Recombinant full-length human CDK2 and CyclinE1 were co-expressed by baculovirus in Sf9 insect cells using an N-terminal GST tag on both proteins. CDK2 is a member of the Cyclin-Dependent Kinase family that is ubiquitously expressed. CDK2 is a catalytic subunit of the cyclin-dependent protein kinase complex, whose activity is restricted to the G1-S phase and essential for cell cycle G1/S phase transition. CDK2 associates with and is regulated by the regulatory subunits of the complex including Cyclin A or E, CDK inhibitor p21Cip1 (CDKN1A) and p27Kip1 (CDKN1B) (1). CDK2 phosphorylates multiple cellular substrates including SMAD3 and FOXO1. Phosphorylation of FOXO1 leads to its inhibition (2).
ADP-Glo™ Kinase Assay is a luminescent kinase assay that measures ADP formed from a kinase reaction; ADP is converted into ATP, which is a substrate in a reaction catalyzed by Ultra-Glo™ Luciferase that produces light. The luminescent signal positively correlates with ADP amount and kinase activity. The assay is well suited for measuring the effects chemical compounds have on the activity of a broad range of purified kinases, making it ideal for both primary screening as well as kinase selectivity profiling. The ADP-Glo™ Kinase Assay can be used to monitor the activity of virtually any ADP-generating enzyme (e.g., kinase or ATPase) using up to 1mM ATP.
Profile More Compounds In-House: ADP-Glo™ Kinase Assay + Kinase Enzyme System is optimized so that you are up and running in no time. Complete Systems: The Kinase Enzyme Systems include a recombinant kinase enzyme, a substrate appropriate for the enzyme, a reaction buffer, DTT and supplemental reagents as needed. Obtain Reliable Results: The broad dynamic range, the ease of use and better sensitivity obtained with ADP-Glo™ Kinase Assay result in less ambiguous data.
Notes
Kinase Enzyme System manufactured by SignalChem.
Bulk quantities available upon request.
References
1.Levkau, B. et al. (1998) Cleavage of p21(Cip1/Waf1) and p27(Kip1) mediates apoptosis in endothelial cells through activation of Cdk2: role of a caspase cascade. Molec. Cell 1, 553–63.
2.Huang, H. et al. (2006) CDK2-dependent phosphorylation of FOXO1 as an apoptotic response to DNA damage. Science 314, 294–7.