GLYCAN BIOSYNTHESIS IN CRYPTOCOCCUS NEOFORMANS

新型隐球菌中的聚糖生物合成

基本信息

批准号：
7580517
负责人：
Tamara L Doering
金额：
$ 31.16万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-06-15 至 2011-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7580517
关键词：
Address Agrobacterium Anabolism Antifungal Agents Automation Award Binding Biochemical Biochemistry Biological Assay Biological Models Biology Biomedical Engineering Budgets Carbohydrates Cell Fractionation Cell Wall Cell surface Cells Cellular Morphology Complex Cryptococcus Cryptococcus neoformans Data Analyses Detection Development Disease Electron Microscopy Encapsulated Enzymes Equilibrium Eukaryota Expenditure Fluorescence Fractionation Funding Galactans Genetic Genetic Screening Genome Glucans Glycobiology Glycolipids Goals Grant Growth Human Image Immunity Immunocompromised Host Immunology Individual Infection Insertional Mutagenesis Investigation Knowledge Lasers Lead Letters Life Methods Microbiology Microscopy Mining Molecular Molecular Analysis Molecular Biology Mutagenesis Mutate North Carolina Opportunistic Infections Organism Pathogenesis Pathway interactions Polymers Polysaccharides Process Prokaryotic Cells Protein Analysis Proteins Proteomics Public Health Publications Research Resources Robotics Role Saints Scanning Schedule Science of genetics Screening procedure Severity of illness Site Structure Text Therapeutic Time Universities Update Virulence Virulence Factors Washington Work Xylose assay development base capsule chemical genetics computer science cost deletion analysis drug discovery experience extracellular factor C fascinate fungus glucuronoxylomannan high throughput screening insight link protein medical schools microbial molecular imaging mutant novel parent project pathogen polypeptide research study xylosyltransferase

项目摘要

The encapsulated fungus Cryptococcus neoformans is responsible for life- threatening disease, particularly in the context of compromised immunity, and current therapy is not adequate. Extracellular glycans define the interface between this opportunistic pathogen and its host; our long-term goal is to determine the biosynthetic pathways of these molecules and discover ways to interfere with them. The main virulence factor of C. neoformans is an extensive polysaccharide capsule composed of two polymers: glucuronoxylomannan (GXM) and glucuronoxylomanno-galactan (GXMGal; previously termed GalXM). We know the structures of these polysaccharides, but major questions remain concerning their synthesis and assembly into the capsule. In the preceding period of this project we purified, cloned, expressed, and mutated a novel xylosyltransferase (Cxt1p), and showed that it acts in synthesis of GXM, GXMGal, and glycolipids; we also discovered a related enzyme (Cxt2p). We further established how capsule is transported to the cell surface and demonstrated that cell wall alpha glucan is required for capsule display. Aim I of the current application is to localize Cxt1p and Cxt2p within the cryptococcal cell and perform targeted experiments on glycan structure and virulence to address their role in cryptococcal biology. Aim II proposes a biochemical approach to identify the machinery of capsule association with the cell surface, followed by phenotypic analysis of strains deficient in candidate proteins. These revised aims can be reasonably performed in two years. We plan creative new strategies to investigate novel features of C. neoformans glycobiology, with emphasis on the capsule. We will capitalize on our prior discoveries to gain greater understanding of cryptococcal biology and pathogenesis, identify targets for anti-fungal drug discovery, and elucidate glycan synthesis in eukaryotes.

封装的真菌Neoformans造成了生命 - 威胁性疾病，特别是在免疫力受损的背景下，当前的治疗不足。细胞外糖定义界面在这种机会性的病原体与其宿主之间；我们的长期目标是确定这些分子的生物合成途径，并发现干扰他们。 Neoformans C. Neoformans的主要毒力因子是广泛的由两个聚合物组成的多糖胶囊：葡萄糖量氧基瘤（GXM）和葡萄糖氧基氧基 - 甘丹（GXMGAL；先前称为GalXM）。我们知道这些多糖的结构，但主要问题仍然存在关于它们的合成和组装到胶囊中。在上述这个项目的时期我们纯化，克隆，表达和突变木糖基转移酶（CXT1P），表明它作用于GXM的合成， GXMGAL和糖脂；我们还发现了相关的酶（CXT2P）。我们进一步确定了如何将胶囊传输到细胞表面和证明胶囊显示需要细胞壁alpha glucan。目的我当前的应用程序是将CXT1P和CXT2P定位并对聚糖结构和毒力进行有针对性的实验以解决它们在隐球菌生物学中的作用。 AIM II提出了一种生化方法识别胶囊与细胞表面关联的机械，然后是缺乏候选蛋白质的菌株的表型分析。这些修订的目标可以在两年内合理地进行。我们计划创新的新策略研究新生梭菌糖生物学的新颖特征，重点是胶囊。我们将利用我们先前的发现以获得更大的了解隐球菌生物学和发病机理，识别抗真菌药物的靶标发现并阐明真核生物中的聚糖合成。