Joint Screening and Identification of Potential Targets of Nitazoxanide by Affinity Chromatography and Label-Free Techniques

文献类型： 外文期刊

第一作者： Zhu, Menghan

作者： Zhu, Menghan;Qi, Dongxia;Li, Juan;Zhu, Menghan;Qi, Dongxia;Chen, Dongliang;Ye, Wenchong;Wang, Xiaoyang;Wang, Chunmei;Zhou, Wen;Zhang, Keyu;Zhou, Bin

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关键词： Nitazoxanide; affinity chromatography; drug target; autophagy

期刊名称：CURRENT DRUG TARGETS （ 影响因子：2.5； 五年影响因子：3.3 ）

ISSN： 1389-4501

年卷期： 2024 年 25 卷 12 期

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收录情况： SCI

摘要： Background Nitazoxanide not only exhibits a broad spectrum of activities against various pathogens infecting animals and humans but also induces cellular autophagy. Currently, the pattern of action and subcellular targets of nitazoxanide-induced cellular autophagy are still unclear.Methods To identify potential targets of nitazoxanide in mammalian cells, we developed an affinity chromatography system using tizoxanide, a deacetyl derivative of nitazoxanide, as a ligand. Affinity chromatography was performed using VERO cell extracts on tizoxanide-biotin, and the isolated binding proteins were identified by mass spectrometry. Candidate target proteins obtained using affinity chromatography were co-analysed with the drug affinity response target stability method. Fluorescent probes obtained by coupling rhodamine B to nitazoxanide were used for intracellular localisation of the binding targets. Solvent-induced protein precipitation profiling and thermal proteome profiling were used to further validate the binding proteins.Results The joint analysis of the drug affinity response target stability method and affinity chromatography resulted in the screening of six possible candidate target proteins. Fluorescent probes localised the nitazoxanide-binding protein around the nuclear membrane. Molecular docking revealed that the binding proteins mainly formed hydrogen bonds with the nitro group of nitazoxanide. Solvent-induced protein precipitation profiling and thermal proteome profiling further validated SEC61A, PSMD12, and PRKAG1 as potential target proteins of nitazoxanide.Conclusion The data supports the idea that nitazoxanide is a multifunctional compound with multiple targets.

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