Allostery in myosins studied at the molecular level

描述(申请人提供)：在典型的真核细胞拥挤的环境中，任何大于50纳米的物体实际上是不能移动的，不能依靠扩散到达目的地。结果，肌球蛋白马达进化成为货物运输、细胞运动和细胞分裂产生力量，这些过程对生命本身至关重要。这些马达通过与肌动蛋白细丝结合时以受控的方式改变形状来转换化学自由能。所有马达蛋白质的一个基本特征是，它们的运动被仔细地协调，因此马达通过一系列特定的生物化学和机械状态的耦合。这种协调的性质仍然不清楚，但它显然是正常运动功能所必需的。最近认识到，每一类肌球蛋白都进化了特定的结构和动力学特征，这使得比较肌球蛋白超家族中不同的协调机制成为可能。这些个体适应现在可能被一系列先进的动力分析所表征。这里提出的工作将确定变构协调机制，使用新的单分子操作来改变马达感受到的分子内应变。通过机械改变肌动蛋白细丝的几何形状，将确定运动轨迹在传递应变中的作用。此外，在电机两个头部分离的条件下，将通过负荷相关的步进测量来确定肌球蛋白V和肌球蛋白VI的通信机制。这些机制将与肌球蛋白X的机制形成对比，肌球蛋白X是一种在体内具有不同寻常的运动性的马达，它暗示了一种新的应变传感形式。这项研究的长期目标是揭示马达和马达系统如何为其细胞贩运任务进行优化。肌球蛋白运动是多种正常和病理细胞功能所必需的，包括极化细胞结构的形成、细胞黏附的形成和组织发育、伤口愈合、细胞内运输和细胞分裂。这项研究将解决与运动相关疾病背后的潜在生物学相关的关键问题，其中包括与感觉毛细胞中立体纤毛的正常发育和维持有关的几种形式的遗传性耳聋。

DESCRIPTION (provided by applicant): In the crowded environment of a typical eukaryotic cell, any object larger than 50 nm is effectively immobile and cannot rely on diffusion to arrive at its destination. As a result, myosin motors evolved to generate force for the transport of cargoes, cell motility, and cell division, processes that are critical for life itself. These motors convert chemical free energy by changing shape in a controlled manner while bound to actin filaments. A fundamental feature of all motor proteins is that their movements are carefully coordinated, so that the motor travels through a specific sequence of coupled biochemical and mechanical states. The nature of this coordination remains obscure, but it is clearly essential for proper motor function. The recent recognition that each myosin class has evolved specific structural and kinetic features allows for the comparison of different coordination mechanisms across the myosin super family. These individual adaptations may now be characterized by a host of advanced motility assays. The work proposed here will determine the allosteric coordination mechanisms using new single-molecule manipulations to alter the intramolecular strain felt by the motor. The role of the motor track in transmitting strain will be established, by mechanically altering the actin filament geometry. In addition, communication mechanisms will be identified for myosin V and myosin VI through load dependent stepping measurements under conditions where the two heads of the motor are uncoupled. These mechanisms will be contrasted with those for myosin X, a motor with unusual in vivo motility that suggests a novel form of strain sensing. The long term goal of this research is to reveal how motors, and systems of motors, have been optimized for their cellular trafficking tasks. Myosin motility is required for various normal and pathological cell functions, including development of polarized cellular structures, formation of cellular adhesions and tissue development, wound healing, intracellular trafficking, and cell division. This research will address critical questions about the underlying biology behind motor-related disease, including, among others, several forms of inherited deafness related to the proper development and maintenance of stereo cilia in sensory hair cells.