed (K-d = 5 nM; EC50 = 1 5 nM). However, when Tyr346, Phe347 and Asn348 of e4 were changed to three alanine residues, ligand binding affinity as well as efficacy in cyclic AMP accumulation were significantly decreased (K-d = 64 nM; EC50 = 32 nM).
- Issue Date
- 1999
- Publisher
- Blackwell Science Ltd
- Journal
- European Journal of Biochemistry
- ISSN
- 0014-2956