Receptor activity-modifying protein 1 determines the species selectivity of non-peptide CGRP receptor antagonists
Receptor activity-modifying protein 1 determines the species selectivity of non-peptide CGRP receptor antagonists
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DOI：
10.1074/jbc.m109661200
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发表时间：
2002-04-19
影响因子：
4.8
通讯作者：
Kane, SA
中科院分区：
文献类型：
--
作者：
Mallee, JJ;Salvatore, CA;Kane, SA
异二聚体CGRP受体需要降钙素受体样受体（CRLR）和称为受体活性修饰蛋白（RAMP）1的辅助蛋白的共表达（McLatchie，L. M.，Fraser，N. J.，美因河J.，怀斯，A.，布朗，J.，汤普森，N.，索拉里河李，M。G.，和Foord，S. M.（1998）Nature 393，333-339）。几种非肽CGRP受体拮抗剂已显示出显著的物种选择性，对人CGRP受体的亲和力比对来自其它物种的受体高> 100倍（Doods，H.，Hallermayer，G.，吴，D.，Entzeroth，M.，Rudolf，K.，Engel，W.，和Eberlein，W.（2000）Br. J. Pharmacol. 129，420-423; Edvinsson，L.，Sams，A.，詹森-奥尔森岛Tajti，J.，凯恩，S。一、拉特利奇河Z.，Koblan，K.美国，希尔河，巴西-地G.，Longmore，J.（2001）Ear. J. Pharmacol. 415，39-44）。这一观察结果提供了绘制由BIBN 4096 BS和截短的类似物化合物1和2表现出的受体亲和力的决定因素的机会。所有三种化合物对人受体（人CRLR/人RAMP 1）的亲和力均高于对大鼠受体（大鼠CRLR/大鼠RAMP 1）的亲和力。我们现在已经证明，这种物种的选择性是专门由RAMP 1。通过产生重组人/大鼠CRLR/RAMP 1受体，我们证明了人CRLR与大鼠RAMP 1的共表达产生了大鼠受体药理学，反之亦然。此外，与大鼠/人RAMP 1嵌合体和定点突变体，我们已经确定了一个单一的氨基酸在位置74的RAMP 1，调节小分子拮抗剂的亲和力CRLR/RAMP 1。用色氨酸（人受体中的同源氨基酸）取代大鼠RAMP 1中的赖氨酸74导致拮抗剂亲和力增加大于或等于100倍，与人受体的Ki值相似。这些观察结果表明，CGRP受体的小分子拮抗剂亲和力的重要决定因素驻留在RAMP 1的细胞外区域，并提供证据表明，这种受体辅助蛋白可能参与拮抗剂结合。
The heterodimeric CGRP receptor requires co-expression of calcitonin receptor-like receptor (CRLR) and an accessory protein called receptor activity-modifying protein (RAMP) 1 (McLatchie, L. M., Fraser, N. J., Main, M. J., Wise, A., Brown, J., Thompson, N., Solari, R., Lee, M. G., and Foord, S. M. (1998) Nature 393, 333-339). Several non-peptide CGRP receptor antagonists have been shown to exhibit marked species selectivity, with > 100-fold higher affinities for the human CGRP receptor than for receptors from other species (Doods, H., Hallermayer, G., Wu, D., Entzeroth, M., Rudolf, K., Engel, W., and Eberlein, W. (2000) Br. J. Pharmacol. 129, 420-423; Edvinsson, L., Sams, A., Jansen-Olesen, I., Tajti, J., Kane, S. A., Rutledge, R. Z., Koblan, K. S., Hill, R. G., and Longmore, J. (2001) Ear. J. Pharmacol. 415, 39-44). This observation provided an opportunity to map the determinants of receptor affinity exhibited by BIBN4096BS and the truncated analogs, Compounds 1 and 2. All three compounds exhibited higher affinity for the human receptor, human CRLR/human RAMP1, than for the rat receptor, rat CRLR/rat RAMP1. We have now demonstrated that this species selectivity was directed exclusively by RAMP1. By generating recombinant human/rat CRLR/RAMP1 receptors, we demonstrated that co-expression of human CRLR with rat RAMP1 produced rat receptor pharmacology, and vice versa. Moreover, with rat/human RAMP1 chimeras and site-directed mutants, we have identified a single amino acid at position 74 of RAMP1 that modulates the affinity of small molecule antagonists for CRLR/RAMP1. Replacement of lysine 74 in rat RAMP1 with tryptophan (the homologous amino acid in the human receptor) resulted in a greater than or equal to100-fold increase in antagonist affinities, similar to the K-i values for the human receptor. These observations suggest that important determinants of small molecule antagonist affinity for the CGRP receptor reside within the extracellular region of RAMP1 and provide evidence that this receptor accessory protein may participate in antagonist binding.