Analysis of functional structure of ribosomal RNA

本研究旨在通过分析核糖体和核糖体RNA(RRNA)的功能结构，阐明翻译系统组成和功能的基本原理。由于翻译系统是生命中最基本的系统，阐明其构成原理将有助于阐明生命的起源。在本研究的开始，我们试图通过用吡啶处理去蛋白核糖体来阐明翻译反应中的每一步，目的是阐明大肠杆菌核糖体和rRNA的功能结构。这类核糖体具有非酶(自发)易位活性，但近两年来，核糖体晶体的X射线分析已经有了很大的进步，通过在原子水平上观察核糖体的三维结构来阐明核糖体的反应机制成为可能。因此，我们的研究目标从大肠杆菌转移到动物线粒体肋骨…更多的糖体，因为线粒体核糖体拥有大约一半缩短的rRNA，而蛋白质加倍放大，而相对分子质量和整体大小与大肠杆菌核糖体相似，这表明rRNA中功能位点的鉴定要容易得多，而且在线粒体进化过程中，RNA的作用可能已经转移到蛋白质上。我们已经从牛的大约70个蛋白质中鉴定出55个核糖体蛋白质，线粒体核糖体(有丝分裂糖体)通过双向凝胶电泳分离每个核糖体蛋白质，然后用LC/MS技术进行鉴定。我们现在正在对这些蛋白质进行表征。研究还发现，只有L7/L12蛋白被大肠杆菌对应物取代的杂交型有丝分裂糖体既可以与一组线粒体tRNA和翻译因子一起发挥作用，也可以与一组大肠杆菌tRNA和翻译因子一起发挥作用，这表明尽管核糖体的大部分核糖体在功能上是相同的，但大肠杆菌和线粒体核糖体的RNA和蛋白质比例是相反的。我们还在研究大肠杆菌和线粒体核糖体之间这种功能互换的起源。较少

This research project aims to elucidate the basal principle by which the translation system is constituted and functions, by analyzings the, functional structures of the ribosome and ribosomal RNA (rRNA). Since the translation system is the most basic system in life, elucidation of its constitution principle would lead to elucidation of origin of life.In the beginning of this research, we tried to elucidate each step in the translation reaction using protein-deprived ribosomes treated with pyridine for the, purpose of elucidating the functional structure of E. coli ribosomes and rRNA. It turned out that such ribosomes possess non-enzymatic (spontaneous) translocation activity.However, in these two years X-ray analysis of ribosome crystals has been greatly improved and it became possible to elucidate the reaction mechanism of ribosomes by inspection of the three-dimensional structure of ribosomes at atomic level. We thus moved our research target from E. coli to animal mitochondrial rib … More osomes, because mitochondrial ribosomes possess about a half-shortened rRNA and instead, doubly enlarged proteins, while the molecular weight and whole size being similar, as compared to those of E. coli ribosomes, suggesting that the identification of functional sites in rRNA is much easier and moreover, the roles of RNA could have transferred to proteins during evolution of mitochondria.We have already identified 55 ribosomal proteins out of about 70 proteins in bovine, mitochondrial ribosome (mitoribosome) by separating each ribosomal, protein, on the two-dimensional gel electrophoresis followed by its identification by LC/MS technique. We are now characterizing these proteins. It also turned out that a hybrid-type mitoribosome in which only L7/L12 protein was replaced by the E. coli counterpart can function either with a set of mitochondrial tRNA and translation factors or with a set of E. coli tRNA and translation factors, suggesting that the most part of the ribosome is functionally equivalent between E. coli and mitochondrial ribosomes, notwithstanding that the RNA-protein ratio is reversed. We are also examining the origin of this functional interchangeability between E. coli and mitochondrial ribosomes. Less