Stepwise Expansion of the Bacteriophage φ6 Procapsid: Possible Packaging Intermediates
Stepwise Expansion of the Bacteriophage φ6 Procapsid: Possible Packaging Intermediates
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DOI：
10.1016/j.jmb.2011.10.004
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发表时间：
2011-11-25
影响因子：
5.6
通讯作者：
Heymann, J. Bernard
中科院分区：
文献类型：
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作者：
Nemecek, Daniel;Cheng, Naiqian;Heymann, J. Bernard
噬菌体416的初始组装产物，即原衣壳，在其三个单链RNA片段的顺序包装期间经历主要的结构转变。原衣壳是一个紧凑的二十面体对称颗粒，具有深深凹陷的顶点，扩展到球形成熟衣壳，使可容纳基因组的体积增加了2.5倍。有人提出，扩增和包装是联系在一起的，扩增的每个阶段都为特定的RNA片段提供一个结合位点。为了研究原衣壳的可转化性，我们通过酸化、加热和提高盐浓度来诱导扩增。所有三种治疗后的冷冻电子显微镜重建产生相同的部分膨胀的颗粒。冷冻电子断层扫描分析表明，一个给定的衣壳的所有顶点要么在一个紧凑的或扩展的状态，表明一个高度合作的过渡。为了对成熟衣壳进行基准测试，我们分析了填充的（体内包装的）衣壳。当这些颗粒被诱导释放它们的RNA时，它们恢复到与扩展的前衣壳相同的中间状态（中间体1）或第二种进一步扩展的状态（中间体2）。这种部分可逆性的扩张表明，成熟的球形衣壳构象，只有当足够的向外压力施加包装RNA。两种中间体的观察结果与拟定的三步包装工艺一致。该模型进一步得到以下观察结果的支持：能够包装第二RNA区段而不预先包装第一区段的突变体具有增强的从原衣壳自发地转换到第一中间状态的易感性。爱思唯尔有限公司出版
The initial assembly product of bacteriophage 416, the procapsid, undergoes major structural transformation during the sequential packaging of its three segments of single-stranded RNA. The procapsid, a compact icosahedrally symmetric particle with deeply recessed vertices, expands to the spherical mature capsid, increasing the volume available to accommodate the genome by 2.5-fold. It has been proposed that expansion and packaging are linked, with each stage in expansion presenting a binding site for a particular RNA segment. To investigate procapsid transformability, we induced expansion by acidification, heating, and elevated salt concentration. Cryo-electron microscopy reconstructions after all three treatments yielded the same partially expanded particle. Analysis by cryo-electron tomography showed that all vertices of a given capsid were either in a compact or an expanded state, indicating a highly cooperative transition. To benchmark the mature capsid, we analyzed filled (in vivo packaged) capsids. When these particles were induced to release their RNA, they reverted to the same intermediate state as expanded procapsids (intermediate 1) or to a second, further expanded state (intermediate 2). This partial reversibility of expansion suggests that the mature spherical capsid conformation is obtained only when sufficient outward pressure is exerted by packaged RNA. The observation of two intermediates is consistent with the proposed three-step packaging process. The model is further supported by the observation that a mutant capable of packaging the second RNA segment without previously packaging the first segment has enhanced susceptibility for switching spontaneously from the procapsid to the first intermediate state. Published by Elsevier Ltd.