MOLECULAR MECHANICS OF MYOSIN FILAMENTS

肌球蛋白丝的分子力学

• 批准号: 6149718
• 负责人: WILLIAM H GUILFORD
• 金额: $ 18.88万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-05 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6149718
• 关键词: Aves; animal tissue; biomechanics; conformation; gel filtration chromatography; intermolecular interaction; microfilaments; muscle proteins; myosins; phosphorylation; protein structure function; smooth muscle; statistics /biometry; striated muscles; ultracentrifugation

The goal of the proposed research is to quantify the individual effects of filament structure and phosphorylation in smooth and skeletal muscle myosins, and to examine their roles in modifying the mechanical behavior of single myosin molecules. Smooth and skeletal muscle myosins polymerize to form two very distinct filament structures. The functional consequence of this structural difference is unknown. Furthermore, there is tremendous disagreement in the literature as to the "step displacement" of myosin, with estimates ranging from 5 - 20 nm. The value obtained appears to correspond to the structural state of the myosin being used in each laboratory. Taken as a whole, these data suggest that the level of structural organization is critical to the overall performance of myosin, and that myosin molecules within a filament may cooperate to generate force and motion. However, intermolecular cooperativity at the level of myosin filaments has never been systematically investigated, and is thus the first major thrust of the proposed research. We will use a laser trap force transducer to measure and compare the single steps in force and displacement generated by smooth and skeletal muscle myosins both in their monomeric and filamentous states. The objective is to understand if the constituent monomers of myosin filaments act cooperatively, and whether smooth and skeletal muscle filaments behave differently in this respect. Phosphorylation of the regulatory light chain of myosin alters the structures of myosin filaments in both smooth and skeletal muscles. Because of this, we are obligated to examine both phosphorylation and filament structure in order to understand either. The structural change that occurs upon phosphorylation appears to be an extension of the myosin heads away from the myosin filament and toward actin. This conformational change may underlie the well-established regulation of contraction in smooth muscle by phosphorylation, as well as the enhancement of force generation in skeletal muscle. However, the mechanical consequences of phosphorylation have yet to be examined at the level of the individual myosin heads. Thus, the second major thrust of the proposed research is to examine the effects of phosphorylation on the forces and displacements generated by these myosins in their monomeric and filamentous states. These data are critical to our understanding of how myosin functions at the molecular level in whole cells and tissues.

拟议研究的目标是量化个体影响 平滑肌和骨骼肌的丝结构和磷酸化 肌球蛋白，并检查它们在改变机械行为中的作用 单个肌球蛋白分子。 平滑肌和骨骼肌肌球蛋白聚合形成两种截然不同的肌球蛋白 细丝结构。 这种结构的功能后果 差异未知。 此外，还有很大的分歧 在文献中关于肌球蛋白的“阶梯位移”， 估计范围为 5 - 20 nm。获得的值似乎 对应于每个中使用的肌球蛋白的结构状态 实验室。 总体而言，这些数据表明 结构组织对于整体绩效至关重要 肌球蛋白，并且丝内的肌球蛋白分子可以合作 产生力和运动。 然而，分子间协同作用 肌球蛋白丝的水平从未被系统地研究过， 因此，这是拟议研究的第一个主要目标。 我们将 使用激光陷阱力传感器来测量和比较单个 平滑肌和骨骼肌产生的力和位移的阶跃 肌球蛋白处于单体和丝状状态。 目标 是为了了解肌球蛋白丝的组成单体是否起作用 合作，以及平滑肌和骨骼肌丝是否表现良好 在这方面有所不同。 肌球蛋白调节轻链的磷酸化改变了 平滑肌和骨骼肌中肌球蛋白丝的结构。 因此，我们有义务检查磷酸化和 以便了解灯丝结构。 结构性变化 磷酸化时发生的情况似乎是 肌球蛋白远离肌球蛋白丝并朝向肌动蛋白。 这 构象变化可能是完善的调控的基础 通过磷酸化使平滑肌收缩，以及 增强骨骼肌的力量产生。 然而， 磷酸化的机械后果尚未得到检验 各个肌球蛋白头的水平。 于是，第二个主推 拟议研究的目的是检查磷酸化的影响 关于这些肌球蛋白在其内部产生的力和位移 单体和丝状状态。 这些数据对我们至关重要 了解肌球蛋白在分子水平上的整体功能 细胞和组织。