Department of Physiological Sciences, Eastern Virginia Medical School, Norfolk, VA 23507, USA.
Department of Cellular and Molecular Medicine, University of Arizona, Tucson, AZ 85724, USA.
J Mol Biol. 2021 Sep 17;433(19):167178. doi: 10.1016/j.jmb.2021.167178. Epub 2021 Jul 27.
Cardiac muscle contraction depends on interactions between thick (myosin) and thin (actin) filaments (TFs). TFs are regulated by intracellular Ca levels. Under activating conditions Ca binds to the troponin complex and displaces tropomyosin from myosin binding sites on the TF surface to allow actomyosin interactions. Recent studies have shown that in addition to Ca, the first four N-terminal domains (NTDs) of cardiac myosin binding protein C (cMyBP-C) (e.g. C0, C1, M and C2), are potent modulators of the TF activity, but the mechanism of their collective action is poorly understood. Previously, we showed that C1 activates the TF at low Ca and C0 stabilizes binding of C1 to the TF, but the ability of C2 to bind and/or affect the TF remains unknown. Here we obtained 7.5 Å resolution cryo-EM reconstruction of C2-decorated actin filaments to demonstrate that C2 binds to actin in a single structural mode that does not activate the TF unlike the polymorphic binding of C0 and C1 to actin. Comparison of amino acid sequences of C2 with either C0 or C1 shows low levels of identity between the residues involved in interactions with the TF but high levels of conservation for residues involved in Ig fold stabilization. This provides a structural basis for strikingly different interactions of structurally homologous C0, C1 and C2 with the TF. Our detailed analysis of the interaction of C2 with the actin filament provides crucial information required to model the collective action of cMyBP-C NTDs on the cardiac TF.
心肌收缩依赖于粗肌丝（肌球蛋白）和细肌丝（肌动蛋白）之间的相互作用。肌丝受细胞内 Ca 水平调节。在激活条件下，Ca 结合到肌钙蛋白复合物上，并将原肌球蛋白从肌丝结合位点上置换下来，从而允许肌球蛋白和肌动蛋白相互作用。最近的研究表明，除了 Ca 之外，心肌肌球蛋白结合蛋白 C（cMyBP-C）的前四个 N 端结构域（NTD）（例如 C0、C1、M 和 C2）也是肌丝活性的有力调节剂，但它们的集体作用机制还不太清楚。以前，我们发现 C1 在低 Ca 下激活肌丝，C0 稳定 C1 与肌丝的结合，但 C2 结合和/或影响肌丝的能力仍然未知。在这里，我们获得了 C2 修饰的肌动蛋白丝的 7.5 Å 分辨率冷冻电镜重建，以证明 C2 以一种不同于 C0 和 C1 与肌动蛋白的多态结合的单一结构模式结合肌动蛋白，而不激活肌丝。C2 与 C0 或 C1 的氨基酸序列比较表明，与 TF 相互作用的残基之间的同一性水平较低，但参与 Ig 折叠稳定的残基的保守性水平较高。这为结构同源的 C0、C1 和 C2 与肌丝的截然不同的相互作用提供了结构基础。我们对 C2 与肌动蛋白丝相互作用的详细分析提供了对 cMyBP-C NTD 对心脏肌丝的集体作用进行建模所需的关键信息。
