Allostery in myosins studied at the molecular level

描述（由申请人提供）：在典型真核细胞的拥挤环境中，任何大于50 nm的物体都是有效不移动的，不能依靠扩散到达目的地。结果，肌凝蛋白运动进化为为货物运输、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。这些马达在与肌动蛋白丝结合时，通过以受控的方式改变形状来转换化学自由能。所有运动蛋白的一个基本特征是，它们的运动是精心协调的，因此，运动通过一个特定的生化和机械状态耦合序列。这种协调的性质尚不清楚，但它显然对正常的运动功能至关重要。最近认识到，每个肌凝蛋白类已经进化出特定的结构和动力学特征，允许跨肌凝蛋白超家族不同的协调机制的比较。这些个体适应现在可以通过一系列先进的运动分析来表征。本文提出的工作将利用新的单分子操作来改变马达感受到的分子内应变，从而确定变构配位机制。通过机械地改变肌动蛋白丝的几何形状，电机轨迹在传递应变中的作用将得到确立。此外，将通过负载相关步进测量确定肌凝蛋白V和肌凝蛋白VI在电机两个磁头不耦合的情况下的通信机制。这些机制将与肌球蛋白X的机制形成对比，肌球蛋白X是一种具有不寻常的体内运动的马达，表明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优化它们的细胞运输任务。肌球蛋白的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发展、细胞粘连和组织发育的形成、伤口愈合、细胞内运输和细胞分裂。这项研究将解决运动相关疾病背后的关键生物学问题，包括与感觉毛细胞中立体纤毛的适当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.