Department of Physics, Université de Montréal, Montréal, QC, Canada.
Department of Pharmacology and Physiology, Université de Montréal, Montréal, QC, Canada.
Biochem Soc Trans. 2022 Oct 31;50(5):1427-1445. doi: 10.1042/BST20220605.
Ion channels undergo major conformational changes that lead to channel opening and ion conductance. Deciphering these structure-function relationships is paramount to understanding channel physiology and pathophysiology. Cryo-electron microscopy, crystallography and computer modelling provide atomic-scale snapshots of channel conformations in non-cellular environments but lack dynamic information that can be linked to functional results. Biophysical techniques such as electrophysiology, on the other hand, provide functional data with no structural information of the processes involved. Fluorescence spectroscopy techniques help bridge this gap in simultaneously obtaining structure-function correlates. These include voltage-clamp fluorometry, Förster resonance energy transfer, ligand binding assays, single molecule fluorescence and their variations. These techniques can be employed to unearth several features of ion channel behaviour. For instance, they provide real time information on local and global rearrangements that are inherent to channel properties. They also lend insights in trafficking, expression, and assembly of ion channels on the membrane surface. These methods have the advantage that they can be carried out in either native or heterologous systems. In this review, we briefly explain the principles of fluorescence and how these have been translated to study ion channel function. We also report several recent advances in fluorescence spectroscopy that has helped address and improve our understanding of the biophysical behaviours of different ion channel families.
离子通道经历了导致通道开放和离子电导的主要构象变化。破译这些结构-功能关系对于理解通道生理学和病理生理学至关重要。低温电子显微镜、晶体学和计算机建模提供了非细胞环境中通道构象的原子尺度快照，但缺乏可以与功能结果相关联的动态信息。另一方面，生物物理技术，如电生理学，提供了功能数据，但没有涉及过程的结构信息。荧光光谱技术有助于在同时获得结构-功能相关性方面弥补这一差距。这些技术包括电压钳荧光法、Förster 共振能量转移、配体结合测定、单分子荧光及其变体。这些技术可用于揭示离子通道行为的几个特征。例如，它们提供了固有于通道特性的局部和全局重排的实时信息。它们还提供了关于离子通道在膜表面上的运输、表达和组装的见解。这些方法的优点是它们可以在天然或异源系统中进行。在这篇综述中，我们简要解释了荧光的原理，以及这些原理如何被转化为研究离子通道功能。我们还报告了荧光光谱学的几项最新进展，这些进展有助于解决和提高我们对不同离子通道家族的生物物理行为的理解。
