Recombinant Human Cathepsin X/Z/P Protein, CF

Catalog # Availability Size / Price Qty
934-CY-010
R&D Systems Recombinant Proteins and Enzymes
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Recombinant Human Cathepsin X/Z/P Protein, CF Summary

Product Specifications

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.
Activity
Measured by its ability to cleave the fluorogenic peptide substrate, Mca-RPPGFSAFK(Dnp)-OH (Catalog # ES005). The specific activity is >800 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Cathepsin X/Z/P protein
Ala21-Val303 (Gly23Val & Ser48Thr), with a C-terminal 10-His tag
Accession #
N-terminal Sequence
Analysis
Ala21
Structure / Form
Pro form
Predicted Molecular Mass
33 kDa
SDS-PAGE
41 kDa, reducing conditions

Product Datasheets

Carrier Free

What does CF mean?

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.

What formulation is right for me?

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.

934-CY

Formulation Supplied as a 0.2 μm filtered solution in Sodium Acetate 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.

Assay Procedure

Materials
  • Activation Buffer: 25 mM Sodium Acetate, pH 3.5
  • Assay Buffer: 25 mM Sodium Acetate, 5 mM Dithiothreitol (DTT), pH 3.5
  • Dithiothreitol (DTT), 1 M stock in deionized water
  • Recombinant Human Cathepsin X/Z/P (rhCathepsin X/Z/P) (Catalog # 934-CY)
  • Substrate: MCA-Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH (Catalog # ES005)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Activate rhCathepsin X/Z/P.
    1. Dilute rhCathepsin X/Z/P to 20 µg/mL in Activation Buffer.
    2. Dilute DTT stock to 10 mM in Activation Buffer.
    3. Mix equal volumes of 20 µg/mL rhCathepsin X/Z/P and 10 mM DTT.
    4. Incubate at room temperature for 5 minutes.
  2. Dilute activated rhCathepsin X/Z/P to 0.4 ng/µL in Assay Buffer.
  3. Dilute Substrate to 20 µM in Assay Buffer.
  4. Load 50 µL of the 0.4 ng/µL rhCathepsin X/Z/P in a black well plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 20 µM Substrate without any rhCathepsin X/Z/P.
  5. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
  6. 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:
  • rhCathepsin X/Z/P: 0.02 µg
  • Substrate: 10 µM
Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: Cathepsin X/Z/P

Cathepsin X (also known as Cathepsin Z and P) is a cysteine protease of the papain family (1-5). Compared to other members of the papain family, Cathepsin X has a short proregion and unique insertions. The cysteine residue in the proregion forms a covalent and reversible bond with the active site cysteine residue (6). Acting as a carboxypeptidase, Cathepsin X displays a unique specificity (7-10). It is ubiquitously expressed in human tissues and conserved in other species such as mouse, nematode and echiuran. The nematode enzyme is apparently involved in molting of third stage larvae (11).

References
  1. Nagler, D.K. and R. Menard (1998) FEBS Lett. 434:135.
  2. Santamaria, I. et al. (1998) J. Biol. Chem. 273:16816.
  3. Deussing, J. et al. (2000) Biochim. Biophys. Acta 1491:93.
  4. Pungercar, J. and G. Ivanovski (2000) Pflugers Arch. Eur. J. Physiol. 439:R116.
  5. Pungercar, J. et al. (2000) Pflugers Arch. Eur. J. Physiol. 439:R119.
  6. Sivaraman, J. et al. (2000) J. Mol. Biol. 295:935.
  7. Menard, R. et al. (2001) Biol. Chem. 382:839.
  8. Therrien, C. et al. (2001) Biochemistry 40:2702.
  9. Klemencic, I. et al. (2000) Eur. J. Biochem. 267:5404.
  10. Guncar, G. et al. (2000) Structure Fold Des. 8:305.
  11. Lustigman, S. et al. (1996) J. Biol. Chem. 271:30181.
Entrez Gene IDs
1522 (Human); 64138 (Mouse)
Alternate Names
Cathepsin P; Cathepsin X/Z/P; Cathepsin Z; CTSX; CTSZ

Citations for Recombinant Human Cathepsin X/Z/P 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.

2 Citations: Showing 1 - 2
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  1. Design of protease-resistant myelin basic protein-derived peptides by cleavage site directed amino acid substitutions.
    Authors: Burster T, Marin-Esteban V, Boehm BO, Dunn S, Rotzschke O, Falk K, Weber E, Verhelst SH, Kalbacher H, Driessen C
    Biochem. Pharmacol., 2007;74(10):1514-23.
    Species: Human
    Sample Types: Cell Lysates
    Applications: Enzyme Assay
  2. An activity-based probe for the determination of cysteine cathepsin protease activities in whole cells.
    Authors: Falgueyret JP, Black WC, Cromlish W, Desmarais S, Lamontagne S, Mellon C, Riendeau D, Rodan S, Tawa P, Wesolowski G, Bass KE, Venkatraman S, Percival MD
    Anal. Biochem., 2004;335(2):218-27.
    Species: Human
    Sample Types: Recombinant Protein
    Applications: Enzyme Assay

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Fluorogenic Peptide Substrates

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