Molecular mechanisms of muscle contraction

背景资料。肌肉收缩是由附着在肌动蛋白细丝上的分子马达肌球蛋白II驱动的。肌球蛋白分子将从三磷酸腺苷(ATP)中提取的化学能转化为机械功。在肌球蛋白和肌动蛋白最初连接后，ATP被水解，磷酸(PI)和随后的ADP从肌球蛋白中释放出来。分子经历了构象变化--产生力量的力量和滑动肌动蛋白细丝的力量。从三磷酸腺苷释放的化学能转化为肌球蛋白动力卒中，并将其放大为肌肉收缩的分子机制尚不清楚。*长期目标。这项研究计划的长期目标是了解肌肉收缩和力量产生的分子和细胞机制。这项建议的具体目标是：*1-确定肌球蛋白结构和肌动蛋白滑动之间的关系*2-确定肌球蛋白负责将化学能转化为机械功的亚结构*3-确定肌球蛋白分子在细丝中协同工作时的新特性。*方法这项研究计划将使用我的实验室开发的各种实验系统和技术。有一些肌球蛋白分子的结构域和亚结构域被假设为定义了力量卒中。在具体目标(1)中，我们将使用天然肌球蛋白分子和肌球蛋白突变体进行实验，在这些突变体中，这些亚域中的一些已经被耗尽。分子将通过一篇运动学论文进行测试，该论文允许测量肌动蛋白滑动的速度。在特定的目标(2)中，将在高速原子力显微镜(HS-AFM)下测试分子(天然的和突变的)，该显微镜允许测量力，同时可视化肌球蛋白-肌动蛋白系统的结构变化。在特定的目标(3)中，分子(天然的和突变的)将被聚合成不同浓度的细丝，以评估肌球蛋白协同工作的新特性。这些细丝将使用纳米级悬臂系统进行测力测试。*意义。了解肌球蛋白动力中风--肌肉收缩的最基本步骤--对于理解肌肉收缩这一生命的基本功能是必不可少的。除了收缩机制外，肌球蛋白II是许多分子马达(如动蛋白、动力蛋白、肌球蛋白V)中的一种，具有多种生理功能。分子马达有几个相似之处，包括动力冲击诱导的运动和力的产生。因此，了解肌球蛋白II的机制将有助于深入了解分子马达的总体功能，对生物学具有深远的意义。

Background. Muscle contraction is driven by the molecular motor myosin II while attached to actin filaments. Myosin molecules transform chemical energy extracted from adenosine tri-phosphate (ATP) into mechanical work. After an initial attachment between myosin and actin, ATP is hydrolyzed, and phosphate (Pi) and subsequently ADP are released from myosin. The molecule undergoes a conformational change - the power stroke - producing force and sliding the actin filament. The molecular mechanisms behind the conversion of chemical energy released from ATP into the myosin power stroke, and its amplification into muscle contraction is not well understood. ***Long-term objective. The long-term objective of this research program is to understand the molecular and cellular mechanisms of muscle contraction and force generation. The specific goals of this proposal are:***1 - To determine the relation between myosin structures and actin sliding ***2 - To determine the sub-structures of myosin responsible for converting chemical energy into mechanical work ***3 - To determine the emerging properties of myosin molecules when working cooperatively in filaments.***Methods. This research program will use a variety of experimental systems and techniques developed in my laboratory. There are domains and sub-domains of the myosin molecule that have been hypothesized to define the power stroke. In specific aim (1) we will experiment with native myosin molecules and also myosin mutants in which some of these sub-domains have been depleted. Molecules will be tested with a motility essay, that allows for measurements of velocity of actin sliding. In specific aim (2) the molecules (native and mutants) will be tested under a High-Speed Atomic Force Microscope (HS-AFM), that allows measurements of force while visualizing structural changes in the myosin-actin system. In specific aim (3) molecules (native and mutants) will be polymerized into filaments in different concentrations, to evaluate the emerging properties of myosins working cooperatively. The filaments will be tested for force measurements using a system of nanofabricated cantilevers. ***Significance. Understanding the myosin power stroke - the most fundamental step of muscle contraction - is essential for understanding muscle contraction, a basic function of life. Beyond the mechanisms of contraction, myosin II is one among many molecular motors (e.g. kinesin, dynein, myosin V), with a variety of physiological functions. Molecular motors share several similarities, including a power-stroke induced motility and force production. Therefore, understanding myosin II mechanisms will provide insight into the functioning of molecular motors in general, with far-reaching implications for biology.**