Kinetics of Cardiac Myofilament Activation

心肌丝激活动力学

• 批准号: 7597232
• 负责人: WEN-JI DONG
• 金额: $ 34.8万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7597232
• 关键词: Actins; Address; Affect; Back; Binding; Binding Sites; Biochemical; Biological Assay; Cardiac; Cardiac Muscle Contraction; Cardiomyopathies; Complex; Contractile Proteins; Coupled; Cyclic AMP-Dependent Protein Kinases; Data; Development; Diagnosis; Disease; Dissociation; Energy Transfer; Equilibrium; Event; Failure; Fiber; Goals; Heart; Heart failure; In Vitro; Individual; Kinetics; Knowledge; Lead; Measurement; Microfilaments; Modification; Molecular; Molecular Conformation; Monitor; Movement; Muscle; Muscle Contraction; Muscle Fibers; Muscle function; Myocardium; Myofibrils; Myosin ATPase; Myosin S-1; N Domain; Nucleic Acid Regulatory Sequences; Phosphorylation; Phosphorylation Site; Play; Post-Translational Protein Processing; Process; Protein Isoforms; Protein Kinase C; Proteins; Regulation; Relaxation; Reporting; Research; Research Personnel; Role; Series; Signal Transduction; Site; Skin; Striated Muscles; Structure; Study models; Surface; System; Testing; Thermodynamics; Thin Filament; Time; Titrations; To specify; Tropomyosin; Troponin C; Troponin I; base; design; feeding; genetic regulatory protein; information processing; mutant; prevent; programs; reconstitution; research study; response; skeletal; transmission process

DESCRIPTION (provided by applicant): Force development during striated muscle contraction is initiated by the binding of Ca2+ to the specific sites in troponin C (TnC), triggering a series of functional structural changes within the thin filament, including opening of the N-domain of TnC, conformational change of the inhibitory region of troponin I (Tnl), and switching interaction between Tnl and actin to Tnl and TnC, which ultimately lead to a cyclic interaction between actin and myosin to form strong force-generating cross-bridges. Full muscle activation requires both Ca2+ binding and feed back modulation of cross-bridge cycling. In cardiac muscle it is also modulated by protein phosphorylation which plays important roles in heart failing/hypertrophic process. To fully understand muscle regulatory mechanism requires structural, thermodynamic and kinetic information on each of these structural transitions during force development. My long-term research goal is to elucidate the kinetics of movements of the thin finlament betweem extremes of contraction/relaxation, and understand how they are modified by cross-bridge cycling and phosphorylation. To achieve the goal, this proposal addresses the following three issues: (1) What is the kinetics of each individual activation/deactivation process of the thin filament? (2) How does the cross-bridge cycling affect these kinetic processes? And (3) what is the role of phsophorylation in modulating these transitions? Newly designed conformational markers based on Forster resonance energy transfer to monitor these structural transitions will be used for stopped-flow kinetic and Ca2+ titration measurements at different activation conditions to acquire the desired information. These markers will be incorporated into reconstituted thin filament, myofibrils and skinned fibers along with/without phosphotylated proteins to specify the time-dependent changes of specific domain movements of the thin filament in response to Ca2+. Results of this study will enhance our understanding of molecular mechanisms of thin filament activation in response of Ca2+ and the role of protein phosphorylation in heart failure.

描述（由申请人提供）：通过CA2+与肌钙蛋白C（TNC）中的特定位点的结合开始，引发了肌肉收缩期间的力发育，从而触发了薄丝内部的一系列功能结构变化，包括TNC的N-域的开放，包括TNC的n-域，与TROPONIN I（TNL）的构象变化（tnl and tnl and tnl）的构象变化（TNL），以及TNL的抑制区域（TNL），TNL和TNL的抑制作用区域（TNL），TNL和TNL的抑制作用区域（TN）（TN）（TN）（TNC）的抑制作用（TNL）和TNL的抑制作用。最终导致肌动蛋白和肌球蛋白之间形成强大的跨桥。全肌肉激活需要CA2+结合和跨桥骑自行车的反馈调制。在心肌中，它也受到蛋白质磷酸化的调节，该蛋白质在心脏失败/肥厚过程中起着重要作用。为了充分理解肌肉调节机制，需要在力开发过程中对这些结构过渡中的每一个结构，热力学和动力学信息。我的长期研究目标是阐明收缩/放松极端的薄鳍片运动动力学，并了解如何通过跨桥循环和磷酸化来修饰它们。为了实现目标，该提案解决了以下三个问题：（1）薄丝细丝的每个个体激活/失活过程的动力学是什么？ （2）跨桥循环如何影响这些动力学过程？ （3）phsophorylation在调节这些过渡中的作用是什么？基于Forster共振能量传递以监测这些结构过渡的新设计的构象标记将用于在不同的激活条件下停止流动动力学和Ca2+滴定测量值，以获取所需的信息。这些标记将被掺入重构的细丝，肌原纤维和皮肤纤维，以及/没有磷酸化蛋白，以指定薄丝特定域运动的时间依赖性变化，以响应Ca2+。这项研究的结果将增强我们对Ca2+反应中细丝激活的分子机制以及蛋白质磷酸化在心力衰竭中的作用。