Recombinant Human Active ERK1 Protein, CF

R&D Systems | Catalog # 1879-KS

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Key Product Details

  • R&D Systems E. coli-derived Recombinant Human Active ERK1 Protein (1879-KS)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

E. coli

Accession Number

NM_002746

Applications

Bioactivity
View all ERK1 Proteins and Enzymes »

Product Specifications

Source

E. coli-derived human ERK1 protein

Purity

>80%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

N-terminal Sequence Analysis

Activated by active MEK1 in vitro

Predicted Molecular Mass

44 kDa

Activity

The specific activity of ERK1 was determined to be 42 nmol/min/mg using a myelin basic protein (MBP) substrate and was equivalent to 382 nmol/min/mg as per radiometric assay.

Scientific Data Images for Recombinant Human Active ERK1 Protein, CF

Recombinant Human Active ERK1 Protein SDS-PAGE

Recombinant Human Active ERK1 Protein SDS-PAGE.

The approximate molecular weight is 44 kDa and the purity is > 80%.

Formulation, Preparation, and Storage

1879-KS
Formulation Supplied in 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.25 mM DTT, 0.1 mM PMSF and 25% glycerol.
Shipping The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage This product is stable at ≤ ‑70°C for up to 1 year from the date of receipt. For optimal storage, aliquot into smaller quantities after centrifugation and store at recommended temperature. Avoid repeated freeze-thaw cycles.

Background: ERK1

ERK1 is a protein Serine/Threonine kinase that is a member of the extracellular signal-regulated kinases (ERKs) which are activated in response to numerous growth factors and cytokines (1). Activation of ERK1 requires both tyrosine and threonine phosphorylation that is mediated by MEK. ERK1 is ubiquitously distributed in tissues with the highest expression in heart, brain, and spinal cord. Activated ERK1 translocates into the nucleus where it phosphorylates various transcription factors.

References

  1. Boulton, T.G. et al. (1991) Biochemistry 30:278.

Long Name

Extracellular Signal-regulated Kinase 1

Alternate Names

MAPK3, P44ERK1, p44mapk, PRKM3

Entrez Gene IDs

5595 (Human); 26417 (Mouse); 50689 (Rat)

Gene Symbol

MAPK3

UniProt

NM_002746 (Human)

Additional ERK1 Products

Product Documents for Recombinant Human Active ERK1 Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Recombinant Human Active ERK1 Protein, CF

For research use only

Citations for Recombinant Human Active ERK1 Protein, CF

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Protocols

View specific protocols for Recombinant Human Active ERK1 Protein, CF (1879-KS):

Materials
  • Active Kinase - Active ERK1 (0.1 μg/μL) diluted with Kinase Dilution Buffer and assayed as outlined in Sample Activity Plot. Note: These are suggested working dilutions. Optimal dilutions should be determined by each laboratory for each application.
  • Kinase Assay Buffer III - 200 mM Tris-HCl, pH 7.4, 100 mM MgCl2, and 0.5 mg/mL BSA. Add fresh DTT prior to use to a final concentration of 250 μM.
  • Kinase Dilution Buffer IX (1x) - Kinase Assay Buffer I diluted at a 1:4 ratio (5-fold dilution) with cold distilled water. Add fresh DTT prior to use to a final concentration of 50 μM.
  • ADP-GloTM Kinase Assay Kit - 10 mM ATP Solution, 10 mM ADP Solution, ADP-GloTM Reagent, Kinase Detection Reagent.
  • Substrate - Myeline basic protein (MBP) diluted in 100 mM MOPS buffer (pH 6.5) to a final concentration of 0.5 mg/mL.
  1. Thaw the Active ERK1, Kinase Assay Buffer III (5x), and Substrate on ice. Prepare a 15 μL enzyme dilution using Kinase Dilution Buffer IX (1x), at the desired concentration, in a pre-chilled 96-well plate.
  2. Prepare a substrate/ATP mixture as follows (25 μM ATP example)
    a. 10 mM ATP Solution: 1 μL
    b. Kinase Assay Buffer III (5x): 79 μL
    c. Substrate at 0.5 mg/mL: 80 μL
  3. Transfer the following reaction components prepared in Step 1 and 2 to a 384-well opaque plate bringing the reaction volume up to 5 μL:
    a. 3 μL of diluted Active ERK1
    b. 2 μL of Substrate/ATP mix as prepared in the Step 2. This initiates the reaction.
  4. Set up the blank control as outlined in step 2, excluding the addition of the kinase. Replace the kinase with an equal volume of Kinase Dilution Buffer IX (1x).
  5. Incubate at ambient temperature for 40 minutes.
  6. After the 40 minute incubation period, terminate the reaction and deplete the remaining ATP by adding 5 μL of ADP-Glo Reagent.  Spin down and shake the 384-well plate.  Then incubate the reaction mixture for another 40 minutes at ambient temperature.
  7. Add 10 μL of the Kinase Detection Reagent to the 384-well plate and incubate the reaction mixture for another 30 minutes at ambient temperature
  8. Read the 384-well reaction plate using the Luminescence Module Protocol on a GloMax®-Multi Microplate Multimode Reader.
  9. Determine the corrected activity (RLU) by removing the blank control value (see step 4) for each sample and calculate the kinase specific activity as outlined below.


    Calculation of Specific Activity of ADP (RLU/pmol)
    From ATP-ADP conversion curve, determine RLU/pmol of ADP

    Kinase Specific Activity (SA) (pmol/minutes/μg or nmol/minutes/mg)
    Corrected RLU from reaction / [(SA of ADP in RLU/pmol) x (Reaction time in minutes) x (Enzyme amount in μg or mg)]

FAQs for Recombinant Human Active ERK1 Protein, CF

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  • Q: What is the activation protocol for this enzyme?

    A: This enzyme is supplied in its activated form, so no activation step is required.

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