title: Profiling of drug binding proteins by monolithic affinity chromatography in combination with liquid chromatography-tandem mass spectrometry.
abstract: 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.
authors: Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, State Key Laboratory of Organic and Natural Products Chemistry, Lingling Road 345, Shanghai 200032, China.
authors: Xuepei
authors: X
authors: Zhang
authors: Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, South Chongqing Road 227, Shanghai 200025, China.
authors: Tongdan
authors: T
authors: Wang
authors: Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, State Key Laboratory of Organic and Natural Products Chemistry, Lingling Road 345, Shanghai 200032, China.
authors: Hanzhi
authors: H
authors: Zhang
authors: Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, South Chongqing Road 227, Shanghai 200025, China.
authors: Bing
authors: B
authors: Han
authors: Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, South Chongqing Road 227, Shanghai 200025, China.
authors: Lishun
authors: L
authors: Wang
authors: Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, State Key Laboratory of Organic and Natural Products Chemistry, Lingling Road 345, Shanghai 200032, China; ShanghaiTech University, Yueyang Road 319, Shanghai 200031, China. Electronic address: jingwu.kang@sioc.ac.cn.
authors: Jingwu
authors: J
authors: Kang
mesh_terms: D002352:Carrier Proteins / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D020104:Immunophilins / Q000737:chemistry*; D007166:Immunosuppressive Agents / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D016559:Tacrolimus / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D002352:Carrier Proteins; D007166:Immunosuppressive Agents; D020104:Immunophilins; D016559:Tacrolimus
keywords: Affinity chromatography; Chemical proteomics; Drug target identification; Monolithic column