Proteomic analysis of the T. gondii vacuolar membrane

弓形虫液泡膜的蛋白质组学分析

基本信息

批准号：
7140472
负责人：
ANTHONY P. SINAI
金额：
$ 17.59万
依托单位：
UNIVERSITY OF KENTUCKY
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-07-15 至 2007-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7140472
关键词：
Toxoplasma gondii bioinformatics biotechnology cell membrane host organism interaction immunoprecipitation intracellular parasitism mass spectrometry membrane activity molecular cloning parasite infection mechanism protein protein interaction protein quantitation /detection proteomics vesicle /vacuole

项目摘要

DESCRIPTION (provided by the applicant): Successful parasitism by the obligate intracellular Toxoplasma gondii is critically dependent on efficient nutrient scavenging and subversion of diverse host functions. Access to nutrients and cellular functions within the host cytoplasm is restricted by the parasitophorous vacuole membrane (PVM), a parasite modified structure that serves as the boundary defining the replication permissive niche. Activities at the PVM are critical in nutrient acquisition, structural integrity and the manipulation of host signaling cascades, including potentially the immune response to the parasite. While the PVM is involved in many facets of infection the molecular basis for these activities remains elusive. The inability to purify the PVM has prevented detailed biochemical analyses of these activities. In this proposal we aim to characterize the proteome of the T. gondii PVM using mass spectroscopic approaches coupled with novel polyclonal antisera as affinity reagents. In addition to conventional mass spectroscopic techniques we will apply multi-dimensional protein identification technology (MudPit) which permits the analysis of complex protein mixtures. These approaches to define the PVM proteome are made feasible by recent advances in the mass spectrometric identification of proteins, open access to the T. gondii genome databases and bioinformatic algorithms to analyze the data. The culmination of this approach is to establish the foundation for the characterization of activities in the PVM by defining its protein composition. These approaches will likely lead to the identification of potentially novel chemotherapeutic targets for Toxoplasma and other apicomplexan including Cryptosporidium parvum and hepatic stages of Plasmodia.

描述（由申请人提供）：专性细胞内弓形虫的成功寄生虫贡献在严重取决于有效的营养清除和颠覆各种宿主功能。寄生虫液态膜（PVM）限制了宿主细胞质内营养和细胞功能的访问，这是一种寄生虫修饰的结构，它是定义复制允许易裂的边界。 PVM的活动对于养分获取，结构完整性和操纵宿主信号级联反应至关重要，包括可能对寄生虫的免疫反应。尽管PVM参与了许多感染方面，但这些活动的分子基础仍然难以捉摸。无法纯化PVM的纯粹是对这些活动的详细生化分析。在此提案中，我们旨在使用与新型多克隆抗血清作为亲和力试剂的质谱方法来表征T. gondii PVM的蛋白质组。除了传统的质谱技术外，我们还将应用多维蛋白质识别技术（MUDPIT），该技术允许对复杂蛋白质混合物进行分析。这些定义PVM蛋白质组的方法是通过蛋白质质谱鉴定，对T. gondii基因组数据库的开放访问和生物信息学算法来分析数据的最新进展。这种方法的顶点是通过定义其蛋白质组成来为PVM中活动的表征建立基础。这些方法可能会导致鉴定出潜在的新型化学治疗靶标的弓形虫和其他Apicomplexan，包括疟原虫的隐孢子虫和肝阶段。