Novel Class of Phospholipases-Molecular Pathogenesis

新一类磷脂酶-分子发病机制

• 批准号: 6726336
• 负责人: Michael L. Vasil
• 金额: $ 44.89万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-06-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6726336
• 关键词: Pseudomonas aeruginosa; betaine compound; cystic fibrosis; cytolysis; enzyme activity; fusion gene; gene mutation; host organism interaction; immunologic receptors; laboratory mouse; laboratory rabbit; molecular cloning; molecular pathology; monoclonal antibody; opportunistic infections; osmotic pressure; phosphatidylcholines; phospholipase C; phosphotransferases; posttranslational modifications; protein structure function; pulmonary surfactants; respiratory infections; site directed mutagenesis; sphingomyelins; virulence

DESCRIPTION (provided by applicant): The diversity of roles that phospholipases C (PLCs) play in biology and medicine is extraordinary. In the past decade this class of phospholipid hydrolyzing enzymes has been shown to be considerably more complex than initially perceived and their impact on a wide range of basic cellular processes in eukaryotes, including oncogenesis, apoptosis, and inflammation has been increasingly appreciated. Likewise, there are many sundry and important functions for PLCs in microbial pathogenesis. We identified and characterized the first member of a novel class of homologous PLCs, the hemolytic phospholipase C (PIcH) of Pseudomonas aeruginosa. Members of this class of PLCs are produced by an array of opportunistic and frank pathogens, including potential bioterrorist agents. The genomes of some of these organisms encode as many as 4 homologs of this class of PLCs. Bacteria carrying genes encoding these PLCs (gene copy number shown in parentheses) include: P. aeruginosa (2), Mycobacterium tuberculosis (4), Francisella tularensis (1), Burkholderia pseudomallei and mallei (3 each) and Bordetella pertussis (1). We, as well as others, provided cogent evidence that members of this novel class of PLCs play significant and diverse roles in the infectious diseases caused by those agents. Although these PLCs share considerable amino acid homology, each member has distinct properties. There are some important differences in their substrate specificities, and many members have unique structural features that probably play a specific functional role in the pathogenesis of the organisms that produce them. This application will mainly focus on the paradigm of this novel class of PLCs (PIcH). In addition to its PLC activity, PIcH is the first prokaryotic or eukaryotic protein yet identified that has Sphingomyelin Synthase activity. The substrates (e.g. phosphatidylcholine & sphingomyelin) of PIcH or the products (e.g. diacylglycerol, ceramide or sphingomyelin) that it generates could have profound biological effects, particularly with respect to signaling processes in eukaryotic cells and the host responses to this infectious agent. We have also provided evidence that PIcH is highly cytotoxic for endothelial cells and probably enters these cells through interaction with integrin receptors. This research project will employ microbiological, genetic, biochemical, structural and cell biology methods to examine how PIcH and other members of this novel class of enzymes affect the virulence of the organisms that produce them. Furthermore, it is likely that information we derive from our efforts will also provide additional insights about the biochemistry and biology of PLCs in general.

描述（由申请人提供）：磷脂酶C（PLC）在生物学和医学中发挥的作用的多样性是非凡的。在过去的十年中，这类磷脂水解酶已被证明是相当复杂的，比最初认为的和它们的影响范围广泛的基本细胞过程中的真核生物，包括肿瘤发生，凋亡和炎症已越来越多地赞赏。同样，PLC在微生物致病机制中也有许多重要的功能。我们确定并表征了一类新型同源PLC的第一个成员，铜绿假单胞菌的溶血性磷脂酶C（PIcH）。这类PLC的成员是由一系列机会主义和坦率的病原体产生的，包括潜在的生物恐怖分子。这些生物体中的一些的基因组编码多达4个这类PLC的同源物。携带编码这些PLC的基因（基因拷贝数显示在括号中）的细菌包括：铜绿假单胞菌（2）、结核分枝杆菌（4）、土拉热弗朗西斯菌（1）、类鼻疽伯克霍尔德菌和鼻疽伯克霍尔德菌（各3）和百日咳博德特氏菌（1）。我们以及其他人提供了令人信服的证据，证明这类新型PLC的成员在这些病原体引起的传染病中发挥着重要而多样的作用。虽然这些PLC共享相当大的氨基酸同源性，但每个成员具有不同的特性。它们的底物特异性存在一些重要差异，许多成员具有独特的结构特征，可能在产生它们的生物体的发病机制中发挥特定的功能作用。本申请将主要集中在这类新型PLC（PIcH）的范例。除了其PLC活性，PIcH是第一个原核或真核蛋白尚未确定的具有鞘磷脂合酶活性。PIcH的底物（例如磷脂酰胆碱和鞘磷脂）或其产生的产物（例如二酰基甘油、神经酰胺或鞘磷脂）可能具有深远的生物学效应，特别是关于真核细胞中的信号传导过程和宿主对这种感染因子的反应。我们还提供了PIcH对内皮细胞具有高度细胞毒性的证据，并可能通过与整合素受体相互作用进入这些细胞。该研究项目将采用微生物学，遗传学，生物化学，结构和细胞生物学方法来研究PIcH和这类新型酶的其他成员如何影响产生它们的生物体的毒力。此外，我们从努力中获得的信息也可能提供有关PLC生物化学和生物学的更多见解。