PLK1 is a member of the Polo-Like Kinase family that localizes to centrosomes or spindle pole bodies and undergoes dramatic subcellular relocation during the cell cycle. Deregulated activities of PLKs often result in abnormalities in centrosome duplication, maturation, and/or microtubule dynamics (1). PLKs also regulate the function of the Golgi complex. Deregulated expression of human PLK1 is strongly correlated with the development of many types of malignancies, and ectopic expression of PLK1 dominant negative protein leads to rapid cell death (2).
Recombinant Human Active PLK1 Protein, CF
R&D Systems | Catalog # 3804-KS
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Key Product Details
- R&D Systems Sf 9 (baculovirus)-derived Recombinant Human Active PLK1 Protein (3804-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
Sf 9 (baculovirus)
Accession Number
Applications
Bioactivity
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Product Specifications
Source
Spodoptera frugiperda, Sf 9 (baculovirus)-derived human PLK1 protein
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
N-terminal Sequence Analysis
Using an N-terminal His tag
SDS-PAGE
70 kDa
Activity
The specific activity of PLK1 was equivalent to 42 nmol/min/mg as per Activity Assay Protocol and was equivalent to 17 nmol/min/mg as per radiometric assay.
Scientific Data Images for Recombinant Human Active PLK1 Protein, CF
Recombinant Human Active PLK1 Protein SDS-PAGE.
The approximate molecular weight is 70 kDa and the purity is > 90%.Formulation, Preparation, and Storage
3804-KS
| Formulation | Supplied in 50 mM Sodium Phosphate (pH 7.0), 300 mM NaCl, 150 mM Imidazole, 0.1 mM PMSF, 0.25 mM DTT, 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: PLK1
References
- Nigg, E.A. et al. (1996) Exp. Cell Res. 229:174.
- Dai, W. et al. (2003) Prog. Cell Cycle Res. 5:327.
Long Name
Polo-like Kinase 1
Alternate Names
STPK13
Gene Symbol
PLK1
UniProt
Additional PLK1 Products
Product Documents for Recombinant Human Active PLK1 Protein, CF
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Human Active PLK1 Protein, CF
For research use only
Related Research Areas
Citations for Recombinant Human Active PLK1 Protein, CF
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Protocols
View specific protocols for Recombinant Human Active PLK1 Protein, CF (3804-KS):
Materials
- Active Kinase - Active PLK1 (0.1 μg/μL) diluted with Kinase Dilution Buffer IX and assayed as outlined in Sample Activity Plot. Note: These are suggested working dilutions and it is recommended that the researcher perform a serial dilution of active PLK1 for optimal results.
- Kinase Assay Buffer III (5X) - 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 III diluted at a 1:4 ratio (5X dilution) with cold distilled or deionized water. Add fresh DTT to the aliquot prior to use to a final concentration of 50 μM.
- ADP-Glo™ Kinase Assay Kit - 10 mM ATP Solution, 10 mM ADP Solution, ADP-Glo™ Reagent, and Kinase Detection Reagent.
- Substrate - PLKtide peptide substrate was diluted in 20 mM Tris-HCl, pH 7.5 solution to a final concentration of 1 mg/mL.
- Thaw the Active PLK1, Kinase Assay Buffer III (5X), and Substrate on ice. Prepare a 15 μL enzyme dilution at the desired concentration, with Kinase Dilution Buffer IX (1X), in a pre-chilled 96-well plate.
- Prepare a Substrate/ATP
mixture as follows (25 μM example):
a. 10 mM ATP Solution: 1 μL
b. Kinase Assay Buffer III (5X): 79 μL
c. Substrate at 1 mg/mL: 80 μL - Transfer the following reaction components prepared in Step 2
to a 384-well opaque plate, bringing the reaction volume up to 5
μL:
a. 3 μL of diluted Active PLK1
b. 2 μL of Substrate/ATP mix as prepared in Step 2. This initiates the reaction. - 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).
- Incubate at ambient temperature for 40 minutes.
- 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.
- 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.
- Read the 384-well reaction plate using the Luminescence Module Protocol on a GloMax®-Multi Microplate Multimode Reader.
- 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 ADP standard curve, determine RLU/pmol of ADP
Kinase Specific Activity (SA) (pmol/min/μg or nmol/min/mg)
Corrected RLU from reaction / [(SA of ADP in RLU/pmol) x (Reaction time in min) x (Enzyme amount in μg or mg)]
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