Understanding the Structure and Function of pre-mRNA Splicing Factor Prp8

了解前体 mRNA 剪接因子 Prp8 的结构和功能

• 批准号: 0718802
• 负责人: Rui Zhao
• 金额: --
• 依托单位: University of Colorado at Denver
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2009-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0718802&HistoricalAwards=false
• 关键词: Understanding; Structure; Function; pre; mRNA

Pre-mRNA splicing is essential for gene expression in all eukaryotes, but our understanding of the molecular mechanisms of pre-mRNA splicing is limited. Splicing of introns is carried out through two transesterification reactions catalyzed by the spliceosome, a large RNA/protein complex composed of five snRNAs and over 100 protein factors. Many lines of evidence point to Prp8 as a key spliceosomal protein that interacts intimately with RNA at the catalytic core, potentially helping the formation and stabilization of the catalytic core. Prp8 is one of the largest and most conserved nuclear proteins known, but it does not have obvious sequence homology with any other known protein. Further structural and biochemical analyses would provide valuable insight into Prp8's function in splicing. However, these studies are hindered by difficulties in obtaining large quantities of full-length Prp8. Identifying, expressing, and purifying domains of Prp8 will provide a valuable alternative approach for characterizing Prp8. This project uses a unique high throughput approach to identify soluble domains of Prp8 and determine structures of these domains. Structures of these domains and comparison with other known structures can provide important information on the function of Prp8 in splicing, directing future mutational/genetic experiments. These soluble domains are also valuable resources for characterizing Prp8's biochemical properties. This research is a critical step toward generating an atomic picture of Prp8 that cannot be obtained otherwise, significantly advancing our understanding of the molecular mechanisms of pre-mRNA splicing. This project focuses on understanding the molecular mechanism of pre-mRNA splicing, one of the most fundamental biological processes critical for all eukaryotic species including humans. In addition, a postdoctoral fellow, as well as Ph.D. and undergraduate students, including minority members, will participate in this work. Results from this project will be published in broad based scientific journals and presented at local and international meetings. This research not only advances the understanding of basic science but also helps train a new generation of scientists.

前信使核糖核酸剪接对于所有真核生物的基因表达都是必不可少的，但我们对前信使核糖核酸剪接的分子机制的了解是有限的。内含子的剪接是通过剪接体催化的两个酯交换反应进行的，剪接体是一个由5个SnRNA和100多个蛋白质因子组成的大型RNA/蛋白质复合体。许多证据表明Prp8是一种关键的剪接体蛋白，它与催化核心的RNA密切相互作用，可能有助于催化核心的形成和稳定。Prp8是已知的最大和最保守的核蛋白之一，但它与任何其他已知的蛋白质没有明显的序列同源性。进一步的结构和生化分析将为深入了解Prp8的S在剪接中的功能提供有价值的信息。然而，这些研究因难以获得大量全长Prp8而受到阻碍。鉴定、表达和纯化Prp8的结构域将为鉴定Prp8提供一种有价值的替代方法。该项目使用一种独特的高通量方法来确定Prp8的可溶性结构域并确定这些结构域的结构。这些结构域的结构以及与其他已知结构的比较可以为Prp8在剪接中的功能提供重要信息，指导未来的突变/遗传学实验。这些可溶性区也是研究Prp8‘S生化特性的宝贵资源。这项研究是朝着产生Prp8的原子图像迈出的关键一步，这是用其他方法无法获得的，大大促进了我们对前mRNA剪接的分子机制的理解。这个项目的重点是了解前mRNA剪接的分子机制，这是包括人类在内的所有真核物种至关重要的最基本的生物学过程之一。此外，还将有一名博士后研究员以及包括少数民族成员在内的博士和本科生参与这项工作。该项目的成果将发表在基础广泛的科学期刊上，并在当地和国际会议上介绍。这项研究不仅增进了对基础科学的理解，而且有助于培养新一代科学家。