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Recombinant Human u-Plasminogen Activator/Urokinase Avi, CF

Catalog #: AVI1310 Datasheet / COA / SDS
His-tag Avi-tag
Catalog # Availability Size / Price Qty
AVI1310-020

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Biotinylated Recombinant Human u‑Plasminogen Activator (uPA)/Urokinase His-tag Avi-tag Protein Enzyme Activity.
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Recombinant Human u-Plasminogen Activator/Urokinase Avi, CF Summary

  • R&D Systems HEK293-derived Recombinant Human u-Plasminogen Activator/Urokinase Avi (AVI1310)
  • 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

Product Specifications

Endotoxin Level
<0.10 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave a peptide substrate, N-carbobenzyloxy-Gly-Gly-Arg-7-amido-4-methylcoumarin (Z-GGR-AMC). The specific activity is >2000 pmol/min/μg, as measured under the described conditions. Measured by its binding ability in a functional ELISA. Biotinylated Recombinant Human u-Plasminogen Activator (uPA)/Urokinase His-tag Avi-tag (Catalog # AVI1310) binds to Human u-Plasminogen Activator (uPA) Antibody (Catalog # MAB1310) with an ED50 of 0.300-4.50 ng/mL.
Source
Human embryonic kidney cell, HEK293-derived human u-Plasminogen Activator (uPA)/Urokinase protein
Met1-Leu431 with C-terminal 6-His and Avi-tag
Accession #
P00749.2
N-terminal Sequence
Analysis
Lys156 & Phe177
Predicted Molecular Mass
18 kDa (long A chain), 3 kDa (short A chain), 31 kDa (B chain)
SDS-PAGE
19-20 kDa & 32-38 kDa, under reducing conditions.

Product Datasheets

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AVI1310

Product Datasheet
COA
SDS

Carrier Free

What does CF mean?

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.

What formulation is right for me?

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.

AVI1310

Formulation Supplied as a 0.2 μm filtered solution in HEPES, NaCl and CaCl2.
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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Assay Procedure

Materials
  • Assay Buffer: 50 mM Tris, 0.01% (v/v) Tween® 20, pH 8.5
  • Biotinylated Recombinant Human u-Plasminogen Activator/Urokinase His-tag Avi-tag (rhuPA) (Catalog # AVI1310)
  • Substrate: Z-GGR-AMC, 10 mM stock in DMSO
  • Black 96-Well Plate
  • Plate Reader with Fluorescence Read Capability
  1. Dilute rhuPA to 0.5 µg/mL in Assay Buffer.
  2. Dilute Substrate to 400 µM in Assay Buffer.
  3. Load 50 µL of the 0.5 µg/mL rhuPA into a black well plate and start the reaction by adding 50 µL of 400 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 400 µM Substrate.
  4. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), 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 (AMC)
Per Well:
  • rhuPA His Avi: 0.025 µg
  • Substrate: 200 µM 

Scientific Data

Enzyme Activity View Larger

Recombinant Human u-Plasminogen Activator (uPA)/Urokinase His-tag Avi-tag (Catalog # AVI1310) is measured by its ability to cleave a peptide substrate, N-carbobenzyloxy-Gly-Gly-Arg-7-amido-4-methylcoumarin (Z-GGR-AMC).

Binding Activity View Larger

In a functional ELISA, Biotinylated Recombinant Human u-Plasminogen Activator (uPA)/Urokinase His-tag Avi-tag Protein (Catalog # AVI1310) binds to Human u-Plasminogen Activator (uPA) Antibody (MAB1310) with an ED50 of 0.300-4.50 ng/mL

Reconstitution Calculator

Reconstitution Calculator

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: u-Plasminogen Activator (uPA)/Urokinase

Urokinase Plasminogen Activator (uPA), also known as u-plasminogen activator or urokinase, is a highly-specific serine protease from the peptidase S1 family that cleaves plasminogen to form plasmin making it a key player in the plasminogen activator (PA) system (1, 2). In cancer, the PA system plays a commanding role in tumor growth, angiogenesis, tumor cell invasion, migration, and metastasis. Expression of uPA is minimal in normal cells but is increased several fold in tumor cells by extracellular stimuli elevated in cancer (3) and corresponds to poor outcomes in several types of cancer (2, 4-7). Therefore, uPA has been identified as an excellent target for therapeutic development through inhibition of protease activity or though inhibition of uPA-dependent signaling while in complex with uPA receptor (uPAR) (2, 7). The pro-enzyme of uPA is synthesized with a N-terminal signal peptide and processed into an active disulfide-linked two-chain molecule (2, 7-10). For human uPA, the B chain starting at Ile179 corresponds to the catalytic domain. Two forms of the A chain exist, one starting at Ser21 (the long form) and the other at Lys156 (the short form). While the B chain is common for both forms, the long and short A chains are unique to expected 49 kDa and 34 kDa two-chain forms, respectively. The long A chain contains an EGF-like domain and the kringle domain. The long A domain is reportedly responsible for the binding of the uPA receptor (uPAR) (2,7). 

References
  1. Ellis, V. (2004) in Handbook of Proteolytic Enzymes. Barrett, A.J. et. al. eds., Academic Press, San Diego, pp.1677. 
  2. Mahmood, N. et. al. (2018) Front Oncol. 8:24.
  3. Nagamine, Y. et. al. (2005) Thromb. Haemost. 93:661.
  4. Duffy, M. and C. Duggan. (2004) Clin. Biochem. 37:541.
  5. Pappot, H. et. al. (2006) Lung Cancer 51:193.
  6. Taubert, H. et. al. (2010) Br. J. Cancer 102:731.
  7. Masucci, M.T. et. al. (2022) Cancers. 14:498.
  8. Riccio, A. et. al. (1985) Nucleic Acids Res. 13:2759.
  9. Nagai, M. et. al. (1985) Gene 36:183.
  10. Jacobs, P. et. al. (1985) DNA 4:139.
Long Name
Urokinase-type Plasminogen Activator
Entrez Gene IDs
5328 (Human); 18792 (Mouse); 102135886 (Cynomolgus Monkey)
Alternate Names
ATF; EC 3.4.21; EC 3.4.21.73; plasminogen activator, urokinase; PLAU; uPA; u-PA; uPlasminogen Activator; u-Plasminogen Activator; urinary; Urokinase; urokinase-type plasminogen activator

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