METABOLIC ASPECTS OF BACTERIAL ECOLOGY IN HOST TISSUE

宿主组织中细菌生态的代谢方面

• 批准号: 3127354
• 负责人: John K Spitznagel
• 金额: $ 14.33万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-06-01 至 1989-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3127354
• 关键词: Escherichia coli; Neisseria gonorrhoeae; Proteus; Salmonella typhimurium; affinity chromatography; antibacterial agents; antibacterial antibody; bacterial antigens; bacterial genetics; bactericidal immunity; cations; density gradient ultracentrifugation; enzyme linked immunosorbent assay; gram negative bacteria; granule; high performance liquid chromatography; host organism interaction; human tissue; immunochemistry; monoclonal antibody; mutant; neutrophil; nuclear magnetic resonance spectroscopy; phagocytosis; tissue /cell culture; virulence

Human neutrophil polymorphonuclear granulocytes (PMN) are in the first line of cellular defense against infection. They phagocytize bacteria and kill them with antimicrobial activities that are oxygen dependent or oxygen independent. The granules of PMN are intrinsic to these activities. Our goal is to continue to define the proteins that account for antimicrobial activities of crude granule extracts. Our main emphasis will be on anti-Proteus activity and purified cationic antimicrobial proteins (CAP), CAP57 and CAP37 that kill Salmonella and E. coli. We will: define genetic and (tissue) environmental factors contributing to sensitivity or resistance of microorganisms; determine the target molecules CAP interact with in the outer membrane of sensitive bacteria. We will complete studies to localize CAPs at the cellular and subcellular levels. We will use techniques required to understand bacterial pathogenesis and host resistance. We will use: cell culture and differential and gradient centrifugation; phagocytosis; assays for antimicrobial action; protein purification, including conventional and high performance liquid chromatography and affinity chromatography; ELISA; monoclonal antibodies; construction of isogenic strains by P22 transduction; and biochemical analysis of lipopolysaccharides, and outer membrane proteins of Salmonella typhimurium which differ in their resistance to CAP. Our long term objective is to define the role of CAP in oxygen independent antimicrobial phagocytosis and to develop monoclonal antibodies for rapid screening of PMN in clinical infectious disease. The research we propose is significant in that it will: a) establish the existence of at least three highly effective Cationic Antimicrobial Proteins (CAPs) whose mode of action in human PMN is oxygen independent; b) provide biochemical evidence for specific target sites on susceptible bacteria to the action of CAP; c) implicate a contribution of low O2 tension to the resistance to CAPs of N. gonorrhoeae; d) provide highly suggestive evidence for the existence of a heretofore unsuspected antimicrobial granule.

人类中性粒细胞多形核粒细胞（PMN）处于第一线 细胞防御感染的能力。 它们吞噬细菌并杀死 它们具有依赖于氧或氧的抗菌活性 独立的。 PMN 颗粒是这些活动所固有的。 我们的 目标是继续定义发挥抗菌作用的蛋白质 粗颗粒提取物的活性。 我们的重点将是 抗变形杆菌活性和纯化的阳离子抗菌蛋白（CAP）， CAP57 和 CAP37 可杀死沙门氏菌和大肠杆菌。 我们将： 定义遗传 和（组织）环境因素导致敏感性或 微生物的抵抗力；确定CAP相互作用的目标分子 位于敏感细菌的外膜上。 我们将完成学业 在细胞和亚细胞水平上定位 CAP。 我们将使用 了解细菌发病机制和宿主所需的技术 反抗。 我们将使用：细胞培养以及差异和梯度 离心；吞噬作用；抗菌作用测定；蛋白质 纯化，包括常规和高性能液体 色谱法和亲和色谱法；酶联免疫吸附试验；单克隆抗体； 通过P22转导构建等基因菌株；和生化 沙门氏菌脂多糖和外膜蛋白分析 鼠伤寒杆菌对 CAP 的抵抗力不同。 我们的长期目标是确定 CAP 在氧独立中的作用 抗菌吞噬作用并开发单克隆抗体以快速 临床感染性疾病中 PMN 筛查. 我们建议的研究 其重要性在于它将： a) 确定至少存在 三种高效阳离子抗菌蛋白（CAP），其模式 人类 PMN 的作用与氧气无关； b) 提供生化证据 针对对 CAP 作用敏感的细菌的特定靶位点； c) 表明低 O2 张力对 N 的 CAP 抗性有贡献。 淋病； d) 提供高度暗示性的证据来证明某项因素的存在 迄今为止未被怀疑的抗菌颗粒。