Structural insight into TRPA1 channel interactionwith agonist and antagonist

描述(申请人提供)：TRPA1是一种钙离子渗透性、非选择性阳离子通道，也是哺乳动物的关键疼痛感受器之一。它已成为镇痛剂和抗炎药的新靶点。TRPA1介导的痛觉反应涉及硫醇活性化合物和炎症介质对通道上N-末端半胱氨酸残基的修饰。在静息状态下，硫醇活性化合物与通道结合，导致通道激活，随后迅速脱敏。虽然电生理学方法已经对通道活动进行了广泛的研究，但对通道激活和脱敏的结构机制知之甚少。此外，两个直接抑制TRPA1的化合物正在进行临床前试验，了解这些化合物抑制TRPA1的机制将有助于我们进一步了解功能相关的TRPA1构象之间的结构差异。使用冷冻-EM、定点突变、有限蛋白分解和质谱学，我们的目标是在这项资助中建立详细的结构 了解TRPA1通道的激活、脱敏和抑制机制，有助于合理设计新型止痛药。

DESCRIPTION (provided by applicant): TRPA1 is a Ca2+-permeable, non-selective cation channel and one of the key pain sensors in mammals. It has emerged as a novel target for analgesics and anti-inflammatory agents. Pain sensation mediated by TRPA1 involves modification of N-terminal cysteine residues on the channel by thiol-reactive compounds and inflammatory mediators. Binding of thiol-reactive compounds to the channel in the resting state leads to channel activation followed rapidly by desensitization. While channel activity has been studied extensively by electrophysiological methods, little is known about the structural mechanisms of channel activation and desensitization. Additionally, two compounds that directly inhibit TRPA1 are in pre-clinical trials, and understanding the mechanism of TRPA1 inhibition by these compounds will be important for advancing our knowledge of the structural differences between functionally relevant TRPA1 conformations. Using cryo-EM, site-directed mutagenesis, limited proteolysis, and mass spectrometry, in this grant we aim to establish a detailed structural understanding of the mechanisms of TRPA1 channel activation, desensitization and inhibition, which will help facilitate rational design of novel analgesics.