Tropomyosin and the Regulation of Muscle Contraction

描述（由申请人提供）：广泛的长期目标是通过确定肌浆化和Ca 2+和肌球蛋白如何调节重要的原肌球蛋白-肌动蛋白相互作用来阐明横纹肌收缩调节的分子细节。健康相关性：每个组件的作用及其相互作用需要了解与某些肌肉疾病相关的功能障碍，例如，家族性肥厚型心肌病（FHC）由Tm、TnT、TnI和肌球蛋白突变引起。Tm在其与肌动蛋白的相互作用中，在确定肌肉细丝的3种状态（封闭、封闭和开放）之间的平衡中起关键作用（收缩发生在开放状态）。通过与肌动蛋白和Tm相互作用，肌钙蛋白（在Ca 2+的辅助下）和肌球蛋白头改变状态之间的平衡，从而打开和关闭收缩。具体目标：（a）确定肌动蛋白上Tm的柔性与激活的肌动蛋白亚基数目之间的关系（B）确定Tn复合物的TnI和TnT组分在肌动蛋白Tm丝上的位置以及在活性状态之间转换时发生的变化;（c）确定组织化肌节中细丝组分的变化的相互作用。研究方法：除了表征的3个国家在溶液中与ATP酶，结合和动力学研究，将广泛使用的高分辨率的距离测量与时间分辨荧光检测能量转移（FRET）在溶液中的组件和在区域的肌节的组件交换成单个肌原纤维之间。特定标记的氨基酸残基之间的这些测量将提供有关参与调节过程的组分之间的变化的动态信息。

DESCRIPTION (provided by applicant): The broad, long-term objective is to elucidate the molecular details of the regulation of striated muscle contraction by determining how trooping and Ca2+ and myosin, regulate the important tropomyosin-actin interaction. Health Relatedness: The role of each component and their interactions are required to understand the malfunctions associated with certain muscle diseases, e.g., familial hypertrophic cardiomyopathy (FHC) resulting from mutations in Tm, TnT, Tnl and myosin. Tm, in its interaction with actin, plays a key role in determining the equilibria between the 3 states of the muscle thin filament, Blocked, Closed and Open (contraction takes place in the Open state). By interacting with actin and Tm, troponin (assisted by Ca2+) and myosin heads change the equilibrium between the states, thereby turning contraction on and off. Specific Aims: (a) to determine the relationship between the flexibility of Tm on actin and the number of actin subunits activated (the cooperative unit size); to determine the altered regulatory properties of the FHC Tm mutants; (b) to determine the location of the Tnl and TnT components of the Tn complex on the actinTm filament and the changes that take place in shifting between activity states; (c) to determine the changing interactions of the thin filament components in the organized sarcomere. Methods: In addition to characterization of the 3 states in solution with ATPase, binding and kinetic studies, extensive use will be made of high resolution distance measurements between components with time-resolved fluorescence-detected energy transfer (FRET) in solution and in regions of the sarcomere of components exchanged into single myofibrils. These measurements between specific labeled amino acid residues will provide information about the dynamics of the changes between the components involved in the regulatory process.