Product Pathways - Kinases
MARK3 Kinase #7354
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 full-length human MARK3 kinase, supplied as a GST fusion protein.
Source / Purification
The GST-Kinase fusion protein was produced using a baculovirus expression system using sf9 cells and a recombinant virus encoding full- length human MARK3 (Met1- Leu713) (GenBank Accession No. NM_002376.4) with an amino-terminal GST tag. The protein was purified by one-step affinity chromatography using GSH-agarose.
Gel Staining
Figure 1. The purity of the GST-MARK3 fusion protein was analyzed using SDS/PAGE followed by Coomassie stain.
Kinase Assay - Radiometric
Figure 2. MARK3 kinase activity was measured in a radiometric assay using the following reaction conditions: 60 mM HEPES-NaOH, pH 7.5, 3 mM MgCl2, 3 mM MnCl2, 3 μM Na-orthovanadate, 1.2 mM DTT, 50 μg/μl PEG20,000, ATP variable, Substrate: CHKtide 5 ng/μL and recombinant MARK3: 200 ng/ 50 μl.
Quality Control
The theoretical molecular weight of the GST-MARK3 fusion protein is 109 kDa. The purity of the kinase was assessed using SDS-PAGE followed by Coomassie stain [Fig.1]. MARK3 kinase activity was determined using a radiometric assay [Fig.2].
Background
Microtubule associated proteins regulate the stability of microtubules and control processes such as cell polarity/differentiation, neurite outgrowth, cell division and organelle trafficking (1). The MARK (MAP/microtubule affinity-regulating kinases) family (MARK1-4) of serine/threonine kinases was identified based on their ability to phosphorylate microtubule-associated proteins (MAPs) including tau, MAP2 and MAP4 (2-6). MARK proteins phosphorylate MAPs within their microtubule binding domains, causing dissociation of MAPs from microtubules and increased microtubule dynamics (2-4). In the case of tau, phosphorylation has been hypothesized to contribute to the formation of neurofibrillary tangles observed in Alzheimer's disease. Overexpression of MARK leads to hyperphosphorylation of MAPs, morphological changes and cell death (4). The tumor suppressor kinase LKB1 phosphorylates MARK and the closely related AMP-kinases within their T-loops, leading to increased activity (7).
- Drubin, D.G. and Nelson, W.J. (1996) Cell 84, 335-344.
- Illenberger, S. et al. (1996) J. Biol. Chem. 271, 10834-10843.
- Drewes, G. et al. (1995) J. Biol. Chem. 270, 7679-7688.
- Drewes, G. et al. (1997) Cell 89, 297-308.
- Kato, T. et al. (2001) Neoplasia 3, 4-9.
- Trinczek, B. et al. (2004) J. Biol. Chem. 279, 5915-5923.
- Lizcano, J. M. et al. (2004) EMBO J. 23, 833-843.
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.
