title: The generality of kinase-catalyzed biotinylation.
abstract: Kinase-catalyzed protein phosphorylation is involved in a wide variety of cellular events. Development of methods to monitor phosphoproteins in normal and diseased states is critical to fully characterize cell signaling. Towards phosphoprotein analysis tools, our lab reported kinase-catalyzed labeling where γ-phosphate modified ATP analogs are utilized by kinases to label peptides or protein substrates with a functional tag. In particular, the ATP-biotin analog was developed for kinase-catalyzed biotinylation. However, kinase-catalyzed labeling has been tested rigorously with only a few kinases, preventing use of ATP-biotin as a general tool. Here, biotinylation experiments, gel or HPLC-based quantification, and kinetic measurements indicated that twenty-five kinases throughout the kinome tree accepted ATP-biotin as a cosubstrate. With this rigorous characterization of ATP-biotin compatibility, kinase-catalyzed labeling is now immediately useful for studying phosphoproteins and characterizing the role of phosphorylation in various biological events.
authors.affiliation: Department of Chemistry, Wayne State University, Detroit, MI 48202, United States.
authors.forename: Chamara
authors.initials: C
authors.lastname: Senevirathne
authors.affiliation: Department of Chemistry, Wayne State University, Detroit, MI 48202, United States.
authors.forename: D Maheeka
authors.initials: DM
authors.lastname: Embogama
authors.affiliation: Department of Chemistry, Wayne State University, Detroit, MI 48202, United States.
authors.forename: Thilani A
authors.initials: TA
authors.lastname: Anthony
authors.affiliation: Department of Chemistry, Wayne State University, Detroit, MI 48202, United States.
authors.forename: Ahmed E
authors.initials: AE
authors.lastname: Fouda
authors.affiliation: Department of Chemistry, Wayne State University, Detroit, MI 48202, United States. Electronic address: pflum@wayne.edu.
authors.forename: Mary Kay H
authors.initials: MK
authors.lastname: Pflum
mesh_terms: D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry; D019921:Biotinylation; D006801:Humans; D007700:Kinetics; D062105:Molecular Docking Simulation; D010766:Phosphorylation; D011494:Protein Kinases / Q000737:chemistry*
chemical_list: C000609801:ATP-biotin; D001710:Biotin; D011494:Protein Kinases
keywords: ATP analogs; Biotinylation; Kinase enzymes; Kinome tree