Advancing cryo-EM technology to address difficult biological questions

摘要 单粒子低温电子显微镜(Cryo-EM)的技术突破使原子 许多具有挑战性的生物大分子的结构测定以前所未有的快速速度进行。它还 促进解决具有挑战性的结构生物学问题，否则这些问题将很难解决。 除了原子结构之外，蛋白质动力学和功能态之间的构象转变也在其中 特别适合用单粒子低温电子显微镜研究的问题。 TRPV1离子通道是多模式伤害性感受器，集成了一系列生化和 生物物理刺激。它在生化上表现良好，在生理和药理上也有很好的特点。 各种内源性刺激和天然外源性化合物以不同的功能捕获通道 我们已经确定了TRPV1在各种稳定构象中的结构。在下一步中，我们将使用 TRPV1作为利用新型低温电子显微镜探测构象状态范围的模型系统 由这个复数信号积分器采样。对于许多其他TRP通道，在这些通道中没有合适的 在特定稳定构象中捕获通道的药理学工具，利用该模型开发的方法 该系统可用于研究通道选通的机理。

ABSTRACT Technological breakthroughs in single particle cryo-electron microscopy (cryo-EM) have enabled atomic structure determinations of many challenging biological macromolecules in an unprecedented rapid pace. It also facilitated addressing challenging structural biology questions that would otherwise be difficult to address. Beyond atomic structures, protein dynamics and conformational transitions between functional states are among questions that are particularly suited to be studied by single particle cryo-EM. TRPV1 ion channel is polymodal nociceptor that integrates signals generated from a range of biochemical and biophysical stimuli. It is biochemically well-behaved and physiologically and pharmacologically well characterized. With various endogenous stimuli and natural exogenous compounds trapping the channel in various functional states, we have determined structures of TRPV1 in various stable conformations. In the next step, we will use TRPV1 as a model system to exploit using novel cryo-EM advances to probe the range of conformational states sampled by this complex signal integrator. For many other TRP channels, in which there is no suitable pharmacological tools to trap the channels in specific stable conformations, methods developed with the model system can be used to study mechanisms of channel gating.