Structural basis for cell surface siganling by a Gram-negative bacteria sigma-regulator

革兰氏阴性菌西格玛调节器细胞表面信号的结构基础

基本信息

  • 批准号:
    10240569
  • 负责人:
  • 金额:
    $ 29万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2018
  • 资助国家:
    美国
  • 起止时间:
    2018-09-20 至 2024-08-31
  • 项目状态:
    已结题

项目摘要

PROJECT SUMMARY/ABSTRACT The CDC recently released a report detailing antibiotic resistant threats in the US. Of particular emphasis in the CDC report is the increased prevalence of multidrug-resistant, Gram-negative bacteria (MDR- GNB) and the need to develop the next generation of antibiotics to combat them. All Gram-negative bacteria rely on a set of homologous, yet highly-specific, outer membrane TonB-dependent transporters (TBDTs) to import critical nutrients from their environment, especially metals like iron, which are bound by high-affinity, metal chelating compounds called siderophores. Recent antibiotic developments have shown that siderophore-antibiotic conjugates can be selectively targeted to specific bacteria, and that this delivery mechanism overcomes several key antibiotic resistance mechanisms. A significant limitation of this delivery system is the low expression levels of the TBDTs. However, a subset of these TBDTs controls their own expression through a cell-surface signaling (CSS) process that up-regulates their own expression. The long- term objective of this research is to understand the CSS regulatory process and manipulate TBDT expression to enhance siderophore-antibiotic conjugate therapy for treatment of MDR-GNB infections. Research outlined in this proposal will help elucidate the structural basis for CSS by a sigma-regulator. As a model system, the pseudobactin BN7/8 transport system of Psuedomonas putida, which consists of the TBDT, PupB, the inner membrane σ-regulator, PupR, and the cytoplasmic σ-factor, PupI, is being used. To accomplish this proposal's objective the following three specific aims will be pursued: 1) establish that PupR anti-σ-factor domain dimerization influences transcriptional activation by PupI, 2) identify the structural determinants and delineate the role of the PupR:PupB periplasmic interactions on the stability of the PupR periplasmic C- terminal CSS domain (CCSSD), and 3) determine changes in the full-length PupB:PupR CCSSD complex in the presence and absence of its cognate siderophore, pseudobactin BN7/8. These aims will be accomplished using a multidisciplinary approach; including X-ray crystallography, small-angle X-ray scattering, molecular biology, cellular assays, and biophysical techniques such as isothermal titration calorimetry and circular dichroism spectroscopy. This research will provide critical structural information about a σ-regulator; explain how it interacts with a σ-factor at the inner membrane, and the extent to which periplasmic conformational changes between the TBDT and σ-regulator lead to proteolytic degradation that is important for controlling transcriptional activation.
项目概要/摘要 疾病预防控制中心最近发布了一份报告,详细介绍了美国的抗生素耐药性威胁。特别是 CDC 报告强调的是多重耐药革兰氏阴性菌(MDR- GNB)以及开发下一代抗生素来对抗它们的需要。所有革兰氏阴性细菌 依赖于一组同源但高度特异性的外膜 TonB 依赖性转运蛋白 (TBDT) 从环境中输入关键的营养物质,特别是像铁这样的金属,它们通过高亲和力结合在一起, 称为铁载体的金属螯合化合物。最近抗生素的发展表明 铁载体-抗生素缀合物可以选择性地靶向特定细菌,并且这种递送 机制克服了几个关键的抗生素耐药机制。这种交付的一个重大限制 系统中TBDTs的表达水平较低。然而,这些 TBDT 的一个子集控制着它们自己的 通过细胞表面信号传导(CSS)过程上调其自身表达。长- 本研究的长期目标是了解 CSS 调控过程并操纵 TBDT 表达 增强铁载体-抗生素结合疗法治疗 MDR-GNB 感染。研究概述 该提案中的内容将有助于通过 sigma 调节器阐明 CSS 的结构基础。作为一个模型系统, 恶臭假单胞菌的假杆菌素BN7/8转运系统,由TBDT、PupB、内 使用膜 σ 调节因子 PupR 和细胞质 σ 因子 PupI。为了实现这一点 提案的目标 将追求以下三个具体目标: 1) 确定 PupR 抗 σ 因子 结构域二聚化影响 PupI 的转录激活,2) 识别结构决定因素和 描述 PupR:PupB 周质相互作用对 PupR 周质 C-稳定性的作用 终端 CSS 域 (CCSSD),以及 3) 确定全长 PupB:PupR CCSSD 复合体的变化 其同源铁载体假杆菌素 BN7/8 的存在和不存在。这些目标将会实现 采用多学科方法;包括X射线晶体学、小角X射线散射、分子 生物学、细胞测定和生物物理技术,例如等温滴定量热法和循环法 二色性光谱。这项研究将提供有关 σ 调节器的关键结构信息;解释 它如何与内膜的 σ 因子相互作用,以及周质构象的程度 TBDT 和 σ 调节剂之间的变化会导致蛋白水解降解,这对于控制蛋白水解很重要 转录激活。

项目成果

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Christopher L Colbert其他文献

Christopher L Colbert的其他文献

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{{ truncateString('Christopher L Colbert', 18)}}的其他基金

Analytical ultracentrifuge with absorbance and interference optics.
具有吸光度和干涉光学器件的分析超速离心机。
  • 批准号:
    10177341
  • 财政年份:
    2021
  • 资助金额:
    $ 29万
  • 项目类别:
Structural basis for cell surface siganling by a Gram-negative bacteria sigma-regulator
革兰氏阴性菌西格玛调节器细胞表面信号的结构基础
  • 批准号:
    9789675
  • 财政年份:
    2018
  • 资助金额:
    $ 29万
  • 项目类别:
Structural basis for cell surface siganling by a Gram-negative bacteria sigma-regulator
革兰氏阴性菌西格玛调节器细胞表面信号的结构基础
  • 批准号:
    10004679
  • 财政年份:
    2018
  • 资助金额:
    $ 29万
  • 项目类别:
Structural basis for cell surface siganling by a Gram-negative bacteria sigma-regulator
革兰氏阴性菌西格玛调节器细胞表面信号的结构基础
  • 批准号:
    10387865
  • 财政年份:
    2018
  • 资助金额:
    $ 29万
  • 项目类别:
Mechanism of inner membrane sigma-regulator function in Gram-negative bacteria
革兰氏阴性菌内膜西格玛调节功能的机制
  • 批准号:
    9316212
  • 财政年份:
    2015
  • 资助金额:
    $ 29万
  • 项目类别:

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