Recombinant West Nile Virus NS3 Protease Protein, CF

Catalog # Availability Size / Price Qty
2907-SE-020
Product Details
Citations (1)
FAQs
Supplemental Products
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Recombinant West Nile Virus NS3 Protease Protein, CF Summary

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 pERTKR-AMC (Catalog # ES013). The specific activity is >750 pmol/min/µg, as measured under the described conditions. 
Source
E. coli-derived viral wnvNS3 Protease protein
MHHHHHH NS2b
(Ser1423-Lys1470)
Accession # ABD85079
GGGGSGGGG

West Nile Virus NS3
(Gly1506-Leu1689)
Accession # ABD85079

GGGGSGGGG West Nile Virus NS3
(Gly1506-Leu1689)
Accession # ABD85079
N-terminus C-terminus
Accession #
N-terminal Sequence
Analysis
Met and Gly
Predicted Molecular Mass
27 kDa & 20 kDa
SDS-PAGE
32 kDa and 22 kDa, reducing conditions

Product Datasheets

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.

2907-SE

Formulation Lyophilized from a 0.2 μm filtered solution in Tris and NaCl.
Reconstitution Reconstitute at 200 μg/mL in sterile, deionized water.
Shipping The product is shipped at ambient temperature. 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 reconstitution.

Assay Procedure

Materials
  • Assay Buffer: 50 mM Tris, 30% (v/v) Glycerol, pH 9.5
  • Recombinant Viral wnvNS3 Protease (Catalog # 2907-SE)
  • Substrate: L-PYROGlu-Arg-Thr-Lys-Arg-AMC (pERTKR-AMC) (Catalog # ES013)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rwnvNS3 Protease to 1 ng/µL in Assay Buffer.
  2. Dilute Substrate to 40 µM in Assay Buffer.
  3. In a plate load 50 µL of 1 ng/µL rwnvNS3 Protease, and start the reaction by adding 50 µL of 40 µM Substrate to wells. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 40 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) (Sigma, Catalog # A-9891).

Per Well:
  • rwnvNS3 Protease: 0.05 µg
  • Substrate: 20 µM
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: wnvNS3 Protease

Infection of mosquito-borne West Nile Virus can cause severe neurological disease and can be epidemic. Two non-structural proteins, NS3 and NS2b, play an essential role in viral replication and are therefore a potential target for treatment and prevention of West Nile Virus disease. NS3 consists of a trypsin-like serine protease with a catalytic triad (His51, Asp75, Ser135) and a putative helicase. Requiring NS2b as the co‑factor, NS3 protease processes viral polyprotein precursor (1, 2). The purified recombinant protein consists of three forms: the full-length fusion protein, the N-terminal NS2b, and the C-terminal NS3 with the G4SG4 linker. NS3 protease has a relatively narrow substrate specificity that prefers Arg in P1 and Lys in P2. The purified recombinant protein has autocatalytic activity that can lead to protein degradation. It is therefore important to store the sample below -20 °C and to keep on ice while working with the sample.

References
  1. Nall, T.A. et al. (2004) J. Biol. Chem. 279:48535.
  2. Chappell, K.J. et al. (2005) J. Biol. Chem. 274:2896.
Long Name
West Nile Virus NS3 Protease
Entrez Gene IDs
912267 (Viral)
Alternate Names
wnvNS3 Protease

Citation for Recombinant West Nile Virus NS3 Protease Protein, CF

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

1 Citation: Showing 1 - 1

  1. Discovery of novel West Nile Virus protease inhibitor based on isobenzonafuranone and triazolic derivatives of eugenol and indan-1,3-dione scaffolds
    Authors: AS de Oliveir, PAR Gazolla, AFCDS Oliveira, WL Pereira, LC de S Viol, AFDS Maia, EG Santos, ÍEP da Silva, TAO Mendes, AM da Silva, RS Dias, CC da Silva, MD Polêto, RR Teixeira, SO de Paula
    PLoS ONE, 2019;14(9):e0223017.
    Species: Virus
    Sample Types: Synthetic Compound
    Applications: Bioassay

FAQs

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Fluorogenic Peptide Substrates

Reconstitution Buffers

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