The second intracellular loop of the calcitonin gene-related peptide receptor provides molecular determinants for signal transduction and cell surface expression
The second intracellular loop of the calcitonin gene-related peptide receptor provides molecular determinants for signal transduction and cell surface expression
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DOI：
10.1074/jbc.m510064200
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发表时间：
2006-01-20
影响因子：
4.8
通讯作者：
Poyner, DR
中科院分区：
文献类型：
--
作者：
Conner, AC;Simms, J;Poyner, DR
降钙素基因相关肽（CGRP）受体是家族B G蛋白偶联受体、降钙素受体样受体（CGRP）和辅助蛋白受体活性修饰蛋白1的异源二聚体。它与G（s）偶联，但尚不清楚哪些细胞内环介导了这一过程。我们已经确定了这个环的边界的基础上的相对位置和长度的质膜跨膜区3和4。已通过将所有残基系统性诱变为丙氨酸、测量cAMP积累、CGRP亲和力和受体表达来分析该环。与视紫红质不同，CGRP受体的ICL 2在激动剂相互作用后的构象转换中起作用。His-216和Lys-227是CGRP诱导的cAMP反应所必需的。（H216 A）突变体受体的作用是由于对细胞表面转运或表面稳定性的破坏。相比之下，（K227 A）β具有野生型表达和激动剂亲和力，表明直接破坏CGRP受体的下游信号转导机制。建模表明，在受体激活过程中，环的位置发生了显着的变化，暴露了潜在的G蛋白结合口袋。Lys-227改变位置以指向口袋，可能使其与结合的G蛋白相互作用。His-216在牛视紫红质中占据与Tyr-136相似的位置，后者是后者受体的DRY基序的一部分。这是第一次全面分析降钙素家族的G蛋白偶联受体内的整个细胞内环。这些数据有助于确定CGRP受体和B家族G蛋白偶联受体的结构和功能特征。
The calcitonin gene-related peptide ( CGRP) receptor is a heterodimer of a family B G-protein-coupled receptor, calcitonin receptor-like receptor (CLR), and the accessory protein receptor activity modifying protein 1. It couples to G(s), but it is not known which intracellular loops mediate this. We have identified the boundaries of this loop based on the relative position and length of the juxtamembrane transmembrane regions 3 and 4. The loop has been analyzed by systematic mutagenesis of all residues to alanine, measuring cAMP accumulation, CGRP affinity, and receptor expression. Unlike rhodopsin, ICL2 of the CGRP receptor plays a part in the conformational switch after agonist interaction. His-216 and Lys-227 were essential for a functional CGRP-induced cAMP response. The effect of (H216A) CLR is due to a disruption to the cell surface transport or surface stability of the mutant receptor. In contrast, (K227A) CLR had wild-type expression and agonist affinity, suggesting a direct disruption to the downstream signal transduction mechanism of the CGRP receptor. Modeling suggests that the loop undergoes a significant shift in position during receptor activation, exposing a potential G-protein binding pocket. Lys-227 changes position to point into the pocket, potentially allowing it to interact with bound G-proteins. His-216 occupies a position similar to that of Tyr-136 in bovine rhodopsin, part of the DRY motif of the latter receptor. This is the first comprehensive analysis of an entire intracellular loop within the calcitonin family of G-protein-coupled receptor. These data help to define the structural and functional characteristics of the CGRP-receptor and of family B G-protein-coupled receptors in general.