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475702 MEK1, GST-Fusion, Human, Recombinant, E. coli

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

Synonyms: MAP Kinase Kinase, ERK Kinase 1, Microtubule-associated protein 2 kinase

475702
  
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Overview

Replacement Information
Description
Overview

This product has been discontinued.



Recombinant, human MEK1 fused to GST at the N-terminus and expressed in E. coli. MEK1 is a member of a dual-specificity protein kinase family that phosphorylates threonine and tyrosine residues on ERK 1 and 2.
Catalogue Number475702
Brand Family Calbiochem®
SynonymsMAP Kinase Kinase, ERK Kinase 1, Microtubule-associated protein 2 kinase
Application Data
Increasing amounts of MEK1 were subjected to SDS-PAGE and the purity analyzed by Coomassie Blue staining.

Increasing amounts of MEK1 were used to activate 10 µg ERK1 and the activity of ERK1 was analyzed as outlined in the activation and activity protocols above.
References
ReferencesAllen, L.F., et al. 2003. Semin. Oncol. 30, 105.
Schramek, H., et al. 1997. J. Biol. Chem. 272, 11426.
Alessandrini, A., et al. 1996. J. Biol. Chem. 271, 31612.
Product Information
Unit of DefinitionOne unit is defined as the amount of enzyme that activates ERK1 (0.3 mg/ml) by 1 unit/min using 100 µM ATP at 30°C. One unit ERK1 activity is defined as that amount of enzyme required to transfer 1 pmol phosphate to myelin basic protein, in the presence of 125 µM ATP, per min at 30°C.
FormLiquid
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≥80% by SDS PAGE
Specific Activity≥ 250,000 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
475702 0

Documentation

MEK1, GST-Fusion, Human, Recombinant, E. coli SDS

Title

Safety Data Sheet (SDS) 

MEK1, GST-Fusion, Human, Recombinant, E. coli Certificates of Analysis

TitleLot Number
475702

References

Reference overview
Allen, L.F., et al. 2003. Semin. Oncol. 30, 105.
Schramek, H., et al. 1997. J. Biol. Chem. 272, 11426.
Alessandrini, A., et al. 1996. J. Biol. Chem. 271, 31612.
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.

Revision02-January-2008 JSW
SynonymsMAP Kinase Kinase, ERK Kinase 1, Microtubule-associated protein 2 kinase
Application Data
Increasing amounts of MEK1 were subjected to SDS-PAGE and the purity analyzed by Coomassie Blue staining.

Increasing amounts of MEK1 were used to activate 10 µg ERK1 and the activity of ERK1 was analyzed as outlined in the activation and activity protocols above.
DescriptionRecombinant, human MEK1 fused to GST at the N-terminus and expressed in E. coli. MEK1 is a member of a dual-specificity protein kinase family that phosphorylates threonine and tyrosine residues on ERK 1 and 2.
FormLiquid
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 ERK1 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 •ERK1 Dilution Buffer: 150 mM NaCl, 50 mM Tris-HCl, 2 mM DTT, pH 8.0 •ERK1 (substrate): in 150 mM NaCl, 50 mM Tris-HCl, 5 mM EDTA, 2 mM DTT, pH 8.0 •MEK1: 10-fold lower concentration that ERK1, 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 ERK1 to be activated. 2. Add 1/5 volume Assay Buffer to a clean microcentrifuge tube on ice. 3. Add ERK1 to a final concentration of 300 µg/ml. 4. Add MEK1 at a concentration that is 1/10 that of ERK1. 5. Add ERK1 Dilution Buffer up to 4/5 volume. 6. Add 1/5 volume Magnesium/ATP Mix. 7. Incubate at 30°C for 30 min. 8. Stop the reaction by placing the tubes on ice. 9. Assay the ERK1 activity using the protocol outlined below. Note: The reaction may be dialyzed against 300 mM NaCl, 100 mM Tris-HCl, 2 mM DTT, pH 8.5, diluted with an equal volume of 99% glycerol, and stored at -70°C until use. This activation protocol can also be used for ERK2. 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 •ERK1, activated: 2.5-10 ng/µl, activated by MEK1 (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 10 µl Assay Buffer to a clean microcentrifuge tube on ice. 2. Add 10 µl Substrate. 3. Add 2.5 µl ERK1 (25-100 ng/assay). 4. Add 10 µl Diluted [γ32P]ATP/Magnesium/ATP Mix. 5. Incubate at 30°C for 10 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≥80% by SDS PAGE
Specific activity≥ 250,000 units/mg protein
Unit definitionOne unit is defined as the amount of enzyme that activates ERK1 (0.3 mg/ml) by 1 unit/min using 100 µM ATP at 30°C. One unit ERK1 activity is defined as that amount of enzyme required to transfer 1 pmol phosphate to myelin basic protein, in the presence of 125 µM ATP, 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
ReferencesAllen, L.F., et al. 2003. Semin. Oncol. 30, 105.
Schramek, H., et al. 1997. J. Biol. Chem. 272, 11426.
Alessandrini, A., et al. 1996. J. Biol. Chem. 271, 31612.