tRNA editing by deamination: Balancing affinity and specificty

通过脱氨基进行 tRNA 编辑：平衡亲和力和特异性

• 批准号: 8858638
• 负责人: Juan D Alfonzo
• 金额: $ 31.57万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8858638
• 关键词: Accounting; Adenosine; Affect; Affinity; Affinity Chromatography; Amino Acids; Anticodon; Binding; Biochemical; Biochemical Reaction; Biological; Biological Assay; Catalysis; Cell Line; Cell Nucleus; Cell physiology; Cells; Cellular biology; Chemicals; Codon Nucleotides; Complex; Coupled; Coupling; Cytidine Deaminase; Cytoplasm; Cytosine; DNA Sequence Alteration; Deaminase; Deamination; Disease; Enzymes; Equilibrium; Eukaryota; Event; Face; Family member; Future; Gene Expression; Genetic; Genome; Genomics; Human; Inosine; Introns; Knowledge; Leishmania; Link; Mammals; Medical; Metabolic Diseases; Metabolism; Methylation; Methyltransferase; Mitochondria; Modeling; Modification; Molecular; Nature; Nuclear; Nucleotides; Organism; Outcome; Parasites; Pathway interactions; Play; Positioning Attribute; Printing; Process; Protein Biosynthesis; Protein Family; Proteins; RNA Interference; RNA Splicing; RNA-Protein Interaction; Reaction; Recombinants; Regulation; Reporter; Research; Role; Series; Site; Specific qualifier value; Specificity; Structure; Substrate Specificity; System; Testing; Therapeutic Intervention; Threonine-Specific tRNA; Transfer RNA; Trypanosoma; Trypanosoma brucei brucei; Uridine; Zinc; base; crosslink; diprotin B; drug development; epigenetic regulation; flexibility; foot; in vitro activity; in vivo; insight; leucyl-alanine; member; mutant; novel; pathogen; reconstitution

DESCRIPTION (provided by applicant): Sequence alterations that change the meaning of a tRNA in decoding are part of a growing number of post-transcriptional changes collectively known as tRNA editing. Editing can be influenced by the structural context of an editing site and in the case of tRNA can be modulated by posttranscriptional modifications. In trypanosomatids, tRNAs are transcribed in the nucleus, exported to the cytoplasm, and later a subset of cytoplasmic tRNAs is actively imported into the mitochondria. However, before tRNAs can be rendered functional in any cellular compartment, they face many enzymatic reactions including end trimming, intron splicing, tRNA editing, and chemical modification. Some of these processes, for example those involved in trimming of extraneous sequences at the tRNA ends, occur in the nucleus, usually preceding cytoplasmic export. Others, like editing and modification, may occur at any point in the tRNA maturation pathway and in any of the tRNA-containing compartments. Despite much progress made in last few years, it is still not clear how editing and modification pathways are integrated both at the molecular and cellular levels. We have proposed that tRNA editing and modification events can be highly coupled and exploited by some organisms to control gene expression at the level of tRNA specificity. This is especially important in single-cell eukaryotes like trypanosomatid parasites where is well accepted that the bulk of the genetic regulation occurs post-transcriptionally. In this proposal, we have continued our studies on the very unique tRNA editing enzyme of T. brucei but now prompted by our newly solved crystal structure we ask new questions of what makes this enzyme so unique. We also focus on a newly discovered set of methyltransferases, which surprisingly target the editing site providing a wonderful testing ground for our hypothesis of the interrelation of editing and modification and what this coupling may mean in terms of cellular function. Significantly, reconstitution of methylation activity in vitro requires addition of the recombinant editing enzyme a finding that is without precedent in the tRNA editing and modification field. As essential steps in tRNA maturation in trypanosomatids (Leishmania and Trypanosoma), these types of editing and modification events also provide very attractive targets for therapeutic intervention against parasites of very major medical importance. Given the link between tRNA maturation and disease, these studies will further expand our knowledge of the role tRNA takes as a central player in cellular metabolism.

描述（由申请人提供）：改变tRNA在解码中的含义的序列改变是越来越多的转录后改变的一部分，这些转录后改变统称为tRNA编辑。编辑可以受到编辑位点的结构背景的影响，并且在tRNA的情况下，可以通过转录后修饰来调节。在锥虫中，tRNA在细胞核中转录，输出到细胞质，随后细胞质tRNA的子集被主动输入到线粒体中。然而，在tRNA可以在任何细胞区室中发挥功能之前，它们面临许多酶促反应，包括末端修剪，内含子剪接，tRNA编辑和化学修饰。其中一些过程，例如涉及在tRNA末端修剪外来序列的过程，发生在细胞核中，通常在细胞质输出之前。其他的，如编辑和修饰，可能发生在tRNA成熟途径的任何一点和任何含有tRNA的隔室。尽管在过去几年中取得了很大进展，但仍然不清楚编辑和修饰途径如何在分子和细胞水平上整合。我们已经提出，tRNA编辑和修饰事件可以高度耦合，并利用一些生物体在tRNA特异性水平上控制基因表达。这在单细胞真核生物如锥虫寄生虫中尤其重要，其中广泛接受大部分遗传调节发生在转录后。在这个提议中，我们继续对T.但是现在我们新解决的晶体结构促使我们提出新的问题，是什么使这种酶如此独特。我们还关注了一组新发现的甲基转移酶，它们令人惊讶地靶向编辑位点，为我们关于编辑和修饰的相互关系以及这种耦合在细胞功能方面可能意味着什么的假设提供了一个很好的测试基础。值得注意的是，体外甲基化活性的重建需要添加重组编辑酶，这在tRNA编辑和修饰领域是没有先例的。作为锥虫（利什曼原虫和锥虫）中tRNA成熟的重要步骤，这些类型的编辑和修饰事件也为针对具有非常重要医学意义的寄生虫的治疗干预提供了非常有吸引力的靶标。鉴于tRNA成熟与疾病之间的联系，这些研究将进一步扩大我们对tRNA在细胞代谢中的核心作用的认识。