Division of Infectious Diseases, Department of Medicine, and Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, MO, USA.
Nat Commun. 2022 Aug 4;13(1):4537. doi: 10.1038/s41467-022-32271-7.
The malaria parasite Plasmodium invades a host erythrocyte, multiplies within a parasitophorous vacuole (PV) and then ruptures the PV and erythrocyte membranes in a process known as egress. Both egress and invasion are controlled by effector proteins discharged from specialized secretory organelles. The aspartic protease plasmepsin X (PM X) regulates activity for many of these effectors, but it is unclear how PM X accesses its diverse substrates that reside in different organelles. PM X also autoprocesses to generate different isoforms. The function of this processing is not understood. We have mapped the self-cleavage sites and have constructed parasites with cleavage site mutations. Surprisingly, a quadruple mutant that remains full-length retains in vitro activity, is trafficked normally, and supports normal egress, invasion and parasite growth. The N-terminal half of the prodomain stays bound to the catalytic domain even after processing and is required for proper intracellular trafficking of PM X. We find that this enzyme cleaves microneme and exoneme substrates before discharge, while the rhoptry substrates that are dependent on PM X activity are cleaved after exoneme discharge into the PV. The data give insight into the temporal, spatial and biochemical control of this unusual but important aspartic protease.
疟原虫 Plasmodium 入侵宿主红细胞，在一个寄生空泡（PV）内繁殖，然后破裂 PV 和红细胞膜，这个过程称为出芽。出芽和入侵都受从专门的分泌细胞器释放的效应蛋白控制。天冬氨酸蛋白酶 plasmepsin X（PM X）调节许多这些效应物的活性，但 PM X 如何访问其位于不同细胞器中的不同底物尚不清楚。PM X 还自我切割以产生不同的同工型。这种处理的功能尚不清楚。我们已经绘制了自我切割位点，并构建了具有切割位点突变的寄生虫。令人惊讶的是，一个保持全长的四重突变体在体外仍具有活性，正常运输，并支持正常的出芽、入侵和寄生虫生长。前肽的 N 端半部分在加工后仍与催化结构域结合，这对于 PM X 的正确细胞内运输是必需的。我们发现这种酶在排出前切割微线体和外显子底物，而依赖 PM X 活性的 rhoptry 底物在排出外显子进入 PV 后被切割。这些数据为这种不寻常但重要的天冬氨酸蛋白酶提供了对其时间、空间和生化控制的深入了解。
