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

前体mRNA剪接是所有真核生物基因表达所必需的，但我们对前体mRNA剪接的分子机制的理解是有限的。 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，剪接体是由5个snRNA和100多个蛋白质因子组成的大型RNA/蛋白质复合物。 许多证据表明Prp 8是一种关键的剪接体蛋白，它在催化核心与RNA密切相互作用，可能有助于催化核心的形成和稳定。 Prp 8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蛋白没有明显的序列同源性。 进一步的结构和生物化学分析将为Prp 8在剪接中的功能提供有价值的见解。 然而，这些研究受到难以获得大量全长Prp 8的阻碍。 鉴定、表达和纯化Prp 8的结构域将为表征Prp 8提供有价值的替代方法。 该项目使用独特的高通量方法来鉴定Prp 8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提供关于Prp 8在剪接中的功能的重要信息，指导未来的突变/遗传实验。 这些可溶性结构域也是表征Prp 8生物化学性质的宝贵资源。 这项研究是产生Prp 8原子图像的关键一步，否则无法获得，显着推进我们对前mRNA剪接分子机制的理解。该项目的重点是了解前mRNA剪接的分子机制，这是对包括人类在内的所有真核生物至关重要的最基本的生物学过程之一。 此外，一名博士后研究员，以及博士。包括少数民族在内的大学生也将参与这项工作。 该项目的成果将在基础广泛的科学期刊上发表，并在当地和国际会议上发表。 这项研究不仅促进了对基础科学的理解，而且有助于培养新一代科学家。

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.