Recombinant Mouse Cathepsin H Protein, CF

R&D Systems | Catalog # 1013-CY

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Key Product Details

  • R&D Systems NS0-derived Recombinant Mouse Cathepsin H Protein (1013-CY)
  • 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

Q3UCD6

Structure / Form

Pro form

Applications

Enzyme Activity
View all Cathepsin H Proteins and Enzymes »

Product Specifications

Source

Mouse myeloma cell line, NS0-derived mouse Cathepsin H protein
Glu22-Val333, with a C-terminal 10-His tag

Purity

>95%, 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

Glu22

Predicted Molecular Mass

36 kDa

SDS-PAGE

40 kDa, reducing conditions

Activity

Measured by its ability to cleave the fluorogenic peptide substrate, Arg-7-amido-4-methylcoumarin (R-AMC).
The specific activity is >400 pmol/min/µg, as measured under the described conditions. See Activity Assay Protocol on www.RnDSystems.com

Formulation, Preparation, and Storage

1013-CY
Formulation Supplied as a 0.2 μm filtered solution in MES and NaCl.
Shipping The product is shipped with polar packs. 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: Cathepsin H

Cathepsin H is a lysosomal cysteine protease of the papain family (1). It is synthesized as a precursor protein, consisting of a signal peptide (residues 1‑20), a propeptide (residues 21‑95), a mini chain (residues 96‑103), a heavy chain (residues 114‑290) and a light chain (residues 291‑333) (2, 3). A truncated form with a 12 amino acid deletion in the signal peptide region is secreted (4). Cathepsin H is the only known mono-aminopeptidase in the papain family (5). Cathepsin H expression is significantly increased in disease states such as in prostate tumors, sera of asthmatic patients, and mucosa of colorectal cancer patients (4, 6, 7).

References

  1. Kirschke, H. (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, et al.) p. 1089, Academic Press, San Diego. 
  2. Lafuse, W.P. et al. (1995) J. Leukoc. Biol. 57:663.
  3. Soderstrom, M. et al. (1999) Biochim. Biophys. Acta 1446:35.
  4. Waghray, A. et al. (2002) J. Biol. Chem. 277:11533.
  5. Guncar, G. et al. (1998) Structure 6:51.
  6. Cimerman, N. et al. (2001) Clin. Chim. Acta 310:113.
  7. del Re, E.C. et al. (2000) Br. J. Cancer. 82:1317.

Alternate Names

CTSH

Entrez Gene IDs

1512 (Human); 13036 (Mouse)

Gene Symbol

CTSH

UniProt

Q3UCD6

Additional Cathepsin H Products

Product Documents for Recombinant Mouse Cathepsin H Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant Mouse Cathepsin H Protein, CF

For research use only

Citations for Recombinant Mouse Cathepsin H Protein, CF

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Protocols

View specific protocols for Recombinant Mouse Cathepsin H Protein, CF (1013-CY):

Materials
  • Activation Buffer: 50 mM MES, 10 mM CaCl2, 150 mM NaCl, pH 6.5
  • Assay Buffer: 50 mM MES, pH 6.5
  • Dithiothreitol (DTT) (Sigma, Catalog # D-0632), 1 M stock in deionized water
  • Recombinant Mouse Cathepsin H (rmCathepsin H) (Catalog # 1013-CY)
  • Bacterial Thermolysin (Catalog # 3097-ZN)
  • Phosphoramidon (Catalog # EI006), 20 mM in methanol
  • Substrate: Arg-7-amido-4-methylcoumarin (R-AMC) (Chem-Impex, Catalog # 5859), 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 rmCathepsin H to 200 µg/mL in Activation Buffer.
  2. Dilute Thermolysin to 100 µg/mL in Activation Buffer.
  3. Mix equal volumes of 100 µg/mL Thermolysin and 200 µg/mL rmCathepsin H.
  4. Incubate at 37 °C for 1 hour.
  5. Stop reaction by adding an equal volume of 2 mM Phosphoramidon in Assay Buffer to reaction mixture.
  6. Incubate at room temperature for 10 minutes.
  7. Then add an equal volume of Assay Buffer containing 20 mM DTT to reaction mixture. The concentration of rmCathepsin H is now 25 ng/µL.
  8. Incubate reaction at room temperature for 30 minutes.
  9. Dilute activated rmCathepsin H to 0.5 ng/µL in Assay Buffer.
  10. Dilute Substrate to 200 µM in Assay Buffer.
  11. In a plate load 50 µL of 0.5 ng/µL rmCathepsin H to wells, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 200 µM Substrate.
  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 (AMC) (Sigma, Catalog # A-9891)

Per Well:

  • rmCathepsin H: 0.025 µg
  • Substrate: 100 µM

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