Separation of Domain Contacts Is Required for Heterotetrameric Assembly of Functional NMDA Receptors
Separation of Domain Contacts Is Required for Heterotetrameric Assembly of Functional NMDA Receptors
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DOI：
10.1523/jneurosci.6041-10.2011
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发表时间：
2011-03-09
影响因子：
5.3
通讯作者：
Nakagawa, Terunaga
中科院分区：
文献类型：
--
作者：
Farina, Anthony N.;Blain, Katherine Y.;Nakagawa, Terunaga
准确了解NMDA受体亚基组装过程对于理解受体结构和通道功能的潜在机制至关重要。由于NMDA-Rs是必需的异源四聚体，需要GluN1亚基，因此研究GluN1和GluN2型亚基如何聚集成四聚体是至关重要的。通过结合细胞生物学、生物化学、单粒子电子显微镜和x射线晶体学等方法，我们报道了GluN1突变亚基在受体成熟过程中存在缺陷的机制和表型。GluN1的n端结构域（NTD）的T110A突变促进GluN1和GluN2的NTDs之间的异二聚，而相邻残基的Y109C突变稳定GluN1的NTD的同二聚体。GluN1的NTD晶体结构揭示了这些突变体生化特性的机制。这些突变对异质NMDA-Rs成熟的影响使用受体运输试验进行了研究。我们的研究结果表明，GluN1亚基的NTDs最初形成同型二聚体，随后的二聚体解离对于形成含有GluN2亚基的异四聚体NMDA-Rs至关重要，这为受体组装确定了分子决定因素。GluN1二聚体NTD的结构域排列在各离子型谷氨酸受体中是独一无二的，这预示着NMDA-Rs在NTD周围的结构和机制与同源AMPA和盐酸盐受体不同。
The precise knowledge of the subunit assembly process of NMDA receptors (NMDA-Rs) is essential to understand the receptor architecture and underlying mechanism of channel function. Because NMDA-Rs are obligatory heterotetramers requiring the GluN1 subunit, it is critical to investigate how GluN1 and GluN2 type subunits coassemble into tetramers. By combining approaches in cell biology, biochemistry, single particle electron microscopy, and x-ray crystallography, we report the mechanisms and phenotypes of mutant GluN1 subunits that are defective in receptor maturation. The T110A mutation in the N-terminal domain (NTD) of the GluN1 promotes heterodimerization between the NTDs of GluN1 and GluN2, whereas the Y109C mutation in the adjacent residue stabilizes the homodimer of the NTD of GluN1. The crystal structure of the NTD of GluN1 revealed the mechanism underlying the biochemical properties of these mutants. Effects of these mutations on the maturation of heteromeric NMDA-Rs were investigated using a receptor trafficking assay. Our results suggest that the NTDs of the GluN1 subunit initially form homodimers and the subsequent dimer dissociation is critical for forming heterotetrameric NMDA-Rs containing GluN2 subunits, defining a molecular determinant for receptor assembly. The domain arrangement of the dimeric NTD of GluN1 is unique among the ionotropic glutamate receptors and predicts that the structure and mechanism around the NTDs of NMDA-Rs are different from those of the homologous AMPA and kainate receptors.