cryoEM Administrative Supplement for Equipment

摘要 请求提供资金，用于从Direct购买革命性的新相机Apollo。 电子有限公司，这将使收集大量的图像数据，目前获得的 以“电影”的形式。收集大量数据的能力将扩大 PI的R35资助中可解决的问题目前集中在获得原子分辨率上 横纹肌的粗丝图像。一般来说，粗丝从所有条纹 肌肉有四个结构不同的区域：（1）在中心的160 nm长的裸露区， 肌球蛋白尾部相互反平行排列，（2）肌球蛋白尾部出现A带， 分子排列成螺旋或准螺旋结构，肌球蛋白尾部包装成 （3）锥形端，在该锥形端处，连接细丝（例如，肌联蛋白）连接 Z盘上有一条粗丝，（4）沿着无脊椎动物的粗丝A带 特别是来自脊椎动物，它们不能遵循肌球蛋白分子排列。的 无脊椎动物粗肌丝A带中肌球蛋白的螺旋排列有助于原子 通过使用所有的肌球蛋白轴向重复序列， 裸露区域的末端到锥形末端的开始。然而，裸露区， 从裸区到螺旋布置的过渡、锥形端部以及附加特征 不遵循肌球蛋白螺旋对称性的蛋白质在厚膜的组装中起重要作用。 细丝。无脊椎动物粗肌丝的副肌球蛋白等特征， 肌球蛋白螺旋对称的A-带，并在cryoEM重建表现不佳。 副肌球蛋白与其他非肌球蛋白的协同作用可能决定肌球蛋白的厚度。 无脊椎动物的细丝长度。肌联蛋白、肌球蛋白结合蛋白C和其他非肌球蛋白 脊椎动物的粗丝也不遵循沿着整个A带的规则模式。 获得一个速度更快的相机，能够在一秒钟内录制60帧电影， 将提供获得这些特征的高分辨率3D图像所需的工具， 如果在通过强制螺旋对称解决的结构中，

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.