Recombinant Mouse Kell Protein, CF

Catalog # Availability Size / Price Qty
1454-ZN-010
R&D Systems Recombinant Proteins and Enzymes
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Recombinant Mouse Kell 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 >0.2 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived mouse Kell protein
Phe50-Trp713, with an N-terminal 7-His tag
Accession #
N-terminal Sequence
Analysis
His
Predicted Molecular Mass
76 kDa
SDS-PAGE
106 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.

1454-ZN

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and ZnCl2.
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
  • Assay Buffer: 0.1 M MES, pH 6.5
  • Recombinant Mouse Kell (rmKell) (Catalog # 1454-ZN)
  • Substrate: MCA-Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH (Catalog # ES005), 2 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 rmKell to 20 ng/µL in Assay Buffer.
  2. Dilute Substrate to 20 µM in Assay Buffer.
  3. Load 50 µL of the 20 ng/µL rmKell 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.
  4. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 1 hour.
  5. 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:
  • rmKell: 1 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: Kell

Kell, a type II membrane glycoprotein, is linked through a single disulfide bond to XK, a putative membrane transporter. The two proteins constitute the Kell blood group antigens (1). Kell is a zinc endopeptidase of the neprilysin (NEP) family which also includes endothelin converting enzymes (ECE-1 and ECE-2), PEX, XCE, DINE and several NEP-like proteins (2). It has been shown to cleave big endothelin-3 (ET-3) at Trp21-Ile22, yielding ET-3, and to a much lesser extent, big ET-1 and big ET-2 at Trp21-Val22, yielding ET-1 and ET-2 (3). Several different molecular defects cause the Kell null phenotype, which has no obvious clinical outcome (4).

References
  1. Lee, S. et al. (2000) Transfus. Med. Rev. 14:93.
  2. Turner, A.J. et al. (2001) BioEssays. 23:261.
  3. Lee, S. et al. (1999) Blood 94:1440.
  4. Lee, S. et al. (2001) J. Biol. Chem. 276:27281.
Long Name
Zinc protease blood group antigen
Entrez Gene IDs
3792 (Human); 23925 (Mouse)
Alternate Names
CD238 antigen; CD238; EC 3.4.24.-; ECE3; KEL; kell blood group antigen; kell blood group glycoprotein; Kell blood group; Kell blood group, metalloendopeptidase; Kell blood group, metallo-endopeptidase; Kell

FAQs

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Fluorogenic Peptide Substrates

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