Molecular determinants of non-competitive antagonist binding to the mouse GPRC6A receptor
Molecular determinants of non-competitive antagonist binding to the mouse GPRC6A receptor
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DOI：
10.1016/j.ceca.2009.09.004
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发表时间：
2009-11-01
期刊：
影响因子：
4
通讯作者：
Ruat, Martial
中科院分区：
文献类型：
--
作者：
Faure, Helene;Gorojankina, Tatiana;Ruat, Martial
GPRC 6A与Ca 2+敏感受体（CaSR）具有高度的序列同源性。在这里，我们报告，钙模拟Calindol和calcilytic NPS 2143拮抗增加磷酸肌醇引起L-鸟氨酸诱导的激活小鼠GPRC 6A后，瞬时共表达G α（qG 66 D）在HEK 293细胞。calcilytic Calhex 231没有调节这种反应。构建了GPRC 6A七个跨膜结构域（TM）的三维模型。它被用来确定七个残基严格保守的CaSR和GPRC 6A变构结合口袋，以前证明与calcilytics或calcimimetics相互作用。突变F666 A（3.32）、F670 A（3.36）、W797 A（6.48）导致L-鸟氨酸激活GPRC 6A突变体的能力丧失。F800 A（6.51）突变体不涉及Calindol或Calindol 2143识别。E816 Q（7.39）突变导致Calindol拮抗剂活性的丧失，但对NPS 2143抑制反应没有影响。总之，这些数据表明Calindol主要通过H-键锚定到TM 7中的E816（7.19），并突出了在CaSR和GPRC 6A变构结合口袋水平上的重要局部差异。我们已经确定了GPRC 6A的第一个拮抗剂，可以代表新的工具来分析GPRC 6A的功能，并作为开发更具体的调节剂的化学线索。(C)2009爱思唯尔有限公司保留所有权利。
GPRC6A displays high sequence homology to the Ca2+-sensing receptor (CaSR). Here we report that the calcimimetic Calindol and the calcilytic NPS2143 antagonize increases in inositol phosphate elicited by L-ornithine-induced activation of mouse GPRC6A after transient coexpression with G alpha(qG66D) in HEK293 cells. The calcilytic Calhex 231 did not modulate this response. A three-dimensional model of the GPRC6A seven transmembrane domains (TMs) was constructed. It was used to identify seven residues strictly conserved within the CaSR and GPRC6A allosteric binding pockets, and previously demonstrated to interact with calcilytics or calcimimetics. The mutations F666A(3.32), F670A(3.36), W797A(6.48) caused a loss Of L-ornithine ability to activate GPRC6A mutants. The F800A(6.51) mutant was not implicated in either Calindol or NPS 2143 recognition. The E816Q(7.39) mutation led to a loss of Calindol antagonist activity but was without effecton NPS2143 inhibitory response. In summary, these data suggest that Calindol is primarily anchored through an H-bond to E816(7.19) in TM7 and highlight important local differences at the level of the CaSR and GPRC6A allosteric binding pockets. We have identified the first antagonists of GPRC6A that could represent new tools to analyze GPRC6A functions and serve as chemical leads for the development of more specific modulators. (C) 2009 Elsevier Ltd. All rights reserved.