Structural mobility of the extracellular ligand-binding core of an ionotropic glutamate receptor. Analysis of NMR relaxation dynamics
Structural mobility of the extracellular ligand-binding core of an ionotropic glutamate receptor. Analysis of NMR relaxation dynamics
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DOI：
10.1021/bi026010p
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发表时间：
2002-08-20
期刊：
影响因子：
2.9
通讯作者：
Oswald, RE
中科院分区：
文献类型：
--
作者：
McFeeters, RL;Oswald, RE
离子型谷氨酸受体在多种神经元突起中发挥重要作用，并与多种神经退行性疾病有关。GluR2亚型的胞外配体结合核心(S1S2)可以在细菌中以可溶的单体蛋白的形式表达，其结合特性与完整受体的结合特性基本相同。这种蛋白质的晶体结构已经在各种激动剂和拮抗剂的存在和不存在的情况下被确定[Armstrong，N.，Sun，Y.，Chen，G.Q，and Gouaux，E.(1998)Natural 395,913-917；Armstrong，N.，and Gouaux，E.(2000)Neuron 28,165-181]。该蛋白由两个叶组成，其中S1片段组成叶1的大部分，S2片段组成叶2的大部分。已经假设了配体结合时结构域的关闭，但还没有研究结构域内运动的细节。本文利用N-15纵向(T-1)和横向(T-2)弛豫测量以及500和600 MHz的{H-1}-N-15核Overhaser效应研究了谷氨酸与GluR2配体结合核心的骨架运动。激动剂结合袋中的残留物表现出两种主要的运动类型。那些接触配体谷氨酸的α-取代基的分子表现出最小的内部运动，而那些接触伽马成分的分子表现出交换动力学，表明结合口袋有两个动态不同的部分。此外，在叶1和叶2之间的跨域连接子中的两个残基显示了交换，这为先前提出的结构域关闭假说提供了新的见解。最后，螺旋F的协同运动为配体解离提供了一条途径，而不需要重新打开结构域。
Ionotropic glutamate receptors play important roles in a variety of neuronal processes and have been implicated in multiple neurodegenerative diseases. The extracellular ligand-binding (S1S2) core of the GluR2 subtype can be expressed in bacteria as a soluble, monomeric protein with binding properties essentially identical to those of the intact receptor. The crystal structure of this protein has been determined in the presence and absence of various agonists and antagonists [Armstrong, N., Sun, Y., Chen, G. Q., and Gouaux, E. (1998) Nature 395, 913-917; Armstrong, N., and Gouaux, E. (2000) Neuron 28, 165-181]. The protein consists of two lobes, with the S1 segment composing the majority of lobe 1 and the S2 segment composing most of lobe 2. A domain closure upon ligand binding has been postulated, but details of intradomain motions have not been investigated. In this paper, the backbone motions of the ligand-binding core of GluR2 bound to glutamate were studied using N-15 longitudinal (T-1) and transverse (T-2) relaxation measurements as well as {H-1}-N-15 nuclear Overhauser effects at 500 and 600 MHz. Residues in the agonist-binding pocket exhibited two main classes of motion. Those contacting the alpha-substituents of the ligand glutamate exhibited minimal internal motion, while those contacting the gamma-constituents exhibited exchange dynamics, indicating two dynamically distinct portions of the binding pocket. Also, two residues in transdomain linkers between lobes 1 and 2 show exchange, lending new insight into the previously proposed domain closure hypothesis. Finally, concerted motion of helix F suggests a pathway for ligand dissociation without the necessity of domain reopening.