Visualization of uncorrelated, tandem symmetry mismatches in the internal genome packaging apparatus of bacteriophage T7
Visualization of uncorrelated, tandem symmetry mismatches in the internal genome packaging apparatus of bacteriophage T7
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DOI：
10.1073/pnas.1215563110
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发表时间：
2013-04-23
影响因子：
11.1
通讯作者：
Jiang, Wen
中科院分区：
文献类型：
--
作者：
Guo, Fei;Liu, Zheng;Jiang, Wen
在大量dsDNA病毒的生命周期中，通过一个独特的门户顶点将dsDNA基因组马达驱动包装到预先形成的蛋白质衣壳中是必不可少的。我们用单粒子电子冷冻显微镜研究了包装中T7DNA的受体噬菌体T7proapsid的入口顶端的多层结构。提出了一种聚焦的非对称重建方法，并应用于选择性地分解门脉顶点对称失配层的相邻对。然而，多层门户顶点的所有层的结构特征不能同时解析。我们的结果表明，对称性不匹配的层可以在几个不同的相对取向上结合在一起，并且不同界面上的取向独立地结合在一起产生结构异构体，这一过程我们称为组合组装异构化。这种异构性解释了以前报道的T7和其他噬菌体门脉顶端的不对称重建中的旋转涂片。组合组装异构化可能代表了结构生物学的一种新机制，在这种机制中，全球不同的结构由一组共同的组件组装而成。我们的重建共同验证了先前提出的T7门户顶点层的对称性、组成和顺序，串联解析了5倍基因产物10(GP10)壳、12倍GP8门脉环和由12倍GP14适配子环、8倍碗形GP15和4倍gp16尖端组成的内部核心堆栈。我们还发现核心堆栈相对于二十面体五倍轴有一个小的倾斜，并认为这种倾斜有助于DNA在包装过程中不缠绕。
Motor-driven packaging of a dsDNA genome into a preformed protein capsid through a unique portal vertex is essential in the life cycle of a large number of dsDNA viruses. We have used single-particle electron cryomicroscopy to study the multilayer structure of the portal vertex of the bacteriophage T7 procapsid, the recipient of T7 DNA in packaging. A focused asymmetric reconstruction method was developed and applied to selectively resolve neighboring pairs of symmetry-mismatched layers of the portal vertex. However, structural features in all layers of the multilayer portal vertex could not be resolved simultaneously. Our results imply that layers with mismatched symmetries can join together in several different relative orientations, and that orientations at different interfaces assort independently to produce structural isomers, a process that we call combinatorial assembly isomerism. This isomerism explains rotational smearing in previously reported asymmetric reconstructions of the portal vertex of T7 and other bacteriophages. Combinatorial assembly isomerism may represent a new regime of structural biology in which globally varying structures assemble from a common set of components. Our reconstructions collectively validate previously proposed symmetries, compositions, and sequential order of T7 portal vertex layers, resolving in tandem the 5-fold gene product 10 (gp10) shell, 12-fold gp8 portal ring, and an internal core stack consisting of 12-fold gp14 adaptor ring, 8-fold bowl-shaped gp15, and 4-fold gp16 tip. We also found a small tilt of the core stack relative to the icosahedral fivefold axis and propose that this tilt assists DNA spooling without tangling during packaging.