Product Pathways - MAPK Signaling
MAPKAPK-2 Kinase #7442
Cell Signaling Technology offers a full line of protein kinases, substrates, antibody detection reagents and HTScan® kits. Browse our "Reagents for High-Throughput Screening" product listing or contact us at drugdiscovery@cellsignal.com.
Description
Purified recombinant human MAPKAPK-2 (Phe46-His400) kinase, supplied as a GST fusion protein.
Source / Purification
The GST-Kinase fusion protein was produced using a baculovirus expression system with a construct expressing human MAPKAPK-2 (Phe46-His400) (GenBank Accession No. NM_032960) with an amino-terminal GST tag. The protein was purified by one-step affinity chromatography using glutathione-agarose.
Gel Staining
Figure 1. The purity of the GST-MAPKAPK-2 fusion protein was analyzed using SDS/PAGE followed by Coomassie stain.
Kinase Assay - Radiometric
Figure 2. MAPKAPK-2 kinase activity was measured in a radiometric assay using the following reaction conditions: 4 mM MOPS, pH 7.2, 2.5 mM β-glycerophosphate, 1 mM EGTA, 0.4 mM EDTA, 4 mM MgCl2, 0.05 mM DTT, 40 ng/μL BSA, 50 μM ATP, Substrate: MAPKAPK-2 substrate peptide 400 ng/μL and recombinant MAPKAPK-2: variable.
Quality Control
The theoretical molecular weight of the GST- MAPKAPK-2 fusion protein is 68 kDa. The purified kinase was quality controlled for purity using SDS-PAGE followed by Coomassie stain [Fig.1]. MAPKAPK-2 kinase activity was determined using a radiometric assay [Fig.2].
Background
In response to cytokines, stress and chemotactic factors, MAP kinase-activated protein kinase 2 (MAPKAPK-2) is rapidly phosphorylated and activated. It has been shown that MAPKAPK-2 is a direct target of p38 MAPK (1). Multiple residues of MAPKAPK-2 are phosphorylated in vivo in response to stress. However, only four residues (Thr25, Thr222, Ser272 and Thr334) are phosphorylated by p38 MAPK in an in vitro kinase assay (2). Phosphorylation at Thr222, Ser272 and Thr334 appears to be essential for the activity of MAPKAPK-2 (2). Thr25 is phosphorylated by p42 MAPK in vitro, but is not required for the activation of MAPKAPK-2 (2).
Application References
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Companion Products
This product is for in vitro research use only and is not intended for use in humans or animals. This product is not intended for use as therapeutic or in diagnostic procedures.
