Functional diversity of myosin VII

描述（由申请人提供）：人类有两种VII类肌球蛋白，myosin VIIa和VIIb，它们在结构域结构上非常相似，除了myosin VIIb在肌动蛋白结合位点具有独特的19个氨基酸插入，并且缺乏假定的卷曲结构域。肌球蛋白VIIa的突变与1b型Usher综合征（人类最常见的耳聋-失明疾病）和另外两种人类听力损失疾病DFNB2和DFNA11有关。Myosin VIIb被认为运输和组织膜和膜相关蛋白。尽管VII类肌凝蛋白在细胞中发挥着关键作用，但由于缺乏关于肌凝蛋白VII的化学和物理特性的信息，对肌凝蛋白VII功能的基础和与肌凝蛋白VIIa相关的耳聋表型的理解受到限制。

DESCRIPTION (provided by applicant): Humans have two class VII myosins, myosin VIIa and VIIb, which are very similar in domain structure except that myosin VIIb possesses a unique 19 amino acid insert at the actin-binding site and lacks a putative coiled-coil domain. Mutations in myosin VIIa have been linked to type 1b Usher syndrome, the most common deafness-blindness disorder in humans, and two other hearing loss disorders in humans, DFNB2 and DFNA11. Myosin VIIb is believed to traffic and organize membranes and membrane-associated proteins. Despite the critical roles class VII myosins play in cells, understanding the basis of myosin VII function and the deafness phenotype associated with myosin VIIa has been limited by a lack of information regarding the chemical and physical properties of myosin VII. Efforts in this proposal focus on understanding the enzymatic and motor properties of myosin VIIa and VIIb. Aim 1 will determine if full-length myosin VIIa and VIIb are monomers or dimers. Aim 2 will provide a complete kinetic characterization of the myosin VIIa and VIIb ATPase cycles, determine the contributions of the 19 amino acid insertion to the ATPase cycle kinetics through the generation of mutants and chimeric proteins, and identify how mutations that generate deafness affect myosin VIIa motor activity. Aim 3 will measure how external loads modulate ADP and actin binding to high duty ratio myosins and evaluate mechanisms of strain-dependent head-head coordination. Aim 4 will employ high-resolution spectroscopic methods to measure cooperative changes in the structural dynamics of actin filaments induced by high duty ratio myosin binding. The two highly conserved isoforms provide a unique opportunity to test the structural and functional effects of the actin-binding loop and of dimerization on myosin function. The proposed detailed analysis of reaction mechanisms and structural dynamics will help develop testable hypotheses regarding the function and mechanism of myosin VII, and illustrate how sequence and structural variations in myosin motors define their enzymatic properties, which ultimately dictates their biological functions. Because mutations and disruption of myosin VIIa function are of clinical relevance, the proposed experiments will advance our understanding of the molecular basis of several human diseases.