Li Weichao, Zhou Yiqing, Tang Guanghui, Xiao Youli
CAS Key Laboratory of Synthetic Biology, CAS Center for Excellence in Molecular Plant Sciences, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences , Shanghai 200032, China.
University of Chinese Academy of Sciences , Beijing 100039, China.
Bioconjug Chem. 2016 Dec 21;27(12):2828-2833. doi: 10.1021/acs.bioconjchem.6b00556. Epub 2016 Nov 18.
Despite the fact that multiple artemisinin-alkylated proteins in Plasmodium falciparum have been identified in recent studies, the alkylation mechanism and accurate binding site of artemisinin-protein interaction have remained elusive. Here, we report the chemical-probe-based enrichment of the artemisinin-binding peptide and characterization of the artemisinin-binding site of P. falciparum translationally controlled tumor protein (TCTP). A peptide fragment within the N-terminal region of TCTP was enriched and found to be alkylated by an artemisinin-derived probe. MS2 fragments showed that artemisinin could alkylate multiple amino acids from Phe12 to Tyr22 of TCTP, which was supported by labeling experiments upon site-directed mutagenesis and computational modeling studies. Taken together, the "capture-and-release" strategy affords consolidated advantages previously unavailable in artemisinin-protein binding site studies, and our results deepened the understanding of the mechanism of protein alkylation via heme-activated artemisinin.
尽管最近的研究已经在恶性疟原虫中鉴定出多种青蒿素烷基化蛋白，但青蒿素与蛋白质相互作用的烷基化机制和精确结合位点仍不清楚。在此，我们报告了基于化学探针富集青蒿素结合肽以及对恶性疟原虫翻译调控肿瘤蛋白（TCTP）的青蒿素结合位点的表征。TCTP N端区域内的一个肽片段被富集，并发现被青蒿素衍生探针烷基化。MS2片段显示青蒿素可烷基化TCTP从Phe12到Tyr22的多个氨基酸，定点诱变后的标记实验和计算建模研究支持了这一点。综上所述，“捕获-释放”策略提供了青蒿素-蛋白质结合位点研究中以前没有的综合优势，我们的结果加深了对血红素激活的青蒿素导致蛋白质烷基化机制的理解。
