Diversity Supplement to Skeletal Myosin-Binding Protein C Regulation and Structural Dynamics

骨骼肌球蛋白结合蛋白 C 调节和结构动力学的多样性补充

• 批准号: 10824055
• 负责人: Brett A Colson
• 金额: $ 5.22万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10824055
• 关键词: Acceleration; Actins; Actomyosin; Address; Affect; Anisotropy; Arthrogryposis; Binding; Binding Proteins; Biochemical; Biological Assay; Biological Models; Biophysical Process; Biophysics; C2 Domain; Cardiac Myosins; Complex; Contracture; Cyclic AMP-Dependent Protein Kinases; DAG/PE-Binding Domain; Deceleration; Detection; Development; Disease; Distal; Drug Screening; Equilibrium; Filament; Fluorescence; Fluorescence Resonance Energy Transfer; Foundations; Frequencies; Gel; Genes; Genetic; Goals; Health; Human; Hypertrophic Cardiomyopathy; In Vitro; Joints; Kinetics; Knowledge; Label; Learning; Live Birth; Location; Lower Extremity; Measures; Mediating; Medical; Methods; Microfilaments; Molecular; Molecular Conformation; Monitor; Muscle; Muscle Cells; Muscle Proteins; Muscle Weakness; Muscle function; Mutation; Myocardial; Myopathy; Myosin ATPase; N-terminal; Outcome; Parents; Pathogenicity; Pathologic; Performance; Pharmaceutical Preparations; Pharmacotherapy; Phosphoproteins; Phosphorylation; Phosphorylation Site; Phosphotransferases; Physiological; Play; Positioning Attribute; Protein Dynamics; Protein Isoforms; Proteins; Publishing; RNA Splicing; Recombinants; Regulation; Reporter; Reporting; Research; Research Project Grants; Research Training; Resolution; Role; Sarcomeres; Scientist; Site; Skeletal Muscle; Slide; Spectrum Analysis; Stains; Structure; Technology; Testing; Therapeutic; Thick Filament; Thin Filament; Time; Training; Tropomyosin; Troponin; Upper Extremity; Variant; Visualization; Work; biophysical tools; career development; design; effective therapy; heart function; improved; innovation; insight; mutant; myosin-binding protein C; nanometer; next generation; novel; novel therapeutics; parent grant; parent project; phosphorescence; prevent; protein complex; protein structure; reconstitution; skeletal; skills; spectroscopic data; spectroscopic survey; therapeutic target; time use; tool

Title of project: DIVERSITY SUPPLEMENT TO SKELETAL MYOSIN-BINDING PROTEIN C REGULATION & STRUCTURAL DYNAMICS. Summary of the diversity supplement to the parent project. Skeletal myosin-binding protein C (MyBP-C) plays a major role in the modulation of cardiac function by its phosphorylation and causes deficits in contractile function due to MyBP-C mutations in distal arthrogryposis (DA) and the role of phosphorylation and DA mutations is not known. Our goal is to understand the molecular biophysics of muscle and to train the next generation of muscle biophysicists, inclusive of diverse trainees. The parent research project and diversity supplement ask fundamental questions about the role of protein interactions and structural dynamics that regulate function in skeletal muscle. To gain insight into the correlation of structure- function involved in MyBP-C mechanisms in physiological and pathological settings, we will probe the actin- myosin-MyBP-C complex of these proteins in solution with varied binding, phosphorylation, DA mutations, and MyBP-C drugs. Our core technology is site-directed spectroscopy, applied to purified MyBP-C and actin/myosin filaments. We will apply innovative complementary methods in site-directed labeling and spectroscopy to correlate protein binding, structural dynamics and function. We will test the central hypothesis that phosphorylation and DA mutations influence N-terminal and central domain skeletal MyBP-C binding with actin and/or myosin in a dynamic equilibrium to modulate contraction. Related to the parent grant, the first period of the diversity supplement focuses on using spectroscopic approaches to accurately measure the structural dynamics within, and adjacent to, the Pro/Ala-rich linker (P/A) of purified skeletal MyBP-C fragment C1-C7, primarily by measuring nanometer distances and molecular disorder. Major emphasis is placed on detection of conformational changes (structure) within and nearby MyBP-C’s P/A due to phosphorylation, DA mutation, actin or myosin binding (function), and drugs. By including the location of probes in P/A and in adjacent C1 and C2 domains, the Candidate will measure structural changes. Fluorescently-labeled MyBP-C will be prepared to acquire fluorescence lifetime using time-resolved methods. Human splice variants containing and missing the phosphorylation site in long and short forms in P/A will be evaluated. In the second period, the Candidate will learn new skills in spectroscopic data fitting analysis to determine probe-to-probe distances and disorder in N- terminal and central domain MyBP-C. The third period will provide molecular details of the structural dynamics upon phosphorylation of P/A in long form sMyBP-C and actin- or myosin- MyBP-C complexes. The Candidate will systematically build in model system complexity, from unbound to actin/myosin-bound MyBP-C, upon phosphorylation. Spectroscopic study of sMyBP-C regulation will determine protein interactions and structural dynamics, providing key insights at the myofilament level to be applied for understanding fundamental mechanisms in the muscle cell. This project is grounded in fundamental biophysics mechanisms, but MyBP-C has emerged as a therapeutic target for skeletal muscle disease. Thus, of our work lays a foundation for testing identified drugs and development of screens for drug therapies using our unique spectroscopic approaches.

项目名称：骨骼肌球蛋白结合蛋白C调控的多样性补充