Department of Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, TN 37212, USA.
Department of Medicine, Division of Clinical Pharmacology, Vanderbilt Center for Arrhythmia Research and Therapeutics, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
Int J Mol Sci. 2022 Feb 17;23(4):2225. doi: 10.3390/ijms23042225.
The cardiac sodium ion channel (Na1.5) is a protein with four domains (DI-DIV), each with six transmembrane segments. Its opening and subsequent inactivation results in the brief rapid influx of Na ions resulting in the depolarization of cardiomyocytes. The neurotoxin veratridine (VTD) inhibits Na1.5 inactivation resulting in longer channel opening times, and potentially fatal action potential prolongation. VTD is predicted to bind at the channel pore, but alternative binding sites have not been ruled out. To determine the binding site of VTD on Na1.5, we perform docking calculations and high-throughput electrophysiology experiments in the present study. The docking calculations identified two distinct binding regions. The first site was in the pore, close to the binding site of Na1.4 and Na1.5 blocking drugs in experimental structures. The second site was at the "mouth" of the pore at the cytosolic side, partly solvent-exposed. Mutations at this site (L409, E417, and I1466) had large effects on VTD binding, while residues deeper in the pore had no effect, consistent with VTD binding at the mouth site. Overall, our results suggest a VTD binding site close to the cytoplasmic mouth of the channel pore. Binding at this alternative site might indicate an allosteric inactivation mechanism for VTD at Na1.5.
心脏钠离子通道（Na1.5）是一种具有四个结构域（DI-DIV）的蛋白质，每个结构域有六个跨膜片段。它的开放和随后的失活导致短暂的快速钠离子内流，导致心肌细胞去极化。神经毒素藜芦碱（VTD）抑制 Na1.5 失活，导致更长的通道开放时间，并可能导致致命的动作电位延长。VTD 预计会结合在通道孔上，但尚未排除替代结合位点。为了确定 VTD 在 Na1.5 上的结合位点，我们在本研究中进行了对接计算和高通量电生理学实验。对接计算确定了两个不同的结合区域。第一个位点在孔内，靠近实验结构中 Na1.4 和 Na1.5 阻断药物的结合位点。第二个位点位于孔的“口”部，在细胞质侧，部分暴露在溶剂中。该位点的突变（L409、E417 和 I1466）对 VTD 结合有很大影响，而孔内更深的残基则没有影响，这与 VTD 在口部的结合一致。总体而言，我们的结果表明 VTD 结合位点靠近通道孔的细胞质口。在该替代位点的结合可能表明 VTD 在 Na1.5 上的变构失活机制。
