Assembly of a functional Machupo virus polymerase complex
Assembly of a functional Machupo virus polymerase complex
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DOI：
10.1073/pnas.1007152107
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发表时间：
2010-11-16
影响因子：
11.1
通讯作者：
Whelan, Sean P. J.
中科院分区：
文献类型：
--
作者：
Kranzusch, Philip J.;Schenk, Andreas D.;Whelan, Sean P. J.
沙粒病毒科的分段负义病毒编码一个大的聚合酶(L)蛋白，该蛋白包含RNA合成所需的所有酶活性。这些活性包括RNA依赖的RNA聚合酶（RdRP）和RNA内切酶，该酶将带帽引物从细胞mrna切割成引物转录。利用纯化的催化活性Machupo病毒L，我们提供了这种多功能聚合酶的整体结构，并在体外用RNA模板重建了复合物的形成。L蛋白含有一个中央环结构域，其外观与dsRNA病毒的RdRP和多个可能负责5'帽形成的附属附件相似。L识别RNA模板需要一个位于病毒基因组3'端2-5位的序列特异性基序。此外，L-RNA复合物的形成依赖于单链RNA，这表明末端间的dsRNA相互作用必须部分中断才能发生复合物组装。我们的研究结果为沙粒病毒聚合酶-模板相互作用提供了一个模型，并揭示了负链RNA病毒L蛋白的结构组织。
Segmented negative-sense viruses of the family Arenaviridae encode a large polymerase (L) protein that contains all of the enzymatic activities required for RNA synthesis. These activities include an RNA-dependent RNA polymerase (RdRP) and an RNA endonuclease that cleaves capped primers from cellular mRNAs to prime transcription. Using purified catalytically active Machupo virus L, we provide a view of the overall architecture of this multifunctional polymerase and reconstitute complex formation with an RNA template in vitro. The L protein contains a central ring domain that is similar in appearance to the RdRP of dsRNA viruses and multiple accessory appendages that may be responsible for 5' cap formation. RNA template recognition by L requires a sequence-specific motif located at positions 2-5 in the 3' terminus of the viral genome. Moreover, L-RNA complex formation depends on single-stranded RNA, indicating that inter-termini dsRNA interactions must be partially broken for complex assembly to occur. Our results provide a model for arenavirus polymerase-template interactions and reveal the structural organization of a negative-strand RNA virus L protein.