Structure and function of 5' to 3' exoribonucleases

5 至 3 核糖核酸外切酶的结构和功能

• 批准号: 8601533
• 负责人: LIANG TONG
• 金额: $ 38.1万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-01-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8601533
• 关键词: Active Sites; Animals; Biochemical; Biological; Biological Assay; C-terminal; Catalysis; Cell Nucleus; Cells; Complex; Cytoplasm; Data; Defect; Development; Dissection; Divalent Cations; Enzymes; Eukaryota; Event; Exoribonucleases; Family; Genes; Genetic Transcription; Goals; Hydrolysis; In Vitro; Knowledge; Link; Mammalian Cell; Mammals; Measures; Metabolism; MicroRNAs; Modeling; Molecular; Mutagenesis; Nonsense-Mediated Decay; Nuclear; Orthologous Gene; Physiological; Play; Polymerase; Process; Proteins; Quality Control; RNA; RNA Interference; RNA Polymerase II; RNA Polymerase III; Research Project Grants; Role; Saccharomyces cerevisiae; Sequence Homologs; Staging; Structure; Testing; Torpedo; Yeasts; analog; base; chromatin immunoprecipitation; comparative; design; enzyme activity; fungus; human disease; interest; mRNA decapping; novel; poly A specific exoribonuclease; public health relevance; research study; trafficking; transcription termination

DESCRIPTION (provided by applicant): The 5'->3' exoribonucleases (XRNs) comprise a large family of conserved enzymes in eukaryotes with important functions in RNA metabolism and RNA interference. In fungi and animals, XRN1 (175 kD) is primarily cytosolic and is involved in degradation of decapped mRNAs, nonsense mediated decay, microRNA decay and is essential for proper development. XRN2 (115 kD) is primarily nuclear, and its ortholog in S. cerevisiae, more commonly known as Rat1, has roles in RNA trafficking from the nucleus to the cytoplasm and transcription of some genes by RNA polymerase III. More recent studies have identified XRN2/Rat1 as the exoribonuclease that is essential for the 'torpedo' model of transcription termination by RNA polymerase II (Pol II). In S. cerevisiae, Rat1 co-purifies with another protein, Rai1 (Rat1 interacting protein 1), which appears to enhance the exoribonuclease activity of Rat1 in vitro. A weak sequence homolog of Rai1, known as DOM3Z, exists in mammals, although it probably does not associate with XRN2. The important functions of the XRNs in RNA metabolism, RNA interference and Pol II termination have generated significant interest in them. However, the molecular mechanism for the functions of these enzymes is currently still poorly understood. As a first step to fill these gaps in our knowledge, we recently determined the crystal structures of the Rat1-Rai1 complex as well as of Rai1 and DOM3Z alone, and carried out preliminary biochemical and functional studies based on the structural information. These results set the stage for further structural, biochemical and biological studies on these proteins. Most interestingly, our initial studies show that Rai1/DOM3Z contains a separate active site, with RNA 5' pyrophosphohydrolase activity, and we hypothesize that Rai1/DOM3Z may have important functions in RNA metabolism or quality control. We will also test this hypothesis in our proposed research project, which will help to define the physiological functions of this novel eukaryotic enzyme.

描述（由申请人提供）：5' - > 3'驱核核酸酶（XRNS）组成了一个在真核生物中具有重要功能在RNA代谢和RNA干扰中的重要功能的大型保守酶。在真菌和动物中，XRN1（175 kd）主要是胞质的，参与了降解的mRNA，胡说八道介导的衰减，microRNA衰变，对于正常发育至关重要。 XRN2（115 kd）主要是核的，其在酿酒酵母中的直系同源物（通常称为Rat1）在从细胞核到细胞质到细胞质的RNA运输中起作用，并通过RNA聚合酶III对某些基因进行转录。最新的研究已将XRN2/Rat1确定为RNA聚合酶II（POL II）的“鱼雷”转录终止模型必不可少的驱虫核酸酶。在酿酒酵母中，Rat1与另一种蛋白RAI1（RAT1相互作用1）共纯化，该蛋白似乎增强了Rat1体外的Exoriboneclease酶活性。 RAI1的弱序列同源物（称为DOM3Z）存在于哺乳动物中，尽管它可能与XRN2相关。 XRN在RNA代谢，RNA干扰和POL II终止中的重要功能引起了对它们的重大兴趣。但是，目前仍然对这些酶功能的分子机制仍然很少了解。作为填补这些空白的第一步，我们最近确定了RAI1-RAI1复合物以及RAI1和DOM3Z的晶体结构，并基于结构信息进行了初步的生化和功能研究。这些结果为这些蛋白质的进一步结构，生化和生物学研究奠定了基础。最有趣的是，我们的初步研究表明，RAI1/DOM3Z包含一个单独的活性位点，具有RNA 5'TOPHOPHOPOPHOHYDROLASE活性，我们假设RAI1/DOM3Z在RNA代谢或质量控制中可能具有重要功能。我们还将在我们提出的研究项目中检验这一假设，这将有助于定义这种新型真核生物酶的生理功能。