Chemical proteomic profiling with photoaffinity labeling strategy identifies antimalarial targets of artemisinin

Gao, Peng; Chen, Jiayun; Sun, Peng; Wang, Jianyou; Tang, Huan; Xia, Fei; Gu, Liwei; Zhang, Huimin; Wang, Chen; Wong, Yin Kwan; Zhu, Yinhua; Xu, Chengchao; Wang, Jigang

doi:10.1016/j.cclet.2023.108296

Cited by 4 publications

(9 citation statements)

References 27 publications

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“…The parasite strains of P. falciparum 3D7 and Dd2 were obtained from the Artemisinin Research Center of the China Academy of Chinese Medical Sciences. The parasite culture was the same as previously described with minor modifications [ 16 , 17 ]. Briefly, the parasites were cultured in 50 mL malaria complete medium containing 10.4 g/L RPMI1640, 0.5% albumin, 0.2 g/L gentamycin, 25 μg/mL hypoxanthine, 0.3 g/L L-glutamine, 25 mmol/L HEPES, 2.5 g/L NaHCO 3 with 2% hematocrit at 37 °C.…”

Section: Methodsmentioning

confidence: 99%

“…To identify the potential antimalarial target proteins of Cel, the pull-down experiments were performed [ 17 ]. The method of collecting parasite proteins after treatment is the same as the in situ fluorescence labeling.…”

Section: Methodsmentioning

confidence: 99%

“…The structure of Cel was downloaded from PubChem (CID:122724), the protein 3D structure of Pf Spdsyn from PDB (2PT9) and Pf EGF1-α from Alphafold (Q8I0P6). The docking simulation was performed by MOE (2019.0102) [ 17 ]. Cysteine in the protein was selected as a potential site and the default method of the software was used for docking.…”

Section: Bio-layer Interferometry (Bli) Assaymentioning

confidence: 99%

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Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

Gao,

Wang,

Tang

et al. 2024

Cell Commun Signal

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Background Malaria remains a global health burden, and the emergence and increasing spread of drug resistance to current antimalarials poses a major challenge to malaria control. There is an urgent need to find new drugs or strategies to alleviate this predicament. Celastrol (Cel) is an extensively studied natural bioactive compound that has shown potentially promising antimalarial activity, but its antimalarial mechanism remains largely elusive. Methods We first established the Plasmodium berghei ANKA-infected C57BL/6 mouse model and systematically evaluated the antimalarial effects of Cel in conjunction with in vitro culture of Plasmodium falciparum. The potential antimalarial targets of Cel were then identified using a Cel activity probe based on the activity-based protein profiling (ABPP) technology. Subsequently, the antimalarial mechanism was analyzed by integrating with proteomics and transcriptomics. The binding of Cel to the identified key target proteins was verified by a series of biochemical experiments and functional assays. Results The results of the pharmacodynamic assay showed that Cel has favorable antimalarial activity both in vivo and in vitro. The ABPP-based target profiling showed that Cel can bind to a number of proteins in the parasite. Among the 31 identified potential target proteins of Cel, PfSpdsyn and PfEGF1-α were verified to be two critical target proteins, suggesting the role of Cel in interfering with the de novo synthesis of spermidine and proteins of the parasite, thus exerting its antimalarial effects. Conclusions In conclusion, this study reports for the first time the potential antimalarial targets and mechanism of action of Cel using the ABPP strategy. Our work not only support the expansion of Cel as a potential antimalarial agent or adjuvant, but also establishes the necessary theoretical basis for the development of potential antimalarial drugs with pentacyclic triterpenoid structures, as represented by Cel.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Bio-layer Interferometry (Bli) Assaymentioning

confidence: 99%

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Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

Gao,

Wang,

Tang

et al. 2024

Cell Commun Signal

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“…LC‐MS/MS analysis for target identification was performed as previously described [ 20 ]. Data acquisition was carried out with Xcalibur (version 4.2.4) in a data‐dependent acquisition mode.…”

Section: Methodsmentioning

confidence: 99%

“…While we and other groups have identified covalently bound proteins that are alkylated by free radicals of heme‐activated ART [ 14 , 15 ], it is likely that certain pharmacological effects of ART may not rely on covalent binding [ 16 , 17 , 18 , 19 ]. Indeed, our recent work suggests that ART could interact noncovalently with a handful of parasite proteins in vitro [ 20 ]. Previous studies have shown that parasites show different sensitivities to ART at different developmental stages (i.e., the ring, trophozoite, and schizont stages) within the IDC, suggesting that the parasite proteins to which ART binds may vary between stages [ 21 , 22 ].…”

Section: Introductionmentioning

confidence: 99%

Photoaffinity probe‐based antimalarial target identification of artemisinin in the intraerythrocytic developmental cycle of Plasmodium falciparum

Gao,

Wang,

Qiu

et al. 2024

iMeta

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Malaria continues to pose a serious global health threat, and artemisinin remains the core drug for global malaria control. However, the situation of malaria resistance has become increasingly severe due to the emergence and spread of artemisinin resistance. In recent years, significant progress has been made in understanding the mechanism of action (MoA) of artemisinin. Prior research on the MoA of artemisinin mainly focused on covalently bound targets that are alkylated by artemisinin‐free radicals. However, less attention has been given to the reversible noncovalent binding targets, and there is a paucity of information regarding artemisinin targets at different life cycle stages of the parasite. In this study, we identified the protein targets of artemisinin at different stages of the parasite's intraerythrocytic developmental cycle using a photoaffinity probe. Our findings demonstrate that artemisinin interacts with parasite proteins in vivo through both covalent and noncovalent modes. Extensive mechanistic studies were then conducted by integrating target validation, phenotypic studies, and untargeted metabolomics. The results suggest that protein synthesis, glycolysis, and oxidative homeostasis are critically involved in the antimalarial activities of artemisinin. In summary, this study provides fresh insights into the mechanisms underlying artemisinin's antimalarial effects and its protein targets.

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Chemical proteomics accelerates the target discovery of natural products

He,

Li,

Zhou

et al. 2024

Biochemical Pharmacology

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Chemical proteomic profiling with photoaffinity labeling strategy identifies antimalarial targets of artemisinin

Cited by 4 publications

References 27 publications

Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

Chemoproteomics-based profiling reveals potential antimalarial mechanism of Celastrol by disrupting spermidine and protein synthesis

Photoaffinity probe‐based antimalarial target identification of artemisinin in the intraerythrocytic developmental cycle of Plasmodium falciparum

Chemical proteomics accelerates the target discovery of natural products

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