Purification and cryo-EM structure determination of Arabidopsis thaliana GLR3.4.
Purification and cryo-EM structure determination of Arabidopsis thaliana GLR3.4.
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DOI：
10.1016/j.xpro.2021.100855
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发表时间：
2021-12-17
期刊：
影响因子：
--
通讯作者：
Sobolevsky AI
中科院分区：
文献类型：
--
作者：
Gangwar SP;Green MN;Yelshanskaya MV;Sobolevsky AI
嗜离子性谷氨酸受体（iGluRs）是一种在中枢神经系统中起重要作用的配体门控离子通道。iGluR同源物被称为谷氨酸受体样通道（GLRs），已在植物中发现。研究iGluRs和GLRs之间的结构和功能关系受到GLR蛋白表达、纯化和结构表征的限制。在此，我们提供了拟南芥GLR3.4 （AtGLR3.4）在哺乳动物细胞系中的表达和纯化的详细方案，并通过低温电子显微镜（cro - em）检测其结构。有关使用和执行本协议的完整细节，请参阅。我们为拟南芥GLR3.4蛋白的表达和纯化提供了详细的方案，该蛋白可用于冷冻电镜结构检测。使用BioRender （https://biorender.com/）创建图像。植物谷氨酸受体样通道（GLR）在HEK细胞中的表达纯化方案产生纯单分散四聚体GLR蛋白纯化蛋白经低温电镜分析产生高分辨率GLR结构离子化谷氨酸受体（iGluRs）是配体门控离子通道，在中枢神经系统中起重要作用。iGluR同源物被称为谷氨酸受体样通道（GLRs），已在植物中发现。研究iGluRs和GLRs之间的结构和功能关系受到GLR蛋白表达、纯化和结构表征的限制。在此，我们提供了拟南芥GLR3.4 （AtGLR3.4）在哺乳动物细胞系中的表达和纯化的详细方案，并通过低温电子显微镜（cro - em）检测其结构。
Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that play crucial roles in the central nervous system. iGluR homologs, termed glutamate receptor-like channels (GLRs), have been found in plants. Investigating the structural and functional relationship between iGluRs and GLRs was limited by GLR protein expression, purification, and structural characterization. Here, we provide a detailed protocol for Arabidopsis thaliana GLR3.4 (AtGLR3.4) expression in a mammalian cell line and purification for structure determination by cryogenic electron microscopy (cryo-EM). For the complete details on the use and execution of this protocol, please refer to. We provide a detailed protocol for expression and purification of Arabidopsis thaliana GLR3.4 protein that can be used for structure determination by cryo-EM. The image has been created using BioRender (https://biorender.com/). Protocol for expression of plant glutamate receptor-like channel (GLR) in HEK cells Purification protocol that yields pure monodisperse tetrameric GLR protein Purified protein subjected to cryo-EM analysis produces high-resolution GLR structure Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that play crucial roles in the central nervous system. iGluR homologs, termed glutamate receptor-like channels (GLRs), have been found in plants. Investigating the structural and functional relationship between iGluRs and GLRs was limited by GLR protein expression, purification, and structural characterization. Here, we provide a detailed protocol for Arabidopsis thaliana GLR3.4 (AtGLR3.4) expression in a mammalian cell line and purification for structure determination by cryogenic electron microscopy (cryo-EM).
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影响因子：
48
作者：
Barad BA;Echols N;Wang RY;Cheng Y;DiMaio F;Adams PD;Fraser JS
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影响因子：
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作者：
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DOI：
10.1107/s0907444904019158
发表时间：
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影响因子：
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作者：
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