Discovery of human cardiac myosin regulatory sites, modulating cross-bridge kinetics in heart muscle

发现人类心肌肌球蛋白调节位点，调节心肌的跨桥动力学

基本信息

批准号：
9098891
负责人：
YURI NESMELOV
金额：
$ 44万
依托单位：
UNIVERSITY OF NORTH CAROLINA CHARLOTTE
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：
2016-04-05 至 2020-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9098891
关键词：
Active Sites Address Affect Animal Model Binding Binding Sites Cardiac Cardiac Myosins Charge Computer Simulation Data Development Electrostatics Exhibits Failure Genes Goals Head Heart Heart failure Human Kinetics Knowledge Lead Learning Left ventricular structure Molecular Molecular Conformation Molecular Motors Muscle Muscle function Mutagenesis Mutation Myocardium Myosin ATPase Myosin Heavy Chains N-terminal Nucleotides Oryctolagus cuniculus Output Performance Positioning Attribute Production Property Protein Isoforms Rattus Recombinants Research Site Site-Directed Mutagenesis Structure System Testing Therapeutic Transgenic Organisms Work abstracting adenoviral-mediated base beta-Myosin crosslink drug development experimental analysis improved insight melting molecular dynamics mutant novel therapeutics public health relevance research study skeletal

项目摘要

DESCRIPTION (provided by applicant): Abstract Healthy human cardiac muscle is composed of two myosin isoforms, alpha and beta, which are significantly different in kinetics of the cross-bridge cycle. The specific myosin isoform composition tunes muscle for maximum performance, force production, power output, and economy. The mechanisms behind this fine muscle's performance tune-up are not known, thus limiting our understanding of a fundamental muscle property. Our long-term goal is to apply insights gained from learning the origin of kinetic differences of cardiac myosin isoforms to help devise ways to restore impaired muscle function. We envision tune-up of failed heart muscle by mimicking the healthy muscle isoform composition. The immediate objective of this proposal is to elucidate the molecular mechanism of kinetic difference of cardiac myosin isoforms. Our central hypothesis is that isoform kinetics i determined by the electrostatic interactions at the regulatory site within myosin head. Our hypothesis is based on a significant body of experimental data, confirming possibility of myosin kinetics modulation. Three specific aims are to validate our hypothesis. The specific aims are focused on potential regulatory sites within different parts of myosin head, Loop1 (Aim 1), upper 50 kDa domain (Aim 2), and the force generating region (Aim 3). According to our working hypothesis, these regulatory sites modulate myosin-nucleotide interaction, including the rate of ADP release from the cross-bridge, which is the rate limiting step of the slow beta myosin isoform. We plan to use site specific mutagenesis, altering electrostatic interactions within proposed regulatory sites. We will prepare mutants of recombinant human cardiac beta myosin and will characterize kinetics of prepared myosin mutants. We will examine our hypothesis on importance of electrostatic interactions within proposed regulatory sites on myosin kinetics modulation. The proposed research is significant because it will provide a detailed understanding of the mechanism by which human cardiac muscle is fine tuned for maximum performance. We will gain new insights into fundamental mechanisms by which produced force and power, and muscle energetics are modulated by muscle isoform composition. Elucidating kinetic tune-up mechanisms may lead to therapeutic ways of restoring or improving muscle function.

描述（由申请人提供）：摘要健康的人类心肌由两种肌球蛋白亚型（α 和 β）组成，它们的跨桥循环动力学显着不同，特定的肌球蛋白亚型成分可调节肌肉以获得最大性能、力量产生、这种精细肌肉的性能调节背后的机制尚不清楚，因此限制了我们对基本肌肉特性的理解，我们的长期目标是应用从学习动力学起源中获得的见解。我们设想通过模仿健康肌肉同种型的组成来调整衰竭的心肌，该提案的直接目标是阐明心肌肌球蛋白动力学差异的分子机制。我们的中心假设是，同种型动力学是由肌球蛋白头部调节位点的静电相互作用决定的。我们的假设基于大量实验数据，证实了肌球蛋白的可能性。三个具体目标是验证我们的假设，具体目标集中在肌球蛋白头、Loop1（目标 1）、上 50 kDa 结构域（目标 2）和力产生区域（目标）内的潜在调节位点。 3）根据我们的工作假设，这些调节位点调节肌球蛋白-核苷酸相互作用，包括ADP从跨桥释放的速率，这是慢β肌球蛋白亚型的限速步骤。计划使用位点特异性诱变，改变拟议的调节位点内的静电相互作用我们将制备重组人心脏β肌球蛋白的突变体，并将表征所制备的肌球蛋白突变体的动力学我们将检查我们关于肌球蛋白上拟议的调节位点内静电相互作用的重要性的假设。所提出的研究意义重大，因为它将提供对人类心肌微调以获得最大性能的机制的详细了解，我们将对产生力和功率以及肌肉的基本机制获得新的见解。能量学是由肌肉亚型组成调节的，阐明动力学调节机制可能会导致恢复或改善肌肉功能的治疗方法。