Recombinant Human ECE-2 Protein, CF

Catalog #: 1645-ZN
3 Citations Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
1645-ZN-010

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Citations (3)

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Summary Product Datasheets Carrier Free Data Images Reconstitution Calculator Background Related Research Areas

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 #

EAW78275

N-terminal Sequence
Analysis

His

Predicted Molecular Mass

79 kDa

SDS-PAGE

113 kDa, reducing conditions

Product Datasheets

1645-ZN

Product Datasheet

COA

SDS

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

Dilute rhECE-2 to 100 µg/mL in Assay Buffer.
Incubate at room temperature for 30 minutes.
Dilute the incubated rhECE-2 to 0.8 ng/µL.
Dilute Substrate to 20 µM in Assay Buffer.
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.
Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
Calculate specific activity:

Specific Activity (pmol/min/µg) =	Adjusted V_max* (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

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

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

Related Research Areas

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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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
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
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