title: Affinity-based probes based on type II kinase inhibitors.
abstract: Protein kinases are key components of most mammalian signal transduction networks and are therapeutically relevant drug targets. Efforts to study protein kinase function would benefit from new technologies that are able to profile kinases in complex proteomes. Here, we describe active site-directed probes for profiling kinases in whole cell extracts and live cells. These probes contain general ligands that stabilize a specific inactive conformation of the ATP-binding sites of protein kinases, as well as trifluoromethylphenyl diazirine and alkyne moieties that allow covalent modification and enrichment of kinases, respectively. A diverse group of serine/threonine and tyrosine kinases were identified as specific targets of these probes in whole cell extracts. In addition, a number of kinase targets were selectively labeled in live cells. Our chemical proteomics approach should be valuable for interrogating protein kinase active sites in physiologically relevant environments.
authors.affiliation: Department of Chemistry, University of Washington, Seattle, Washington 98195, USA.
authors.forename: Pratistha
authors.initials: P
authors.lastname: Ranjitkar
authors.forename: B Gayani K
authors.initials: BG
authors.lastname: Perera
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Swaney
authors.forename: Daniel L
authors.initials: DL
authors.lastname: Swaney
authors.forename: Sanjay B
authors.initials: SB
authors.lastname: Hari
authors.forename: Eric T
authors.initials: ET
authors.lastname: Larson
authors.forename: Ratika
authors.initials: R
authors.lastname: Krishnamurty
authors.forename: Ethan A
authors.initials: EA
authors.lastname: Merritt
authors.forename: Judit
authors.initials: J
authors.lastname: Villén
authors.forename: Dustin J
authors.initials: DJ
authors.lastname: Maly
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D008958:Models, Molecular; D019920:Photoaffinity Labels; D047428:Protein Kinase Inhibitors / Q000737:chemistry*; D040901:Proteomics
chemical_list: D019920:Photoaffinity Labels; D047428:Protein Kinase Inhibitors; D000255:Adenosine Triphosphate