Structural Dynamics of the Myosin Super Relaxed State in Aging Muscle and Metabolic Disease

衰老肌肉和代谢疾病中肌球蛋白超松弛状态的结构动力学

• 批准号: 10223178
• 负责人: Lien Ai Phung
• 金额: $ 2.24万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10223178
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Actins; Address; Adipose tissue; Affect; Age; Aging; Animals; Attenuated; Basal metabolic rate; Binding; Biochemistry; Biological Assay; Biological Process; Biophysics; Body Temperature; Body Weight; Calcium; Chemicals; Cues; Data; Development; Diabetes Mellitus; Diabetic mouse; Disease; Disease model; Doctor of Philosophy; Elderly; Energy Metabolism; Estradiol; Fellowship; Female; Fluorescence; Fluorescence Resonance Energy Transfer; Fostering; Functional disorder; Future; Goals; Gonadal Steroid Hormones; Health Status; Hormonal; Immunochemistry; Impairment; In Vitro; Incidence; Individual; Insulin; Kinetics; Knowledge; Life; Light; Link; Maps; Measures; Mechanics; Medical; Metabolic; Metabolic Diseases; Microscopy; Mission; Molecular; Molecular Mechanisms of Action; Mus; Muscle; Muscle Contraction; Muscle Fibers; Muscle Weakness; Muscle function; Myopathy; Myosin ATPase; Nature; Nucleotides; Obese Mice; Obesity; Optics; Phosphorylation; Physicians; Physiological; Physiology; Play; Population; Postmenopause; Predisposition; Prevalence; Problem Solving; Process; Production; Proteins; Regulation; Research Personnel; Rest; Role; Scientist; Site; Skeletal Muscle; Skin; Specificity; Spectrum Analysis; Structure; Technology; Technology Transfer; Temperature; Testing; Therapeutic; Thermogenesis; Thick Filament; Time; Training; Translational Research; Vertebral column; Woman; Work; age related; aged; attenuation; base; design; genetic regulatory protein; improved; innovation; insight; kinetosome; lens; link protein; male; men; metabolic rate; molecular dynamics; mouse model; muscle aging; muscle physiology; novel; novel therapeutics; obesity treatment; physically handicapped; response; sarcopenia; sex; skeletal; small molecule therapeutics; therapeutic target; tool; training opportunity

Project Summary The myosin super-relaxed (SRX) state, first described six years ago, is emerging as an important player in muscle regulation and function, yet its mechanistic structure and dynamics have not been resolved. The goal of this proposal is to incorporate time-resolved fluorescence resonance energy transfer (TR-FRET) with quantitative epifluorescence microscopy of fluorescent nucleotides, to detect and quantitate the SRX in chemically skinned skeletal muscle fibers from different disease mouse models. The long-term goal is to establish the role of SRX in age-related attenuation of muscle function, and use these insights to design therapeutic approaches. The population of SRX myosin in resting skeletal muscle is as much as 50%. These molecules are sequestered on the myosin thick-filament backbone and are thus auto-inhibited, catalyzing ATP hydrolysis about 10 times slower than purified myosin in solution, while generating no force. My proposal focuses on the role of SRX in the attenuation of muscle function with age, focusing on three independent but inter-related Aims: concerning the roles of sex-specificity (Aim 1), muscle thermogenesis and thus control of whole body basal metabolic rate (Aim 2), and metabolic diseases such as obesity and diabetes (Aim 3). Thus, the myosin SRX provides a platform for potential innovative treatments for obesity and diabetes. However, the molecular mechanism by which this SRX state affects aging muscle physiology and pathophysiology is unresolved. To solve this problem, my effort will be focused on applying site-directed spectroscopic probes on myosin to elucidate its structural changes as it undergoes transitions between the super-relaxed state and the normal relaxed state. The SRX can be rapidly mobilized into the normal relaxed or the active state in response to regulatory allosteric cues such as increased in cellular calcium, regulatory light chain (RLC) phosphorylation, and decreased temperature. These perturbations will be thoroughly tested on each proposed mouse model (Aims 1-3). This will be the first time that the molecular dynamics of SRX myosin will be directly studied through the physiological lens of aging and its associated metabolic disorders. Results from this work will provide crucial insights into the nature of the aging process on skeletal myosin and map a path toward novel therapies to treat diseases and extend healthy, active years of life by targeting myosin. The present fellowship proposal will also provide extensive training opportunity in muscle biochemistry, biophysics, and physiology, contributing to a meaningful MD/PhD degree. The training will prepare me to become an independent biomedical researcher with the means to apply optical spectroscopy to translational science, to both appreciate the molecular basis of disease and subsequent development of new treatments. This work aligns strongly with the mission statement of the NIA, by increasing the molecular understanding of the biological processes on aging along with fostering the development of a physician-scientist.

项目摘要 六年前首次描述的肌球蛋白超级省（SRX）状态正在成为重要的参与者 肌肉调节和功能，但其机械结构和动力学尚未解决。目标 该建议的内容是结合时间分辨的荧光共振能量传递（TR-FRET） 荧光核苷酸的定量表荧光显微镜，以检测和定量SRX中的SRX 来自不同疾病小鼠模型的化学皮肤骨骼肌纤维。长期目标是 确定SRX在与年龄相关的肌肉功能衰减中的作用，并使用这些见解来设计 治疗方法。在静止的骨骼肌中，SRX肌球蛋白的种群高达50％。这些 分子在肌球蛋白厚丝骨架上隔离，因此自动抑制，催化ATP 水解在溶液中的纯肌球蛋白慢约10倍，而没有产生力。我的建议 专注于SRX在随着年龄的年龄的肌肉功能衰减中的作用，专注于三个独立但 相互关联的目的：涉及性别特异性的作用（AIM 1），肌肉热发生，从而控制 全身基础代谢率（AIM 2）以及肥胖和糖尿病等代谢疾病（AIM 3）。因此， 肌球蛋白SRX为肥胖和糖尿病的潜在创新治疗提供了一个平台。但是， 该SRX状态影响衰老肌肉生理和病理生理学的分子机制是 未解决。为了解决这个问题，我的努力将专注于在现场定向的光谱探针上 肌球蛋白阐明其结构变化，因为它经历了超省略状态和 正常的放松状态。 SRX可以迅速动员到正常的放松状态或活跃状态以响应 调节性变构线索，例如细胞钙，调节轻链（RLC）磷酸化的增加， 并降低温度。这些扰动将在每个提出的鼠标模型上进行彻底测试 （目标1-3）。这将是第一次直接研究SRX肌球蛋白的分子动力学 通过衰老及其相关的代谢性疾病的生理视角。这项工作的结果将 提供对骨骼肌球蛋白衰老过程的性质的关键见解，并绘制通向新颖的路径 通过靶向肌球蛋白来治疗疾病并延长健康活跃的生活的疗法。目前的奖学金 建议还将在肌肉生物化学，生物物理学和生理学方面提供广泛的培训机会， 促进有意义的MD/PHD学位。培训将使我成为一个独立的人 生物医学研究人员的手段将光谱法应用于转化科学，两者都欣赏 疾病的分子基础以及随后的新疗法发展。这项工作与 NIA的使命陈述，通过提高对生物学过程的分子理解 衰老，并促进医师科学家的发展。