Global kinase screening. Applications of frontal affinity chromatography coupled to mass spectrometry in drug discovery.

Utilizing frontal affinity chromatography with mass spectrometry detection (FAG-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. Ibis ability of FAGMS 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 FAGMS can be used as a "global kinase binding assay".