Assigning function of unknown genes of Acinetobacter baumanii by cell envelope

这项提议将定义革兰氏阴性细菌功能未知的基因(GUF) 对细胞膜的结构和屏障功能有贡献的鲍曼氏杆菌(Ab)。革兰氏阴性杆菌 细菌包膜是一种有序的大分子复合体，其独特之处在于它由两个 膜。外膜(OM)，一个独特的结构，由不对称的双层组成，包括 由脂多糖(LPS)(外叶)和甘油磷脂(内叶)组成的第一行 通过起渗透屏障的作用来防御防腐剂和抗菌剂。AB有一种新的电池 对抗生素相对不能渗透的包膜和对粘菌素有抵抗力的抗体已经被分离出来，而不能 从粘菌素治疗的患者体内合成内毒素。这些新的性质表明，对抗体的研究 包膜应该揭示临床相关的重要新知识。我们的初步数据并公布了 对AbLPS零突变体的转录图谱的研究导致了生物信息学的分析和选择 O作为参与维护受调节的渗透性屏障的候选的一组AbGUF。 这些GUF将通过构建突变菌株来表征，这些突变菌株将被分析是否发生了改变 对化学探头的渗透性。被膜渗透性改变的菌株将受到敏感性的影响。 抗菌肽和抗生素的检测，磷脂和类脂A的化学分析，以及何时 适当，在疾病的动物模型中进行测试。在定义新GUF的迭代过程中 在未来几年，我们将使用基于FACS的细胞分类的特定基因筛查， 药物和温度敏感筛选，以确定有助于细胞被膜结构和 功能。最后，我们将整合来自Manoil和Harwood博士项目的数据，并利用 布鲁斯博士的技术核心，以定义更多用于表征的GUF。这项提案将考验 调节未被发现的革兰氏阴性包膜结构改变的假说 GUF是病原体抵抗宿主天然免疫、抗菌和消毒剂的组成部分 进攻。

This proposal will define genes of unknown function (GUF) of the Gram-negative bacteria Acinetobacter baumannii (Ab) that contribute to the structure and barrier function of the cell envelope. The Gram-negative bacterial envelope is an ordered complex of macromolecules that is unique in being comprised of two membranes. The outer membrane (OM), a unique structure composed of an asymmetrical bilayer consisting of lipopolysaccharide (LPS) (outer leaflet) and glycerophospholipids (inner leaflet), forms the first line of defense against antiseptics and antimicrobials by functioning as a permeability barrier. Ab has a novel cell envelope that is relatively impermeable to antibiotics, and colistin resistant Ab have been isolated that cannot synthesize LPS from patients treated with colistin. These novel properties suggest that the study of the Ab envelope should reveal important new knowledge of clinical relevance. Our preliminary data and published studies of transcriptional profiles of the Ab LPS null mutant have led to bioinformatics analysis and selection o f a set of Ab GUF that are candidates for participation in maintenance o f a regulated permeability barrier. These GUF will be characterized by construction of mutant strains that will be analyzed for altered permeability to chemical probes. Strains with altered envelope permeability will be subjected to susceptibility testing to antimicrobial peptides and antibiotics, chemical analysis of phospholipids and lipid A, and, when appropriate, testing in animal models of disease. In an iterative process to define new GUF for characterization in further years, we will employ specific genetic screens using FACS-based cell sorting, drug, and temperature sensitivity screens to identify GUF that contribute to cell envelope structure and function. Lastly, we will integrate data from the projects of Drs. Manoil and Harwood and utilize the technological core of Dr. Bruce to define additional GUF for characterization. This proposal will test the hypothesis that regulated undiscovered structural alterations of the Gram-negative envelope encoded by GUF are a component of pathogen defense against host innate immune, antimicrobial, and disinfectant attack.