Legumain is a lysosomal cysteine protease whose activity is found in several tissues tested (1, 2). Legumain plays a pivotal role in the endosomal/lysosomal degradation system because the Legumain deficiency causes the accumulation of pro cathepsins B, H and L, another group of lysosomal cysteine proteases (3). Over-expression of Legumain in tumors is significant for invasion/metastasis (4). Also known as Asparaginyl Endopeptidase, it specifically cleaves peptide bonds with Asn at the P1 position. Nevertheless, it also cleaves peptide bonds with Asp at the P1 position. Auto-activation of pro Legumain involves both types of the cleavage, which result in the removal of the pro peptides in both C- and N-termini (5). In addition, Legumain activates pro MMP-2 and processes bacterial antigens for MHC class II presentation and pro thymosin alpha to thymosin alpha 1 and thymosin alpha 11, two acidic peptides with immunoregulatory properties (6‑8). Human Legumain is synthesized as a 433 amino acid precursor with a signal peptide (residues 1‑17). The pro enzyme (residues 18‑433) was expressed with an N-terminal His tag. This activity of Legumain can be inhibited by recombinant human Cystatins C and E/M and recombinant mouse Cystatin C (Catalog # http://www.rndsystems.com/product_results.aspx?k=1196-PI">1196-PI, http://www.rndsystems.com/product_results.aspx?k=1286-PI">1286-PI and http://www.rndsystems.com/product_results.aspx?k=1238-PI">1238-PI, respectively).
Recombinant Human Legumain/Asparaginyl Endopeptidase, CF
R&D Systems | Catalog # 2199-CY
Loading...
Key Product Details
- R&D Systems NS0-derived Recombinant Human Legumain/Asparaginyl Endopeptidase (2199-CY)
- Quality control testing to verify active proteins with lot specific assays by in-house scientists
- All R&D Systems proteins are covered with a 100% guarantee
Source
NS0
Accession Number
Structure / Form
Pro form
Applications
Enzyme Activity
Loading...
Product Specifications
Source
Mouse myeloma cell line, NS0-derived human Legumain/Asparaginyl Endopeptidase protein
Ile18-Tyr433, with an N-terminal 7-His tag
Ile18-Tyr433, with an N-terminal 7-His tag
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.
N-terminal Sequence Analysis
His
Predicted Molecular Mass
49 kDa
SDS-PAGE
60 kDa, reducing conditions
Activity
Measured by its ability to cleave the fluorogenic peptide substrate, N-carbobenzyloxy-Ala-Ala-Asn-7-amido-4-methylcoumarin (Z-AAN-AMC).
The specific activity is >250 pmol/min/µg, as measured under the described conditions.
The specific activity is >250 pmol/min/µg, as measured under the described conditions.
Reviewed Applications
Read 2 reviews rated 5 using 2199-CY in the following applications:
Formulation, Preparation, and Storage
2199-CY
| Formulation | Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol. |
| 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.
|
Background: Legumain/Asparaginyl Endopeptidase
References
- Chen, J.-M. et al. (1997) J. Biol. Chem. 272:8090.
- Tanaka, T. et al. (1996) Cytogenet. Cell Genet. 74:120.
- Shirahama-Noda, K. et al. (2003) J. Biol. Chem. 278:33194.
- Liu, C. et al. (2003) Cancer Res. 63: 2957.
- Li D.N. et al. (2003) J. Biol. Chem. 278:38980.
- Chen, J.M. et al. (2001) Biol. Chem. 382:777.
- Schwarz, G. et al. (2002) Biol. Chem. 383:1813.
- Sarndeses, C.S. et al. (2003) J. Biol. Chem. 278:13286.
Alternate Names
AEP, Asparaginyl Endopeptidase, LGMN
Gene Symbol
LGMN
UniProt
Additional Legumain/Asparaginyl Endopeptidase Products
Product Documents for Recombinant Human Legumain/Asparaginyl Endopeptidase, CF
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Human Legumain/Asparaginyl Endopeptidase, CF
For research use only
Related Research Areas
Citations for Recombinant Human Legumain/Asparaginyl Endopeptidase, CF
Powered by Bioz
Customer Reviews for Recombinant Human Legumain/Asparaginyl Endopeptidase, CF (2)
5 out of 5
2 Customer Ratings
Have you used Recombinant Human Legumain/Asparaginyl Endopeptidase, CF?
Submit a review and receive an Amazon gift card!
$25/€18/£15/$25CAN/¥2500 Yen for a review with an image
$10/€7/£6/$10CAN/¥1110 Yen for a review without an image
Submit a review
Customer Images
Showing
1
-
2 of
2 reviews
Showing All
Filter By:
-
Application: In vitro bioactivity in cell cultureVerified Customer | Posted 12/04/2020
-
Application: In vitro bioactivity in cell cultureVerified Customer | Posted 12/04/2020
There are no reviews that match your criteria.
Protocols
View specific protocols for Recombinant Human Legumain/Asparaginyl Endopeptidase, CF (2199-CY):
Materials
- Activation Buffer: 50 mM Sodium Acetate, 100 mM NaCl, pH 4.0
- Assay Buffer: 50 mM MES, 250 mM NaCl, pH 5.0
- Recombinant Human Legumain/Asparaginyl Endopeptidase (rhLegumain) (Catalog # 2199-CY)
- Substrate: Z-Ala-Ala-Asn-AMC (Bachem, Catalog # I-1865), 10 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhLegumain to 100 µg/mL in Activation Buffer.
- Incubate for 2 hours at 37 °C.
- Dilute rhLegumain to 1 ng/µL in Assay Buffer.
- Dilute Substrate to 200 µM in Assay Buffer.
- Load into a black well plate 50 µL of 1 ng/µL rhLegumain 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.
- Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), 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 (AMC) (Sigma, Catalog # A-9891).
Per Well:- rhLegumain: 0.050 µg
- Substrate: 100 µM