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 抗生素耐药性的日益增加和新抗菌药物发现的减少威胁着全球 卫生保健。特别值得关注的是革兰氏阴性病原体，即具有额外外膜的生物体 (OM) 对多种抗生素具有内在耐药性。与内膜 (IM) 不同的是 OM 仅由甘油磷脂 (GPL) 组成，与内叶中发现的 GPL 不对称 和脂多糖（LPS）位于外叶。这种独特的膜不对称性使得 防止大极性分子以及亲脂性化合物的侵害，形成不透水的屏障。自从 OM 至关重要，其组装所需的途径是抗菌设计的关键目标。现在， 临床上尚无直接针对 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。 在过去的几十年里，我们扩展了对 OM 组装的理解，揭示了新的目标 抗菌设计。然而，仍然存在一个重大差距。 GPL 如何从 IM 传输到 OM 穿过水性周质？最近，我们发现 AsmA 样家族的关键成员（YhdP、 TamB 和 YdbH）对于 OM 完整性至关重要并参与 GPL 传输。我们发现 YhdP、TamB 和 YdbH 在 OM 脂质稳态中的作用是多余的；然而，它们并不等同。值得注意的是，这三个 蛋白质与真核 GPL 转运蛋白具有同源性和结构特征，并且能够跨越 周质。该应用的总体目标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，我们将描述路径 使用大肠杆菌作为模型系统在细胞膜上运输 GPL 是必需的。在当前 应用程序中，我们将 (i) 描述主要 GPL 传输程序（YhdP、TamB 和 YdbH）的特征，(ii) 识别附件 GPL 跨双层运动所需的蛋白质，以及 (iii) 确定这些系统的丢失如何影响 OM 脂质稳态和抗生素耐药性。