• 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, Leu-AMC. The specific activity is >100 pmol/min/µg, as measured under the described conditions. See Activity Assay Protocol on www.RnDSystems.com

• Source

  Mouse myeloma cell line, NS0-derived Ser27-Leu930, with a C-terminal 10-His tag

• Accession #

• N-terminal Sequence    
  Analysis

  Ser27

• Predicted Molecular Mass

  105 kDa

• SDS-PAGE

  111 kDa, reducing conditions

Assay Procedure

Materials

• Assay Buffer: 25 mM Tris, pH 8.0

• Recombinant Mouse Aminopeptidase PILS/ARTS1 (rmARTS1) (Catalog # 2500-ZN)

• Substrate: Leu-AMC (Bachem, Catalog # I-1240)

• F16 Black Maxisorp Plate (Nunc, Catalog # 475515)

• Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

1. Dilute rmARTS1 to 2 µg/mL in Assay Buffer.

2. Dilute Substrate to 200 µM in Assay Buffer.

3. Load 50 µL of the 2 µg/mL rmARTS1 into a plate, and start the reaction by adding 50 µL of 200 µM Substrate. Include a Substrate Blank containing 50 µL 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:

     *Adjusted for Substrate Blank
     **Derived using calibration standard 7-amino, 4-Methyl Coumarin AMC (Sigma, Catalog # A-9891).

Per Well:

• rmARTS1: 0.1 µg

• Substrate: 100 µM

Background: Aminopeptidase PILS/ARTS1

The mouse ARTS1 gene encodes aminopeptidase PILS (Puromycin-Insensitive Leucyl-Specific), which is also known as adipocyte-derived leucine aminopeptidase, type 1 tumor necrosis factor receptor shedding aminopeptidase regulator and ERAAP (the aminopeptidase associated with antigen processing in the endoplasmic reticulum) (1-3). The deduced amino acid sequence of mouse ARTS1 consists of a signal peptide and a large ectodomain. Widely expressed and releasing an N‑terminal amino acid (Leu), the enzyme may play a role in many processes such as antigen processing and angiogenesis.

• References:

1. Miyashita, H. et al. (2002) Blood 99:3241.

2. Serwold, T. et al. (2002) Nature 419:480.

3. Schomburg, L. (2004) in Handbook of Proteolytic Enzymes. Barrett, et al. eds. p. 311, Academic Press, San Diego.

• Long Name:

  Aminopeptidase Puromycin-insensitive Leucyl-specific

• Entrez Gene IDs:

  51752 (Human); 80898 (Mouse)

• Alternate Names:

  Adipocyte-derived leucine aminopeptidase; A-LAP; A-LAPALAP; Aminopeptidase PILS; ARTS1; ARTS1aminopeptidase regulator of TNFR1 shedding; ARTS-1PILSAP; EC 3.4.11; EC 3.4.11.-; EC 3.4.11.1; endoplasmic reticulum aminopeptidase 1; endoplasmic reticulum aminopeptidase associated with antigen processing; ERAAP1; ERAP1; KIAA0525APPILS; PILS-AP; PILS-APERAAP; Puromycin-insensitive leucyl-specific aminopeptidase; Type 1 tumor necrosis factor receptor shedding aminopeptidase regulator