Profiling of drug binding proteins by monolithic affinity chromatography in combination with liquid chromatography-tandem mass spectrometry
Profiling of drug binding proteins by monolithic affinity chromatography in combination with liquid chromatography-tandem mass spectrometry
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通过整体式亲和层析结合液相色谱-串联质谱分析药物结合蛋白
DOI：
10.1016/j.chroma.2014.07.020
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发表时间：
2014
影响因子：
4.1
通讯作者：
Kang Jingwu
中科院分区：
文献类型：
--
作者：
Zhang Xuepei;Wang Tongdan;Zhang Hanzhi;Han Bing;Wang Lishun;Kang Jingwu
报道了一种利用整体毛细管亲和色谱结合高效液相色谱-串联质谱对药物结合蛋白进行全蛋白质组分析的新方法。两种免疫抑制药物，即FK 506和环孢菌素A，被用作概念验证的实验模型。通过甲基丙烯酸缩水甘油酯和二甲基丙烯酸乙二醇酯与药物衍生物的一步共聚反应制备了毛细管整体柱。采用亲和整体柱的毛细管色谱法有利于从细胞裂解物中纯化药物结合蛋白。结合毛细管亲和柱纯化和鸟枪蛋白质组学分析，共鉴定出33个FK 506和32个CsA结合蛋白，包括所有文献报道的这两种药物的靶蛋白。其中，两个蛋白，即电压依赖性阴离子选择性通道蛋白1和丝氨酸/苏氨酸蛋白磷酸酶PGAM 5，通过使用重组蛋白进行验证。该结果支持整体毛细管亲和色谱可能成为一种有价值的工具，小分子药物以及生物活性化合物的结合蛋白的分析。
A new approach for proteome-wide profiling drug binding proteins by using monolithic capillary affinity chromatography in combination with HPLC–MS/MS is reported. Two immunosuppresive drugs, namely FK506 and cyclosporin A, were utilized as the experimental models for proof-of-concept. The monolithic capillary affinity columns were prepared through a single-step copolymerization of the drug derivatives with glycidyl methacrylate and ethylene dimethacrylate. The capillary chromatography with the affinity monolithic column facilitates the purification of the drug binding proteins from the cell lysate. By combining the capillary affinity column purification and the shot-gun proteomic analysis, totally 33 FK506- and 32 CsA-binding proteins including all the literature reported target proteins of these two drugs were identified. Among them, two proteins, namely voltage-dependent anion-selective channel protein 1 and serine/threonine-protein phosphatase PGAM5 were verified by using the recombinant proteins. The result supports that the monolithic capillary affinity chromatography is likely to become a valuable tool for profiling of binding proteins of small molecular drugs as well as bioactive compounds.