Characterization of translation elongation mechanisms

基因表达是将DNA解码为RNA并转化为蛋白质的基本过程。将RNA解码为蛋白质，称为翻译，涉及一个称为核糖体的大型复杂大分子机器。核糖体读取并沿着RNA序列移动，从而将氨基酸顺序添加到新生蛋白质中。很明显，翻译是高度调节的，并且翻译的速率和保真度可以在不同的生物学情况下受到影响，例如细胞应激和病毒感染。目前，影响核糖体解码和核糖体运动的确切调控和机制知之甚少。由于它的根本重要性，我们必须了解翻译机制和规则的全部内容。为了直接解决这些问题，我们已经开发了一种新的最小重构翻译系统，以描绘所涉及的机制和因素。这个系统将允许我们一次系统地添加一个因素，以阐明每个步骤的功能。此外，该系统允许独特的机会来研究起始下游的事件，例如RNA的运动和解码。使用生物化学方法，我们将描述核糖体如何准确地解码RNA以及因子如何介导核糖体的运动。 最后，由于RNA可以折叠并形成稳定的结构，我们还将测试核糖体如何相互作用并解开这些RNA结构。通过利用这个最小的系统，这些实验将提高我们的基本基因表达机制的知识。

Gene expression is the fundamental process of decoding DNA to RNA into protein. Decoding RNA into protein, called translation, involves a large complex macromolecular machine called the ribosome. The ribosome reads and moves along the RNA sequence to sequentially add amino acids to the nascent protein. It is apparent that translation is highly regulated and the rates and fidelity of translation can be influenced under different biological situations such as cellular stress and viral infection. Currently, the exact regulations and mechanisms affecting ribosome decoding and movement of the ribosome is poorly understood. Because of its fundamental importance, it is imperative that we understand the full repertoire of translation mechanisms and regulations. To directly address these issues, we have developed a novel minimal reconstituted translation system in order to delineate the mechanisms and factors involved. This system will allow us to systematically add back factors one at a time in order to elucidate the function of each step. Moreover, this system allows the unique opportunity to study events downstream of initiation such as the movement and decoding of RNA. Using biochemical approaches, we will characterize how the ribosome accurately decodes RNA as well as how factors are mediating the movement of the ribosome. Finally, as RNA can fold and form stable structures, we will also test how the ribosome interacts and unwinds these RNA structures. By taking advantage of this minimal system, these experiments will enhance our knowledge of basic gene expression mechanisms.