Dissecting the structural origin of relaxation in skeletal muscle

剖析骨骼肌松弛的结构起源

• 批准号: 10567284
• 负责人: Raul Padron
• 金额: $ 58.37万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2028-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10567284
• 关键词: ATP phosphohydrolase; Actins; Animals; Area; Binding; Biochemical; Bone structure; Cardiac; Collaborations; Consumption; Contracts; Cryoelectron Microscopy; DNA Sequence Alteration; Disease; Drug Targeting; Energy consumption; Equilibrium; Exhibits; Fiber; Filament; H-Meromyosin; Head; Human; Impairment; Laboratories; Life; MLL gene; Measures; Microscope; Modeling; Molecular; Molecular Conformation; Molecular Motors; Motor; Movement; Mus; Muscle; Muscle Contraction; Muscle function; Muscle relaxation phase; Myopathy; Myosin ATPase; Oryctolagus cuniculus; Pharmaceutical Preparations; Pharmacotherapy; Proteins; Relaxation; Resolution; Role; Side; Skeletal Muscle; Slide; Structure; Tail; Testing; Therapeutic; Thick; Thick Filament; Thin Filament; Work; X ray diffraction analysis; antagonist; connectin; insight; particle; repaired; skeletal

How muscle contracts has been a long-standing question. Despite major advances in this area, how muscle relaxes is still not fully understood. Contraction occurs by the sliding of myosin-containing thick past actin-containing thin filaments, powered by myosin heads, motors that produce sliding force, fueled by ATP. Relaxation occurs when thin filaments are switched off so heads cannot bind to produce force, leaving the idling heads to organize themselves helically in the thick filament. What is currently known about the role of thick filaments in relaxation? On the structural side, low-resolution models of cardiac (mouse, human) and skeletal (tarantula) thick filaments have been achieved, but their atomic structure remains unsolved. On the energetics side, the energy consumption of relaxed skeletal muscle revealed a surprising phenomenon, so-called super- relaxation (SRX) that greatly reduces ATP consumption. A widely accepted view associates this ubiquitous and fundamental energy-saving state with the unique way myosin’s two heads fold together in the relaxed tarantula filament—the so-called interacting-heads motif (IHM), found across the animal kingdom, which structurally inhibits both heads, switching off their activity. Regardless of its appeal, this SRX=IHM hypothesis has not been proved, and recent ATP turnover results suggest, instead, association of SRX with a specific myosin head conformation. Elucidating this puzzle is crucial to understanding how muscle relaxes, how it malfunctions in disease and how therapeutic drug treatments work. The solution requires determination by cryo-EM of the atomic structures of the thick filament and myosin molecules from muscle. Here, we propose to determine the structures of skeletal myosin molecules and filaments, far less studied than cardiac. This will allow us to dissect how key IHM interactions constrain activity of the two heads, shutting them off, thus conserving ATP in relaxation. We will use single particle EM and cryo-EM to define the structural basis of the SRX state at near-atomic level in thick filaments and myosin molecules from rabbit skeletal muscle. By comparing with tarantula, which shows tenfold- greater energy-saving (hyper-relaxation, HRX), we will gain deeper insight into the mechanism of ATPase inhibition. And we will use EM and X-ray diffraction to investigate how therapeutic drugs alter the IHM. Aim 1 will define the structural basis of SRX in skeletal thick filaments by revealing their near-atomic cryo-EM structures. Aim 2 will define the structural basis of SRX in skeletal myosin heads and heavy meromyosin molecules by assessing: (A) if SRX results from a specific head conformation, and (B) if the IHM correlates with the SRX state. Aim 3 will reveal the structural impact of drugs on skeletal thick filaments and myosin molecules. Despite the vital role of SRX in skeletal muscle relaxation, its structural basis and relation to the IHM and to other thick filament proteins (MyBP-C, titin) remains unknown. Our studies will reveal the IHM structure in skeletal thick filaments and myosin molecules, clarify its association with the SRX state and with MyBP-C and titin, and provide critical insights into the molecular basis of relaxation and the influence of therapeutic drugs.

肌肉如何收缩一直是一个长期存在的问题。尽管在这方面取得了重大进展，但如何 肌肉松弛的机制还没有完全被了解。收缩是由含有肌球蛋白的厚浆 含有肌动蛋白的细丝，由肌球蛋白头提供动力，肌球蛋白头是产生滑动力的马达，由ATP提供燃料。 当细丝被切断时，松弛发生，这样头就不能产生力，留下空转 头部在粗丝中螺旋状排列。目前所知的厚的作用是什么 松弛的细丝在结构方面，心脏（小鼠，人类）和骨骼的低分辨率模型 （狼蛛）粗丝已经实现，但它们的原子结构仍然没有解决。关于能量学 另一方面，放松骨骼肌的能量消耗揭示了一个令人惊讶的现象，即所谓的超- 放松（SRX），大大减少ATP的消耗。一种被广泛接受的观点将这种无处不在的现象联系起来， 在放松的狼蛛中，肌球蛋白的两个头以独特的方式折叠在一起， 在动物王国中发现的所谓的相互作用头基序（IHM），在结构上 抑制了两个头的活动不管它的吸引力，这个SRX=IHM假说还没有被 最近的ATP周转结果表明，相反，SRX与一个特定的肌球蛋白头的关联 构象阐明这一谜题对于理解肌肉如何放松，肌肉如何在运动中发生故障至关重要。 疾病和治疗药物治疗如何工作。该解决方案需要通过原子的低温-EM测定 粗丝和肌球蛋白分子的结构。在这里，我们建议确定结构 骨骼肌球蛋白分子和肌丝，远不如心脏研究。这将使我们能够剖析 IHM相互作用限制了两个头部的活动，关闭它们，从而在放松中保存ATP。我们将 使用单粒子EM和cryo-EM来定义厚层中近原子水平的SRX态的结构基础。 纤维和肌球蛋白分子。与狼蛛相比，狼蛛表现出十倍的- 更大的节能（超松弛，HRX），我们将获得更深入的了解ATP酶的机制 抑制作用我们将使用EM和X射线衍射来研究治疗药物如何改变IHM。 目的1将通过揭示骨架粗纤维的近原子结构， 冷冻电镜结构目的2将确定骨骼肌球蛋白头和重肌球蛋白中SRX的结构基础 通过评估：（A）SRX是否由特定的头部构象产生，以及（B）IHM是否与 SRX国家。目的3将揭示药物对骨骼粗丝和肌球蛋白分子的结构影响。 尽管SRX在骨骼肌松弛中起着重要作用，但其结构基础及其与IHM和 与其他粗丝蛋白（MyBP-C、肌联蛋白）的关系尚不清楚。我们的研究将揭示IHM的结构， 骨骼粗丝和肌球蛋白分子，阐明其与SRX状态和MyBP-C的关系， 肌联蛋白，并提供关键的见解放松的分子基础和治疗药物的影响。