Recombinant Human Aminopeptidase A/ENPEP Protein, CF Summary
Arg41-Gly957, with an N-terminal 6-His tag
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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.
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.
|Formulation||Supplied as a 0.2 μm filtered solution in MES and NaCl.|
|Shipping||The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.|
|Stability & Storage:||Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
- Assay Buffer: 25 mM Tris, 50 mM CaCl2, 0.2 M NaCl, pH 8.0
- Recombinant Human Aminopeptidase A/ENPEP (rhENPEP) (Catalog # 2499-ZN)
- Substrate: Glu-AMC (Bachem, Catalog # I-1180), 10 mM stock in 30 mM HCl
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhENPEP to 0.4 ng/µL in Assay Buffer.
- Dilute Substrate Glu-AMC to 200 µM in Assay Buffer.
- Load into plate 50 µL of 0.4 ng/µL rhENPEP, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 200 µM Substrate.
- Read at excitation and emission wavelengths of 380 nm and 460 nm, respectively, in kinetic mode for 5 minutes.
- 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 7-amino, 4-Methyl Coumarin (Sigma, Catalog # A-9891).
- rhENPEP: 0.020 μg
- Substrate: 100 µM
Background: Aminopeptidase A/ENPEP
The human ENPEP gene encodes aminopeptidase A (APA), which is also known as glutamyl aminopeptidase (EAP), aspartate aminopeptidase, angiotensinase A, Ca2+-activated glutamate aminopeptidase, membrane aminopeptidase II and the BP-1/6C3 antigen (1). The various names reflect its substrate specificity (with a preference for Glu, Asp and angiotensin II as one of the physiological substrates), Ca2+-dependence, cellular location (type II membrane protein) and immunological reactivity to the antibody BP-1/6C3. The deduced amino acid sequence of human ENPEP consists of a short cytoplasmic tail (residues 1 to 17), a transmembrane region (residues 18 to 40), and a long ectodomain (residues 41 to 957) (2, 3). In addition to the N-terminal zinc metalloprotease domain, the ectodomain also contains the C-terminal region, which functions as an intramolecular chaperone required for the correct folding, cell surface expression and activity (4). The purified recombinant human ENPEP consists of the entire ectodomain and is active in the assay as described in Activity Assay Protocol.
- Wang, J. and M.D. Cooper (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, A.J. et al.) p. 299, Academic Press, San Diego.
- Nanus, D.M. et al. (1993) Proc. Natl. Acad. Sci. USA 90:7069.
- Li, L. et al. (1993) Genomics 17:657.
- Rozenfeld, R. et al. (2004) J. Biol. Chem. 279:43285.
Citation for Recombinant Human Aminopeptidase A/ENPEP 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.
1 Citation: Showing 1 - 1
Aminopeptidase A contributes to the N-terminal truncation of amyloid beta-peptide.
Authors: Sevalle J, Amoyel A, Robert P, Fournie-Zaluski MC, Roques B, Checler F
J. Neurochem., 2009;109(1):248-56.
Sample Types: Recombinant Protein
Applications: Enzyme Assay
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