Structural analysis of macromolecular assemblies by electron microscopy.
Structural analysis of macromolecular assemblies by electron microscopy.
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DOI：
10.1021/cr100353t
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发表时间：
2011-12-14
期刊：
影响因子：
62.1
通讯作者：
Saibil, H. R.
中科院分区：
文献类型：
--
作者：
Orlova, E. V.;Saibil, H. R.
为了充分理解从细菌代谢到人脑运作的生物过程，有必要了解组成分子的三维（3D）空间排列和动力学，它们如何组装成复杂的分子机器，以及它们如何形成功能性细胞器，细胞和组织。X射线晶体学和核磁共振波谱学方法可以提供分子结构和动力学的详细信息。在细胞水平上，光学显微镜揭示了荧光标记分子的空间分布和动力学。电子显微镜（EM）与这些方法重叠，涵盖了从原子到细胞结构的广泛范围。低温方法的发展使得EM成像能够提供捕获在接近天然水合状态的生物分子和细胞的快照。（1，2）由于大分子组装在活细胞机械中的重要性以及EM和图像处理方法的进步，EM已成为分子至细胞尺寸范围内的结构生物学的主要工具。由于光学和电子显微镜的发展，在理解细胞和组织中大分子及其组装体的3D空间组织方面取得了巨大的进展。在光学显微镜中，超分辨率和单分子方法已经将荧光图像的分辨率推到了50 nm，利用分子生物学的力量将感兴趣的分子与荧光标记蛋白融合。(3)X射线低温层析成像是一种用于较厚（10 μm）水合样品的三维重建方法，分辨率达到15 nm分辨率范围。(4)在EM中，仪器和方法的主要发展已经推进了玻璃化溶液中的单个颗粒（孤立的大分子复合物）的研究，以及通过对不规则物体（如细胞或亚细胞结构）进行断层扫描进行3D重建。（1，5-7）冷冻切片可用于制备细胞和组织的玻璃化切片，否则其将太厚而不能通过透射EM（TEM）成像。（八、九）
To fully understand biological processes from the metabolism of a bacterium to the operation of a human brain, it is necessary to know the three-dimensional (3D) spatial arrangement and dynamics of the constituent molecules, how they assemble into complex molecular machines, and how they form functional organelles, cells, and tissues. The methods of X-ray crystallography and NMR spectroscopy can provide detailed information on molecular structure and dynamics. At the cellular level, optical microscopy reveals the spatial distribution and dynamics of molecules tagged with fluorophores. Electron microscopy (EM) overlaps with these approaches, covering a broad range from atomic to cellular structures. The development of cryogenic methods has enabled EM imaging to provide snapshots of biological molecules and cells trapped in a close to native, hydrated state.(1, 2)Because of the importance of macromolecular assemblies in the machinery of living cells and progress in the EM and image processing methods, EM has become a major tool for structural biology over the molecular to cellular size range. There have been tremendous advances in understanding the 3D spatial organization of macromolecules and their assemblies in cells and tissues, due to developments in both optical and electron microscopy. In light microscopy, super-resolution and single molecule methods have pushed the resolution of fluorescence images to∼ 50 nm, using the power of molecular biology to fuse molecules of interest with fluorescent marker proteins.(3) X-ray cryo-tomography is developing as a method for 3D reconstruction of thicker (10 μm) hydrated samples, with resolution reaching the 15 nm resolution range.(4) In EM, major developments in instrumentation and methods have advanced the study of single particles (isolated macromolecular complexes) in vitrified solution as well as in 3D reconstruction by tomography of irregular objects such as cells or subcellular structures.(1, 5-7) Cryo-sectioning can be used to prepare vitrified sections of cells and tissues that would otherwise be too thick to image by transmission EM (TEM).(8, 9)
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影响因子：
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作者：
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通讯作者：
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