Identification of nucleolus ⁃ related proteins using proximate biotin labeling technique combined with mass spectrometry
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    Abstract:

    Objective:In mouse embryonic stem cells(mESC),proximity labeling technology based on engineered ascorbate peroxidase(APEX2)and mass spectrometry were used to identify nucleolar related proteins. Methods:An APEX2-FBL overexpression stable cell line was constructed,biotin-labeled protein was obtained by APEX2 proximity labeling technology,the biotin-labeled protein was enriche with magnetic beads coated with streptavidin,and then liquid chromatography-tandem mass spectrometry(LC-MS/MS),and bioinformatics analysis was used to screen differential proteins. Results:Highly expressed APEX2-FBL stable cell lines were suuessfully constructed;Western blot results showed that a large number of biotin-labeled proteins were obtained by APEX2 proximity labeling technology;837 differential proteins were identified by mass spectrometry,and the biological processes were mostly related to nucleoli. Conclusion:The use of adjacent biotin labeling technology combined with mass spectrometry is a feasible method to identify nucleolar related proteins.

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臧春燕,姚礼萍,邢 鸣,张 军.利用邻近生物素标记技术结合质谱分析鉴定核仁相关蛋白[J].南京医科大学学报(自然科学版英文版),2020,(12):1750-1755.

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  • Received:May 19,2020
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  • Online: January 04,2021
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