Slon-Usakiewicz Jacek J, Dai Jin-Rui, Ng William, Foster J Estelle, Deretey Eugen, Toledo-Sherman Leticia, Redden Peter R, Pasternak Andrew, Reid Neil
Lead Discovery and Optimization Division, Protana Inc., 251 Attwell Drive, Toronto, Ontario, M9W 7H4 Canada.
Anal Chem. 2005 Mar 1;77(5):1268-74. doi: 10.1021/ac048716q.
Utilizing frontal affinity chromatography with mass spectrometry detection (FAC-MS), we have identified novel applications in the discovery of small-molecule hits to protein targets that are difficult if not impossible to accomplish using traditional assays. We demonstrate for the first time an ability to distinguish between competitive ligands for the ATP and substrate sites of protein kinase C independently in the same experiment and show that ATP competitive ligands using a functionally inactive receptor tyrosine kinase can be identified. This ability of FAC-MS to simultaneously monitor binding at the ATP and substrate binding sites, as well as measure ligand binding to both active and inactive kinases, suggests that FAC-MS can be used as a "global kinase binding assay".
利用具有质谱检测功能的前沿亲和色谱法(FAC-MS),我们在发现针对蛋白质靶点的小分子命中物方面确定了新的应用,而这些应用使用传统检测方法即便不是不可能完成,也是很难实现的。我们首次证明了在同一实验中能够独立区分蛋白激酶C的ATP位点和底物位点的竞争性配体,并表明使用功能失活的受体酪氨酸激酶可以鉴定出ATP竞争性配体。FAC-MS能够同时监测ATP和底物结合位点的结合情况,以及测量配体与活性和失活激酶的结合,这表明FAC-MS可用作“全局激酶结合测定法”。