Cell Division Cycle 25B (Cdc25B) phosphatase removes inorganic phosphate groups covalently attached to tyrosine, serine and threonine residues in proteins (1). Breast cancer patients bearing tumors containing high levels of Cdc25B have been found to have a greater incidence of aggressive, high‑grade tumors than those with low Cdc25B levels (2). In cells, the levels of Cdc25B activity are highest during the G2/M transition of the cell cycle, where it is suspected to be involved in “checkpoint” control of cell cycle progression (3). Overexpression of Cdc25B reduces the G2/M cell cycle block caused by ionizing radiation (4). Although activated by phosphorylation, Ser323 phosphorylation causes the enzyme to bind the protein 14-3-3, preventing substrate access to the catalytic site (5). One of the major substrates of Cdc25B is Cdc2, a kinase that is activated by dephosphorylation (6). The recombinant protein is truncated to remove the N-terminal regulatory domains and is fully active.
Recombinant Human CDC25B (aa 377-566) Protein, CF
R&D Systems | Catalog # 1649-CD
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Key Product Details
- R&D Systems E. coli-derived Recombinant Human CDC25B (aa 377-566) Protein (1649-CD)
- 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
Structure / Form
Based on mass spectrometric analysis, a carboxyl-terminal truncated peptide with a molecular mass of approximately 22.1 kDa may also be present in the recombinant protein preparation.
Applications
Enzyme Activity
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Product Specifications
Source
E. coli-derived human CDC25B protein
Glu377-Gln566, with an N-terminal Met and 6-His tag
Glu377-Gln566, with an N-terminal Met and 6-His tag
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
Met
Predicted Molecular Mass
23 kDa
SDS-PAGE
23-25 kDa, reducing conditions
Activity
Measured by its ability to cleave a substrate, p-Nitrophenyl phosphate (pNPP).
The specific activity is >70 pmol/min/μg, as measured under the described conditions.
The specific activity is >70 pmol/min/μg, as measured under the described conditions.
Formulation, Preparation, and Storage
1649-CD
| Formulation | Supplied as a 0.2 μm filtered solution in Tris, NaCl, EDTA, DTT and Glycerol. |
| Shipping | The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Background: CDC25B
References
- Draetta, G. and J. Eckstein (1997) Biochim. Biophys. Acta 1332:M53.
- Galaktionov, K. et al. (1995) Science 269:1575.
- Lammer, C. et al. (1998) J. Cell Sci. 111:2445.
- Miyata, H. et al. (2001) Cancer Res. 61:3188.
- Forrest, A. and B. Gabrielli (2001) Oncogene 20:4393.
- Gautier, J. et al. (1991) Cell 67:197.
Long Name
Cell Division Cycle 25B
Alternate Names
CDC25HU2, cell division cycle 25 homolog B (S. cerevisiae), cell division cycle 25 homolog B (S. pombe), cell division cycle 25B, Dual specificity phosphatase Cdc25B, EC 3.1.3.48, M-phase inducer phosphatase 2
Entrez Gene IDs
994 (Human)
Gene Symbol
CDC25B
UniProt
Additional CDC25B Products
Product Documents for Recombinant Human CDC25B (aa 377-566) 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 CDC25B (aa 377-566) Protein, CF
For research use only
Related Research Areas
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Protocols
View specific protocols for Recombinant Human CDC25B (aa 377-566) Protein, CF (1649-CD):
Materials
- Assay Buffer: 50 mM HEPES, 10 mM DTT, pH 7.5
- Recombinant Human CDC25B aa 377‑566 (rhCDC-25B) (Catalog # 1649-CD)
- Substrate: p-Nitrophenyl phosphate (pNPP), 10 mM stock in deionized water
- Sodium Hydroxide (NaOH), 0.2 M stock in deionized water
- Clear 96-well Plate (Catalog # DY990)
- Plate Reader with absorbance read capability
- Dilute rhCDC-25B to 20 µg/mL in Assay Buffer.
- Dilute Substrate to 6 mM in Assay Buffer.
- Prepare reaction mixtures by combining equal volumes of dilute rhCDC-25B and dilute Substrate in microtubes. Include an Enzyme Control by combining dilute rhCDC-25B with twice the volume of 0.2 M NaOH, mix briefly; then add a volume of dilute Substrate equal to the volume of dilute rhCDC-25B. The Enzyme Control will have 2x the volume of the reaction mixture.
- Incubate Reactions and Enzyme Control at 37 °C for 4 hours.
- Load 100 µL of Reactions into a plate and stop the reactions by adding 100 µL 0.2 M NaOH.
- Load 200 µL of Enzyme Controls into plate.
- Read plate at 410 nm (absorbance) in endpoint mode.
- Calculate specific activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted Abs* (OD) x Conversion Factor** (pmol/OD) |
| Incubation time (min) x amount of enzyme (µg) |
*Adjusted for Enzyme Controls
**Derived using calibration standard p-Nitrophenol
Per Well:
- rhCDC-25B: 1 µg
- pNPP: 1.5 mM
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