Molecular mechanisms of nucleic acid machines

摘要 DNA复制是所有生物精确复制遗传物质之前的一个基本过程 细胞分裂。这个过程的中心是一种解旋酶，它使用ATP水解酶来分离碱基对 DNA允许聚合酶获得合成互补链的途径，并驱动 DNA上的复制机制。在人类和其他真核细胞中，解旋酶引擎是6- 蛋白质MCM复合体。拟议的研究将通过提供原子水平来填补知识空白 MCM蛋白质与三磷酸腺苷化合物和DNA相互作用的机制图。这些将是 通过X射线结构研究的协调方法在分子水平上进行研究 结晶学和低温电子显微镜以及体外方法研究它们的功能和 互动。DEAD-box RNA解旋酶DDX3X与RNA的许多方面有关 并与几种人类疾病有关，包括艾滋病毒、丙型肝炎病毒和髓母细胞瘤。这个 拟议的研究将通过提供DDX3X蛋白质相互作用的详细图片来填补知识空白 使RNA和ATP化合物处于功能状态。已经有相当多的初步数据 该项目包括鉴定含有RNA的DDX3X的最小片段- 刺激的ATPase活性及其与相关底物结合的晶体结构。

ABSTRACT DNA replication is a fundamental process for all organisms to precisely duplicate genetic material prior to cell division. Central to the process is a helicase enzyme that uses ATP-hydrolysis to separate base-paired DNA to allow polymerases to gain access to synthesize complementary strands and also to drive the replication machinery along the DNA. In human and other eukaryotic cells, the helicase engine is the 6- protein MCM complex. The proposed research will fill a knowledge gap by providing atomic level mechanism pictures of MCM proteins as they interact with ATP compounds and with DNA. These will be studied at the molecular level by a coordinated approach involving structural studies by X-ray crystallography and cryo-electron microscopy and also in vitro methods to study their functions and interactions. The DEAD-box RNA helicase DDX3X has been implicated in many aspects of RNA processing and is associated with several human diseases, including HIV, HCV, and medulloblastoma. The proposed research will fill a knowledge gap by providing detailed pictures of DDX3X proteins interacting with RNA and ATP compounds in the functional state. A considerable body of preliminary data has been obtained for this project that includes the identification of a minimal fragment of DDX3X that has RNA- stimulated ATPase activity and its crystal structure bound to relevant substrates.