Activation and desensitization of heat-sensor TRPV1

项目摘要 热激活和辣椒素激活的离子通道TRPV 1是维持细胞增殖的重要细胞传感器 正常体温和检测有害的周围环境，因此是重要的药物靶标。 TRPV 1也是理解多种TRP多模式激活的原型模型 渠道TRPV 1的结构已经通过冷冻电子显微镜（cyro-EM）在高达2.9-2.9 ° C下解析。 尽管有许多解决方案，但对通道激活（特别是热激活）的机械理解仍然存在， 初步的一个主要的限制是缺乏有效的工具来探测结构动力学， 功能渠道。我们最近从毒液中鉴定出两种结构相关的肽毒素 中国红头蜈蚣的RhTx 1和RhTx 2，具有高度特异性， 与TRPV 1的外孔区亲和结合。有趣的是，虽然RhTx 1强烈激活 RhTx 2在其N-末端仅具有四个额外的氨基酸，是一种有效的抑制剂。我们 初步研究进一步表明，RhTx 1特异性地促进热激活。因为这些 小而紧凑的肽毒素可以很容易地合成和修饰，它们提供了独特的机会， 探索TRPV 1激活的分子机制，其快速的ON和OFF动力学 特别有利于生物物理研究。拟议的研究将利用 这些新的毒素，以获得更好地了解TRPV 1的激活机制，揭示其 使用多学科方法的结构动力学结合了结构，电生理学， 光学和计算建模方法。

Project Summary Heat- and capsaicin-activated ion channel TRPV1 is an important cellular sensor for maintaining normal body temperature and detecting noxious ambient environment, hence an important drug target. TRPV1 also serves as a prototypical model for understanding polymodal activation of diverse TRP channels. Structures of TRPV1 have been solved by cryo-electron microscopy (cyro-EM) at up-to-2.9- Å resolutions, yet mechanistic understanding of channel activation, in particular by heat, remains preliminary. A major limitation has been the lack of effective tools for probing structural dynamics in functional channels. We have recently identified two structurally related peptide toxins from the venom of Chinese red-headed centipede, termed RhTx1 and RhTx2, that exhibit highly specific and sub-μM affinity binding to the outer pore region of TRPV1. Interestingly, while RhTx1 strongly activates the channel, RhTx2, having just four additional amino acids at its N-terminus, is a potent inhibitor. Our preliminary study further suggested that RhTx1 specifically promotes heat activation. Because these small, compact peptide toxins can be readily synthesized and modified, they offer unique opportunities to probe the molecular mechanism underlying TRPV1 activation, with their rapid ON and OFF kinetics being particularly favorable for biophysical investigation. The proposed study will take advantage of these novel toxins to gain a better understanding of TRPV1 activation mechanism by revealing its structural dynamics using a multidisciplinary approach combines structural, electrophysiological, optical, and computational modeling methods.