Recombinant Human Chymase/CMA1 Protein, CF

R&D Systems | Catalog # 4099-SE

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Key Product Details

  • R&D Systems NS0-derived Recombinant Human Chymase/CMA1 Protein (4099-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

NS0

Accession Number

P23946

Structure / Form

Pro form

Applications

Enzyme Activity
View all Chymase/CMA1 Proteins and Enzymes »

Product Specifications

Source

Mouse myeloma cell line, NS0-derived human Chymase/CMA1 protein
Met1-Asn247, with a C-terminal 10-His tag

Purity

>90%, by SDS-PAGE under reducing conditions and visualized by silver stain.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Gly20

Predicted Molecular Mass

27 kDa

SDS-PAGE

38 kDa, reducing conditions

Activity

Measured by its ability to cleave the fluorogenic peptide substrate, SUC-Ala-Ala-Pro-Phe-AMC.
The specific activity is >80 pmol/min/µg, as measured under the described conditions. 

Formulation, Preparation, and Storage

4099-SE
Formulation Supplied as a 0.2 μm filtered solution in HEPES 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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Background: Chymase/CMA1

Chymases are a group of chymotrypsin-like serine proteases secreted by mast cells (1). They are synthesized as inactive precursors containing a 2-residue propeptide, which needs to be removed by dipeptidyl peptidase I/cathepsin C for the enzymatic activity (2). Human Chymase encoded by the CMA1 gene is known to be involved in hypertention and heart failure through its ability to convert angiotensin I (Ang I) to angiotensin II (Ang II), which plays a key role in the regulation of arterial pressure (3). In addition, it is also important in physiological and pathological conditions including inflammation, fibrosis and processing of cytokines (4). Therefore, designing a specific inhibitor for Chymase activity has been a pharmacologic strategy to develop therapeutic agents.

References

  1. Caughey, G.H. (2004) in Handbook of Proteolytic Enzymes. Barrett, A.J. et al. ed. p. 1531, Academic Press, San Diego.
  2. Murakami, M. et al. (1995) J. Biol. Chem. 270:2218.
  3. Miyazaki, M. and S. Takai (2006) J. Pharmacol. Sci. 100:391.
  4. Nakajima, M. and N. Naya (2002) Jpn. J. Pharmacol. 90:206.

Alternate Names

CMA1, CYH, MCP3P, Mcpt5, MCT1

Entrez Gene IDs

1215 (Human); 17228 (Mouse); 25627 (Rat)

Gene Symbol

CMA1

UniProt

P23946

Additional Chymase/CMA1 Products

Product Documents for Recombinant Human Chymase/CMA1 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 Chymase/CMA1 Protein, CF

Triton is a registered trademark of Union Carbide Corp.

For research use only

Citations for Recombinant Human Chymase/CMA1 Protein, CF

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Protocols

View specific protocols for Recombinant Human Chymase/CMA1 Protein, CF (4099-SE):

Materials
  • Maturation Buffer: 50 mM MES, pH 5.5
  • Cathepsin Buffer: 50 mM MES, 50 mM NaCl, 5 mM DTT, pH 5.5
  • Assay Buffer: 20 mM Tris, 2 M KCl, 0.02% (v/v) Triton® X-100, pH 9.0
  • Recombinant Human Chymase/CMA1 (rhChymase) (Catalog # 4099-SE)
  • Recombinant Mouse Active Cathepsin C/DPPI (rmCathepsin C) (Catalog # 2336-CY)
  • Heparin (Sigma, Catalog # H3393), 20 mg/mL stock in deionized water
  • N-Ethylmaleimide (NEM) (Sigma, Catalog # E1271), 50 mM stock in deionized water
  • Substrate: SUC-Ala-Ala-Pro-Phe-AMC (Bachem, Catalog # I-1465), 10 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhChymase to 20 µg/mL in Maturation Buffer.
  2. Dilute rmCathepsin C to 20 µg/mL in Cathepsin Buffer.
  3. Dilute Heparin to 1.5 mg/mL in deionized water.
  4. Combine 15 µL of 20 µg/mL rhChymase and 15 µL of 20 µg/mL rmCathepsin C.
  5. Add 1 µL of Heparin and mix well.
  6. Incubate at room temperature for 1 hour.
  7. Stop maturation by adding 1.98 µL of 50 mM stock of NEM for a final concentration of 3 mM.
  8. Dilute rhChymase to 2 µg/mL in Assay Buffer.
  9. Incubate at room temperature for 5 minutes.
  10. Dilute Substrate to 200 µM in Assay Buffer.
  11. Load 50 µL of 2 µg/mL rhChymase in a plate, and start the reaction by adding 50 µL of 200 µM Substrate. As a Substrate Blank combine 50 μL of Substrate with 50 μL of Assay Buffer.
  12. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in kinetic mode for 5 minutes.
  13. 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 7-amino, 4-Methyl Coumarin (Sigma, Catalog # A-9891).

Per Well:

  • rhChymase: 0.100 µg
  • Substrate: 100 µM

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