Recombinant Rat ACE-2 Protein, CF

• Reactivity: Rt
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Rat ACE-2 Protein, CF Summary

• Details of Functionality: Measured by its ability to cleave a fluorogenic peptide substrate, Mca-YVADAPK(Dnp)-OH (Catalog # ES007). The specific activity is >300 pmol/min/µg, as measured under the described conditions.
• Source: Mouse myeloma cell line, NS0-derived rat ACE-2 protein
  Met1-Thr740, with a C-terminal 10-His tag
• Accession #: NP_001012006
• N-terminal Sequence: No results obtained: Gln18 predicted
• Protein/Peptide Type: Recombinant Enzymes
• Gene: Ace2
• 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: 85 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: 120 kDa, reducing conditions

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, ZnCl₂ and Glycerol.
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain
• Assay Procedure:
  • Assay Buffer: 50 mM Tris, 1 M NaCl, pH 7.5
  • Recombinant Rat ACE-2 (rrACE-2) (Catalog # 4516-ZN)
  • Substrate: MCA-Tyr-Val-Ala-Asp-Ala-Pro-Lys(Dnp)-OH (Catalog # ES007), 2 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rrACE-2 to 0.5 ng/μL in Assay Buffer.
  2. Dilute Substrate 20 μM in Assay Buffer.
  3. In plate load 50 μL of 0.5 ng/μL rrACE-2. In a separate well load 50 µL Assay Buffer to be used as a Substrate Blank.
  4. Start the reaction by adding 50 μL of 20 μM Substrate to all wells.
  5. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
  6. 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:
  • rrACE-2: 0.025 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 Rat ACE-2 Protein, CF

• ACE2
• ACE-2
• ACEH
• ACEHangiotensin I converting enzyme 2
• ACE-related carboxypeptidase
• angiotensin I converting enzyme (peptidyl-dipeptidase A) 2
• angiotensin-converting enzyme 2
• Angiotensin-converting enzyme homolog
• DKFZp434A014
• EC 3.4.17
• EC 3.4.17.23
• Metalloprotease MPROT15

Background

Angiotensin I Converting Enzyme 2 (ACE-2), also called ACEH (ACE homologue), is an integral membrane protein and a zinc metalloprotease of the ACE family that also includes somatic and germinal ACE (1). The predicted rat ACE-2 protein sequence consists of 805 amino acids, including an N-terminal signal peptide, a single catalytic domain, a C-terminal membrane anchor, and a short cytoplasmic tail. ACE-2 cleaves Angiotensins I and II as a carboxypeptidase. ACE-2 mRNA is found at high levels in testis, kidney and heart and at moderate levels in colon, small intestine and ovary. Classical ACE inhibitors such as captopril and lisinopril do not inhibit ACE-2 activity. Novel peptide inhibitors of ACE-2 do not inhibit ACE activity (2). Genetic data from Drosophila, mice and rats show that ACE-2 is an essential regulator of cardiovascular function in vivo (3). In addition, ACE-2 has a critical function in acute lung injury (4).

1. Tipnis, S.R. et al. (2000) J. Biol. Chem. 275:33238.
2. Crackower, M.A. et al. (2002) Nature 417:822.
3. Huang, L. et al. (2003) J. Biol. Chem. 278:15532.
4. Kuba, K. et al. (2006) Curr. Opinion Pharmacol. 6:271.

Bioinformatics

• Gene Symbol: Ace2
• Uniprot:
  • NP_001012006()