Recombinant Human Tryptase epsilon/BSSP-4 Protein, CF Summary
Ala33-Ser317 & Val50-Ser317, both with a C-terminal 10-His tag
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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.
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.
|Formulation||Supplied as a 0.2 μm filtered solution in Sodium Acetate, NaCl and Glycerol.|
|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.
- Assay Buffer: 50 mM Tris, 0.05% (w/v) Brij-35, pH 8.0
- Recombinant Human Tryptase epsilon /BSSP‑4 (rhBSSP-4) (Catalog # 5047-SE)
- Substrate: Z-Arg-SBzl (SM Biochemicals, Catalog # SMSB01)
- DTNB (5,5’-dithibis(2-nitrobenzoic acid)) (Sigma, Catalog # D-8130)
- 96-well Clear Plate (Costar, Catalog # 92592)
- Plate reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
- Dilute rhBSSP-4 to 0.5 ng/µL in Aassay Buffer.
- Dilute Substrate to 200 µM in Assay Buffer containing 200 µM DTNB.
- Load 50 µL of 0.5 ng/µL rhBSSP-4 into a plate, and start the reaction by adding 50 µL of 200 µM Substrate to wells. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of Substrate.
- Read at an absorbance of 405 nm for 5 minutes in kinetic mode.
- Calculate specific activity:
Specific Activity (pmol/min/µg) =
|Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/M|
|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 mODPer Well:
- rhBSSP-4: 0.025 µg
- Substrate: 100 µM
Background: Tryptase epsilon/BSSP-4
Tryptase epsilon, also known as brain-specific protease-4 (BSSP-4) and prosemin, is expressed in the brain and epithelium-rich tissues such as the lung and eye (1, 2). BSSP-4 is a serine protease related to trypsin, preferentially hydrolyzing substrates after arginine and lysine residues. However, Tryptase epsilon is less susceptible to inhibition by common trypsin inhibitors such as aprotinin, alpha 1-antitrypsin and secretory leukocyte protease inhibitor (3). Tryptase epsilon efficiently converts pro‑urokinase‑type plasminogen activator to its mature, active form (3).
- Wong, G.W. et al. (2004) J. Biol. Chem. 279:2438.
- Wong, G.W. et al. (2001) J. Biol. Chem. 276:49169.
- Yasuda, S. et al. (2005) Blood 105:3893.
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