Outer Membrane Proteins of Pathogenic Leptospira Species

致病性钩端螺旋体物种的外膜蛋白

• 批准号: 8039966
• 负责人: DAVID A HAAKE
• 金额: $ 29.83万
• 依托单位: BRENTWOOD BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-05-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8039966
• 关键词: Adherence; Adhesions; Adhesives; Amino Acids; Animals; Antibodies; Antigens; Area; Bacterial Adhesins; Binding; Binding Proteins; Bioinformatics; Biological Assay; Cell Fractionation; Culture Media; Data; Development; Disease; Environment; Expression Library; Family; Fibrinogen; Fibronectins; Fractionation; Genes; Goals; Growth; Hamsters; Health; Hemorrhage; Histocytochemistry; Human; Immune response; Immunity; Immunization; Immunoglobulins; Infection; Kidney Failure; Laboratories; Laminin; Lead; Leptospira; Leptospirosis; Life; Lipopolysaccharides; Lipoproteins; Liver Dysfunction; Lung; Mediating; Membrane; Membrane Proteins; Modeling; Molecular; O Antigens; Order Spirochaetales; Osmolar Concentration; Pathogenesis; Patients; Peripheral; Phenotype; Physiological; Play; Potassium Chloride; Prevention strategy; Process; Proteins; Rattus; Recombinants; Regulation; Research; Research Design; Reverse Transcriptase Polymerase Chain Reaction; Role; Screening procedure; Serum; Signal Transduction; Sodium Chloride; Sphingomyelinase; Structure; Sucrose; Surface; Tertiary Protein Structure; Tissues; Urbanization; Virulent; bactericide; induced pluripotent stem cell; novel; novel vaccines; porin; promoter; protein protein interaction; research study; sodium sulfate

DESCRIPTION (provided by applicant): Leptospirosis is a common zoonotic disease, typically transmitted to humans by rat exposure. Leptospiral infection in humans frequently results in fulminant, life-threatening illness characterized by liver dysfunction, kidney failure, and pulmonary hemorrhage. Leptospirosis is emerging in areas of the world undergoing rapid urbanization. Research is urgently needed to understand how these pathogenic spirochetes interact with their mammalian hosts. The focus of this proposal is to elucidate the molecular mechanisms of pathogenesis and immunity mediated by outer membrane proteins (OMPs). Our laboratory, a leader in this field, has developed strategies for membrane fractionation and identification of surface-exposed OMPs. We have identified a family of leptospiral immunoglobulin-like repeat (Lig) proteins that are targeted by the human immune response to leptospirosis. When virulent leptospires are exposed to host-physiologic levels of osmolarity (-300 mOsm/liter), expression of LigA and LigB is dramatically increased, while LipL36 and IPS expression are downregulated, changes known to be associated with infection of the mammalian host. Regulation by osmolarity indicates that Lig expression occurs early in the infectious process. We have recently shown that the LigA and LigB are fibronectin- and fibrinogen-binding proteins, indicating that these proteins are involved in host tissue colonization. In both LigA and LigB, the fibronectin- and fibrinogen-binding domains overlap, indicating a novel mechanism of protein-protein interaction. Specific Aim 1 describes studies designed to determine the role of Lig proteins in leptospiral binding to adhesive matrix molecules including fibronectin, fibrinogen, and laminin. We will examine the hypothesis that LigA and LigB are promiscuous adhesins able to bind multiple adhesive matrix molecules using a single adhesion domain. The goal of Specific Aim 2 is to identify osmo-upregulated OMPs exposed on the leptospiral surface during infection using immuno-histochemistry, reverse-transcriptase PCR, membrane fractionation, and surface-exposure analysis. Specific Aim 3 will determine which osmo-upregulated OMPs are protective immunogens in the hamster model of leptospirosis. We will examine the mechanism(s) by which antibodies to osmo-upregulated OMPs mediate protection using assays of bactericidal activity, growth inhibition, opsonophagocytosis, and adherence inhibition.

描述（由申请人提供）：钩端螺旋体病是一种常见的人畜共患病，通常通过大鼠暴露传播给人类。人类的钩端螺旋体感染通常会导致暴发性，威胁生命的疾病，其特征是肝功能障碍，肾衰竭和肺出血。在世界快速城市化的世界地区，钩端螺旋体病正在出现。迫切需要研究这些病原螺旋体如何与哺乳动物宿主相互作用。该建议的重点是阐明由外膜蛋白（OMP）介导的发病机理和免疫的分子机制。 我们的实验室是该领域的领导者，已经制定了膜分馏和鉴定表面暴露OMP的策略。我们已经确定了一个由人类免疫反应靶向钩端螺旋体病的靶向诱导性免疫球蛋白样重复蛋白（LIG）蛋白的家族。当有毒的钩端螺旋会暴露于渗透压的宿主 - 生理水平（-300 MOSM/L）时，LIGA和LIGB的表达大大增加，而LIPL36和IPS表达下调，已知与哺乳动物宿主感染相关的变化。渗透压调节表明LIG表达发生在传染过程的早期。我们最近表明，LIGA和LIGB是纤连蛋白质和纤维蛋白原结合蛋白，表明这些蛋白质参与宿主组织定植。在LIGA和LIGB中，纤连蛋白原和纤维蛋白原结合结构域重叠，表明蛋白质 - 蛋白质相互作用的新机制。 具体目标1描述了旨在确定LIG蛋白在钩端螺旋中与粘合剂基质分子（包括纤连蛋白，纤维蛋白原和层粘连蛋白）中的作用的研究。我们将研究LIGA和LIGB的假设是混杂的粘附蛋白能够使用单个粘附域结合多个粘附基质分子。特定目标2的目的是使用免疫 - 归化学，反向转移酶PCR，膜分馏和表面暴露分析在感染过程中识别在感染过程中暴露于钩端螺旋表面上的OSMO升级的OMP。具体目标3将确定哪些OSMO升级的OMP是钩端螺旋体病模型中的保护性免疫原子。我们将使用杀菌活性，生长抑制作用，调查性吞噬作用和粘附抑制的测定方法来研究OSMO升级OMPS抗体介导的保护的机制。