Recombinant Mouse Kallikrein 5 Protein, CF

• Reactivity: Mu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Mouse Kallikrein 5 Protein, CF Summary

• Details of Functionality: Measured by its ability to cleave the fluorogenic peptide substrate Boc-VPR-AMC (Catalog # ES011). The specific activity is >1,350 pmol/min/μg, as measured under the described conditions. www.RnDSystems.com.
• Source: Mouse myeloma cell line, NS0-derived mouse Kallikrein 5 protein
  Val23-Asn293, with a C-terminal 10‑His tag
• Accession #: NP_081082
• N-terminal Sequence: Val23
• Protein/Peptide Type: Recombinant Enzymes
• Gene: Klk5
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 31 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 40-50 kDa, reducing conditions

Packaging, Storage & Formulations

• 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.
• Buffer: Supplied as a 0.2 μm filtered solution in Sodium Acetate, NaCl and Glycerol.
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
• Assay Procedure:
  • Assay Buffer: 50 mM Tris, 0.005% (w/v) Brij-35, pH 8.0
  • Recombinant Mouse Kallikrein 5 (rmKLK5) (Catalog # 7236-SE)
  • Recombinant Human Kallikrein 5 (rhKLK5) (Catalog # 1108-SE)
  • Substrate: Boc-Val-Pro-Arg-AMC (Catalog # ES011)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rmKLK5 to 200 µg/mL in Assay Buffer.
  2. Dilute rhKLK5 to 2.02 µg/mL in Assay Buffer.
  3. Combine equal volumes of 200 µg/mL rmKLK5 and 2.02 µg/mL rhKLK5. For a control, combine equal volumes of 2.02 µg/mL rhKLK5 and Assay Buffer.
  4. Incubate at 37 °C for 16 - 20 hours.
  5. Dilute activated rmKLK5 to 1 µg/mL in Assay Buffer. Perform same dilution on control.
  6. Dilute Substrate to 800 µM in Assay Buffer.
  7. In a plate, load 50 µL of 1 µg/mL rmKLK5 and control, and start the reaction by adding 50 µL of 800 µM Substrate to all wells.
  8. Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in kinetic mode for 5 minutes.
  9. 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 # A9891).

  Per Well:
  • rmKLK5: 0.050 µg
  • Substrate: 400 µM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Mouse Kallikrein 5 Protein, CF

• EC 3.4.21
• EC 3.4.21.4
• Kallikrein 5
• Kallikrein-like protein 2
• kallikrein-related peptidase 5
• KLK5
• KLKL2
• KLK-L2
• KLK-L2EC 3.4.21.-
• SCTE
• SCTEkallikrein-5
• Stratum corneum tryptic enzyme

Background

Tissue Kallikrein 5 (KLK5) is a secreted serine protease with trypsin‑like substrate specificity. KLK5 is highly expressed in the skin, and is also known as stratum corneum tryptic enzyme due to its high expression level in this tissue. KLK5 is thought to play an important role in the process of desquamation (1). KLK5 is a potential biomarker for ovarian and breast cancers (2). It may also be a physiological activator of KLK7 in the stratum corneum (3). Recombinant mouse KLK5 is purified as the proform, and can undergo autoactivation under the appropriate conditions. KLK5 enzymatic activity can be inhibited by the following recombinant human serpins: Serpin‑A1, ‑A5, ‑F2, and ‑G1 (Catalog # 1268-PI, 1266-PI, 1470-PI, and 2488-PI.

1. Brattsand M. and T. Egelrud (1999) J. Biol. Chem. 274:30033.
2. Yousef G.M. et al. (2003) Cancer Res. 63:3958.
3. Kim H. et al. (2001) Br. J. Cancer 84:543.

Bioinformatics

• Gene Symbol: Klk5
• Uniprot:
  • NP_081082()