用于冷冻电镜研究的人源 TRPA1 离子通道的样品制备。
Sample preparation of the human TRPA1 ion channel for cryo-EM studies.
机构信息
Department of Biochemistry, Duke University School of Medicine, Durham, NC, United States.
Department of Biochemistry, Duke University School of Medicine, Durham, NC, United States.
出版信息
Methods Enzymol. 2021;653:75-87. doi: 10.1016/bs.mie.2020.12.018. Epub 2021 Jan 27.
The transient receptor potential ankyrin 1 (TRPA1) ion channel is a member of the TRP channel family that is involved in sensing noxious stimuli that elicit pain and inflammation. Because of its critical physiological role and therapeutic importance, great efforts have been made to understand the structure and mechanism of TRPA1. Several human TRPA1 structures have been reported using single particle cryo-electron microscopy (cryo-EM) over the last 6 years. Here, we present a protocol for the heterologous expression, large-scale purification, and nanodisc reconstitution of the human TRPA1 channel for cryo-EM and biochemical studies.
瞬时受体电位锚蛋白 1（TRPA1）离子通道是 TRP 通道家族的一员，参与感知引起疼痛和炎症的有害刺激。由于其关键的生理作用和治疗重要性，人们做出了巨大努力来理解 TRPA1 的结构和机制。在过去的 6 年中，使用单颗粒冷冻电子显微镜（cryo-EM）已经报道了几种人类 TRPA1 结构。在这里，我们提供了一个用于冷冻电镜和生化研究的异源表达、大规模纯化和纳米盘重建人类 TRPA1 通道的方案。
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