Recombinant Human ECE-2 Protein, CF

Catalog # Availability Size / Price Qty
1645-ZN-010
R&D Systems Recombinant Proteins and Enzymes
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Citations (3)
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Recombinant Human ECE-2 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 > 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
Analysis
His
Predicted Molecular Mass
79 kDa
SDS-PAGE
113 kDa, reducing conditions

Product Datasheets

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1645-ZN

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.

1645-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, 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
Reconstitution Calculator

Reconstitution Calculator

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Background: ECE-2

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.

References
  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.
Long Name
Endothelin-converting Enzyme-2
Entrez Gene IDs
9718 (Human)
Alternate Names
EC 3.4.24; EC 3.4.24.71; ECE2; ECE-2; endothelin converting enzyme 2; endothelin-converting enzyme 2; KIAA0604MGC2408; MGC17664; MGC78487

Citations for Recombinant Human ECE-2 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.

3 Citations: Showing 1 - 3
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  1. Endothelin-converting enzymes degrade ?-synuclein and are reduced in dementia with Lewy bodies
    Authors: JS Miners, S Love
    J. Neurochem, 2017-03-29;0(0):.
    Applications: Bioassay
  2. Endothelin-converting enzyme-2 is increased in Alzheimer's disease and up-regulated by Abeta.
    Authors: Palmer JC, Baig S, Kehoe PG, Love S
    Am. J. Pathol., 2009-06-18;175(1):262-70.
    Applications: ELISA (Standard), Western Blot
  3. Endothelin-converting enzyme-1 degrades internalized somatostatin-14.
    Authors: Roosterman D, Kempkes C, Cottrell GS, Padilla BE, Bunnett NW, Turck CW, Steinhoff M
    Endocrinology, 2008-02-14;149(5):2200-7.
    Species: Human
    Sample Types: Recombinant Protein
    Applications: Enzyme Assay

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