Impact of cardiomyopathy mutations on cardiac myosin structure and function

心肌病突变对心肌肌球蛋白结构和功能的影响

基本信息

批准号：
9028146
负责人：
CHRISTOPHER M YENGO
金额：
$ 48.15万
依托单位：
PENNSYLVANIA STATE UNIV HERSHEY MED CTR
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-02-15 至 2020-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9028146
关键词：
ATP Hydrolysis ATP phosphohydrolase Actins Actomyosin Actomyosin Adenosinetriphosphatase Binding Biochemical Biological Process Cardiac Myocytes Cardiac Myosins Cardiomyopathies Cell division Classification Cleaved cell Communication Congenital cardiomyopathy Coupled Coupling Data Defect Depressed mood Development Disease Dissociation Drug Targeting Fluorescence Fluorescence Resonance Energy Transfer Foundations Functional disorder Generations Goals Heart Heart failure Human Impairment In Vitro Intracellular Transport Kinetics Label Lead Light Link Measures Mediating Microfilaments Modeling Molecular Molecular Motors Motion Motor Motor Activity Movement Muscle Contraction Mutation Myocardium Myosin ATPase Myosin Type V Nucleotides Pathogenesis Pathology Pathway interactions Patients Pharmaceutical Preparations Phenotype Point Mutation Power stroke Property Proteins Research Site Staging Structural defect Structure Sudden Death System Therapeutic Thermodynamics Time Ventricular arm base beta-Myosin cell motility heart function inorganic phosphate insight mutant novel novel therapeutics programs public health relevance research study small molecule

项目摘要

DESCRIPTION (provided by applicant): The ability of myosin to generate force and motion through its interaction with actin filaments is essential to many biological processes including muscle contraction, cell division, and intracellular transport. The atomic level structures of myosin in various stages of its enzymatic cycle have provided a framework of the molecular mechanism of force generation utilized by myosin. These structures as well as other biochemical and structural data suggest that myosin generates force by coupling small conformational changes in the nucleotide-binding region to a large swing of the light-chain binding region (lever arm) while myosin is strongly bound to actin. Mutations in human beta cardiac myosin are associated with several forms of cardiomyopathies, while it is unclear how the mutations lead to different disease pathologies. We propose the mutations alter the conserved structural mechanism of force generation by disrupting the subdomain coordination necessary for actin to activate the release of the products of ATP hydrolysis (phosphate and ADP) and trigger the force generating swing of the lever arm. We will investigate how the mutations impact specific conformational changes in the actin-binding, nucleotide-binding, and lever arm regions. Novel extrinsic fluorescence probes will be strategically placed to measure conformational changes in these three critical regions using fluorescence resonance energy transfer (FRET). In addition, transient kinetic experiments will be used to correlate the conformational changes with specific biochemical steps in the actomyosin ATPase cycle. The shift in the ensemble of structural states during key biochemical transitions will be examined by transient time resolved FRET. We will also investigate how the mutations alter the enzymatic and force generating properties of myosin, which will allow us to develop detailed models of how the mutations impair motor structure and function. We will determine how the cardiac myosin activator drug alters the conformational dynamics of human beta cardiac myosin and determine if it can rescue the altered motor structure-function in the cardiomyopathy mutants. Overall, our studies will be instrumental in developing therapeutic drugs that target myosin motor activity in heart failure and establishing the structural defects associated with cardiomyopathy mutations in myosin.

描述（由适用提供）：肌球蛋白通过与肌动蛋白丝相互作用通过其与肌动蛋白丝相互作用产生力和运动的能力对于许多生物学过程至关重要，包括肌肉收缩，细胞分裂，细胞分裂和细胞内转运。肌球蛋白在其酶促周期的各个阶段的原子水平结构提供了肌球蛋白使用的力产生的分子机理的框架。这些结构以及其他生物化学和结构数据表明，肌球蛋白通过将核刺接结构区域的小会议变化与轻链结合区域（杠杆臂）的大挥杆耦合而产生力，而肌球蛋白则与肌动蛋白密切相关。人β心脏肌球蛋白中的突变与几种形式的心肌病有关，而尚不清楚突变如何导致不同的疾病病理。我们提出的突变通过破坏肌动蛋白激活ATP水解产物（磷酸盐和ADP）并触发杠杆臂挥杆的力所需的亚域的配位来改变力产生的结构机制。我们将调查突变如何影响肌动蛋白结合，核震准结合和杠杆部门区域的特定会议变化。新型外部荧光问题将在战略上放置，以使用荧光共振能量转移（FRET）来衡量这三个关键区域的会议变化。另外，将使用瞬态动力学实验将构象变化与肌动蛋白ATPase周期中的特定生化步骤相关联。在关键生物化学转变过程中结构状态的合奏的变化将通过瞬时时间解析的fret进行检查。我们还将研究突变如何改变肌球蛋白的酶促和力产生特性，这将使我们能够开发出突变如何损害运动结构和功能的详细模型。我们将确定心脏肌球蛋白激活剂药物如何改变人β心脏肌球蛋白的构象动力学，并确定它是否可以挽救心肌病突变体中改变的运动结构功能。总体而言，我们的研究将有助于开发靶向肌球蛋白运动活性在心力衰竭中并确定与肌球蛋白中心肌病突变相关的结构缺陷的治疗药物。