PROTEIN RECOGNITION OF RNA MOTIFS: STRUCTURAL STUDIES

描述：这项工作的长期目标是确定原则 管理蛋白质-RNA识别的基因。蛋白质和RNA之间的相互作用是 生物过程的中心，从调控基因表达到 引导细胞死亡。对这些原理的了解很少， 这在很大程度上是由于缺乏蛋白质-RNA复合结构。这些 原理对于理解蛋白质-RNA机器很重要，例如 核糖体和剪接体，以及蛋白质-RNA结构和功能 将军。此外，它们对于发展以知识为基础的 针对依赖RNA的感染性病原体的治疗方法，如艾滋病毒。 作为迈向长期目标的一步，这项提议侧重于如何 核糖体失活蛋白限制性识别两个常见的基序(a 四环和G-凸起的交叉链A堆)在一个重要的部分 核糖体RNA。值得注意的是，具有86%序列同源性的限制性催产素 与其功能同源的sarcin，只裂解一个磷酸二酯键 在真核细胞核糖体RNA中发现的大约7000个。这个强大而有力的 特定的毒素识别核糖体RNA的一个保守区域，称为 Sarcin/ricin结构域(SRD)是蛋白质合成所必需的。 X射线结晶学、动力学和能量学研究将按顺序合并。 为了更深入地了解限制性内毒素是如何识别 SRD核糖核酸。第二系列的结晶学研究将集中在突变体上。 在SRD RNA中发现的两个基序中。这些研究将提供 对蛋白质识别原理的全新见解 这两个基序中，因为没有相关的蛋白质-RNA复合体 结构。 限制素及相关核糖体失活蛋白的临床研究 已经因为它们在“神奇子弹”疗法中的潜在应用而受到鼓舞 通过将毒素与肿瘤特异性物质联系起来，将毒素定向到肿瘤细胞 抗体。确定RNA底物识别表面和接触 对限制药物的作用至关重要的因素将有助于设计或选择 未来以核毒素为基础的疗法。

DESCRIPTION: The long-term goal of this work is to determine the principles that govern protein-RNA recognition. Interaction between protein and RNA is central to biological processes ranging from regulating gene expression to directing cell mortality. Knowledge of these principles is poorly understood, due in large part to a paucity of protein-RNA complex structures. These principles are important for understanding protein-RNA machines, such as the ribosome and the spliceosome, and protein-RNA structure and function in general. In addition, they are important for developing knowledge-based therapies against RNA-dependent infectious agents such as HIV. As a step toward the long-term goal, this proposal focuses on how the ribosome-inactivating protein restrictocin recognizes two common motifs (a tetraloop and a G-bulged cross-strand A stack) in an essential piece of ribosomal RNA. Remarkably, restrictocin, which shares 86% sequence identity with its functional homolog sarcin, cleaves only one phosphodiester bond out of the approximately 7000 found in eukaryotic ribosomal RNA. This potent and specific toxin recognizes a conserved region of ribosomal RNA called the sarcin/ricin domain (SRD) that is essential for protein synthesis. X-ray crystallographic, kinetic and energetic studies will be combined in order to obtain a deeper understanding of how restrictocin recognizes both motifs in the SRD RNA. A second series of crystallographic studies will focus on mutants in the two motifs found in the SRD RNA. These studies will provide fundamentally new insights into the principles that govern protein recognition of these two motifs, because there is an absence of related protein-RNA complex structures. Clinical interest in restrictocin and related ribosome-inactivating proteins has been invigorated by their potential use in "magic bullet" therapies that direct toxins to tumor cells by linking them to tumor-specific agents such as antibodies. Determining the RNA substrate recognition surface and the contacts that are critical for restrictocin action will aid the design or selection of future ribotoxin-based therapies.