Crystal structure of the dengue virus RNA-dependent RNA polymerase catalytic domain at 1.85-Angstrom resolution
Crystal structure of the dengue virus RNA-dependent RNA polymerase catalytic domain at 1.85-Angstrom resolution
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DOI：
10.1128/jvi.02283-06
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发表时间：
2007-05-01
影响因子：
5.4
通讯作者：
Lescar, Julien
中科院分区：
文献类型：
--
作者：
Yap, Thai Leong;Xu, Ting;Lescar, Julien
登革热是一种被忽视的新发疾病，目前尚无疫苗或抗病毒药物，由登革热病毒引起，登革热病毒是黄病毒属的成员，该属包括几种重要的人类病原体，例如黄热病和西尼罗河病毒。登革热病毒的 NS5 蛋白具有双功能，含有 900 个氨基酸。 S-腺苷甲硫氨酸转移酶活性位于其 N 末端结构域内，残基 270 至 900 形成 RNA 依赖性 RNA 聚合酶 (RdRp) 催化结构域。病毒复制开始于从登革热病毒正链RNA基因组合成负链RNA，随后将其用作合成额外正链RNA基因组的模板。 NS5 RdRp 催化了这一产生新病毒颗粒的基本功能。在这里，我们提出了一种高通量体外测定，部分概括了这种活性以及以 1.85 埃分辨率精制的登革热病毒 RdRp 酶活性片段的晶体结构。 NS5 核定位序列以前被认为折叠成一个单独的结构域，但现在形成了聚合酶子结构域的一个组成部分。该结构还揭示了两个锌离子结合基序的存在。在不存在模板链的情况下，链终止核苷类似物与引发环位点结合。这些结果应该为针对登革热病毒的抗病毒化合物的基于结构的设计提供信息并加速。
Dengue fever, a neglected emerging disease for which no vaccine or antiviral agents exist at present, is caused by dengue virus, a member of the Flavivirus genus, which includes several important human pathogens, such as yellow fever and West Nile viruses. The NS5 protein from dengue virus is bifunctional and contains 900 amino acids. The S-adenosyl methionine transferase activity resides within its N-terminal domain, and residues 270 to 900 form the RNA-dependent RNA polymerase (RdRp) catalytic domain. Viral replication begins with the synthesis of minus-strand RNA from the dengue virus positive-strand RNA genome, which is subsequently used as a template for synthesizing additional plus-strand RNA genomes. This essential function for the production of new viral particles is catalyzed by the NS5 RdRp. Here we present a high-throughput in vitro assay partly recapitulating this activity and the crystallographic structure of an enzymatically active fragment of the dengue virus RdRp refined at 1.85-angstrom resolution. The NS5 nuclear localization sequences, previously thought to fold into a separate domain, form an integral part of the polymerase subdomains. The structure also reveals the presence of two zinc ion binding motifs. In the absence of a template strand, a chain-terminating nucleoside analogue binds to the priming loop site. These results should inform and accelerate the structure-based design of antiviral compounds against dengue virus.