Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences , Saint Petersburg, Russia.
Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Canada.
J Gen Physiol. 2023 Nov 6;155(11). doi: 10.1085/jgp.202313418. Epub 2023 Sep 20.
Opposite effects of 1,4-dihydropyridine (DHP) agonists and antagonists on the L-type calcium channels are a challenging problem. Cryo-EM structures visualized DHPs between the pore-lining helices S6III and S6IV in agreement with published mutational data. However, the channel conformations in the presence of DHP agonists and antagonists are virtually the same, and the mechanisms of the ligands' action remain unclear. We docked the DHP agonist S-Bay k 8644 and antagonist R-Bay k 8644 in Cav1.1 channel models with or without π-bulges in helices S6III and S6IV. Cryo-EM structures of the DHP-bound Cav1.1 channel show a π-bulge in helix S6III but not in S6IV. The antagonist's hydrophobic group fits into the hydrophobic pocket formed by residues in S6IV. The agonists' polar NO2 group is too small to fill up the pocket. A water molecule could sterically fit into the void space, but its contacts with isoleucine in helix S6IV (motif INLF) would be unfavorable. In a model with π-bulged S6IV, this isoleucine turns away from the DHP molecule and its position is occupied by the asparagine from the same motif INLF. The asparagine provides favorable contacts for the water molecule at the agonist's NO2 group but unfavorable contacts for the antagonist's methoxy group. In our models, the DHP antagonist stabilizes entirely α-helical S6IV. In contrast, the DHP agonist stabilizes π-bulged helix S6IV whose C-terminal part turned and rearranged the activation-gate region. This would stabilize the open channel. Thus, agonists, but not antagonists, would promote channel opening by stabilizing π-bulged helix S6IV.
1,4-二氢吡啶（DHP）激动剂和拮抗剂对 L 型钙通道的相反作用是一个具有挑战性的问题。冷冻电镜结构显示 DHP 位于 S6III 和 S6IV 孔衬螺旋之间，与已发表的突变数据一致。然而，DHP 激动剂和拮抗剂存在时通道构象几乎相同，配体作用的机制仍不清楚。我们将 DHP 激动剂 S-Bay k 8644 和拮抗剂 R-Bay k 8644 对接在具有或不具有 S6III 和 S6IV 螺旋 π 凸起的 Cav1.1 通道模型中。DHP 结合 Cav1.1 通道的冷冻电镜结构显示 S6III 螺旋中有一个 π 凸起，但 S6IV 中没有。拮抗剂的疏水区适合由 S6IV 残基形成的疏水区。激动剂的极性 NO2 基团太小，无法填满口袋。一个水分子可以占据这个空位，但它与 S6IV 螺旋中异亮氨酸（ motif INLF）的接触是不利的。在具有 π 凸起 S6IV 的模型中，这个异亮氨酸远离 DHP 分子，其位置被来自相同 motif INLF 的天冬酰胺占据。天冬酰胺为水分子提供了与激动剂的 NO2 基团有利的接触，但与拮抗剂的甲氧基基团不利的接触。在我们的模型中，DHP 拮抗剂完全稳定了 α-螺旋 S6IV。相比之下，DHP 激动剂稳定了 π 凸起的 S6IV，其 C 端部分转动并重新排列了激活门区域。这将稳定开放的通道。因此，激动剂而不是拮抗剂通过稳定 π 凸起的 S6IV 促进通道开放。
