Quantitative Chemical Proteomics for Identifying Candidate Drug Targets

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• 作者：Yoshiya Oda ; Takashi Owa ; Toshitaka Sato ; Brian Boucher ; Scott Daniels ; Hidenori Yamanaka ; Yasuhiro Shinohara ; Akira Yokoi ; Junro Kuromitsu ; and Takeshi Nagasu
• 刊名：Analytical Chemistry
• 出版年：2003
• 出版时间：May 1, 2003
• 年：2003
• 卷：75
• 期：9
• 页码：2159 - 2165
• 全文大小：265K
• 年卷期：v.75,no.9(May 1, 2003)
• ISSN：1520-6882

文摘

We have developed a systematic strategy for drug targetidentification. This consists of the following sequentialsteps: (1) enrichment of total binding proteins using twodifferential affinity matrixes upon which are immobilizedpositive and negative chemical structures for drug activity,respectively; (2) covalent labeling of the proteins with anew cleavable isotope-coded affinity tag (ICAT) reagent,followed by proteolysis of the combined proteins; (3)isolation, identification, and relative quantification of thetagged peptides by liquid chromatography-mass spectrometry; (4) array-based transcription profiling to selectcandidate proteins; and (5) confirmation of direct interaction between the activity-associated structure and theselected proteins by using surface plasmon resonance. Wepresent a typical application to identify the primarybinding protein of a novel class of anticancer agentsexemplified by E7070. Our results suggest that thisapproach provides a new aspect of quantitative proteomicsto find specific binding proteins from protein mixture andshould be applicable to a wide variety of biologically activesmall molecules with unidentified target proteins.