Recombinant Human ECE-2 Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

Order Details

Recombinant Human ECE-2 Protein, CF Summary

Details of Functionality
Measured by its ability to cleave the fluorogenic peptide substrate, Mca-RPPGFSAFK(Dnp)-OH (Catalog # ES005). The specific activity is > 250 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human ECE-2 protein
Gly104-Trp787, with an N-terminal 10-His tag and Val
Accession #
N-terminal Sequence
His
Protein/Peptide Type
Recombinant Enzymes
Gene
ECE2
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
79 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
SDS-PAGE
113 kDa, reducing conditions
Publications
Read Publications using
1645-ZN in the following applications:

Packaging, Storage & Formulations

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.
Buffer
Supplied as a 0.2 μm filtered solution in Tris, NaCl and ZnCl2.
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Assay Procedure
  • Assay Buffer: 0.1 M MES, 0.25 M NaCl, pH 5.75
  • Recombinant Human ECE-2 (rhECE-2) (Catalog # 1645-ZN)
  • 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. Dilute rhECE-2 to 100 µg/mL in Assay Buffer.
  2. Incubate at room temperature for 30 minutes.
  3. Dilute the incubated rhECE-2 to 0.8 ng/µL.
  4. Dilute Substrate to 20 µM in Assay Buffer.
  5. Load 50 µL of 0.8 ng/µL rhECE-2 into a plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing Assay Buffer and 20 µM Substrate.
  6. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
  7. 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:
  • rhECE-2: 0.040 µg
  • Substrate: 10 µM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human ECE-2 Protein, CF

  • EC 3.4.24
  • EC 3.4.24.71
  • ECE2
  • ECE-2
  • endothelin converting enzyme 2
  • endothelin-converting enzyme 2
  • KIAA0604MGC2408
  • MGC17664
  • MGC78487

Background

ECE-2 is a zinc protease of the neprilysin (NEP) family, which also includes ECE-1, PEX, XCE, DINE, Kell and several NEP-like proteins (1). ECE-2 is a type II transmembrane protein with a short cytoplasmic tail and a large ectodomain. Three alternatively spliced forms differ in their cytoplasmic tail (2). In addition to big endothelin-1, ECE-2 cleaves a variety of bioactive peptides such as bradykinin, neurotensin, angiotensin I, substance P, dynorphin B, proenkephalin-derived peptides such as peptide E, BAM 22 and BAM18, and PEN-LEN, an endogenous inhibitor of prohormone convertase 1 (3). Together with ECE-1, it is also involved in degradation of beta -amyloid peptide (4). The ectodomain of human ECE-2 was expressed with an N-terminal His tag and purified.

  1. Turner, A.J. et al. (2001) BioEssays 23:261.
  2. Lorenzo, M.N. et al. (2001) Biochim. Biophys. Acta 1522:46.
  3. Mzhavia, N. et al. (2003) J. Biol. Chem. 278:14704.
  4. Eckman, E.A. et al. (2003) J. Biol. Chem. 278:2081.

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Publications for ECE-2 (1645-ZN)(3)

We have publications tested in 2 confirmed species: Human, N/A.

We have publications tested in 3 applications: Bioassay, ELISA (Standard), Enzyme Assay.


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Bioassay
(1)
ELISA (Standard)
(1)
Enzyme Assay
(1)
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Human
(1)
N/A
(1)
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Bioinformatics

Gene Symbol ECE2
Uniprot