VERY HIGH RESOLUTION STRUCTURES OF THE FFH GTPASE

所有细胞都含有蛋白质靶向不同细胞区室的机制。 蛋白质向细胞膜的共翻译靶向由信号识别颗粒（SRP）介导，其识别新生多肽的氨基末端信号序列并将翻译核糖体靶向膜受体。 SRP核糖核蛋白的结构和功能元件在遗传学上高度保守。 该颗粒的一种蛋白质成分SRP 54或Ffh（在原核生物中）特别令人感兴趣，因为它提供对疏水性信号肽的特异性识别，并且也是一种直接与膜受体相互作用的GTPase。 GTP酶是一个普遍存在的蛋白质家族，其利用GTP的结合和水解来引发各种细胞功能-研究充分的GTP酶参与翻译、细胞调节和信号转导。 这个建议是针对详细的蛋白质结构的理解NG GT3结构域的Ffh。 这些结果将有助于我们理解这种特定的GTPases在SRP途径中的作用，并且更普遍地，深入了解不同的GTPases如何建立在共同的核心蛋白结构上以引发不同的细胞功能。该项目有三个晶体学目标。 从现有晶体中获得apo和GDP结合的NG GTdR的高度精制的超高（1.0埃）分辨率结构模型。 为了改进现有的MgGDP形式的晶体，以便我们可以在类似的细节水平上描述该结构。 并且，使用不可水解的核苷酸类似物或GTP使MgGTP结合的NG GTP酶结晶，并解析其晶体结构。的结构数据，跨越的GTdR的配体状态，将提供的基础，详细和准确的分析蛋白质，水，镁和核苷酸之间的相互作用。 它们还应该揭示在较低分辨率的结构中未解决的结构现象-蛋白质立体化学的偏差，包装缺陷和构象底物-是否可能与GTdR功能相关。分析将解决的GTP酶和蛋白质的设计原则，促进动员不同的结构基序在GTP结合和水解的周期中的化学。

All cells contain mechanisms by which proteins are targeted to different cellular compartments. Co-translational targeting of proteins to cell membranes is mediated by the signal recognition particle (SRP), which recognizes the amino-terminal signal sequence of the nascent polypeptide and targets the translating ribosome to a membrane receptor. Structural and functional elements of the SRP ribonucleoprotein are highly phylogenetically conserved. One protein component of the particle, SRP54 or Ffh (in prokaryotes), is of particular interest because it provides specific recognition of the hydrophobic signal peptide and is also a GTPase which interacts directly with the membrane receptor. GTPases are a ubiquitous family of proteins which use the binding and hydrolysis of GTP to elicit various cellular functions - well studied GTPases are involved in translation, cell regulation, and signal transduction. This proposal is directed towards detailed protein structural understanding of the NG GTPase domain of Ffh. The results will contribute to our understanding of the role of this specific GTPase in the SRP pathway, and, more generally, give insight into how different GTPases build on a common core protein structure to elicit different cellular function. The project has three crystallographic objectives. To obtain highly refined ultra-high (1.0 Angstroms) resolution structural models of the apo- and GDP-bound NG GTPase from crystals now in hand. To improve existing MgGDP-form crystals so that we can describe that structure at a similar level of detail. And, to crystallize the MgGTP-bound NG GTPase using nonhydrolyzable nucleotide analogs or GTP, and to solve its crystal structure as well. The structural data, spanning the ligand states of the GTPase, will provide the basis for detailed and accurate analysis of the interaction between the protein, water, magnesium, and nucleotide. They should also reveal whether structural phenomena not resolved in structures at lower resolution - deviations of the protein stereochemistry, packing imperfections, and conformational substrates - may be relevant to GTPase function. The analyses will address the chemistry of the GTPase and the protein design principles which facilitate mobilization of different structural motifs during the cycle of GTP binding and hydrolysis.