Quantification of Protein Kinase Enzymatic Activity in Unfractionated Cell Lysates Using CSox‐Based Sensors
Author(s)Beck, Jon R.; Peterson, Laura B; Imperiali, Barbara; Stains, Cliff I.
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Defining perturbations in protein kinase activity within biological samples can provide insight into disease mechanisms as well as potential targets for drug development. In this article, we present a method that utilizes a phosphorylation-sensitive amino acid, termed CSox, to afford kinase-selective biosensors capable of reporting on enzymatic activity directly in biological samples. These sensors produce an increase in fluorescence in response to phosphorylation of an amino acid residue adjacent to CSox. Probes can be designed for either serine/threonine or tyrosine kinases, and analysis can be performed using standard fluorescence equipment. The procedures provided herein represent our optimized protocols for the design, validation, and application of CSox-based protein kinase activity sensors.
DepartmentMassachusetts Institute of Technology. Department of Biology; Massachusetts Institute of Technology. Department of Chemistry
Beck, John R. et al. "Quantification of Protein Kinase Enzymatic Activity in Unfractionated Cell Lysates Using CSox‐Based Sensors." Current Protocols 6, 3 (September 2014): 135-156 © 2014 John Wiley & Sons, Inc.
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