Role of Proteases in Cryptosporidium-Host Cell Interactions

蛋白酶在隐孢子虫-宿主细胞相互作用中的作用

• 批准号: 7878271
• 负责人: Honorine D Ward
• 金额: $ 1.91万
• 依托单位: TUFTS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-15 至 2009-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7878271
• 关键词: Acquired Immunodeficiency Syndrome; Antibodies; Apical; CD7 gene; Cell Communication; Cells; Cleaved cell; Cryptosporidiosis; Cryptosporidium; Cryptosporidium parvum; Development; Disease; Enzyme Inhibitor Drugs; Enzyme Inhibitors; Enzymes; Epithelial Cells; Essential Genes; Future; Genes; Genome; Glycopeptides; Glycoproteins; Goals; Immunocompromised Host; In Vitro; Infection; Intestines; Investigation; Mediating; Modality; Mucins; Parasites; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Plasmodium falciparum; Play; Process; Proprotein Convertases; Proteins; Proteolytic Processing; Recombinants; Role; Serine Protease; Staging; Structure; Subtilisins; Surface; System; Testing; Toxoplasma gondii; base; cell motility; design; drug development; effective therapy; in vivo; inhibitor/antagonist; mRNA Expression; nitazoxanide; novel; prevent; protein complex

The apicomplexan parasite Cryptosporidium is a significant cause of diarrheal disease worldwide, particularly in immunocompromised hosts such as AIDS patients. The overall goal of this project is to investigate the role of subtilisin-like serine proteases (subtilases) of C. parvum in host-parasite interactions. Two genes encoding putative subtilases (designated CpSUB1 and CpSUB2) have been identified in the C. parvum genome but have not yet been investigated. Preliminary studies show that both genes are expressed during C. parvum infection in vitro suggesting that the proteins they encode are likely to be important in host-parasite interactions. The hypothesis is that CpSUB1 and/or CpSUB2 process surface and apical complex proteins such as gp40/15 and mediate C. parvum infection in vitro. In the first specific aim we will quantify mRNA expression of CpSUB1 and 2, identify the precursor, processed and mature forms of the CpSUB1 and 2 proteins expressed in C. parvum, investigate their post-translational processing and determine their subcellular localization. In addition, we will express enzymatically active recombinant CpSUB1 and CpSUB2 and characterize their enzymatic activity. In the second specific aim we will determine whether either subtilase processes recombinant gp40/15. In addition we will determine whether propeptide inhibitors or antibodies to the CpSUBs inhibit C. parvum infection of intestinal epithelial cells in vitro. The long term goal is the rational, structure-based design of inhibitors of these enzymes as drugs to prevent or treat cryptosporidiosis. Project Narrative/Relevance The intestinal parasite Cryptosporidium is a significant cause of diarrheal disease worldwide, particularly in immunocompromised hosts such as AIDS patients. In this project we will study enzymes that are important for infection of host cells by the parasite, with the long term goal of developing inhibitors of these enzymes as drugs for cryptosporidiosis. Since there is no consistently effective treatment available for this disease, these studies are important for development of new drugs.

Apicomplexan寄生虫隐孢子虫是全球腹泻疾病的重要原因， 特别是在免疫功能低下的宿主中，例如艾滋病患者。该项目的总体目标是 研究白梭菌在宿主 - 寄生虫中的枯草素样丝氨酸蛋白酶（枯草酶）的作用 互动。编码假定枯草酶（指定的cpsub1和cpsub2）的两个基因已 在C. parvum基因组中鉴定，但尚未进行研究。初步研究表明 两种基因在体外表达期间表达在体外感染期间，表明它们编码的蛋白质 在宿主 - 寄生虫相互作用中可能很重要。假设是cpsub1和/或cpsub2 过程表面和顶端复合物蛋白（例如GP40/15）在体外介导了圆孢菌感染。 在第一个特定目的中，我们将量化CPSUB1和2的mRNA表达，确定前体， CPSUB1和2蛋白的加工和成熟形式在C. parvum中表达，研究它们的 翻译后处理并确定其亚细胞定位。此外，我们将表达 酶促活性重组CPSUB1和CPSUB2，并表征其酶活性。在 第二个特定目的我们将确定任何一种牙酶过程是否重组GP40/15。在 此外，我们还将确定丙肽抑制剂或对CPSUB的抗体是否抑制parvum C. parvum 体外感染肠上皮细胞。长期目标是理性的，基于结构的设计 这些酶的抑制剂是预防或治疗隐孢子虫病的药物。项目叙述/相关性 肠道寄生虫隐孢子虫是腹泻病的重要原因 全球，特别是在艾滋病患者等免疫功能低下的宿主中。在这个 项目我们将研究对宿主细胞感染至关重要的酶 寄生虫，长期目标是将这些酶的抑制剂作为药物的抑制剂 隐孢子虫病。由于没有一贯有效的治疗 疾病，这些研究对于开发新药很重要。