Voltage-gated ion channels are intricate molecular machines that detect changes in transmembrane voltage and respond by rapid opening and closing of their ion-selective pore. Numerous types of voltage gated channels work together to orchestrate the electrical signaling of neurons as they signal, sense, and learn. The voltage-gated K+ channels (Kv channels) have served as a prototype system for understanding the moving parts involved in channel gating. Beyond the fundamental interest in learning about these moving parts, this work provides practical understanding of the mechanisms by which the channels can be modulated biologically by other cellular components, and pharmacologically by therapeutic drugs used, for instance, in epilepsy. Our previous work has characterized the key gating motions of the pore in Shaker Kv channels -- activation and two forms of inactivation -- using a combination of electrophysiology, site-directed mutagenesis, and chemical modification. At the same time, work revealing the structure of related bacterial K+ channels has provided a physical framework for understanding and interpreting our results. Together the structural and functional work provides an increasingly clear picture of the gating motions. ? Our present and future work is focused on resolving some critical inconsistencies between the functional and structural work, and on advancing our understanding of the dynamics of gating which are beyond the reach of the current crystallographic methods. Specifically, we have found that both open and closed Shaker Kv channels have specific properties that appear incompatible with the crystal structures of bacterial channels that provide the best framework for understanding the gating motions. We will test a specific hypothesis that dynamic fluctuations of open channels can reconcile the two sets of observations. We will also look at intermediate conformations of the channel protein as it passes from the resting closed state to the final open state. Finally, by measuring the energetic effects of site-directed mutations at different points in the protein, we will learn about the conformational wave involved in the opening and closing process, and about the functional role of two possible hinges in gating. ? ? ?
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