Structure and function analysis of nucleocapsid protein of tomato spotted wilt virus interacting with RNA using homology modeling.
Structure and function analysis of nucleocapsid protein of tomato spotted wilt virus interacting with RNA using homology modeling.
复制标题
使用同源模型分析番茄斑萎病毒核衣壳蛋白与 RNA 相互作用的结构和功能
DOI：
10.1074/jbc.m114.604678
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发表时间：
2015-02-13
期刊：
影响因子：
--
通讯作者：
Tao X
中科院分区：
文献类型：
--
作者：
Li J;Feng Z;Wu J;Huang Y;Lu G;Zhu M;Wang B;Mao X;Tao X
背景：包括番茄斑点枯萎病毒(TSWV)在内的许多植物病毒都不具备病毒蛋白的晶体结构。结果：通过同源模建，我们绘制了TSWV核衣壳(N)的RNA结合位点图，发现了TSWV核衣壳(N)的保护作用。结论：同源模建为研究TSWV N与RNA相互作用的功能奠定了基础。意义：这种方法可能适用于其他植物病毒。番茄斑枯病病毒(TSWV)核衣壳蛋白(N)在基因组RNA合成核糖核蛋白(RNP)中起着关键作用，RNP是病毒基因转录和基因组复制的模板。然而，TSWV N如何与基因组RNA相互作用的分子机制却知之甚少。在这项研究中，我们证明了TSWV N蛋白形成了一系列更高顺序的寡聚体。对N蛋白的RNA结合行为的分析表明，没有特定的低聚物优先与RNA结合，而是每种类型的N低聚体都能与RNA结合。为了更好地表征N蛋白与RNA相互作用的结构和功能，我们构建了TSWV N和N-RNA复合体的同源模型。基于这些同源模型，我们证明了TSWV N表面裂隙中的正电荷和极性氨基酸是RNA结合的关键。此外，通过N-RNA同源建模，我们发现RNA组分深深嵌入到预测的蛋白质裂解中；一致地，TSWV N-RNA复合体相对抵抗RNase的消化。总之，利用同源模型，我们确定了N蛋白上的RNA结合位点，并发现了N蛋白的新的保护功能。我们的发现也为TSWV N与RNA组分相互作用的分子细节提供了新的见解。
Background: The crystal structure of viral proteins is not available for many plant viruses including tomato spotted wilt virus (TSWV). Results: By homology modeling, we mapped the RNA binding sites and discovered a protective feature of TSWV nucleocapsid (N). Conclusion: Homology modeling provided a basis for functional analysis of TSWV N interacting with RNA. Significance: This approach might be applicable for other plant viruses. The nucleocapsid (N) protein of tomato spotted wilt virus (TSWV) plays key roles in assembling genomic RNA into ribonucleoprotein (RNP), which serves as a template for both viral gene transcription and genome replication. However, little is known about the molecular mechanism of how TSWV N interacts with genomic RNA. In this study, we demonstrated that TSWV N protein forms a range of higher ordered oligomers. Analysis of the RNA binding behavior of N protein revealed that no specific oligomer binds to RNA preferentially, instead each type of N oligomer is able to bind RNA. To better characterize the structure and function of N protein interacting with RNA, we constructed homology models of TSWV N and N-RNA complexes. Based on these homology models, we demonstrated that the positively charged and polar amino acids in its predicted surface cleft of TSWV N are critical for RNA binding. Moreover, by N-RNA homology modeling, we found that the RNA component is deeply embedded in the predicted protein cleft; consistently, TSWV N-RNA complexes are relatively resistant to digestion by RNase. Collectively, using homology modeling, we determined the RNA binding sites on N and found a new protective feature for N protein. Our findings also provide novel insights into the molecular details of the interaction of TSWV N with RNA components.