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首页> 外文期刊>Journal of Biomolecular Structure and Dynamics >Structure Elucidation of the Hepatitis B Virus Encapsidation Signal by NMR on Selectively Labeled RNAs.
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Structure Elucidation of the Hepatitis B Virus Encapsidation Signal by NMR on Selectively Labeled RNAs.

机译：通过NMR在选择性标记的RNA上阐明乙型肝炎病毒衣壳信号的结构。

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摘要

Hepatitis B virus (HBV) HBV is DNA virus with a unique replication strategy, which involves reverse transcription of its pregenomic RNA. Essential for this reverse transcription are the 5 - and 3 -ends of its pregenomic RNA (5 -RT-RNA and 3 -RT-RNA, respectively) which form conserved bulged stem-loop structures. The 5 -RT-RNA consists of a 67 nucleotide bulged stem-loop structure, epsilon, which constitutes the signal for encapsidation of the pregenomic RNA and subsequent reverse transcription. The reverse transcriptase (RT) initially binds to the completely conserved apical loop of epsilon and a 4-nucleotide primer is synthesized from the adjacent 6-nucleotide bulge. Structural studies of epsilon can provide important parameters required for the design of RNA targeted anti- viral drugs directed against Hepatitis B virus. NMR studies of large RNA systems (> ca. 50 nucleotides) require novel approaches, e.g., different labeling schemes and reduction of the system into separate structural building blocks. Recently, a new method of synthesizing (13)C/(15)N/(2)H labeled nucleotides has been developed based on converting specifically labeled glucose and bases into nucleotides by using enzymes from the pentose phosphate pathway and nucleotide and salvage pathways. These NTPs give a large freedom in designing different labeling patterns in in vitro synthesized RNAs under study for NMR. This opens up the way for NMR studies of RNAs that are considerably above the present size limit (up to 150 nucleotides). Here this new technique is applied for structural studies on 27, 36 and 61 nucleotides long RNA fragments, mimicking different regions of epsilon.
机译：乙型肝炎病毒（HBV）HBV是具有独特复制策略的DNA病毒，涉及其前基因组RNA的逆转录。对于这种逆转录而言，必不可少的是其前基因组RNA的5和3端（分别为5 -RT-RNA和3 -RT-RNA），形成保守的鼓出的茎环结构。 5 -RT-RNA由67个核苷酸突出的茎环结构epsil构成，构成了前基因组RNA衣壳化和随后的反转录信号。逆转录酶（RT）最初与ε的完全保守的顶端环结合，并从相邻的6-核苷酸凸起合成4-核苷酸引物。 ε的结构研究可以提供设计针对B型肝炎病毒的RNA靶向抗病毒药物所需的重要参数。大型RNA系统（>大约50个核苷酸）的NMR研究需要新颖的方法，例如不同的标记方案以及将系统还原为单独的结构构件。最近，已经开发出一种新的合成（13）C /（15）N /（2）H标记核苷酸的新方法，该方法是通过使用戊糖磷酸途径以及核苷酸和挽救途径的酶，将经过特殊标记的葡萄糖和碱基转化为核苷酸。这些NTP具有很大的自由度，可以在NMR研究的体外合成RNA中设计不同的标记模式。这为大大超过当前大小限制（最多150个核苷酸）的RNA的NMR研究开辟了道路。在这里，这项新技术被用于对27、36和61个核苷酸长的RNA片段进行结构研究，模仿了epsilon的不同区域。

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