Tryptases are trypsin-like serine proteases, and beta tryptases appear to be the main isoenzymes expressed in mast cells (1). Mast Cell Protease-6 (MCP-6, Mcpt6, or Tpsb2) is one of beta tryptases produced from mouse mast cells. It is stored in secretory granules of mast cells, where it forms active tetramers with heparin proteoglycan. Because of the unique arrangement of the active sites in the tetramer, which are facing a narrow central pore, MCP-6 is resistant to macromolecular protease inhibitors (2). When mast cells are activated, MCP-6 is released along with other proteins in secretory granules, participating in provoking inflammatory conditions (3). MCP-6 has been implicated as a mediator in the pathogenesis of asthma and other allergic disorders in mouse models.
Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF
R&D Systems | Catalog # 3736-SE
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Key Product Details
- R&D Systems NS0-derived Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein (3736-SE)
- 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
Accession Number
Structure / Form
Applications
Product Specifications
Source
Ala22-Ser276, with a C-terminal 10-His tag
Purity
Endotoxin Level
N-terminal Sequence Analysis
Predicted Molecular Mass
SDS-PAGE
Activity
Measured by its ability to cleave the fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002).
The specific activity is >500 pmol/min/µg, as measured under the described conditions.
Reviewed Applications
Read 1 review rated 5 using 3736-SE in the following applications:
Formulation, Preparation, and Storage
3736-SE
| Formulation | Supplied as a 0.2 μm filtered solution in Tris and NaCl. |
| 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: Mast Cell Protease-6/Mcpt6
References
- Caughey, G. H. (2004) in Handbook of Proteolytic Enzymes, Barrett, A.J. et al. eds. p. 1535 .
- Sommerhoff, C.P. et al. (1999) Proc. Natl. Acad. Sci. USA. 96:10984.
- Hallgren, J. and G. Pejler, (2006) FEBS J. 273:1871.
Alternate Names
Gene Symbol
UniProt
Additional Mast Cell Protease-6/Mcpt6 Products
Product Documents for Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF
Certificate of Analysis
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Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF
For research use only
Related Research Areas
Citations for Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF
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Protocols
View specific protocols for Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF (3736-SE):
- Maturation Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
- Assay Buffer: 0.05 M MES, 0.1 M NaCl, pH 6.5
- Recombinant Mouse Mast Cell Protease‑6/Mcpt6 (rmMCP-6) (Catalog # 3736-SE)
- Substrate: MCA-Arg-Pro-Lys-Pro-Val-Glu-NVAL-Trp-Arg-Lys(DNP)-NH2 (Catalog # ES002), 2 mM stock in DMSO
- Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
- Heparin (Sigma, Catalog # H3393), 20 mg/mL deionized water
- 1,10-Phenanthroline (Sigma, Catalog # 320056), 0.6 M stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rmMCP-6 to 200 µg/mL in Maturation Buffer.
- Dilute Thermolysin to 0.2 µg/mL in Maturation Buffer.
- Combine 25 µL of 200 µg/mL rmMCP-6 with 25 µL of 0.2 µg/mL Thermolysin.
- Incubate at room temperature for 15 minutes.
- Dilute 1,10-Phenanthroline to 10 mM in Assay Buffer.
- Stop maturation by adding 50 µL of 10 mM 1,10-phenanthroline to reactions.
- Dilute Heparin to 1 mg/mL in Assay Buffer.
- Add 50 µL of Heparin and 350 µL of Assay Buffer to the stopped reaction vials.
- Incubate for 30 minutes at room temperature.
- Dilute reaction vials to a final enzyme concentration of 2 ng/µL in Assay Buffer.
- Dilute Substrate to 20 µM in Assay Buffer.
- Load 50 µL of 2 ng/µL rmMCP-6 mixture into wells of a plate and start the reaction by adding 50 µL of Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of Substrate.
- Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
- Calculate Specific Activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU) |
| amount of enzyme (µg) |
*Adjusted for Substrate Blank
**Derived using calibration standard MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).
Per Well:
- rmMCP-6: 0.100 µg
- Substrate: 10 µM
FAQs for Recombinant Mouse Mast Cell Protease-6/Mcpt6 Protein, CF
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Q: After the maturation step, how long is Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) active? After maturation, can the product be stored, or must it be used immediately?
A: Stability of Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) has only been evaluated prior to maturation and tested for activity as outlined in the Assay Procedure section of the product-specific page. We anticipate end-users use the product immediately after maturation. End-users will need to evaluate stability of the matured enzyme under long-term storage conditions to determine if there are any deletarious effect on their application.
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Q: Is Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3536-SE) supplied as the mature form?
A: Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) requires a maturation step involving Thermolysin prior to measuring activity using our routine in vitro testing protocol. For more details, please see the Assay Procedure step on the product specific page.
-
Q: After the maturation step, how long is Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) active? After maturation, can the product be stored, or must it be used immediately?
A: Stability of Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) has only been evaluated prior to maturation and tested for activity as outlined in the Assay Procedure section of the product-specific page. We anticipate end-users use the product immediately after maturation. End-users will need to evaluate stability of the matured enzyme under long-term storage conditions to determine if there are any deletarious effect on their application.
-
Q: Is Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3536-SE) supplied as the mature form?
A: Recombinant Mouse Mast Cell Protease-6/Mcpt6 (Catalog # 3736-SE) requires a maturation step involving Thermolysin prior to measuring activity using our routine in vitro testing protocol. For more details, please see the Assay Procedure step on the product specific page.