title: Fluorescent peptide assays for protein kinases.
abstract: Protein kinases are enzymes that regulate many cellular events in eukaryotic cells, such as cell-cycle progression, transcription, metabolism, and apoptosis. Protein kinases each have a conserved ATP-binding site, as well as one or more substrate-binding site(s) that exhibit recognition features for a protein substrate. Thus, by bringing ATP and a substrate into close proximity, each protein kinase can modify its substrate by transferring the gamma phosphate of the ATP molecule to a serine, threonine, or tyrosine residue on the substrate. In such a way, signaling pathways downstream from the substrate can be regulated, dependent on the phosphorylated versus dephosphorylated forms of the substrate. This unit describes an assay employing a fluorescent peptide substrate to measure the incorporation of non-radiolabeled phosphate. The assay is based on the principle that the phosphorylation of the peptide substrate leads to an increase in the fluorescence emission intensity of an appended fluorophore.
authors.affiliation: University of Texas at Austin, Austin, Texas, USA.
authors.forename: Ashwini K
authors.initials: AK
authors.lastname: Devkota
authors.forename: Tamer S
authors.initials: TS
authors.lastname: Kaoud
authors.forename: Mangalika
authors.initials: M
authors.lastname: Warthaka
authors.forename: Kevin N
authors.initials: KN
authors.lastname: Dalby
mesh_terms: D005453:Fluorescence; D005456:Fluorescent Dyes / Q000494:pharmacology*; D010455:Peptides / Q000378:metabolism*; D010758:Phosphorus / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D005456:Fluorescent Dyes; D010455:Peptides; D010758:Phosphorus; D011494:Protein Kinases