INTRINSIC PROBES OF SMOOTH MUSCLE MYOSIN DYNAMICS

平滑肌肌球蛋白动力学的内在探针

基本信息

批准号：
7373654
负责人：
CHRISTOPHER L. BERGER
金额：
$ 36.92万
依托单位：
UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-02-01 至 2010-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7373654
关键词：
ATP Hydrolysis ATP phosphohydrolase ATP-G-actin Actins Active Sites Address Affinity Binding Biochemical Biological Clocks Cardiovascular Diseases Catalytic Domain Chemicals Cleaved cell Coupled Cytokinesis DNA Sequence Rearrangement Disease Engineering Event F-Actin Familial Hypertrophic Cardiomyopathy Fluorescence Fluorescent Probes Goals Hydrolysis Inherited Kinetics Lead Link Location Mediating Molecular Molecular Motors Monitor Motion Motor Movement Muscle Muscle Contraction Mutation Myosin ATPase Nucleotides Organelles Point Mutation Positioning Attribute Process Production Protein Isoforms Proteins Relative (related person)Reporting Research Research Personnel Series Signal Transduction Site Skeletal system Smooth Muscle Myosins Solutions Staging Structure Testing Thinking Time Tryptophan Upper arm actin 2 cell motility design desire inorganic phosphate interest programs research study response

项目摘要

The ultimate goal of this proposal is to examine conformational changes at specific locations within smooth muscle myosin using intrinsic tryptophan fluorescence. Smooth muscle myosin constructs will be genetically engineered to contain either a single tryptophan or a pair of tryptophans at the desired site of interest, which will provide a unique intrinsic fluorescence signal reporting local conformational and structural changes in response to nucleotide binding, ATP hydrolysis, actin-binding, and lever arm movement. These experiments are complementary to the ongoing structural studies in the field, allowing us to explicitly test predictions about domain movements and structural rearrangements during critical steps in the contractile cycle smooth muscle myosin. Thus we will be able to correlate structural changes in myosin with functional consequences, which relates directly to certain cardiovascular disease. For example, FHC (familial hypertrophic cardiomyopathy) is an inherited, often lethal disease caused by point mutations at key structures within myosin critical to its proper functioning as a molecular motor. We will be examining structural changes in myosin in regions of the molecule directly impacted by mutations that underlie FHC. This will lead to a better understanding of the disease, and thus to better treatment options as well. Therefore, this proposal offers a unique opportunity to critically test fundamental questions about the molecular mechanism of muscle contraction that have not been previously accessible by other spectroscopic probe studies, and the results will have important implications for serious disease states such as FHC.

该提案的最终目标是检查平滑内特定位置的构象变化使用内在色氨酸荧光的肌肉肌球蛋白。平滑肌肌球蛋白结构将通过遗传被设计为在所需的感兴趣位点包含单个色氨酸或一对色氨酸，这将提供独特的内在荧光信号，报告局部构象和结构变化对核苷酸结合、ATP 水解、肌动蛋白结合和杠杆臂运动的反应。这些实验与该领域正在进行的结构研究相辅相成，使我们能够明确地测试预测关于收缩周期平滑关键步骤期间的域运动和结构重排肌肉肌球蛋白。因此，我们将能够将肌球蛋白的结构变化与功能后果联系起来，与某些心血管疾病有直接关系。例如，FHC（家族性肥大症）心肌病）是一种遗传性的、通常致命的疾病，由体内关键结构的点突变引起肌球蛋白对其作为分子马达的正常功能至关重要。我们将研究结构性变化分子区域中的肌球蛋白直接受 FHC 突变的影响。这将带来更好的结果了解该疾病，从而获得更好的治疗方案。因此，本提案提供了一个批判性测试有关肌肉分子机制的基本问题的独特机会以前其他光谱探针研究无法获得的收缩，以及结果将对 FHC 等严重疾病状态产生重要影响。