VERY HIGH RESOLUTION STRUCTURES OF THE FFH GTPASE

FFH GTPASE 的极高分辨率结构

• 批准号: 6151226
• 负责人: Douglas M. Freymann
• 金额: $ 23.62万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-01 至 2002-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6151226
• 关键词: X ray crystallography; bioimaging /biomedical imaging; conformation; crystallization; enzyme structure; guanosine triphosphate; guanosinetriphosphatases; intermolecular interaction; physical model; protein sequence; protein structure function; ribonucleoproteins; stereochemistry; structural biology; water

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.

所有细胞都含有蛋白质靶向不同细胞区室的机制。 蛋白质共翻译靶向细胞膜是由信号识别颗粒 (SRP) 介导的，该颗粒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系统发育上高度保守。 颗粒的一种蛋白质成分 SRP54 或 Ffh（在原核生物中）特别令人感兴趣，因为它提供了疏水信号肽的特异性识别，并且也是直接与膜受体相互作用的 GTP 酶。 GTP 酶是一个普遍存在的蛋白质家族，它利用 GTP 的结合和水解来引发各种细胞功能 - 经过充分研究的 GTP 酶涉及翻译、细胞调节和信号转导。 该提案旨在详细了解 Ffh 的 NG GTPase 结构域的蛋白质结构。 这些结果将有助于我们了解这种特定 GTP 酶在 SRP 途径中的作用，更广泛地说，有助于深入了解不同的 GTP 酶如何在共同的核心蛋白结构上构建以引发不同的细胞功能。该项目具有三个晶体学目标。 从现有晶体中获得高度精炼的超高（1.0 埃）分辨率的 apo 和 GDP 结合 NG GTPase 结构模型。 改进现有的 MgGDP 形式晶体，以便我们能够以类似的细节水平描述该结构。 并且，使用不可水解的核苷酸类似物或GTP使MgGTP结合的NG GTPase结晶，并解析其晶体结构。涵盖 GTP 酶配体状态的结构数据将为详细、准确地分析蛋白质、水、镁和核苷酸之间的相互作用提供基础。 他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 GTP 酶功能相关。分析将涉及 GTP 酶的化学性质和蛋白质设计原理，这些原理有助于在 GTP 结合和水解循环中调动不同的结构基序。