A cyanobacterial serine protease of Plasmodium falciparum is targeted to the apicoplast and plays an important role in its growth and development
A cyanobacterial serine protease of Plasmodium falciparum is targeted to the apicoplast and plays an important role in its growth and development
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DOI：
10.1111/j.1365-2958.2010.07251.x
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发表时间：
2010-08
影响因子：
3.6
通讯作者：
S. Rathore;D. Sinha;M. Asad;T. Böttcher;F. Afrin;Virander S. Chauhan;D. Gupta;S. Sieber;A. Mohmme
中科院分区：
文献类型：
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作者：
S. Rathore;D. Sinha;M. Asad;T. Böttcher;F. Afrin;Virander S. Chauhan;D. Gupta;S. Sieber;A. Mohmme
原核生物依赖于ATP的蛋白酶机制，如疟原虫中的ClpQY和ClpAP，可能是潜在的药物靶点。在本研究中，我们发现恶性疟原虫蓝细菌ClpP蛋白(PfClpP)的同源基因(PfClpP)在无性血阶段表达，并具有丝氨酸蛋白酶活性。PfClpP通过绿色荧光蛋白靶向、免疫电子显微镜和免疫荧光方法定位于质外体。用PfClpP的体外蛋白水解酶活性测定法筛选出一组能与原核生物β活性部位特异结合的细胞通透性ClpP-内酯。在筛选中鉴定出一种PfClpP特异性的蛋白水解酶抑制物，标记为U1-内酯。U1-内酯处理显著抑制了无性期寄生虫的体外生长。U1处理的寄生虫在裂殖体晚期表现出发育停滞。我们进一步表明，U1-内酯处理导致了不能在寄生虫后代中生长和分离的异常质外体的形成；这些影响也明显地表现在对质外体基因组的复制的阻断上。总体而言，我们的数据表明，PfClpP蛋白水解酶已经在质外体中定位，它在功能质外体的发育中起着重要的作用。
The prokaryotic ATP‐dependent protease machineries such as ClpQY and ClpAP in the malaria parasite may represent potential drug targets. In the present study, we show that the orthologue of cyanobacterial ClpP protease in Plasmodium falciparum (PfClpP) is expressed in the asexual blood stages and possesses serine protease activity. The PfClpP was localized in the apicoplast using a GFP‐targeting approach, immunoelectron microscopy and by immunofluorescence assays. A set of cell permeable β‐lactones, which specifically bind with the active site of prokaryotic ClpP, were screened using an in vitro protease assay of PfClpP. A PfClpP‐specific protease inhibitor was identified in the screen, labelled as U1‐lactone. In vitro growth of the asexual stage parasites was significantly inhibited by U1‐lactone treatment. The U1‐treated parasites showed developmental arrest at the late‐schizont stage. We further show that the U1‐lactone treatment resulted in formation of abnormal apicoplasts which were not able to grow and segregate in the parasite progeny; these effects were also evident by blockage in the replication of the apicoplast genome. Overall, our data show that the PfClpP protease has confirmed localization in the apicoplast and it plays important role in development of functional apicoplasts.