Mutations in the hepatitis C virus polymerase that increase RNA binding can confer resistance to cyclosporine A.
Mutations in the hepatitis C virus polymerase that increase RNA binding can confer resistance to cyclosporine A.
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DOI：
10.1002/hep.22987
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发表时间：
2009-07
期刊：
影响因子：
13.5
通讯作者：
Tang, Hengli
中科院分区：
文献类型：
--
作者：
Liu, Zhe;Robida, John M.;Chinnaswamy, Sreedhar;Yi, Guanghui;Robotham, Jason M.;Nelson, Heather B.;Irsigler, Andre;Kao, C. Cheng;Tang, Hengli
丙型肝炎病毒（HCV）感染导致急性和慢性肝脏疾病，需要新型的抗HCV治疗药物。环孢素A （Cyclosporine A, CsA）可抑制丙型肝炎病毒复制，缺乏免疫抑制功能的CsA衍生物目前正作为抗丙型肝炎病毒的候选药物进行临床试验。在这里，我们描述了几个独立衍生的HCV复制子，这些复制子由于HCV编码的聚合酶非结构蛋白5B （NS5B）突变而具有不同水平的CsA抗性。携带这些突变的突变型HCV复制子显示出对CsA的抗性。突变存在于聚合酶的两个不同的斑块中：模板通道和模板通道后面凹表面的一面。在CsA存在的情况下，表达HCV复制子的细胞产生的突变体NS5B与RNA的结合能力增强。含有突变的纯化重组NS5B蛋白在从头启动RNA合成方面优于野生型对照。此外，突变蛋白能够以大约8倍高的亲和力结合RNA。最后，模板通道附近的突变减轻了凹斑P540A突变的致死性表型。这种通过不同结构域氨基酸变化对HCV复制酶功能的分子内补偿在感染性细胞培养衍生的病毒系统中得到进一步证实。CsA耐药水平的增加与NS5B基因的不同突变有关，该基因在CsA存在时增加RNA结合，并且拇指残基和c端结构域之间的分子内通信对HCV复制酶功能很重要。
Hepatitis C virus (HCV) infection leads to acute and chronic liver diseases, and new classes of anti-HCV therapeutics are needed. Cyclosporine A (CsA) inhibits HCV replication, and CsA derivatives that lack the immunosuppressive function are currently in clinical trials as candidate anti-HCV drugs. Here, we characterize several independently derived HCV replicons with varying levels of CsA resistance due to mutations in nonstructural protein 5B (NS5B), the HCV-encoded polymerase. Mutant HCV replicons engineered with these mutations showed resistance to CsA. The mutations reside in two distinct patches in the polymerase: the template channel and one face of a concave surface behind the template channel. Mutant NS5B made by cells expressing the HCV replicon had increased ability to bind to RNA in the presence of CsA. Purified recombinant NS5B proteins containing the mutations were better at de novo initiated RNA synthesis than the wild-type control. Furthermore, the mutant proteins were able to bind RNA with approximately eightfold higher affinity. Last, mutation near the template channel alleviated the lethal phenotype of a mutation in the concave patch, P540A. This intramolecular compensation for the HCV replicase function by amino-acid changes in different domains was further confirmed in an infectious cell culture-derived virus system. Increased level of CsA resistance is associated with distinct mutations in the NS5B gene that increase RNA binding in the presence of CsA, and the intramolecular communications between residues of the thumb and the C-terminal domains are important for HCV replicase function.
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影响因子：
14.8
作者：
Kato, Takanobu;Date, Tomoko;Wakita, Takaji
通讯作者：
Wakita, Takaji
影响因子：
4.6
作者：
Firpi, RJ;Zhu, HZ;Nelson, DR
通讯作者：
Nelson, DR
影响因子：
5.4
作者：
Shim, JH;Larson, G;Hong, Z
通讯作者：
Hong, Z
影响因子：
13.5
作者：
Flisiak, Robert;Horban, Andrzej;Scalfaro, Pietro
通讯作者：
Scalfaro, Pietro
DOI：
10.1073/pnas.081077198
发表时间：
2001-04-24
影响因子：
11.1
作者：
Kim, MJ;Kao, C
通讯作者：
Kao, C