cryoEM Administrative Supplement for Equipment

冷冻电镜设备管理补充文件

• 批准号: 10389992
• 负责人: KENNETH ALLEN TAYLOR
• 金额: $ 24.96万
• 依托单位: FLORIDA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10389992
• 关键词: Administrative Supplement; Collection; Cryoelectron Microscopy; Data; Defect; Electrons; Equipment; Filament; Funding; Grant; Head; Image; Invertebrates; Length; Mechanics; Muscle; Muscle function; Mutation; Myopathy; Myosin ATPase; Paratropomyosin; Pattern; Proteins; Resolution; Role; Sarcomeres; Striated Muscles; Structure; Tail; Thick Filament; Thin Filament; Three-Dimensional Image; Three-Dimensional Imaging; Ursidae Family; Vertebral column; Vertebrates; connectin; disease-causing mutation; movie; myosin-binding protein C; plasma protein Z; reconstruction; sensor; tool

ABSTRACT Funds are requested for the purchase of a revolutionary new camera, the Apollo, from Direct Electron Ltd. that would enable the collection of large amounts of image data, currently obtained in the form of “movies”. The ability to collect large amounts of data would broaden the types of questions addressable in the PI's R35 grant currently focused on obtaining atomic resolution images of thick filaments from striated muscle. Generally, thick filaments from all striated muscles have four structurally distinct regions: (1) a 160 nm long bare zone in the center where myosin tails are packed antiparallel to each other, (2) an A-band follows where myosin molecules are arranged in a helical or quasi helical structure with the myosin tails packed in a parallel arrangement, (3) a tapered end where the connecting filament, e.g. titin, connects the thick filament to the Z-disk, and (4) features along the thick filament A-band from invertebrates in particular, but also from vertebrates, that fail to follow the myosin molecule arrangement. The helical arrangement of myosin in the A-band of invertebrate thick filaments facilitates an atomic structure of the myosin tail and head arrangement by using all of the myosin axial repeats from the end of the bare zone to the beginning of the tapered ends. However, the bare zone, the transition from bare zone to helical arrangement, the tapered ends as well as additional features that do not follow the myosin helical symmetry perform important roles in the assembly of thick filaments. Features, such as the paramyosin of invertebrate thick filaments fail to follow the myosin helical symmetry in the A-band and are poorly represented in cryoEM reconstructions. Paramyosin in concert with other non-myosin proteins may function to determine the thick filament length in invertebrates. Titin, myosin binding protein C and other non-myosin proteins of vertebrate thick filaments also do not follow a regular pattern along the entire A-band. Acquisition of a much faster camera capable of recording a 60-frame movie in a single second, will provide the tool needed to obtain high resolution 3-D images of these features that are not well represented if at all in structures solved by enforcing helical symmetry.

抽象的 需要资金从 Direct 购买革命性的新相机 Apollo Electron Ltd. 将能够收集目前获得的大量图像数据 以“电影”的形式收集大量数据的能力将拓宽类型。 PI R35 资助中可解决的问题目前集中于获得原子分辨率 来自横纹肌的粗肌丝的图像 一般来说，来自所有横纹肌的粗肌丝。 肌肉有四个结构不同的区域：(1) 中心有一个 160 nm 长的裸露区域，其中 肌球蛋白尾部彼此反平行排列，(2) A 带紧随肌球蛋白的位置 分子排列成螺旋或准螺旋结构，肌球蛋白尾部堆积在 平行排列，（3）连接细丝（例如肌动蛋白）的锥形末端 粗丝到 Z 盘，以及 (4) 沿着无脊椎动物粗丝 A 带的特征 特别是，但也来自无法遵循肌球蛋白分子排列的脊椎动物。 无脊椎动物粗丝 A 带中肌球蛋白的螺旋排列有利于原子 通过使用来自的所有肌球蛋白轴向重复来观察肌球蛋白尾部和头部排列的结构 裸露区域的末端到锥形末端的起点。 从裸露区域到螺旋排列的过渡、锥形端部以及附加功能 不遵循肌球蛋白螺旋对称性的细胞在厚的组装中发挥重要作用 细丝的特征，例如无脊椎动物粗细丝的副肌球蛋白无法遵循。 A 波段的肌球蛋白螺旋对称性，在冷冻电镜重建中表现不佳。 副肌球蛋白与其他非肌球蛋白协同作用可能决定厚度 无脊椎动物中肌联蛋白、肌球蛋白结合蛋白 C 和其他非肌球蛋白的丝长度。 脊椎动物的粗丝也不沿着整个 A 波段遵循规则的模式。 购买速度更快的相机，能够在一秒钟内录制 60 帧的电影， 将提供获取这些特征的高分辨率 3D 图像所需的工具，这些特征不是 如果有的话，在通过强制螺旋对称性解决的结构中得到很好的体现。