Specificity of Plasmodium falciparum protein export

恶性疟原虫蛋白输出的特异性

• 批准号: 10632093
• 负责人: Daniel E. Goldberg
• 金额: $ 19.46万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10632093
• 关键词: Address; Adherence; Africa South of the Sahara; Alanine; Aspartic Endopeptidases; Biochemical; Biotinylation; Brefeldin A; Cells; Child; Development; Disease; Elements; Erythrocytes; Future; Immune response; Immunoprecipitation; Impairment; Interruption; Investigation; Malaria; Mass Spectrum Analysis; Mediating; Modeling; Molecular; Molecular Chaperones; Molecular Conformation; Mutation; Nutrient; Parasites; Pathogenesis; Pathway interactions; Persons; Phenotype; Plasmodium falciparum; Process; Protein Export Pathway; Proteins; Proteomics; Reporter; Specificity; Streptavidin; System; Vacuole; Vascular Endothelium; Work; biophysical properties; comparative; experimental study; insight; knock-down; mutant; novel therapeutics; plasmepsin; prevent; reverse genetics; secretory protein; therapeutic target; uptake

The malaria parasite Plasmodium falciparum exports several hundred proteins into its host cell to modulate nutrient uptake, infected RBC biophysical properties, adherence to vascular endothelium and host immune response. These exported protein effectors are synthesized in the parasite ER, traverse the secretory system and are then translocated from the parasitophorous vacuole (PV) into the host RBC. Most have a protein export element (PEXEL) that is essential for the export process but is cleaved co-translationally by the aspartic protease plasmepsin V. How the processed protein is recognized for export is not at all clear. Certain mutations in the mature N-terminus of PEXEL-containing reporter constructs can prevent export, but changing the mature N-terminus to all alanines does not impair export. We hypothesize that export is the default pathway for secretory proteins and that specific mutations block export or promote PV retention. We posit that these mutations dictate recognition by a chaperone. To establish this model, we must identify the chaperone(s) that confer specificity. To address these questions, Aim 1 will focus on the protein machinery that mediates export specificity. Export-destined and PV retained proteins will be pulled down from the PV and from the ER. Comparative proteomic analysis will be employed to identify key components of the export pathway. Proximity biotinylation will comprise a second approach to this analysis. Candidate interacting proteins will be validated in reciprocal pulldowns. This will pave the way for future reverse genetic and biochemical assessments. We anticipate that the proposed studies will yield great insight into the mechanism of protein export that is so important to the pathogenesis of malaria. We hope that this work will point the way to new therapies for this devastating disease.

疟疾寄生虫恶性疟原虫出口数百个 蛋白质进入其宿主细胞以调节养分吸收，感染的RBC生物物理 特性，遵守血管内皮和宿主免疫反应。这些 导出的蛋白质效应子在寄生虫ER中合成，遍历 分泌系统，然后从寄生虫液泡（PV）中易位 进入主机RBC。大多数都有必不可少的蛋白质导出元件（PEXEL） 对于出口过程，但由天冬氨酸蛋白酶共裂解 纤溶酶V.如何识别出导出的加工蛋白如何清楚。 含Pexel的记者成熟的N末端的某些突变 结构可以防止出口，但将成熟的N末端更改为所有丙氨酸 不会损害出口。我们假设导出是 分泌蛋白和特定突变阻止出口或促进PV保留。 我们认为这些突变决定了伴侣的识别。建立这个 模型，我们必须确定赋予特异性的伴侣。 为了解决这些问题，AIM 1将重点放在蛋白质机械上 介导出口特异性。将拉出出口目录和光伏保留的蛋白质 从光伏和急诊室下降。比较蛋白质组学分析将是 用于识别出口途径的关键组件。接近生物素化 将构成该分析的第二种方法。候选蛋白质相互作用 将在相互倾诉中进行验证。这将为将来的逆转铺平道路 遗传和生化评估。我们预计拟议的研究 将对蛋白质出口机理的深入了解，这对 疟疾的发病机理。我们希望这项工作将指向新的道路 这种毁灭性疾病的疗法。