Neuberger Arthur, Nadezhdin Kirill D, Sobolevsky Alexander I
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, United States.
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY, United States.
Methods Enzymol. 2021;652:31-48. doi: 10.1016/bs.mie.2021.02.006. Epub 2021 Mar 12.
The transient receptor potential vanilloid-superfamily member 3 (TRPV3) channel is implicated in a variety of physiological processes, including temperature sensing, nociception and itch, maintenance of the skin barrier, wound healing, hair growth, and embryonic development. TRPV3 is also associated with various skin diseases, including Olmsted syndrome, atopic dermatitis, and rosacea. Studies of TRPV3 are of fundamental importance for structural pharmacology aimed at the design of drugs targeting this channel and for understanding the molecular basis of temperature sensing. Here we describe a detailed protocol for expression and purification of chemically pure and stable TRPV3 protein that is suitable for structural and functional characterization of this channel, in particular for cryo-EM sample preparation and high-resolution 3D reconstruction.
瞬时受体电位香草酸超家族成员3（TRPV3）通道参与多种生理过程，包括温度感知、伤害感受和瘙痒、皮肤屏障的维持、伤口愈合、毛发生长以及胚胎发育。TRPV3还与多种皮肤疾病相关，包括奥姆斯特德综合征、特应性皮炎和酒渣鼻。对TRPV3的研究对于旨在设计靶向该通道的药物的结构药理学以及理解温度感知的分子基础至关重要。在此，我们描述了一种详细的方案，用于表达和纯化化学纯且稳定的TRPV3蛋白，该蛋白适用于该通道的结构和功能表征，特别是用于冷冻电镜样品制备和高分辨率三维重建。
