Protein-RNA Interactions in the Ribosome

9818051所有细胞中遗传信息的表达取决于蛋白质和RNA分子之间特异性的非共价相互作用。没有什么比核糖体更能说明这种相互作用的重要性了，核糖体是复杂的核糖核蛋白颗粒，根据信使RNA编码的遗传信息进行细胞蛋白质的生物合成。核糖体颗粒由大约55种不同的蛋白质，两个大RNA分子和一个小RNA分子组成，由于非共价蛋白质-RNA相互作用的复杂网络，它们组装成紧凑的功能结构。核糖体蛋白和核糖体RNA之间的特异性关联对于核糖体的组装和结构完整性都是必不可少的，并且是该颗粒在蛋白质合成中的功能活性的基础。在大肠杆菌中，许多与核糖体RNA相关的蛋白质也通过与相应信使RNA的特异性相互作用来调节核糖体蛋白基因的表达。这些复合物为研究蛋白质-RNA相互作用的分子基础提供了丰富的材料来源。本研究旨在阐明介导小亚基蛋白S8与其16 S核糖体RNA结合位点和大亚基蛋白L1与其23 S核糖体RNA结合位点之间关联的特定结构特征。每个蛋白质结合位点已被缩小到约30个核苷酸，蛋白质S8的结合位点已通过NMR解析。通过定点诱变、蛋白质-RNA交联、官能团分析、SELEX、NMR和X射线晶体学，将进一步深入了解S8-和L1-RNA相互作用的性质。关于核糖体蛋白-rRNA复合物结构的精确信息不仅对于理解几十种独特的核糖体组分自发组装成功能性核糖核蛋白颗粒的方式很重要，而且对于理解许多不同的蛋白-RNA相互作用的基础原理也很重要，这些蛋白-RNA相互作用在通过DNA合成表达遗传信息中起着至关重要的作用，RNA和蛋白质分子。储存在DNA中的遗传信息的表达最终导致蛋白质的合成，这些蛋白质定义了单个细胞的特性。值得注意的是，这一过程的机制在所有生物体中非常相似，从细菌到人类。蛋白质的合成是由被称为核糖体的紧凑的球状颗粒进行的，核糖体由大量蛋白质和RNA分子以复杂和高度特异性的方式排列组成。核糖体颗粒引导许多其他细胞组分在其表面上的组装，其方式允许根据DNA中编码的遗传计划将氨基酸有效地连接成蛋白质链。核糖体在蛋白质合成中的功能关键取决于这些粒子所呈现的精确的三维结构。核糖体的结构反过来又由其蛋白质和RNA成分之间的无数化学相互作用决定。本研究的主要目的是通过确定构成核糖体颗粒三维结构的一些蛋白质-RNA复合物的分子结构来更好地理解这种相互作用。这些结果将有助于解释核糖体的结构如何影响其在蛋白质合成中的功能。

9818051The expression of genetic information in all cells depends on specific, noncovalent interactions between protein and RNA molecules. Nowhere is the importance of such interactions better illustrated than in ribosomes, the complex ribonucleoprotein particles that carry out the biosynthesis of cellular proteins according to genetic information encoded in messenger RNA. Ribosomal particles are composed of roughly 55 different proteins, two large RNA molecules, and one small RNA molecule which assemble into a compact, functional structure owing to an intricate network of noncovalent protein-RNA interactions. Specific associations between ribosomal proteins and ribosomal RNAs are essential both for the assembly and for the structural integrity of the ribosome, and underlie the functional activity of this particle in protein synthesis. In the bacterium Escherichia coli, many of the proteins that associate with ribosomal RNA also regulate the expression of the ribosomal protein genes through specific interactions with the corresponding messenger RNAs. These complexes provide a rich source of material for investigating the molecular bases of protein-RNA interaction. The present research is directed toward elucidating the specific structural features that mediate the association of small-subunit protein S8 with its binding site in 16S ribosomal RNA and of large-subunit protein L1 with its binding site in 23S ribosomal RNA. Each of the protein binding sites has been narrowed down to about 30 nucleotides and that for protein S8 has been solved by NMR. Further insight into the nature of the S8- and L1-RNA interactions will be gained through site-directed mutagenesis, protein-RNA cross-linking, functional group analysis, SELEX, NMR and X-ray crystallography. Precise information about the structure of ribosomal protein-rRNA complexes is important not only for understanding the way in which several dozen unique ribosomal components spontaneously assemble into functional ribonucleoprotein particles, but also for comprehending the principles that underlie the many different protein-RNA interactions that play crucial roles in the expression of genetic information via the synthesis of DNA, RNA and protein molecules.The expression of the genetic information stored in the DNA culminates in the synthesis of the proteins that define the properties of individual cells. Remarkably, the mechanism of this process is quite similar in all organisms, from bacteria to humans. Protein synthesis is carried out by compact globular particles known as ribosomes which consist of a large number of protein and RNA molecules arranged in an intricate and highly specific manner. The ribosomal particles guide the assembly of many other cellular components on its surface in a way that permits the efficient linkage of amino acids into protein chains according to the genetic plans encoded in the DNA. Ribosome function in protein synthesis depends critically on the precise three-dimensional structure that these particles assume. The structure of the ribosome is in turn determined by a myriad of chemical interactions among its protein and RNA components. The main objective of this research is to gain a better understanding of such interactions by determining the molecular architecture of some of the protein-RNA complexes that underlie the three-dimensional structure of the ribosomal particle. The results will help to explain the manner in which the structure of the ribosome contributes to its function in protein synthesis.