Shu Fanfan, Li Yu, Chu Wenlun, Chen Xuehua, Zhang Ziding, Guo Yaqiong, Feng Yaoyu, Xiao Lihua, Li Na
Center for Emerging and Zoonotic Diseases, College of Veterinary Medicine, South China Agricultural University, Guangzhou, China.
State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing, China.
Front Microbiol. 2022 Apr 25;13:883674. doi: 10.3389/fmicb.2022.883674. eCollection 2022.
Calcium-dependent protein kinases (CDPKs) are important in calcium influx, triggering several biological processes in spp. As they are not present in mammals, CDPKs are considered promising drug targets. Recent studies have characterized CDPK1, CDPK3, CDPK4, CDPK5, CDPK6, and CDPK9, but the role of CPK2A remains unclear. In this work, we expressed recombinant CDPK2A encoded by the cgd2_1060 gene in and characterized the biologic functions of CDPK2A using qRT-PCR, immunofluorescence microscopy, immuno-electron microscopy, and neutralization. The results revealed that CDPK2A protein was highly expressed in the apical region of sporozoites and merozoites and in macrogamonts. Monoclonal or polyclonal antibodies against CDPK2A failed to block the invasion of host cells. Among the 44 candidate inhibitors from molecular docking of CDPK2A, one inhibitor was identified as having a potential effect on both growth and CDPK2A enzyme activities. These data suggest that CDPK2A may play some roles during the development of and might be a potential drug target against cryptosporidiosis.
钙依赖性蛋白激酶(CDPKs)在钙内流中起重要作用,触发了隐孢子虫属物种的多种生物学过程。由于它们在哺乳动物中不存在,CDPKs被认为是有前景的药物靶点。最近的研究已经对CDPK1、CDPK3、CDPK4、CDPK5、CDPK6和CDPK9进行了表征,但CPK2A的作用仍不清楚。在这项工作中,我们在毕赤酵母中表达了由cgd2_1060基因编码的重组CDPK2A,并使用qRT-PCR、免疫荧光显微镜、免疫电子显微镜和中和实验对CDPK2A的生物学功能进行了表征。结果显示,CDPK2A蛋白在子孢子和裂殖子的顶端区域以及大配子体中高度表达。抗CDPK2A的单克隆或多克隆抗体未能阻断宿主细胞的入侵。在来自CDPK2A分子对接的44种候选抑制剂中,一种抑制剂被确定对隐孢子虫生长和CDPK2A酶活性均有潜在影响。这些数据表明,CDPK2A可能在隐孢子虫发育过程中发挥某些作用,并且可能是抗隐孢子虫病的潜在药物靶点。