Five Amino Acids of the Xenopus laevis CRF (Corticotropin-Releasing Factor) Type 2 Receptor Mediate Differential Binding of CRF Ligands in Comparison with Its Human Counterpart

• Published in: Molecular pharmacology 2002-05, Vol.61 (5), p.1132-1139
• Main Authors: Dautzenberg, Frank M, Higelin, Jacqueline, Brauns, Olaf, Butscha, Brigitte, Hauger, Richard L
• Format: Article
• Language: English
• Subjects: Amino Acid Motifs; Amino Acid Sequence; Animals; Binding Sites; Cells, Cultured; Corticotropin-Releasing Hormone - metabolism; Cyclic AMP - metabolism; Humans; Molecular Sequence Data; Point Mutation; Protein Conformation; Receptors, Corticotropin-Releasing Hormone - chemistry; Receptors, Corticotropin-Releasing Hormone - genetics; Receptors, Corticotropin-Releasing Hormone - metabolism; Transfection; Xenopus laevis - metabolism
• ISSN: 0026-895X; 1521-0111
• DOI: 10.1124/mol.61.5.1132

The ligand selectivity of human (hCRF 2A ) and Xenopus laevis (xCRF 2 ) forms of the corticotropin-releasing factor type 2 (CRF 2 ) receptor differs. The purpose of this study was to identify amino acids in these two CRF 2 receptors conferring these differences. An amino acid triplet in the third extracellular domain (Asp 262 Leu 263 Val 264 in hCRF 2A or Lys 264 Tyr 265 Ile 266 in xCRF 2 ) was found to diverge between both receptors. When binding and signaling characteristics of receptor mutants hR2KYI and xR2DLV were assessed, the tri-amino acid motif replacement produced receptors with binding properties resembling the xCRF 2 receptor. The converse mutation created a mutant receptor with a binding pharmacology identical to the profile of the hCRF 2A receptor. This effect was most notable for xR2DLV, which possessed a binding affinity for astressin ∼15-fold greater for astressin than sauvagine. In contrast, the binding profiles of the hCRF 2A receptor and hR2KYI did not differ. These data indicate that another domain of the xCRF 2 receptor mediated low-affinity binding of astressin. Two amino acids in the first extracellular domain differ in xCRF 2 (Asp 69 Ser 70 ) and hCRF 2A (Glu 66 Tyr 67 ) receptors. The hCRF 2A receptor mutant (hR2DS-KYI) bound astressin with a low affinity indistinguishable from the xCRF 2 receptor. Therefore, these data demonstrate that ligand selectivity differences between amphibian and human forms of the CRF 2A receptor are governed by these two motifs of the extracellular domains of the xCRF 2 receptor.