MOLECULAR BIOPHYSICS OF POTASSIUM CHANNEL INACTIVATIONS

失活是一种常见的分子记忆形式 大多数钾离子通道。时间进程可以从慢到慢 (秒)到超快，可以恢复；而多个分子 机制已经确定，那些潜在的失活 心脏中的钾通道仍未完全被了解。 该项目将以我们开发的初步数据为基础 测试与这些机制有关的假设。我们发现了 Kv1.4的N末端链中的二级结构是一个 决定其失活的关键因素~球链~ 机制。在具体目标1中，我们将使用基于结构的 删除策略来验证假设Kv4.3，最近- 被确认为人类ITO的主要成分，通过以下方式失活 类似的机制。在具体目标2中，实验相结合 离子电流修饰通道的荧光成像 测量结果将被用来检验复苏的假设 从失活开始是由S4的运动启动的。我们有 在N端发现了一种超快速失活的形式 Kv1.4的突变体。在具体目标3中，我们将检验假设 这种形式的超快速失活也发生在Kv4.3和 在IKRα亚基中，HERG。此外，这些突变体 通道将被用来检验这样的假设：增强 预先存在的停用过程(而不是球和链 机制)是钾通道β-的作用基础。 亚单位。通过阐明失活机制的范围 存在于心脏的钾通道中，这些实验将 提供对决定因素的进一步了解 健康人心脏的异质性电生理行为 和疾病。

Inactivation is a form of molecular memory that is common to most potassium channels. The time course can range from slow (seconds) to ultra-fast, as can recovery; while multiple molecular mechanisms have been identified, those underlying inactivation of potassium channels in the heart remain incompletely understood. This Project will build on preliminary data we have developed to test hypotheses with respect to these mechanisms. We have found that secondary structure in the N-terminus chain of Kv1.4 is an crucial determinant of its inactivation by a ~ball-and-chain~ mechanism. In Specific Aim 1, we will use a structure-based deletion strategy to test the hypothesis that Kv4.3, recently- identified as a major component of human Ito, inactivates by similar mechanisms. In Specific Aim 2, experiments combining fluorescent imaging of modified channels with ion current measurements will be used to test the hypothesis that recovery from inactivation is initiated by movement of S4. We have identified a form of ultra-rapid inactivation in an N-terminus mutant of Kv1.4. In Specific Aim 3, we will test the hypothesis that this form of ultra-rapid inactivation also occurs in Kv4.3 and in the Ikr alpha-subunit, HERG. In addition, these mutant channels will be used to test the hypothesis that enhancement of pre-existing inactivation processes (rather than a ball-and-chain mechanism) underlies the effect of potassium channel beta- subunits. By elucidating the range of inactivation mechanisms present in cardia potassium channels, these experiments will provide further understanding of the determinants of the heterogeneous electrophysiologic behavior of the heart in health and disease.