Author:
Date:
2020-06-20
[Abstract] Transfer RNAs (tRNAs) are heavily decorated with post-transcriptional modifications during their biosynthesis. To fulfil their functions within cells, tRNAs undergo a tightly controlled biogenesis process leading to the formation of mature tRNAs. In addition, functions of tRNAs are often modulated by their modifications. Although the biological importance of post-transcriptional RNA modifications is widely appreciated, methods to directly detect their introduction during RNA biosynthesis are rare and do not easily provide information on the temporal nature of events. To obtain information on the tRNA maturation process, we have developed a methodology, using NMR as a tool to monitor tRNA maturation in a non-disruptive and continuous fashion in cellular extracts. By following the ...
[摘要] [摘要 ] 转移RNA(tRNA )在其生物合成过程中大量修饰有转录后修饰。为了在细胞内履行其功能,tRNA 经历了严格控制的生物生成过程,导致了成熟的tRNA 的形成。此外,tRNA的功能通常是虽然转录后修饰RNA的生物学重要性被广泛理解,方法直接检测它们的RNA生物合成过程中引入是罕见的,并且不容易提供上events.To的时间特性信息获取的信息的tRNA 成熟 在此过程中,我们开发了一种方法,使用NMR作为监测细胞提取物中tRNA 成熟的无中断和连续方式。通过模型酵母tRNA 的时间分辨NMR 成熟,我们发现修饰是该方法的实施需要对具有不同修饰状态的tRNA 样品进行NMR光谱学分析,以鉴定各个修饰的NMR特征。此处将介绍用于NMR光谱分析修饰途径的tRNA 样品的生产,并在酵母tRNA Phe 上进行例证,但可以通过更改构建体的序列扩展到其他tRNA 。该方案描述了未修饰的生产通过体外转录获得tRNA 样品,并通过在大肠杆菌中重组表达tRNA 产生修饰的tRNA 样品。大肠杆菌。
[背景 ] 在生活的各个领域,合成和RNA的成熟包括在特定地点的核苷酸的转录后的化学修饰。在不同的RNA家族,tRNA基因不仅显示最高多种化学修饰,而且密度最高每转录修饰(〜中经修饰的核苷酸8-25%的tRNA 各种生物体的)(Boccaletto ...
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