低温电子断层扫描分子和亚细胞结构的选择性标记技术

IF 2.3 4区 生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY
Evgeny P. Kazakov, Igor I. Kireev, Sergei A. Golyshev
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引用次数: 0

摘要

电子显微镜(EM)是研究细胞精细结构最有效的方法之一,其分辨率比可见光显微镜高数千倍。电子显微镜在生物学中最先进的应用是通过冷冻而非水结晶(cryoET)稳定样品的电子显微镜断层扫描。通过避免化学固定和脱水的缺点,该技术可以在分子水平上三维研究细胞结构,直至分解单个蛋白质及其子结构域。然而,有效识别和定位感兴趣目标的问题尚未得到解决,从而将目标范围限制在易于识别或丰富的亚细胞成分上。标记技术提供了在显微镜图像中定位调查对象的唯一途径。CryoET对标记系统提出了相互矛盾的要求,包括需要将由细胞化学外物质组成的颗粒引入活细胞,这些颗粒必须与感兴趣的分子结合,而不破坏其重要功能和细胞生理。本文综述了现有的和有前景的选择性标记蛋白质和亚细胞结构的方法,目的是使它们能够在低温低温成像中定位。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Techniques for Selective Labeling of Molecules and Subcellular Structures for Cryo-Electron Tomography

Electron microscopy (EM) is one of the most efficient methods for studying the fine structure of cells with a resolution thousands of times higher than that of visible light microscopy. The most advanced implementation of electron microscopy in biology is EM tomography of samples stabilized by freezing without water crystallization (cryoET). By circumventing the drawbacks of chemical fixation and dehydration, this technique allows investigating cellular structures in three dimensions at the molecular level, down to resolving individual proteins and their subdomains. However, the problem of efficient identification and localization of objects of interest has not yet been solved, thus limiting the range of targets to easily recognizable or abundant subcellular components. Labeling techniques provide the only way for locating the subject of investigation in microscopic images. CryoET imposes conflicting demands on the labeling system, including the need to introduce into a living cell the particles composed of substances foreign to the cellular chemistry that have to bind to the molecule of interest without disrupting its vital functions and physiology of the cell. This review examines both established and prospective methods for selective labeling of proteins and subcellular structures aimed to enable their localization in cryoET images.

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来源期刊
Biochemistry (Moscow)
Biochemistry (Moscow) 生物-生化与分子生物学
CiteScore
4.70
自引率
3.60%
发文量
139
审稿时长
2 months
期刊介绍: Biochemistry (Moscow) is the journal that includes research papers in all fields of biochemistry as well as biochemical aspects of molecular biology, bioorganic chemistry, microbiology, immunology, physiology, and biomedical sciences. Coverage also extends to new experimental methods in biochemistry, theoretical contributions of biochemical importance, reviews of contemporary biochemical topics, and mini-reviews (News in Biochemistry).
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