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475701 MKK6, GST-Fusion, Human, Recombinant, E. coli

MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.

Synonyms: MEK6, Mitogen Activated Protein Kinase kinase 6, SAPKK3

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      Overview

      Replacement Information
      Description
      Overview

      This product has been discontinued.



      Recombinant, human MKK6 fused to GST at the N-terminus to GST and expressed in E. coli. MKK6 is a member of the dual specificity protein kinase family, which functions as a mitogen-activated protein (MAP) kinase kinase. It is suitable for labeling MAP kinase substrates.
      Catalogue Number475701
      Brand Family Calbiochem®
      SynonymsMEK6, Mitogen Activated Protein Kinase kinase 6, SAPKK3
      Application Data
      Increasing amounts of MKK6 were subjected to SDS-PAGE and the purity analyzed by Coomassie Blue staining.

      Increasing amounts of MKK6 were used to activate 10 µg p38α/SAPK2a and the activity of p38α/SAPK2a was measured as outlined in the protocols above.
      References
      ReferencesChabaud-Riou, M. and Firestein, G.S. et al. 2004. Am. J. Pathol. 164, 177.
      Zhu, X., et al. 2001. J. Neurochem. 79, 311.
      Product Information
      Unit of DefinitionOne unit is defined as the amount of enzyme that will activate p38alpha/SAPK2α (0.5 mg/ml) by 1 unit/min using 125 µM ATP at 30°C. One unit of p38α/SAPK2α activity is defined as the amount of enzyme that will transfer 1 pmol phosphate to myelin basic protein per min at 30°C.
      FormulationIn 270 mM sucrose, 150 mM NaCl, 50 mM Tris-HCl, 1 mM DTT, 0.1 mM EGTA, pH 7.5.
      Quality LevelMQ100
      Applications
      Biological Information
      Purity≥ 90% by SDS PAGE
      Specific Activity≥ 50,000 units/mg protein
      Concentration Label Please refer to vial label for lot-specific concentration
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Ship Code Dry Ice Only
      Toxicity Standard Handling
      Storage ≤ -70°C
      Avoid freeze/thaw Avoid freeze/thaw
      Do not freeze Ok to freeze
      Special InstructionsFollowing initial thaw, aliquot and freeze (-70°C).
      Packaging Information
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Catalogue Number GTIN
      475701 0

      Documentation

      MKK6, GST-Fusion, Human, Recombinant, E. coli SDS

      Title

      Safety Data Sheet (SDS) 

      MKK6, GST-Fusion, Human, Recombinant, E. coli Certificates of Analysis

      TitleLot Number
      475701

      References

      Reference overview
      Chabaud-Riou, M. and Firestein, G.S. et al. 2004. Am. J. Pathol. 164, 177.
      Zhu, X., et al. 2001. J. Neurochem. 79, 311.
      Data Sheet

      Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

      Revision08-August-2008 RFH
      SynonymsMEK6, Mitogen Activated Protein Kinase kinase 6, SAPKK3
      Application Data
      Increasing amounts of MKK6 were subjected to SDS-PAGE and the purity analyzed by Coomassie Blue staining.

      Increasing amounts of MKK6 were used to activate 10 µg p38α/SAPK2a and the activity of p38α/SAPK2a was measured as outlined in the protocols above.
      DescriptionRecombinant, human MKK6 fused to GST at the N-terminus and expressed in E. coli. MKK6 is a member of the dual specificity protein kinase family, which functions as a mitogen-activated protein (MAP) kinase kinase. It is suitable for labeling MAP kinase substrates.
      FormulationIn 270 mM sucrose, 150 mM NaCl, 50 mM Tris-HCl, 1 mM DTT, 0.1 mM EGTA, pH 7.5.
      Concentration Label Please refer to vial label for lot-specific concentration
      Recommended reaction conditions
      Kinase Activity Assay Activation of p38α/SAPK2a Required Materials •Assay Buffer: 25 mM β-glycerophosphate, 20 mM Tris-HCl, 5 mM EGTA, 1 mM sodium orthovanadate, 1 mM DTT, pH 7.5 • p38&alpha/SPK2a (substrate): in 300 mM NaCl, 50 mM Tris-HCl, 2 mM Dtt, pH 8.5 •MKK6: 100-fold lower concentration than p38α/SAPK2, diluted in Assay Buffer •Magnesium/ATP Mix: 75 mM MgCl2, 500 µM ATP Activation Protocol 1. Determine the reaction volume based on the concentration of p38α/SAPK2a and MKK6. 2. Add 1/8 volume Assay Buffer to a clean microcentrifuge tube on ice. 3. Add 3/8 volume H2O. 4. Add 1/8 volume MKK6. 5. Add 1/8 volume p38α/SAPK2a. 6. Add 1/4 volume Magnesium/ATP Mix. 7. Incubate at 30°C for 1 h. 8. Stop the reaction by placing the tubes on ice. 9. Assay the p38α/SAPK2a activity using the protocol outlined below. Note: The reaction may be dialyzed against 300 mM NaCl, 50 mM Tris-HCl, 2 mM DTT, pH 8.5, diluted with an equal volume of 99% glycerol, and stored at -70°C until use. Activity Assay Required Materials •Assay Buffer: 25 mM β-glycerophosphate, 20 mM Tris-HCl, 5 mM EGTA, 1 mM sodium orthovanadate, 1 mM DTT, pH 7.5 •Substrate: myelin basic protein, 2 mg/ml •p38α/SAPK2a, activated: 2.5-10 ng/µl, activated by MKK6 (above), diluted in Assay Buffer just prior to use •Magnesium/ATP Mix: 75 mM MgCl2, 500 µM ATP •Diluted [γ32P]ATP/Magnesium/ATP Mix: 197 µl Magnesium/ATP Mix, 3 µl (30 Ci) [γ32P]ATP (3000 Ci/mmol) Activity Protocol 1. Add 2.5 µl Assay Buffer to a clean microcentrifuge tube on ice. 2. Add 10 µl Substrate. 3. Add 2.5 µl p38α/SAPK2a (25-100 ng/assay). 4. Add 5 µl Diluted [γ32P]ATP/Magnesium/ATP Mix. 5. Incubate at 30°C for 20 min. 6. Stop the reaction by placing the tubes on ice. 7. Spot 10 µl of each reaction on P81 paper and allow to bind for 30 s. 8. Wash by immersing the paper in 0.75% phosphoric acid and gently shaking. Repeat for a total of 3 washes. 9. Wash as in step 8 using acetone. 10. Dry using an infrared light. 11. Determine the cpm using a scintillation counter or imager.
      Purity≥ 90% by SDS PAGE
      Specific activity≥ 50,000 units/mg protein
      Unit definitionOne unit is defined as the amount of enzyme that will activate p38alpha/SAPK2α (0.5 mg/ml) by 1 unit/min using 125 µM ATP at 30°C. One unit of p38α/SAPK2α activity is defined as the amount of enzyme that will transfer 1 pmol phosphate to myelin basic protein per min at 30°C.
      Storage Avoid freeze/thaw
      ≤ -70°C
      Do Not Freeze Ok to freeze
      Special InstructionsFollowing initial thaw, aliquot and freeze (-70°C).
      Toxicity Standard Handling
      ReferencesChabaud-Riou, M. and Firestein, G.S. et al. 2004. Am. J. Pathol. 164, 177.
      Zhu, X., et al. 2001. J. Neurochem. 79, 311.