Synthesis and transport of outer membrane components across the Gram-negative cell envelope

外膜成分的合成和跨革兰氏阴性细胞包膜的运输

• 批准号: 10680968
• 负责人: Michael Stephen Trent
• 金额: $ 57.98万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-27 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10680968
• 关键词: Antibiotic Resistance; Antibiotics; Antimicrobial Resistance; Attention; Bacteria; Bacterial Infections; Biochemical; Biogenesis; Biological Models; Biology; Cells; Cessation of life; Clinical; Complex; Cytoplasm; Data; Dedications; Development; Encapsulated; Energy-Generating Resources; Escherichia coli; Family; Future; Genetic; Glycerophospholipids; Gram-Negative Bacteria; Healthcare; Homeostasis; Hydrophobicity; Infection; Lipids; Lipopolysaccharides; Lipoproteins; Maintenance; Membrane; Membrane Lipids; Membrane Proteins; Methodology; Molecular; Molecular Chaperones; Monitor; Movement; Multi-Drug Resistance; Organism; Pathway interactions; Peptidoglycan; Proteins; Reporting; Resistance; Role; Side; Surface; System; Systems Analysis; antimicrobial; aqueous; beta barrel; cell envelope; design; economic cost; fighting; global health; lipid transport; lipophilicity; member; membrane assembly; membrane biogenesis; new therapeutic target; novel; novel therapeutics; pathogen; periplasm; prevent; protein complex

Abstract The increasing rise in antibiotic resistance and the diminished discovery of new antimicrobials threatens global healthcare. Of particular concern are Gram-negative pathogens, organisms with an additional outer membrane (OM) that provides intrinsic resistance to multiple classes of antibiotics. Unlike the inner membrane (IM) that is composed solely of glycerophospholipids (GPLs), the OM is asymmetrical with GPLs found in the inner leaflet and lipopolysaccharide (LPS) localized to the outer leaflet. This unique membrane asymmetry affords protection from large polar molecules, as well as lipophilic compounds, creating an impervious barrier. Since the OM is essential, pathways required for its assembly are key targets for antimicrobial design. Currently, there are no antibiotics that directly target OM biogenesis in clinical use. Thus, it remains critical to investigate cell envelope biogenesis for future and current antimicrobial design. Over the last few decades, we have expanded our understanding of OM assembly revealing new targets for antimicrobial design. However, one major gap remained. How are GPLs transported from the IM to the OM across the aqueous periplasm? Recently, we discovered that key members of the AsmA-like family (YhdP, TamB, and YdbH) are critical for OM integrity and involved in GPL transport. We found that YhdP, TamB, and YdbH are redundant in their role in OM lipid homeostasis; however, they are not equivalent. Notably, all three proteins share homology and structural features with eukaryotic GPL transporters and are capable of spanning the periplasm. The overall objective of this application is to investigate the molecular mechanisms required for the assembly and maintenance of the Gram-negative OM. More specifically, we will characterize pathways required to transport GPLs across the cell envelope using E. coli as the model system. In the current application we will (i) characterize the major GPL transporters (YhdP, TamB, and YdbH), (ii) identify accessory proteins required for GPL transbilayer movement, and (iii) determine how loss of these systems impact OM lipid homeostasis and antibiotic resistance.

抽象的 抗生素耐药性的增加以及新抗菌剂的发现减少威胁着全球 卫生保健。特别关注的是革兰氏阴性病原体，具有附加外膜的生物 （OM）对多种类别的抗生素具有内在的抗性。与内膜（IM）不同 OM仅由甘油磷脂（GPLS）组成，与内部小叶中发现的GPL是不对称的 和脂多糖（LPS）位于外部小叶中。这种独特的膜不对称提供 防止大型极性分子以及亲脂化合物，从而产生不透水的屏障。自从 OM是必不可少的，其组装所需的途径是抗菌设计的关键目标。现在， 在临床用途中，没有直接靶向OM生物发生的抗生素。因此，调查仍然至关重要 用于未来和当前抗菌设计的细胞包膜生物发生。 在过去的几十年中，我们扩展了对OM集会的理解，揭示了新的目标 抗菌设计。但是，仍然存在一个主要差距。 GPLS如何从IM运输到OM 穿过骨质水性？最近，我们发现了类似Asma的家庭的主要成员（YHDP， TAMB和YDBH）对于OM完整性至关重要，并且参与GPL运输。我们发现YHDP，Tamb和 YDBH在OM脂质稳态中的作用是多余的。但是，它们不是等效的。值得注意的是，这三个 蛋白质与真核GPL转运蛋白共享同源性和结构特征，并且能够跨越 周期。该应用的总体目的是研究所需的分子机制 用于革兰氏阴性OM的组装和维护。更具体地说，我们将表征途径 需要使用大肠杆菌作为模型系统将GPLS运输在细胞包膜上。在电流中 应用程序我们将（i）表征主要的GPL转运蛋白（YHDP，TAMB和YDBH），（ii）标识附件 GPL Transbilayer运动所需的蛋白质，以及（iii）确定这些系统丢失如何影响OM 脂质稳态和抗生素耐药性。