Targeting cell separation systems of gram-negative bacteria.

针对革兰氏阴性细菌的细胞分离系统。

• 批准号: 9238648
• 负责人: Thomas G Bernhardt
• 金额: $ 50.85万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9238648
• 关键词: Acinetobacter baumannii; Antibiotic Resistance; Antibiotics; Bacteria; Bacterial Infections; Biogenesis; Biological Models; Cell Membrane Permeability; Cell Separation; Cell Wall; Cell division; Cells; Cellular biology; Chemicals; Chemosensitization; Cleaved cell; Clinical; Complex; Detergents; Development; Drug Targeting; Drug resistance; Enterobacter; Enzymes; Escherichia coli; Foundations; Generations; Genetic; Gram-Negative Bacteria; Growth; Hypersensitivity; Incidence; Infection; Klebsiella pneumonia bacterium; Knowledge; Laboratories; Lead; Logic; Mammalian Cell; Membrane; Methods; Modeling; Molecular Genetics; Morphology; Multi-Drug Resistance; Mutation; Mutation Analysis; Nature; New Agents; Newborn Infant; Organism; Pathway interactions; Peptidoglycan; Permeability; Pharmaceutical Preparations; Phase; Process; Proteobacteria; Pseudomonas aeruginosa; Pump; Resistance; Role; Site; System; Therapeutic; Toxic effect; Toxin; United States; amidase; antimicrobial peptide; bacterial resistance; base; cell envelope; combat; constriction; daughter cell; design; drug sensitivity; human disease; inhibitor/antagonist; killings; mutant; new therapeutic target; novel; novel therapeutics; pathogen; programs; public health relevance; scale up; screening; small molecule; small molecule libraries; therapeutic target; tool

DESCRIPTION (provided by applicant): Gram-negative bacteria are the causative agents of a variety of important human diseases. Successful treatment of infections with these organisms is hampered by the complex cell envelope that surrounds them. This envelope includes a second (outer) membrane layer that is difficult for drugs to penetrate. Gram-negative bacteria thus have a high intrinsic resistance to antibiotics and are completely insensitive to many drugs that are effective against bacteria lacking an outer membrane. In this project, we intend to validate new targets for antibiotics that when inactivated will compromise the permeability barrier of the outer membrane to either kill gram-negative bacteria or eliminate their intrinsic resistance to approved therapeutics. The project is based on our studies of cell division in Escherichia coli. In this model gram-negative bacterium, we recently identified factors required for cell envelope remodeling during division and have developed tools to study them. During division, new cell wall material produced by the cytokinetic machinery must be processed by hydrolytic enzymes for daughter cells to separate. We have discovered that this processing is carried out by enzymes called amidases that are activated by factors with LytM domains. Once the cell wall is processed, a second set of factors called the Tol-Pal system catalyzes the constriction of the outer membrane to complete the division process. Inactivation of either the amidases or the Tol-Pal system in E. coli and other gram-negative bacteria leads to the formation of long chains of cells that cannot separate. These cell chains have been shown to have a compromised outer membrane permeability barrier and are hypersensitive to many drugs. Despite their role in maintaining the outer membrane permeability barrier, the importance of these cell separation systems for drug resistance has not been investigated in problematic gram-negative pathogens. Therefore, in this project, we will validate cell separation systems as potential drug targets in te pathogen Pseudomonas aeruginosa. Molecular genetics will be used to investigate the effect of inactivating these systems on the growth and drug resistance of this organism, and chemical screens will be implemented to identify inhibitors of the process. Although we focus on P. aeruginosa, the systems chosen for targeting are highly conserved in gram-negative bacteria. Thus, our results will be broadly relevant to the development of novel treatments for gram-negative infections.

描述（由申请人提供）：革兰氏阴性细菌是多种重要人类疾病的病原体。这些生物体周围复杂的细胞包膜阻碍了对这些生物体感染的成功治疗。该包膜包括药物难以渗透的第二（外）膜层。因此，革兰氏阴性细菌对抗生素具有高度的内在耐药性，并且对许多有效对抗缺乏外膜的细菌的药物完全不敏感。在这个项目中，我们打算验证抗生素的新靶标，这些靶标在失活时会损害外部的渗透屏障。 膜可以杀死革兰氏阴性细菌或消除其对批准的治疗方法的内在耐药性。该项目基于我们对大肠杆菌细胞分裂的研究。在这个革兰氏阴性细菌模型中，我们最近确定了分裂过程中细胞包膜重塑所需的因素，并开发了研究它们的工具。在分裂过程中，细胞分裂机制产生的新细胞壁物质必须经过水解酶的处理，以便子细胞得以分离。我们发现这种加工过程是由一种称为酰胺酶的酶进行的，这种酶被具有 LytM 结构域的因子激活。一旦细胞壁被处理，第二组称为 Tol-Pal 系统的因子就会催化外膜收缩以完成分裂过程。大肠杆菌和其他革兰氏阴性细菌中酰胺酶或 Tol-Pal 系统的失活会导致形成无法分离的长链细胞。这些细胞链已被证明具有受损的外膜渗透性屏障，并且对许多药物过敏。尽管它们在维持外膜通透性屏障方面发挥作用，但这些细胞分离系统对于耐药性的重要性尚未在有问题的革兰氏阴性病原体中进行研究。因此，在这个项目中，我们将验证细胞分离系统作为病原体铜绿假单胞菌的潜在药物靶点。分子遗传学将用于研究失活这些系统对该生物体的生长和耐药性的影响，并将实施化学筛选来识别该过程的抑制剂。尽管我们关注的是铜绿假单胞菌，但选择用于靶向的系统在革兰氏阴性细菌中高度保守。因此，我们的结果将与革兰氏阴性感染新疗法的开发广泛相关。