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7615
HTScan® PRK1 Kinase Assay Kit
Cellular Assay Kits

HTScan® PRK1 Kinase Assay Kit #7615

This product is discontinued

Kinase Assay - Radiometric Image 1

Figure 1. PRK1 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, ATP, variable, 2.5 µg/50 µl PEG20,000, Substrate: Histone H2B, 1.5 µg/50 µl and Recombinant PRK1: 200 ng/50 µl.

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Kinase Assay - DELFIA Image 2

Figure 5. Staurosporine inhibition of PRK1 kinase activity: DELFIA® data generated using Phospho-PKA Substrate (RRXS/T) (100G7) Rabbit mAb #9624 to detect phosphorylation of PRK1 substrate peptide (#1331) by PRK1 kinase. In a 50 µl reaction, 50 ng PRK1, 1.5 µM substrate peptide, 20 µM ATP and increasing amounts of staurosporine were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 3

Figure 3. Dose dependence curve of PRK1 kinase activity: DELFIA® data generated using Phospho-PKA Substrate (RRXS/T) (100G7) Rabbit mAb #9624 to detect phosphorylation of substrate peptide (#1331) by PRK1 kinase. In a 50 µl reaction, increasing amounts of PRK1 and 1.5 µM substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 4

Figure 4. Peptide concentration dependence of PRK1 kinase activity: DELFIA® data generated using Phospho-PKA Substrate (RRXS/T) (100G7) Rabbit mAb #9624 to detect phosphorylation of substrate peptide (#1331) by PRK1 kinase. In a 50 µl reaction, 50 ng of PRK1 and increasing concentrations of substrate peptide were used per reaction at room temperature for 30 minutes. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Kinase Assay - DELFIA Image 5

Figure 2. Time course of PRK1 kinase activity: DELFIA® data generated using Phospho-PKA Substrate (RRXS/T) (100G7) Rabbit mAb #9624 to detect phosphorylation of PRK1 substrate peptide (#1331) by PRK1 kinase. In a 50 µl reaction, 50 ng PRK1 and 1.5 µM substrate peptide were used per reaction. (DELFIA® is a registered trademark of PerkinElmer, Inc.)

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Product Includes Quantity (with Count) Solution Color
Kinase Buffer (10X) 9802 1 x 15 ml
ATP (10 mM) 9804 1 x 1 ml

The kit provides a means of performing kinase activity assays with recombinant human PRK1 kinase. It includes active PRK1 kinase (supplied as a GST fusion protein), a biotinylated peptide substrate and a phospho-serine/threonine antibody for detection of the phosphorylated form of the substrate peptide.

Molecular Formula:

Peptide substrate, Biotin-CREB (Ser133): 2,326 Daltons. GST-PRK1 Kinase: 137 kDa.

Peptide Core Sequence:

RRPS*YRK

The protein kinase C-related kinases (PRKs) are a subfamily of Ser/Thr-specific kinases with a catalytic domain highly homologous to the PKC family (1-3). They are effectors of Rho family GTPases (4-6) and are activated by fatty acids and phospholipids in vitro (7,8). Activation in vitro and in vivo involves the activation loop phosphorylation of PRK1 (Thr774)/PRK2 (Thr816) by PDK1 (9,10).

  1. Mukai, H. et al. (1994) Biochem. Biophys. Res. Commun. 199, 897-904.
  2. Morrice, N.A. et al. (1994) J. Biol. Chem. 269, 20040-20046.
  3. Palmer, R.H. et al. (1994) FEBS Lett. 356, 5-8.
  4. Watanabe, G. et al. (1996) Science 271, 645-648.
  5. Amano, M. et al. (1996) Science 271, 648-650.
  6. Vincent, S. and Settleman, J. (1997) Mol. Cell. Biol. 17, 2247-2256.
  7. Morrice, N.A. et al. (1994) FEBS Lett. 351, 171-175.
  8. Palmer, R.H. et al. (1995) J. Biol. Chem. 270, 22412-22416.
  9. Flynn, P. et al. (2000) J Biol Chem 275, 11064-70.
  10. Dong, L.Q. et al. (2000) Proc. Natl. Acad. Sci. USA 97, 5089-94.
Entrez-Gene Id
5585
Swiss-Prot Acc.
Q16512
For Research Use Only. Not For Use In Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
DELFIA is a registered trademark of PerkinElmer, Inc.

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