title: Proteome-wide discovery of unknown ATP-binding proteins and kinase inhibitor target proteins using an ATP probe.
abstract: ATP-binding proteins, including protein kinases, play essential roles in many biological and pathological processes and thus these proteins are attractive as drug targets. Acyl-ATP probes have been developed as efficient probes for kinase enrichment, and these probes have also been used to enrich other ATP-binding proteins. However, a robust method to identify ATP-binding proteins with systematic elimination of nonspecific binding proteins has yet to be established. Here, we describe an ATP competition assay that permitted establishment of a rigorous ATP-binding protein list with virtual elimination of nonspecific proteins. A total of 539 ATP-binding protein candidates were identified, including 178 novel candidates. In informatics analysis, ribosomal proteins were overrepresented in the list of novel candidates. We also found multiple ATP-competitive sites for several kinases, including epidermal growth factor receptor, serine/threonine-protein kinase PRP4 homologue, cyclin-dependent kinase 12, eukaryotic elongation factor 2 kinase, ribosomal protein S6 kinase alpha-1, and SRSF protein kinase 1. Using our cataloged ATP-binding protein list, a selectivity profiling method that covers the kinome and ATPome was established to identify off-target binding sites of ATP-competitive kinase inhibitors, staurosporine and crizotinib.
authors.affiliation: Laboratory of Proteome Research, National Institute of Biomedical Innovation , Ibaraki, Osaka 567-0085, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Adachi
authors.forename: Marina
authors.initials: M
authors.lastname: Kishida
authors.forename: Shio
authors.initials: S
authors.lastname: Watanabe
authors.forename: Yuuki
authors.initials: Y
authors.lastname: Hashimoto
authors.forename: Kazuna
authors.initials: K
authors.lastname: Fukamizu
authors.forename: Takeshi
authors.initials: T
authors.lastname: Tomonaga
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000378:metabolism*; D002853:Chromatography, Liquid; D000077547:Crizotinib; D006367:HeLa Cells; D006801:Humans; D015335:Molecular Probes / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000378:metabolism / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D011720:Pyrazoles / Q000378:metabolism / Q000494:pharmacology; D011725:Pyridines / Q000378:metabolism / Q000494:pharmacology; D019311:Staurosporine / Q000378:metabolism / Q000494:pharmacology; D053719:Tandem Mass Spectrometry
chemical_list: D002352:Carrier Proteins; D015335:Molecular Probes; D047428:Protein Kinase Inhibitors; D020543:Proteome; D011720:Pyrazoles; D011725:Pyridines; D000077547:Crizotinib; D000255:Adenosine Triphosphate; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D019311:Staurosporine
keywords: ATP-binding proteins; ATP-competitive kinase inhibitor; Chemical proteomics