Global kinase screening. Applications of frontal affinity chromatography coupled to mass spectrometry in drug discovery.
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".
Slon-Usakiewicz
Jacek J
JJ
Lead Discovery and Optimization Division, Protana Inc., 251 Attwell Drive, Toronto, Ontario, M9W 7H4 Canada.
Dai
Jin-Rui
JR
Ng
William
W
Foster
J Estelle
JE
Deretey
Eugen
E
Toledo-Sherman
Leticia
L
Redden
Peter R
PR
Pasternak
Andrew
A
Reid
Neil
N
D000470:Alkaloids / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D053119:Benzophenanthridines / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D020134:Catalytic Domain; D002846:Chromatography, Affinity / Q000379:methods*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D006801:Humans; D007093:Imidazoles / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D015394:Molecular Structure; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D051571:Protein Kinase C-alpha / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D011725:Pyridines / Q000737:chemistry / Q000378:metabolism; D011799:Quinazolines / Q000737:chemistry / Q000378:metabolism; D036183:Receptor, EphB2 / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D000470:Alkaloids; D053119:Benzophenanthridines; D007093:Imidazoles; D010455:Peptides; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D011799:Quinazolines; D011994:Recombinant Proteins; C118361:WHI P180; C016299:chelerythrine; D011494:Protein Kinases; D036183:Receptor, EphB2; D051571:Protein Kinase C-alpha; C093642:SB 203580; C088860:4-((3-bromophenyl)amino)-6,7-dimethoxyquinazoline