Konno Keisuke, Nishikawa Satoshi, Hasegawa Tsunemi, Fukuda Kotaro
Department of Material and Biological Chemistry, Faculty of Science, Yamagata University, Yamagata 990-8560, Japan.
Nucleic Acids Symp Ser (Oxf). 2007(51):393-4. doi: 10.1093/nass/nrm197.
The minus-IRES ((-)IRES), corresponding to the 3'-terminal end of the negative strand of hepatitis C virus (HCV) RNA, is well conserved among HCV subtypes. The higher order structure of (-)IRES is essential for HCV replication, because the viral RNA dependent RNA polymerase, NS5B, recognizes it as the initiation site for plus-strand synthesis of the HCV genome. To inhibit the "de novo" synthesis of plus-strand RNA molecules, we performed an in vitro selection procedure that is specific for the (-)IRES domain I. After confirming the binding convergence in the ninth RNA pool, 42 RNA clones were sequenced and analyzed. Of these, 25 clones (Family-I) had the consensus sequence, 5'-UGGAUC-3', which is complementary to the apical loop of SL-E1, an important region for NS5B recognition. Another 13 clones (Family-II) had the consensus sequence, 5'-GAGUAC-3', which is complementary to the apical loop of SL-D1. Biochemical analyses are in progress to evaluate whether these RNA aptamers have the ability to inhibit HCV replication.
负内部核糖体进入位点（(-)IRES），对应丙型肝炎病毒（HCV）RNA负链的3'末端，在HCV各亚型中高度保守。(-)IRES的高级结构对HCV复制至关重要，因为病毒RNA依赖性RNA聚合酶NS5B将其识别为HCV基因组正链合成的起始位点。为了抑制正链RNA分子的“从头”合成，我们针对(-)IRES结构域I进行了体外筛选程序。在确认第九个RNA文库中的结合趋同后，对42个RNA克隆进行了测序和分析。其中，25个克隆（家族I）具有一致序列5'-UGGAUC-3'，它与SL-E1的顶端环互补，SL-E1是NS5B识别的重要区域。另外13个克隆（家族II）具有一致序列5'-GAGUAC-3'，它与SL-D1的顶端环互补。目前正在进行生化分析，以评估这些RNA适体是否具有抑制HCV复制的能力。
