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Casein Kinase II (CK2)

Catalog # Concentration Size List Price Quantity Your Price
P6010L 500000 units/ml 50000 units $1,035.00
$931.50
P6010S 500000 units/ml 10000 units $260.00
$234.00
Catalog # Size List Price Your Price
P6010L 50000 units $1,035.00
$931.50
P6010S 10000 units $260.00
$234.00
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca

Casein Kinase II (CKII) is a constitutively active serine/threonine protein kinase composed of two 44 kDa catalytic α-subunits and two 26 kDa regulatory β-subunits in an α2β2 configuration to form stable heterotetramers. The general recognition motif for phsophorylation by CKII is SXXE/D.

  • Recombinant enzyme with no detectable protease or phosphatase contaminating activities
  • Optimal activity and stability for up to 12 months
Casein Kinase II (CK2) is a constitutively active serine/threonine protein kinase composed of two 44 kDa catalytic α-subunits and two 26 kDa regulatory β-subunits in an α2β2 configuration to form stable heterotetramers. CK2 holoenzyme undergoes autophosphorylation at two serine residues (S2/S3) of its β-subunit. Recently it has been shown that CK2 α-subunits undergo intermolecular tyrosine-autophosphorylation at Y182, which may represent a specific regulatory mechanism. Also, CK2 is able to phosphorylate, under special circumstances, tyrosyl residues in proteins. CK2 is implicated in a variety of cellular functions (1,2).
Product Source
Isolated from a strain of E. coli expressing both α and β CK2 subunits derived from a human glioblastoma cDNA library (kindly provided by Dr. D. Marshak) (3).
Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  Casein Kinase II (CK2) P6010SVIAL -80 1 x 0.02 ml 500,000 units/ml
  NEBuffer™ for Protein Kinases (PK) B6022SVIAL -20 1 x 1 ml 10 X
  Casein Kinase II (CK2) P6010LVIAL -80 1 x 0.1 ml 500,000 units/ml
  NEBuffer™ for Protein Kinases (PK) B6022SVIAL -20 1 x 1 ml 10 X

Properties & Usage

Unit Definition

One unit is defined as the amount of CK2 required to catalyze the transfer of 1 pmol of phosphate to CK2 Peptide Substrate, RRRADDSDDDDD (100 µM), in 1 minute at 30°C in a total reaction volume of 25 µl (4,5).

Reaction Conditions

1X NEBuffer™ for Protein Kinases (PK)
Supplement with 200 µM ATP
Incubate at 30°C

1X NEBuffer™ for Protein Kinases (PK)
50 mM Tris-HCl
10 mM MgCl2
0.1 mM EDTA
2 mM DTT
0.01% Brij 35
(pH 7.5 @ 25°C)

Storage Notes

  • Avoid repeated freeze/thaw cycles.


Companion Products
Materials Sold Separately
Notes
  • Molecular Weight: α-subunit (45 kDa), β-subunit (25 kDa). The apparent molecular weight of the α-subunit estimated by SDS-PAGE is about 42 kDa.
  • General notes:
    • For short term storage (two weeks or less) CK2 can be stored at -20°C. 
    • If the source of protein to be phosphorylated is a crude extract of cells or tissue, it is very important to include the appropriate protease and protein phosphatase inhibitors in the lysis buffer and to use shorter incubation time for phosphorylation. 
    • Reaction Conditions: 1X NEBuffer for Protein Kinases (PK), supplement with 200 µM ATP and gamma-labeled ATP to a final specific activity of 100-500 µCi/µmol. (NEBuffer for Protein Kinases (PK) will also accept GTP as a phosphoryl donor in place of ATP).
  • Usage notes:
    • Optimal incubation times and enzyme concentrations must be determined empirically for each particular substrate. 
    • If possible, the ATP concentration should be at or near saturation (5 -10-fold over Km). Apparent Km values of ATP for most protein kinases are below 100 μM. 
    However, if the objective is to measure enzyme activity using gamma-labeled ATP, it is best to use 100-200 μM ATP in order to have higher specific activity of gamma-labeled ATP (100-500 cpm/pmol). Also, an excess of substrate should be used, and the level of phosphorylation should not exceed 10% for determination of the initial rate.

    To phosphorylate a protein or peptide substrate to completion, the ATP concentration should be about 5-fold over the limited substrate concentration. Higher enzyme concentration and prolonged incubation times should be employed.

    Recommended reference:Protein Phosphorylation: A Practical Approach (1993) ed. Hardie, D.G. IRL Press.
References
  • Chester, N. and Marshak, D.R. (1993). Anal. Biochem. 209, 284-290.
  • Donella-Deana, A. et al. (2001). Biochem J. 357, 563-567.
  • Marin, O. et al. (1999). J. Biol. Chem. 274, 29260-29265.
  • Marin, O. et al. (1994). BBRC. 198, 898-905.
  • Sarno, S. et al. (1996). J. Biol. Chem. 271, 10595-10601.
FAQs
Tech Tips
  • Make sure to use the appropriate protease and protein phosphatase inhibitors in the lysis buffer and to use shorter incubation time for phosphorylation if the substrate is a crude lysate.
  • If possible, the ATP concentration should be at or near saturation (5 -10-fold over Km). Apparent Km values of ATP for most protein kinases are below 100 µM.
  • 1-2 µl of enzyme is a good starting point for a 1 hr incubation of 1µg of protein.
  • The consensus sequence is SXXE/D
  • Heparin inhibits CKII at a Ki of 1.4 nM
Quality Control Assay
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
Certificate of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety Data Sheets
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This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

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