Calcitonin Receptor N-Glycosylation Enhances Peptide Hormone Affinity by Controlling Receptor Dynamics
Calcitonin Receptor N-Glycosylation Enhances Peptide Hormone Affinity by Controlling Receptor Dynamics
复制标题
降钙素受体 N-糖基化通过控制受体动力学增强肽激素亲和力
DOI：
10.1096/fasebj.2020.34.s1.00704
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Lee S
中科院分区：
文献类型：
--
作者：
Lee S
B类G蛋白偶联受体(GPCR)降钙素受体(CTR)是治疗骨质疏松症和糖尿病的药物靶点。天冬酰胺130胞外区(ECD)的N-糖基化增强了降钙素激素与近端GlcNAc残基的亲和力，通过未知的机制介导这一作用。在这里，我们提出了鲑鱼降钙素结合的GlcNAc-CTR ECD在1.78和2.85分辨率下的两种晶体结构，并分析了其作用机制。N130 GlcNAc不接触荷尔蒙。令人惊讶的是，这些结构与激素结合、无N-糖链的ECD结构几乎相同，这表明GlcNAc可能影响静态晶体快照中没有观察到的CTR动力学。氢-氚交换质谱仪和分子动力学模拟表明，糖基化稳定了N130GlcNAc附近的β-折叠和多肽结合部位附近的N-末端α-螺旋，同时增加了多肽结合部位转塔环的灵活性。N-糖基化引起的这些变化增加了配体的开启速率，降低了配体的关闭速率。这种多糖效应延伸到RAMP-CTR胰淀素受体复合体，在相关的CGRP受体中也是保守的。这些结果表明，N-糖基化可以通过改变受体动力学来调节GPCR的功能。
The class B G protein-coupled receptor (GPCR) calcitonin receptor (CTR) is a drug target for osteoporosis and diabetes. N-glycosylation of asparagine 130 in its extracellular domain (ECD) enhances calcitonin hormone affinity with the proximal GlcNAc residue mediating this effect through an unknown mechanism. Here, we present two crystal structures of salmon calcitonin-bound, GlcNAc-bearing CTR ECD at 1.78 and 2.85 Å resolutions and analyze the mechanism of the glycan effect. The N130 GlcNAc does not contact the hormone. Surprisingly, the structures are nearly identical to a structure of hormone-bound, N-glycan-free ECD, which suggested that the GlcNAc might affect CTR dynamics not observed in the static crystallographic snapshots. Hydrogen-deuterium exchange mass spectrometry and molecular dynamics simulations revealed that glycosylation stabilized a β-sheet adjacent to the N130 GlcNAc and the N-terminal α-helix near the peptide-binding site while increasing flexibility of the peptide-binding site turret loop. These changes due to N-glycosylation increased the ligand on-rate and decreased its off-rate. The glycan effect extended to RAMP-CTR amylin receptor complexes and was also conserved in the related CGRP receptor. These results reveal that N-glycosylation can modulate GPCR function by altering receptor dynamics.