LptA-mediated transport of LPS

LptA 介导的 LPS 转运

• 批准号: 8919418
• 负责人: CANDICE S KLUG
• 金额: $ 29.07万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8919418
• 关键词: Affinity; Alanine; Amino Acids; Antibiotics; Bacteria; Binding; Binding Sites; Biological Assay; C-terminal; Calorimetry; Carrier Proteins; Cell surface; Cells; Cessation of life; Characteristics; Data; Development; Disease; Drug Design; Drug Targeting; Electron Spin Resonance Spectroscopy; Employee Strikes; Endotoxins; Environment; Escherichia coli; Foundations; Future; Genes; Genetic Screening; Genetic study; Gram-Negative Bacteria; Growth; Health; Human; In Vitro; Infection; Inflammatory; Knowledge; Lasers; Lead; Libraries; Lipid Binding; Lipopolysaccharides; Measurement; Measures; Mediating; Membrane; Membrane Proteins; Modeling; N-terminal; Names; Periplasmic Proteins; Physiologic pulse; Plasmids; Process; Proteins; Pseudomonas aeruginosa; Role; Salmonella typhimurium; Septic Shock; Site; Spectrum Analysis; Staging; Structure; Techniques; Temperature; Time; Titrations; Transport Process; biophysical techniques; cell growth; in vivo; innovation; insight; light scattering; mutant; novel; pathogen; pathogenic bacteria; periplasm; pressure; protein protein interaction; protein transport; research study; tool

DESCRIPTION (provided by applicant): Lipopolysaccharide (LPS) is the major component of the outer leaflet of the outer membrane (OM) of Gram-negative bacteria such as Escherichia coli, Salmonella typhimurium and many other important pathogens. LPS, also referred to as endotoxin, is essential for survival in this large class of bacteria and serves as a first line of defense against hostile environments encountered during host infection. Given the essential role of LPS in the survival of Gram-negative bacteria - i.e., the bacterial cells die if any step o LPS transport does not occur - and the unique cell surface it creates, a detailed understanding of the proteins and mechanisms involved in LPS synthesis and transport will be the foundation on which to develop novel antibiotics against these promising new drug targets. Many of the proteins involved in LPS transport have been identified through recent genetics studies, suggesting that a set of seven inner membrane (IM), periplasmic, and OM proteins (named LptA, LptB, LptC, LptD, LptE, LptF, and LptG) are directly involved in moving LPS from the IM to the OM. However, the mechanism of how this group of proteins transports LPS to the OM is yet unknown. One of the most striking questions about this process is how the hydrophobic domain of LPS crosses the periplasm. Therefore, the proposed studies will focus on how the periplasmic protein LptA receives LPS from the IM-associated protein LptC, how LptA protects the hydrophobic acyl chains of LPS as it crosses the periplasm, and how LptA delivers LPS to LptDE at the OM. The successful completion of the proposed studies will include the development of a novel functional assessment tool for LptA, the creation of a comprehensive library of in vivo growth assay results to identify LptA amino acids critical for its structure or function, the identification of the specific LptA sites and conformational changes involved in LPS binding, and the characterization of the interactions between LptA and its binding partners LptC, LptDE, and LPS. The results of the novel genetic screenings, the laser light scattering analyses, the innovative electron paramagnetic resonance (EPR) spectroscopy studies, and the isothermal titration calorimetry measurements will provide detailed insights into the mechanism of LPS transport across the periplasm of Gram-negative bacteria. This unique knowledge will greatly enhance our growing understanding of LPS transport in bacteria and set the stage for future studies on the other Lpt proteins of unknown structure and function.

描述（由申请人提供）：脂多糖（LPS）是革兰氏阴性细菌外膜（OM）外叶的主要组成部分，例如大肠杆菌，鼠伤寒沙门氏菌和许多其他重要病原体。 LPS（也称为内毒素）对于在这种大类细菌中生存至关重要，并且是针对宿主感染期间遇到的敌对环境的第一道防线。鉴于LP在革兰氏阴性细菌的存活中的重要作用 - 即，如果没有发生任何步骤OLPS运输的任何步骤，细菌细胞会死亡 - 并且它产生的独特细胞表面，对LPS合成和运输中涉及的蛋白质和机制的详细理解将是抗这些有影响力的新药物的基础。 通过最近的遗传学研究鉴定了许多参与LPS转运的蛋白质，这表明一组七个内膜（IM），周质和OM蛋白（称为LPTA，LPTA，LPTB，LPTC，LPTC，LPTD，LPTD，LPTE，LPTE，LPTF和LPTG）直接参与从IM到IM到OM的LPS。但是，这组蛋白质如何将LP传输到OM的机制尚不清楚。关于此过程的最引人注目的问题之一是LPS的疏水结构域如何越过周期。因此，拟议的研究将重点关注周质蛋白LPTA如何从IM相关蛋白LPTC中接收LPS，LPTA如何保护LPS的疏水性酰基链时，它如何越过周期张力，以及LPTA如何将LPS传递到OM的LPTDE。 The successful completion of the proposed studies will include the development of a novel functional assessment tool for LptA, the creation of a comprehensive library of in vivo growth assay results to identify LptA amino acids critical for its structure or function, the identification of the specific LptA sites and conformational changes involved in LPS binding, and the characterization of the interactions between LptA and its binding partners LptC, LptDE, and LPS.新型遗传筛查，激光光散射分析，创新的电子顺磁共振（EPR）光谱研究的结果以及等温滴定量热法测量结果将提供详细的见解，以了解LPS在跨粒子性细胞质的periplymas的跨层中的机制。这种独特的知识将大大增强我们对细菌中LPS运输的不断增长，并为未来对未知结构和功能的LPT蛋白的研究奠定了基础。