Recombinant Mouse Cathepsin C/DPPI Protein, CF Summary
Product Specifications
Asp25-Leu462, with a C-terminal 10-His tag
Accession # P97821
Analysis
Product Datasheets
Carrier Free
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
1034-CY
| Formulation | Supplied as a 0.2 μm filtered solution in MES 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.
|
Assay Procedure
- Activation Buffer: 25 mM MES, 5 mM DTT, pH 5.5
- Assay Buffer: 25 mM MES, 50 mM NaCl, 5 mM DTT, pH 6.0
- Recombinant Mouse Cathepsin C/DPPI (rmCathepsin C) (Catalog # 1034-CY)
- Recombinant Human Cathepsin L (rhCathepsin L) (Catalog # 952-CY)
- Substrate: Gly-Arg-AMC (Bachem, Catalog # I-1215)
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rmCathepsin C to 20 µg/mL in Activation Buffer.
- Dilute rhCathepsin L to 2 µg/mL in Activation Buffer.
- Combine equal volumes of diluted rmCathepsin C and diluted rhCathepsin L for final concentrations of 10 µg/mL rmCathepsin C and 1 µg/mL for rhCathepsin L.
- Incubate at room temperature for 20 minutes to activate rmCathepsin C.
- Dilute activated rmCathepsin C to 0.04 ng/µL in Assay Buffer.
- Dilute Substrate to 20 µM in Assay Buffer.
- Load 50 µL of the 0.04 ng/µL rmCathepsin C in a plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 20 µM Substrate.
- Read at excitation and emission wavelengths of 380 nm and 460 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 7-Amino, 4-Methyl Coumarin (Sigma, Catalog # A9891).
- rmCathepsin C: 0.002 µg
- Substrate: 10 µM
Reconstitution Calculator
Background: Cathepsin C/DPPI
Cathepsin C is a cysteine protease of the papain family (1). Cathepsin C sequentially removes dipeptides from the free N-termini of proteins and peptides. It has broad specificity except that it does not cleave a basic amino acid (Arg or Lys) in the N-terminal position or Pro on either side of the scissle bond. It requires halide ions for activity. The pro form contains a pro peptide and a catalytic region, which can be further processed into heavy/ alpha and light/ beta chains that are linked by a disulfide bond. It is broadly distributed. Cathepsin C plays a role in the lysosomal degradation. It also functions as a key enzyme in the activation of granule serine proteases in cytotoxic T lymphocytes and natural killer cells (granzymes A and B), mast cells (tryptase and chymase), and neutrophils (Cathepsin G and elastase) by removing their N-terminal activation dipeptides (2).
- Turk, B. et al. (2004) in Handbook of Proteolytic Enzymes. Barrett, et al. eds. p. 1192, Academic Press, San Diego.
- Dahl, S.W. et al. (2001) Biochemistry 40:1671.
Citation for Recombinant Mouse Cathepsin C/DPPI Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
1 Citation: Showing 1 - 1
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Coxiella burnetii-infected NK cells release infectious bacteria by degranulation
Authors: S Matthiesen, L Zaeck, K Franzke, R Jahnke, C Fricke, M Mauermeir, S Finke, A Lührmann, MR Knittler
Infect. Immun., 2020-10-19;0(0):.
Species: Bacteria
Sample Types: Whole Cells
Applications: Bioassay
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