Comparative Structural and Functional Analysis of Bunyavirus and Arenavirus Cap-Snatching Endonucleases.
Comparative Structural and Functional Analysis of Bunyavirus and Arenavirus Cap-Snatching Endonucleases.
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DOI：
10.1371/journal.ppat.1005636
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发表时间：
2016-06
期刊：
影响因子：
6.7
通讯作者：
Cusack S
中科院分区：
文献类型：
--
作者：
Reguera J;Gerlach P;Rosenthal M;Gaudon S;Coscia F;Günther S;Cusack S
竞技场病毒、布尼亚病毒和正粘病毒科的分段负链RNA病毒通过夺帽机制独特地进行病毒mRNA转录。这涉及通过位于病毒聚合酶N-末端区域的内切核酸酶（EN）结构域切割宿主mRNA接近其加帽的5′端。本文报道了汉坦病毒（一种属于汉坦病毒属的本扬病毒）的夺帽EN的结构。汉滩EN具有活性位点构型，包括金属配位组氨酸，和与先前报道的拉克罗斯病毒和流感病毒EN（分别为正布尼亚病毒和正粘病毒）相似的核酸酶活性，但在切割双链RNA底物方面更有活性。相反，拉沙沙粒病毒EN仅具有酸性金属配位残基。我们提出了三个高分辨率结构的拉沙病毒EN与不同的结合离子配置，并显示在比较生物物理和生物化学实验与汉滩，拉克罗斯和流感EN的分离拉沙EN基本上是无活性的。结果进行了讨论，根据EN激活机制揭示了最近的全长流感病毒聚合酶的结构。节段负链病毒（sNSV）如流感病毒、拉沙病毒或汉滩病毒是导致大量严重人类传染病的原因。目前，有疫苗和抗病毒治疗可用于流感，但没有其他sNSV引起的感染。所有这些都通过夺帽机制进行转录，这需要金属离子依赖性核酸内切酶（EN）的作用，EN是它们的大型病毒聚合酶内的一个结构域。在这里，我们提供了晶体结构的汉滩病毒（布尼亚病毒科）和拉沙病毒（沙粒病毒科）帽抢夺EN复杂的锰和比较功能的研究，他们的催化活性。尽管两个EN之间的高度结构同源性的活性位点的一些变化，涉及催化组氨酸，导致不同的结合的金属离子与显着的后果，其体外活性。汉坦EN与甲型流感病毒（正粘病毒科）和LACV（布尼亚病毒科）EN结合金属离子，这三种EN在体外都有活性。相反，拉沙病毒EN在相同的实验条件下是无活性的。我们现在可以将sNSV分为两个功能不同的组（His+和His-EN），提供了sNSV夺帽EN性质的广泛观点，这将是广谱抗病毒药物的全面开发的决定性因素。这些结果也有影响的病毒转录调控在最近的研究全长sNSV聚合酶。
Segmented negative strand RNA viruses of the arena-, bunya- and orthomyxovirus families uniquely carry out viral mRNA transcription by the cap-snatching mechanism. This involves cleavage of host mRNAs close to their capped 5′ end by an endonuclease (EN) domain located in the N-terminal region of the viral polymerase. We present the structure of the cap-snatching EN of Hantaan virus, a bunyavirus belonging to hantavirus genus. Hantaan EN has an active site configuration, including a metal co-ordinating histidine, and nuclease activity similar to the previously reported La Crosse virus and Influenza virus ENs (orthobunyavirus and orthomyxovirus respectively), but is more active in cleaving a double stranded RNA substrate. In contrast, Lassa arenavirus EN has only acidic metal co-ordinating residues. We present three high resolution structures of Lassa virus EN with different bound ion configurations and show in comparative biophysical and biochemical experiments with Hantaan, La Crosse and influenza ENs that the isolated Lassa EN is essentially inactive. The results are discussed in the light of EN activation mechanisms revealed by recent structures of full-length influenza virus polymerase. Segmented negative strand viruses (sNSV) such as Influenza, Lassa or Hantaan viruses are responsible for a large number of severe human infectious diseases. Currently, there are vaccines and antiviral treatments available for influenza but none for the infections caused by other sNSV. All carry out transcription by the cap-snatching mechanism, which requires the action of a metal ion dependent endonuclease (EN), a domain within their large viral polymerases. Here we provide the crystal structure of the Hantaan virus (family Bunyaviridae) and Lassa virus (family Arenaviridae) cap-snatching ENs in complex with manganese and a comparative functional study of their catalytic activity. Despite the high structural homology between the two ENs a few changes in the active site, involving a catalytic histidine, cause a different binding of the metal ions with dramatic consequences for their in vitro activity. Hantaan EN binds the metal ions as Influenza A (family Orthomyxoviridae) and LACV (family Bunyaviridae) ENs and all three are active in vitro. In contrast Lassa virus EN is inactive in the same experimental conditions. We can now classify sNSV into two functionally distinct groups (His+ and His- ENs), providing a broad view of the sNSV cap-snatching ENs properties that will be determinant for the comprehensive development of broad-spectrum antiviral drugs. These results also have implications for the viral transcription regulation in the light of recent studies on full-length sNSV polymerases.