title: Functional interrogation of the kinome using nucleotide acyl phosphates.
abstract: The central role of protein kinases in signal transduction pathways has generated intense interest in targeting these enzymes for a wide range of therapeutic indications. Here we report a method for identifying and quantifying protein kinases in any biological sample or tissue from any species. The procedure relies on acyl phosphate-containing nucleotides, prepared from a biotin derivative and ATP or ADP. The acyl phosphate probes react selectively and covalently at the ATP binding sites of at least 75% of the known human protein kinases. Biotinylated peptide fragments from labeled proteomes are captured and then sequenced and identified using a mass spectrometry-based analysis platform to determine the kinases present and their relative levels. Further, direct competition between the probes and inhibitors can be assessed to determine inhibitor potency and selectivity against native protein kinases, as well as hundreds of other ATPases. The ability to broadly profile kinase activities in native proteomes offers an exciting prospect for both target discovery and inhibitor selectivity profiling.
authors.affiliation: ActivX Biosciences, 11025 North Torrey Pines Road, La Jolla, California 92037, USA..
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Patricelli
authors.forename: A Katrin
authors.initials: AK
authors.lastname: Szardenings
authors.forename: Marek
authors.initials: M
authors.lastname: Liyanage
authors.forename: Tyzoon K
authors.initials: TK
authors.lastname: Nomanbhoy
authors.forename: Min
authors.initials: M
authors.lastname: Wu
authors.forename: Helge
authors.initials: H
authors.lastname: Weissig
authors.forename: Arwin
authors.initials: A
authors.lastname: Aban
authors.forename: Doris
authors.initials: D
authors.lastname: Chun
authors.forename: Stephen
authors.initials: S
authors.lastname: Tanner
authors.forename: John W
authors.initials: JW
authors.lastname: Kozarich
mesh_terms: D000227:Adenine Nucleotides / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D017124:Conserved Sequence; D006801:Humans; D008958:Models, Molecular; D015336:Molecular Probe Techniques; D011487:Protein Conformation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020543:Proteome; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology
chemical_list: D000227:Adenine Nucleotides; D047428:Protein Kinase Inhibitors; D020543:Proteome; D011494:Protein Kinases; D019311:Staurosporine