GluK1 配体结合结构域与凯恩酸盐和全跨度正异构调节剂 BPAM538 的晶体结构。

IF 3 3区 生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY
Yasmin Bay , Federico Javier Miguez Cabello , Chloe C. Koens , Stine M. Frantsen , Darryl S. Pickering , Karla Frydenvang , Pierre Francotte , Bernard Pirotte , Anders S. Kristensen , Derek Bowie , Jette Sandholm Kastrup
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引用次数: 0

摘要

凯恩酸盐受体通过突触前机制介导突触后兴奋性神经递质并调节抑制性神经递质 GABA 的释放,在中枢神经系统中发挥着重要作用。迄今为止,通过 X 射线晶体学方法确定的凯纳特受体亚基 GluK1 的配体结合域(LBD)与正异位调节剂复合物的结构只有三个,均属于第二类调节剂。在这里,我们报告了 GluK1-LBD 与凯恩酸盐和 BPAM538 复合物的高分辨率结构。一个 BPAM538 分子与 GluK1 二聚体界面结合,从而同时占据了两个异位结合位点。BPAM538 能稳定受体的活性构象,受体只需发生微小的构象变化。利用钙敏感荧光测定法,在 100 μM BPAM538 存在的情况下,GluK1(Q)b 的凯氏反应(100 μM)会增强 5 倍,而 GluK2(VCQ)a 则不会增强。通过对从 HEK293 细胞中切除的外向型贴片进行电生理学记录,BPAM538 使与 NETO2 共同表达的 GluK1(Q)b 对快速施加 10 mM L-谷氨酸的峰值反应增加了 130 ± 20 %,并使根据稳态/峰值反应比确定的脱敏率从 23 ± 2 % 降至 90 ± 4 %。根据 GluK1(Q)b 的剂量-反应关系实验,BPAM538 的 EC50 值估计为 57.5 ± 29.2 μM。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Crystal structure of the GluK1 ligand-binding domain with kainate and the full-spanning positive allosteric modulator BPAM538

Crystal structure of the GluK1 ligand-binding domain with kainate and the full-spanning positive allosteric modulator BPAM538

Kainate receptors play an important role in the central nervous system by mediating postsynaptic excitatory neurotransmission and modulating the release of the inhibitory neurotransmitter GABA through a presynaptic mechanism. To date, only three structures of the ligand-binding domain (LBD) of the kainate receptor subunit GluK1 in complex with positive allosteric modulators have been determined by X-ray crystallography, all belonging to class II modulators. Here, we report a high-resolution structure of GluK1-LBD in complex with kainate and BPAM538, which belongs to the full-spanning class III. One BPAM538 molecule binds at the GluK1 dimer interface, thereby occupying two allosteric binding sites simultaneously. BPAM538 stabilizes the active receptor conformation with only minor conformational changes being introduced to the receptor. Using a calcium-sensitive fluorescence-based assay, a 5-fold potentiation of the kainate response (100 μM) was observed in presence of 100 μM BPAM538 at GluK1(Q)b, whereas no potentiation was observed at GluK2(VCQ)a. Using electrophysiology recordings of outside-out patches excised from HEK293 cells, BPAM538 increased the peak response of GluK1(Q)b co-expressed with NETO2 to rapid application of 10 mM L-glutamate with 130 ± 20 %, and decreased desensitization determined as the steady-state/peak response ratio from 23 ± 2 % to 90 ± 4 %. Based on dose–response relationship experiments on GluK1(Q)b the EC50 of BPAM538 was estimated to be 58 ± 29 μM.

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来源期刊
Journal of structural biology
Journal of structural biology 生物-生化与分子生物学
CiteScore
6.30
自引率
3.30%
发文量
88
审稿时长
65 days
期刊介绍: Journal of Structural Biology (JSB) has an open access mirror journal, the Journal of Structural Biology: X (JSBX), sharing the same aims and scope, editorial team, submission system and rigorous peer review. Since both journals share the same editorial system, you may submit your manuscript via either journal homepage. You will be prompted during submission (and revision) to choose in which to publish your article. The editors and reviewers are not aware of the choice you made until the article has been published online. JSB and JSBX publish papers dealing with the structural analysis of living material at every level of organization by all methods that lead to an understanding of biological function in terms of molecular and supermolecular structure. Techniques covered include: • Light microscopy including confocal microscopy • All types of electron microscopy • X-ray diffraction • Nuclear magnetic resonance • Scanning force microscopy, scanning probe microscopy, and tunneling microscopy • Digital image processing • Computational insights into structure
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