Quantification of protein kinase activities by LC-MS.
Measuring the enzymatic activity of protein kinases in cell and tissue extracts represents a difficult task owing to the complex regulation and dynamics of such enzymes. Here we describe a sensitive and specific approach for the quantitative analysis of PI3K-dependent protein kinase activity based on the mass spectrometry measurement of reaction products. The principle of this method can be applied to develop other kinase assays and thus should contribute to the understanding of processes controlled by protein kinases. Because of the enhanced sensitivity of this technique, it may be applied to the multiplex measurement of pathway activities when sample amounts are limiting.
Alcolea
Maria P
MP
Analytical Signalling Group, Centre for Cell Signalling, Institute of Cancer, Bart's and the London School of Medicine, Queen Mary University of London, London, UK.
Cutillas
Pedro R
PR
D053000:Analytic Sample Preparation Methods; D045744:Cell Line, Tumor; D002852:Chromatography, Ion Exchange; D002853:Chromatography, Liquid / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D015398:Signal Transduction; D013379:Substrate Specificity
D010455:Peptides; D011494:Protein Kinases; D019869:Phosphatidylinositol 3-Kinases