Tryptase-5 is encoded by Prss32, one of 13 genes on mouse chromosome 17A3.3 that correspond to functional trypsin-like serine proteases (1). The deduced amino acid sequence of mouse Tryptase-5 consists of 331 residues with a signal peptide (residues 1 to 19), a pro region (residue 20 to 53), a catalytic domain (54 to 304), and a C-terminal hydrophobic peptide (residues 305 to 331). The mRNA is expressed in smooth muscle, eye, stomach, uterus and lymph node. Apparently, Prss32 does not seem to have a counterpart in the human genome. The mouse Tryptase-5 (residues 20 to 304) was expressed in the NS0 cells with a foreign signal peptide. The protein was secreted and purified. After being treated by thermolysin, the enzyme is active against a peptide substrate as described in the Activity Assay Protocol.
Recombinant Mouse Tryptase-5/Prss32 Protein, CF
R&D Systems | Catalog # 2634-SE
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Key Product Details
- R&D Systems NS0-derived Recombinant Mouse Tryptase-5/Prss32 Protein (2634-SE)
- 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
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Product Specifications
Source
Mouse myeloma cell line, NS0-derived mouse Tryptase-5/Prss32 protein
Ser20-Arg304, with a C-terminal 10-His tag
Ser20-Arg304, with a C-terminal 10-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
Ser20
Predicted Molecular Mass
32 kDa
SDS-PAGE
40 kDa doublet, reducing conditions
Activity
Measured by its ability to cleave a colorimetric peptide substrate, N-carbobenzyloxy-Arg-ThioBenzyl ester (Z-R-SBzl), in the presence of 5,5’Dithio-bis (2-nitrobenzoic acid) (DTNB). Edwards, K.M. et al. (1999) J. Biol. Chem. 274:30468.
The specific activity is >400 pmol/min/µg, as measured under the described conditions.
The specific activity is >400 pmol/min/µg, as measured under the described conditions.
Formulation, Preparation, and Storage
2634-SE
| Formulation | Lyophilized from a 0.2 μm filtered solution in MES and NaCl. |
| Reconstitution | Reconstitute at 200 μg/mL in sterile 50 mM Tris, 10 mM CaCl2, 150 mM NaCl and 0.05% Brij-35 pH 7.5. |
| 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.
|
Calculators
Background: Tryptase-5/Prss32
References
- Wong, G. W. et al. (2004) J. Biol. Chem. 279:2438.
Alternate Names
Prss32, Tryptase5
Entrez Gene IDs
69814 (Mouse)
Gene Symbol
PRSS32
UniProt
Additional Tryptase-5/Prss32 Products
Product Documents for Recombinant Mouse Tryptase-5/Prss32 Protein, CF
Certificate of Analysis
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Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Mouse Tryptase-5/Prss32 Protein, CF
For research use only
Related Research Areas
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Protocols
View specific protocols for Recombinant Mouse Tryptase-5/Prss32 Protein, CF (2634-SE):
Materials
- Activation Buffer: (TCNB) 50 mM Tris, 0.15 M NaCl, 10 mM CaCl2, 0.05% Brij-35, pH 7.5
- Assay Buffer: 50 mM Tris, 0.25 M NaCl, pH 8.0
- Recombinant Mouse Tryptase‑5/Prss32 (rmTPS-5) (Catalog # 2634-SE)
- Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
- 1,10 Phenanthroline (Sigma, Catalog # 320056), 0.6 M stock in DMSO
- Substrate: Z-Arg-SBzl (SM Biochemicals, Catalog # SMSB01), 10 mM stock in DMSO
- 5,5’Dithio-bis(2-nitrobenzoic acid) (DTNB) (Sigma Catalog # D-8130)
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute rmTPS-5 to 100 µg/mL with Activation Buffer.
- Dilute Thermolysin to 0.4 µg/mL with Activation Buffer.
- Mix equal volumes of 100 µg/mL rmTPS-5 and 0.4 µg/mL Thermolysin for final concentrations of 50 µg/mL and 0.2 µg/mL, respectively.
- Incubate at 37 °C for 1 hour.
- Stop the reaction with 10 mM 1,10 Phenanthroline.
- Dilute activated rmTPS-5 to 4 ng/µL in Assay Buffer.
- Dilute Substrate to 200 µM in Assay Buffer with 200 µM of DTNB.
- Load 50 µL of the 4 ng/µL rmTPS-5 in a clear plate and start the reaction by adding 50 µL of the Substrate/DTNB mixture to wells. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL Substrate mix without any rmTPS-5.
- Read at 405 nm (Absorbance), in kinetic mode for 5 minutes.
- Calculate specific activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol |
| ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg) |
*Adjusted for Substrate Blank
**Using the extinction coefficient 13260 M-1cm-1
***Using the path correction 0.32 cm
Note: the output of many spectrophotometers is in mOD Per Well:
- rmTPS-5: 0.20 µg
- DTNB: 100 µM
- Substrate: 100 µM
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