CRYO-DIFFRACTION OF INSECT FLIGHT MUSCLE

昆虫飞行肌的冷冻衍射

• 批准号: 7601606
• 负责人: MICHAEL REEDY
• 金额: $ 0.55万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7601606
• 关键词: Computer Retrieval of Information on Scientific Projects Database; Condition; Fiber; Filament; Freezing; Funding; Grant; Image; Insecta; Institution; KLK3 gene; Lead; Length; Microfilaments; Microtomy; Motor; Muscle; Myosin ATPase; Pattern; Purpose; Radiation; Rana; Research; Research Personnel; Resolution; Resources; Roentgen Rays; Sampling; Sarcomeres; Scheme; Skeletal system; Source; Specimen; Striated Muscles; Structure; System; Thick Filament; United States National Institutes of Health; Work; X ray diffraction analysis; X-Ray Diffraction; detector; novel; programs; research study; tomography

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our program combines & compares X-ray diffraction (glycerinated native fibers) and thin-section EM tomography (fibers quick-frozen & freeze-substituted) to characterize structure, arrangement and dynamic choreography of myosin crossbridges (motor molecules of muscle) in a waterbug insect flight muscle (IFM) of unexcelled crystalline regularity. All X-ray experiments proposed will, whenever revealing, be matched by parallel EM experiments on fibers quick-frozen during the same state or maneuver, to directly image the structural changes. (IFM gives the clearest EM images of myosin crossbridges of any muscle.) X-ray patterns from IFM differ in important ways from those of other striated muscles (frog skeletal) studied by XRD, owing to differences in filament lattice packing, in thick filament helical geometry, and a full-overlap sarcomere scheme limited to ~3% length changes. X-ray reflections (to ~5 nm) attributable to myosin filaments, actin filaments, and both jointly, are as well worked out as in frog muscle, but not yet so well studied in dynamic (contraction) states. Thanks to the beam quality, intensity and detector systems at the APS, much of the catch-up with frog muscle studies can be done in 2-4 years. The present proposal is to continue our novel recent work on crossbridge braking actions. The purpose of this rapid access proposal is to evaluate freezing conditions that can enhance sample lifetime so that much higher resolution diffraction patterns can be obtained from small specimens before radiation damage sets in. Because of the weakness of the diffraction from the muscle system this study must be done on a synchrtrron source. As far as we know this has not been successfully achieved in muscle before and may be expected to lead to novel findings.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 我们的项目结合并比较了 X 射线衍射（甘油天然纤维）和薄片 EM 断层扫描（快速冷冻和冷冻替代的纤维），以表征具有无与伦比的晶体规律性的水蝽昆虫飞行肌 (IFM) 中肌球蛋白横桥（肌肉运动分子）的结构、排列和动态编排。 所有提出的 X 射线实验，无论何时揭示，都将与在相同状态或操作期间快速冷冻的纤维上的并行 EM 实验相匹配，以直接对结构变化进行成像。 （IFM 给出了任何肌肉的肌球蛋白横桥的最清晰的 EM 图像。）IFM 的 X 射线模式与 XRD 研究的其他横纹肌（青蛙骨骼）的 X 射线模式在重要方面有所不同，这是由于细丝晶格堆积、粗细丝螺旋几何形状和全重叠肌节方案限制为约 3% 长度变化的差异。 与青蛙肌肉一样，肌球蛋白丝、肌动蛋白丝以及两者共同产生的 X 射线反射（约 5 nm）也得到了很好的研究，但在动态（收缩）状态下尚未得到很好的研究。 得益于 APS 的光束质量、强度和探测器系统，大部分青蛙肌肉研究可以在 2-4 年内完成。 目前的建议是继续我们最近在横桥制动作用方面的新颖工作。 这项快速访问建议的目的是评估可以延长样品寿命的冷冻条件，以便在辐射损伤发生之前从小样品中获得更高分辨率的衍射图案。由于肌肉系统的衍射较弱，这项研究必须在同步源上进行。据我们所知，以前在肌肉中尚未成功实现这一点，并且有望带来新的发现。