CORRELATION OF MUSCLE PHYSIOLOGY AND BIOCHEMISTRY

这项研究的目的是将生理、结构和 从蛋白质测量看肌肉收缩的生化理论 肌动球蛋白ATPase的运动和生化速率常数 具有生理活性的骨骼肌和心肌制剂。蛋白 肌肉的结构变化将在电子显微镜下被检测到 纤维在超速冷冻过程中开始拉伸发展后 笼状三磷酸腺苷或笼状钙离子的激光脉冲光解活化。关在笼子里 分子是惰性的，但不能发光的底物或信号的前体 在脉冲脉冲照射下释放活性化合物的分子 近乎UV。激光。蛋白质的旋转运动将用 一种新的时间分辨荧光偏振方法，将允许 微观空间分辨率。荧光肌动蛋白和肌球蛋白灯 链将被微量注射到活的、分离的心脏或培养的心脏中 骨骼肌细胞。标记的肌动蛋白或肌球蛋白光的方向 链条将在自发或 电刺激的收缩。具体的反应步骤 ATPase在工作肌肉纤维中的作用机制将通过 测定稳定氧同位素在溶剂和溶剂之间的交换 底物和ATPase反应产物中的磷酸基团。 同位素的分布将用气相色谱测定。 与磷酸盐化合物的质谱学联用。这些氧气 交换技术探索了细菌中的基本生化反应。 完整的灯丝格子。细丝滑动对这些参数的影响 将确定反应步骤。 这个项目的结果应该会大大促进人们对 收缩过程，从而更好地了解两者的正常 以及骨骼肌和心肌以及其他类型的 细胞死亡率。

The goal of this research is to relate physiological, structural and biochemical theories of muscle contraction by measurement of protein motions and biochemical rate constants of the actomyosin ATPase in physiologically active skeletal and heart muscle preparations. Protein structural changes will be detected in electron micrographs of muscle fibers ultra-rapidly frozen during the onset of tension development after activation by laser pulse photolysis of caged ATP or caged Ca2+. Caged molecules are inert, but photolabile precursors of substrate or signaling molecules that release the active compound on irradiation by a pulse of near-u.v. laser light. Protein rotational motions will be measured using a novel time-resolved fluorescence polarization method that will allow microscopic spatial resolution. Fluorescent actin and myosin light chains will be micro-injected into live, isolated cardiac or cultured skeletal myocytes. The orientation of the labeled actin or myosin light chains will be monitored during the time course of spontaneous or electrically stimulated contractions. Specific reaction steps of the ATPase mechanism will be investigated in working muscle fibers by measuring exchange of stable oxygen isotopes between the solvent and phosphate groups in the substrate and products of the ATPase reaction. The distribution of isotopes will be determined by gas chromatography coupled to mass spectrometry of the phosphate compounds. This oxygen exchange techniques probes the elementary biochemical reactions in the intact filament lattice. The influence of filament sliding on these reaction steps will be determined. Results from this project should significantly advance knowledge of the contractile process and thus bring a greater understanding of both normal and pathological states of skeletal and heart muscle and other types of cell mortality.