Mesbahi-Vasey Samaneh, Veras Lea, Yonkunas Michael, Johnson Jon W, Kurnikova Maria G
Department of Chemistry, Carnegie Mellon University, Pittsburgh, Pennsylvania, United States of America.
Department of Neuroscience and Center for Neuroscience, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
PLoS One. 2017 Jun 5;12(6):e0177686. doi: 10.1371/journal.pone.0177686. eCollection 2017.
N-methyl-d-aspartate receptors (NMDARs) are members of the ionotropic glutamate receptor family that mediate excitatory synaptic transmission in the central nervous system. The channels of NMDARs are permeable to Ca2+ but blocked by Mg2+, distinctive properties that underlie essential brain processes such as induction of synaptic plasticity. However, due to limited structural information about the NMDAR transmembrane ion channel forming domain, the mechanism of divalent cation permeation and block is understood poorly. In this paper we developed an atomistic model of the transmembrane domain (TMD) of NMDARs composed of GluN1 and GluN2A subunits (GluN1/2A receptors). The model was generated using (a) a homology model based on the structure of the NaK channel and a partially resolved structure of an AMPA receptor (AMPAR), and (b) a partially resolved X-ray structure of GluN1/2B NMDARs. Refinement and extensive Molecular Dynamics (MD) simulations of the NMDAR TMD model were performed in explicit lipid bilayer membrane and water. Targeted MD with simulated annealing was introduced to promote structure refinement. Putative positions of the Mg2+ and Ca2+ ions in the ion channel divalent cation binding site are proposed. Differences in the structural and dynamic behavior of the channel protein in the presence of Mg2+ or Ca2+ are analyzed. NMDAR protein conformational flexibility was similar with no ion bound to the divalent cation binding site and with Ca2+ bound, whereas Mg2+ binding reduced protein fluctuations. While bound at the binding site both ions retained their preferred ligand coordination numbers: 6 for Mg2+, and 7-8 for Ca2+. Four asparagine side chain oxygens, a back-bone oxygen, and a water molecule participated in binding a Mg2+ ion. The Ca2+ ion first coordination shell ligands typically included four to five side-chain oxygen atoms of the binding site asparagine residues, two water molecules and zero to two backbone oxygens of the GluN2B subunits. These results demonstrate the importance of high-resolution channel structures for elucidation of mechanisms of NMDAR permeation and block.
N-甲基-D-天冬氨酸受体（NMDARs）是离子型谷氨酸受体家族的成员，介导中枢神经系统中的兴奋性突触传递。NMDARs的通道对Ca2+通透，但被Mg2+阻断，这些独特特性是诸如突触可塑性诱导等基本脑过程的基础。然而，由于关于NMDAR跨膜离子通道形成结构域的结构信息有限，二价阳离子通透和阻断的机制了解甚少。在本文中，我们构建了由GluN1和GluN2A亚基组成的NMDARs跨膜结构域（TMD）的原子模型（GluN1/2A受体）。该模型是使用（a）基于NaK通道结构和AMPA受体（AMPAR）部分解析结构的同源模型，以及（b）GluN1/2B NMDARs的部分解析X射线结构生成的。在明确的脂质双分子层膜和水中对NMDAR TMD模型进行了优化和广泛的分子动力学（MD）模拟。引入了带有模拟退火的靶向MD以促进结构优化。提出了Mg2+和Ca2+离子在离子通道二价阳离子结合位点的假定位置。分析了在存在Mg2+或Ca2+的情况下通道蛋白的结构和动力学行为的差异。NMDAR蛋白构象灵活性在没有离子结合到二价阳离子结合位点和有Ca2+结合时相似，而Mg2+结合减少了蛋白波动。当结合在结合位点时，两种离子都保持其优选的配体配位数：Mg2+为6，Ca2+为7 - 8。四个天冬酰胺侧链氧原子、一个主链氧原子和一个水分子参与结合一个Mg2+离子。Ca2+离子的第一配位层配体通常包括结合位点天冬酰胺残基的四到五个侧链氧原子、两个水分子和GluN2B亚基的零到两个主链氧原子。这些结果证明了高分辨率通道结构对于阐明NMDAR通透和阻断机制的重要性。
