Enzymatic and motor properties of myosin III

描述（由申请人提供）：本研究项目的长期目标是了解肌球蛋白III的酶和运动特性，使其能够在光感受器的光转导过程中发挥直接作用。肌球蛋白III是一种基于肌动蛋白的分子马达，包含肌球蛋白超家族特有的保守马达结构域，但也包含N-末端激酶结构域以及其C-末端尾部的额外肌动蛋白结合基序。我们建议，激酶结构域上的肌球蛋白结构域的自磷酸化位点的能力降低了肌球蛋白III的酶和运动特性，但当激酶结构域被灭活或绑定到另一个基板的肌球蛋白III电机被激活。我们将研究肌球蛋白III结构与激酶结构域删除或失活，以及在细胞底物的激酶结构域的存在下的酶和运动特性。肌球蛋白III的激酶调节可能提供了一种机制，从而肌球蛋白III可以响应特定的细胞信号被激活/抑制。我们建议，额外的肌动蛋白结合位点的尾巴允许肌球蛋白III交联肌动蛋白丝，协同激活肌球蛋白III酶循环，并通过增加肌球蛋白的整体亲和力肌动蛋白增强肌球蛋白III的能动性。我们将直接检查肌动蛋白结合亲和力的尾部肌动蛋白结合基序，并确定它在介导肌球蛋白III的酶和运动特性的作用。因此，额外的肌动蛋白结合位点可能允许肌球蛋白III定位于肌动蛋白丝网络，并帮助稳定肌动蛋白丝网络，这是组装参与光转导信号复合物的蛋白质所必需的。总的来说，研究肌球蛋白III的酶促、马达和调节特性可能揭示光转导信号复合物如何在光感受器中形成和稳定的潜在机制。

DESCRIPTION (provided by applicant): The long-term goal of this research project is to understand the enzymatic and motor properties of myosin III that allow it to play a direct role in the phototransduction process in photoreceptors. Myosin III is an actinbased molecular motor that contains a conserved motor domain characteristic of the myosin superfamily, but also contains an N-terminal kinase domain as well as an additional actin-binding motif in its C-terminal tail. We propose that the ability of the kinase domain to autophosphorylate sites on the myosin domain reduces the enzymatic and motile properties of myosin III, but when the kinase domain is inactivated or bound to another substrate the myosin III motor is activated. We will examine the enzymatic and motile properties of myosin III constructs with the kinase domain deleted or inactivated, as well as in the presence of cellular substrates for the kinase domain. The kinase regulation of myosin III may provide a mechanism whereby myosin III can be activated/inhibited in response to specific cellular signals. We propose that the additional actin binding site in the tail allows myosin III to cross-link actin filaments, cooperatively activate the myosin III enzymatic cycle, and enhance the motile properties of myosin III by increasing the overall affinity of myosin for actin. We will directly examine the actin binding affinity of the tail actin-binding motif, and determine the role that it plays in mediating the enzymatic and motile properties of myosin III. Thus, the additional actin-binding site may allow myosin III localize to and help stabilize actin filament networks, which are necessary for assembling the proteins involved in the phototransduction signaling complex. Overall, studying the enzymatic, motor, and regulatory properties of myosin III may reveal potential mechanisms of how the formation and stabilization of the phototransduction signaling complex occurs in photoreceptors.