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Recombinant Human Aminopeptidase A/ENPEP Protein, CF

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Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

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Recombinant Human Aminopeptidase A/ENPEP Protein, CF Summary

Details of Functionality
Measured by its ability to cleave the fluorogenic peptide substrate, Glu-7-amido-4-methylcoumarin (Glu-AMC). The specific activity is > 2,000 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Aminopeptidase A/ENPEP protein
Arg41-Gly957, with an N-terminal 6-His tag
Accession #
N-terminal Sequence
His
Protein/Peptide Type
Recombinant Enzymes
Gene
ENPEP
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

Dilutions
  • Enzyme Activity
Theoretical MW
106 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
145 kDa, reducing conditions
Publications
Read Publication using
2499-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 MES and NaCl.
Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Assay Procedure
  • 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
  1. Dilute rhENPEP to 0.4 ng/µL in Assay Buffer.
  2. Dilute Substrate Glu-AMC to 200 µM in Assay Buffer.
  3. 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.
  4. Read at excitation and emission wavelengths of 380 nm and 460 nm, respectively, in kinetic mode for 5 minutes.
  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 7-amino, 4-Methyl Coumarin (Sigma, Catalog # A-9891).

Per Well:
  • rhENPEP: 0.020 μg
  • Substrate: 100 µM

Notes

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

Alternate Names for Recombinant Human Aminopeptidase A/ENPEP Protein, CF

  • Aminopeptidase A
  • APA
  • AP-A
  • CD249 antigen
  • CD249
  • Differentiation antigen gp160
  • EAP
  • EC 3.4.11
  • EC 3.4.11.7
  • ENPEP
  • glutamyl aminopeptidase (aminopeptidase A)
  • glutamyl aminopeptidase
  • gp160

Background

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.

  1. Wang, J. and M.D. Cooper (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, A.J. et al.) p. 299, Academic Press, San Diego.
  2. Nanus, D.M. et al. (1993) Proc. Natl. Acad. Sci. USA 90:7069.
  3. Li, L. et al. (1993) Genomics 17:657.
  4. Rozenfeld, R. et al. (2004) J. Biol. Chem. 279:43285.

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Recombinant Human
Aminopeptidase A/ENPE ...
2499-ZN
Recombinant Human Aminopeptidase A/ENPEP Protein, CF
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Publications for Aminopeptidase A/ENPEP (2499-ZN)(1)

We have publications tested in 1 confirmed species: Human.

We have publications tested in 1 application: Enzyme Assay.


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Bioinformatics

Gene Symbol ENPEP
Uniprot