Specificity of Plasmodium falciparum protein export

疟疾寄生虫恶性疟原虫出口数百 蛋白质进入宿主细胞调节营养吸收，感染红细胞生物物理 性质、对血管内皮细胞的黏附和宿主免疫反应。这些 输出的蛋白质效应器在寄生虫的内质网中合成，穿过 分泌系统，然后从寄生性液泡(PV)转移。 进入宿主红细胞。大多数都含有蛋白质输出元件(PEXEL)，这是必不可少的 用于出口过程，但被天冬氨酸蛋白酶共翻译切割 纤溶酶V。加工后的蛋白质是如何被识别出口的，目前还完全不清楚。 含PEXEL报告基因成熟N端的某些突变 构造体可以阻止输出，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 分泌蛋白和特定的突变阻止或促进光伏病毒的保留。 我们假设，这些突变决定了伴侣的识别。要确立这一点， 模型，我们必须确定赋予特异性的监护人(S)。 为了解决这些问题，目标1将重点放在蛋白质机制上 调解出口专用性。出口和光伏保留的蛋白质将被拉出 从PV和急诊室下来。比较蛋白质组学分析将是 用来确定出口途径的关键组成部分。邻位生物素化 将包括对这种分析的第二种方法。候选相互作用蛋白 将在互惠的下拉列表中进行验证。这将为未来的逆转铺平道路 遗传和生化评估。我们预期拟议的研究 将对蛋白质输出的机制有很大的洞察力，这对 疟疾的发病机制。我们希望这项工作将为新的 治疗这种毁灭性疾病的方法。

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.