Perrin M H, Sutton S, Bain D L, Berggren W T, Vale W W
The Clayton Foundation Laboratories for Peptide Biology, The Salk Institute, La Jolla, California 92037-1099, USA.
Endocrinology. 1998 Feb;139(2):566-70. doi: 10.1210/endo.139.2.5757.
The CRF receptors are members of a 7-transmembrane receptor family that includes GH-releasing hormone (GRF), calcitonin, vasoactive intestinal peptide (VIP), secretin, and PTH receptors. To determine the structural features of the CRF receptor that may influence ligand recognition, a series of mutant receptors was analyzed for binding to astressin, a CRF antagonist, and to urocortin, a CRF agonist. Mutant receptors included chimeras between the CRF-R1 and GRF-R or Activin IIB-R, a single membrane spanning receptor serine/threonine kinase. Binding to the mutant receptors was assessed using 125I-[DTyr1] astressin (Ast*) and 125I-[Tyr0]-rat urocortin (Ucn*). There was no binding to a chimeric receptor in which the first extracellular domain (E1c) (i.e. the N-terminal region) of the CRF-R1 was replaced by that of the GRF-R. The complementary chimera in which E1 domain of the GRF-R was replaced by that of the CRF-R1 bound astressin and urocortin with Ki values approximately 10 nM, compared with inhibitory binding dissociation constant (Ki) values of approximately 2-4 nM for the wild-type CRF-R1. The chimera in which E1 of the activin IIB receptor was replaced by E1 of the CRF-R1 bound astressin with a Ki approximately 4 nM. A chimera in which both the first and fourth extracellular domains of the CRF-R1 replaced the corresponding domains of the GRF-R bound astressin with Ki approximately 4 nM and urocortin with a Ki approximately 2 nM. A chimera in which all four extracellular domains of the CRF receptor replaced those of the GRF-R bound astressin and urocortin with Ki values approximately 4 nM and approximately 1 nM, respectively. In conclusion, the major determinants for high affinity binding of CRF agonists and antagonists to CRF-R1 are found in the first extracellular domain of the receptor.
促肾上腺皮质激素释放因子（CRF）受体是一个7次跨膜受体家族的成员，该家族还包括生长激素释放激素（GRF）、降钙素、血管活性肠肽（VIP）、促胰液素和甲状旁腺激素受体。为了确定可能影响配体识别的CRF受体的结构特征，分析了一系列突变受体与CRF拮抗剂阿斯特辛和CRF激动剂尿皮质素的结合情况。突变受体包括CRF-R1与GRF-R或激活素IIB-R（一种单次跨膜受体丝氨酸/苏氨酸激酶）之间的嵌合体。使用125I-[DTyr1]阿斯特辛（Ast*）和125I-[Tyr0]-大鼠尿皮质素（Ucn*）评估与突变受体的结合。CRF-R1的第一个细胞外结构域（E1c，即N端区域）被GRF-R的相应结构域取代的嵌合受体没有结合。将GRF-R的E1结构域替换为CRF-R1的互补嵌合体与阿斯特辛和尿皮质素结合，其抑制性结合解离常数（Ki）值约为10 nM，而野生型CRF-R1的Ki值约为2-4 nM。激活素IIB受体的E1被CRF-R1的E1取代的嵌合体与阿斯特辛结合，Ki约为4 nM。CRF-R1的第一个和第四个细胞外结构域均被GRF-R的相应结构域取代的嵌合体与阿斯特辛结合，Ki约为4 nM，与尿皮质素结合，Ki约为2 nM。CRF受体的所有四个细胞外结构域均被GRF-R的相应结构域取代的嵌合体与阿斯特辛和尿皮质素结合，Ki值分别约为4 nM和约1 nM。总之，CRF激动剂和拮抗剂与CRF-R1高亲和力结合的主要决定因素存在于受体的第一个细胞外结构域中。
