Drug therapy targeted to the voltage-gated sodium channel

该应用程序的总体目标是了解局部 与心脏电压门控钠（Na）通道（NaV1.5）的麻醉/抗心律失常药物（LA）。的 实验将使用野生型（WT）和定点突变的Na通道异源表达， 哺乳动物细胞和卵母细胞结合Na通道孔的分子建模。目标1。洛杉矶 孔中的结合位点。我们假设LA与DIV-Phe 1759的结合直接与DIIIS 6相互作用， 或间接通过开放/失活通道中DIIIS 6和DIVS 6之间的紧密堆积相互作用 该配置导致DIIIS的稳定4。为此，我们建议进一步探索药物 与DIV和DIII中S6之间的Na通道内孔的相互作用及其稳定性 对应的S4。根据这一假设，我们将确定DIII-Leu 1461在偶联中的作用 从DIV到DIII。目的2：快速失活与LA亲和力之间的联系。我们假设禁食的结合 灭活盖对于稳定高亲和力药物/通道相互作用是必需的。为此，我们建议： 通过将LA的大小和柔性相关联，确定快速失活如何有助于高亲和力LA阻断 和抗惊厥药物在突变通道中的EC 50，通过诱变去除快速失活， 失活盖和调查的基础上，最近报道的利多卡因引起的Brugada综合征 NaV1.5通道病，V232 I + L1308 F。目标3. LA与闭合通道的相互作用以及与早期步骤的相互作用 activation.我们假设S6节段形成的Na通道孔的开放是被动的， 并且药物与封闭通道的结合适度地稳定了该构型。依据本 假设我们将通过交联S6之间的Cys取代来确定束交叉处的残基 片段

The overall aim of the application is to understand the fundamental basis of the interaction of local anesthetic/antiarrhythmic drugs (LA) with the cardiac voltage-gated sodium (Na) channel (NaV1.5). The experiments will use wild-type (WT) and site-directed mutant Na channels heterologously expressed in mammalian cells and oocytes combined with molecular modeling of the Na channel pore. Aim 1. The LA binding site in the pore. We hypothesize that binding of LA to DIV-Phe1759 directly interacts with DIIIS6 residues or indirectly by close-packed interactions between DIIIS6 and DIVS6 in the open/inactivated channel configuration that results in stabilization of the DIIIS4. In this aim we propose to further explore drug interactions with the Na channel inner pore between S6's in DIV and DIII and stabilization of their corresponding S4's. In pursuance of this hypothesis we will determine the role of DIII-Leu1461 in the coupling of DIV to DIII. Aim 2: The link between fast inactivation and LA affinity. We hypothesize that binding of the fast inactivation lid is obligatory for stabilizing the high-affinity drug/channel interaction. In this aim we propose to determine how fast inactivation contributes to high affinity LA block by correlating the size and flexibility of LA and anticonvulsant drugs to their EC50's in mutant channels with fast inactivation removed by mutagenesis of the inactivation lid and by investigating the basis of the recently reported lidocaine-induced Brugada Syndrome NaV1.5 channelopathy, V232I+L1308F. Aim 3. LA interactions with the closed channel and with early steps in activation. We hypothesize that the opening of the Na channel pore formed by the S6 segments is passive, and that drug binding to the closed channel modestly stabilizes this configuration. In pursuance of this hypothesis we will determine the residues at the bundle crossing by crosslinking Cys substitutions between S6 segments.