A chemical proteomics survey of Plasmodium gametocyte development

摘要 疟疾寄生虫通过按蚊媒介的传播维持了高水平的再感染。 这是疟疾流行地区的一个严重问题，严重阻碍了疟疾控制。因此，消除疟疾的努力必须包括 防止寄生虫传播的策略。因为只有寄生虫的无性血液阶段才会引起 疾病，它们是大多数抗疟疾药物的主要靶点。相比之下，负责的发育阶段 对于配子体的传播，人们知之甚少，而且大多数有效治疗无性生殖的药物 血液阶段对配子体无效。在这里，我们将直接测量和识别一个 恶性疟原虫天然蛋白质组中的丝氨酸水解酶家族 (Pf)裂殖体和配子体阶段。我们将使用基于活性的蛋白质图谱(ABPP)和丝氨酸 水解酶活性探针法与定量质谱仪联用研究 从无性向有性转变的过程中酶活性的一部分，蚊子可传播。 由于ABPP是一种基于机制和顺序独立的方法，这些研究也将捕获 功能未知的Pf蛋白的活性。我们将进一步鉴定命中的丝氨酸蛋白酶， 并定义已知和以前未知的蛋白酶的生理底物。定义SH活动 与无性向有性性转变相关的变化将为PF配子体提供新的和广泛的 这一发展转型的基于活动的观点，确定潜在的传播阻断目标，以及 为大量未知功能的蛋白质提供实验注释。

Abstract The spread of malaria parasites by the Anopheles mosquito vector sustains high levels of reinfection in endemic regions and severely impedes malaria control. As a result, efforts to eliminate malaria must include strategies that prevent parasite transmission. Because only the asexual blood stages of the parasite cause disease, they are the main target of most antimalarials. In contrast, the developmental stage responsible for transmission, the gametocyte, is only poorly understood, and most drugs that are effective against the asexual blood stages are ineffective against gametocytes. Here, we will directly measure and identify the activity of a particularly druggable enzyme family, the serine hydrolases, in the native proteome of Plasmodium falciparum (Pf) schizont and gametocyte stages. We will use activity-based protein profiling (ABPP) with a serine hydrolase-directed activity probe in combination with quantitative mass spectrometry to survey a broad cross- section of enzyme activity during the transition from the asexual to the sexual, mosquito-transmissible form. Because ABPP is a mechanism-based and sequence-independent approach, these studies will also capture the activity of Pf proteins with unknown function. We will further identify the serine proteases among the hits, and define the physiologic substrates of known and previously unknown proteases. Defining the SH activity changes associated with the transition from asexual to sexual Pf gametocytes will provide a new and broad activity-based view of this developmental transition, identify potential transmission blocking targets, and provide experimental annotation for a large number of proteins with unknown function.