- Other Names
- Kallikrein Related Peptidase 5 (KLK5), Kallikrein-Like Protein 2 (KLK-L2), KLKL2, Stratum Corneum Tryptic Enzyme (SCTE)
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Human KLK5 belongs to the kallikrein gene family of serine proteases that includes 15 members and colocalize in tandem in chromosome 19. Kallikreins are translated inactive preproenzymes. Human KLK5 possesses a 29-amino acid signal peptide, a 37-amino acid activation peptide, and 237 amino acids containing the mature enzyme. KLK5 was initially identified in the normal human epidermis as the stratum corneum tryptic enzyme, where it takes part in the degradation of corneodesmosomes in the most external layer of human skin, giving as a result skin desquamation. KLK5 has been associated with breast, ovarian, prostate, and testicular tumors. Human KLK5 induces shedding of DPP4 from CD4+ Th17 cells from patients with Type 2 diabetes mellitus (T2DM). CD4+ T cells from patients with T2DM are the source of KLK5 that acts in the paracrine and autocrine systems. KLK5 is induced by cytokines in neutrophils, and it has been suggested that neutrophil could be associates to the shedding of DPP4 from Th17 cells.Product Details
- Human KLK5, amino acid (Ile67- Ser293) (Accession: # Q9Y337), with a C-terminal GGS-8H tag was expressed in CHO cells.
- Molecular Mass
- The 236 amino acid recombinant protein has a predicted molecular mass of approximately 26.1 kD. The DTT-reduced and non-reduced protein migrates at approximately 60 and 50 kD by SDS-PAGE respectively. The predicted N-terminal amino acid is Ile.
- > 95%, as determined by Coomassie stained SDS-PAGE
- 0.22 µm filtered protein solution is in 25 mM MES, 150 mM NaCl, 20% glycerol pH 5.5
- Endotoxin Level
- Less than 0.1 EU per µg cytokine as determined by the LAL method
- 10 and 25 µg sizes are bottled at 200 µg/mL.
- Storage & Handling
- Unopened vial can be stored at -20°C or -70°C for six months. For maximum results, quick spin vial prior to opening. Avoid repeated freeze/thaw cycles.
- Recombinant human KLK5 activity is measured by its ability to cleave the fluorogenic peptide substrate Boc-Val-Pro-Arg-AMC. The specific activity is > 200 pmol/min/µg in the presence of 0.1 µg of recombinant human KLK5.
- Application Notes
Human KLK5 Activity Assay
Recombinant Human KLK5 activity is measured by its ability to cleave the fluorogenic peptide substrate Boc-Val-Pro-Arg-AMC. The increase of the product is monitored by an increase in the intensity of fluorescence at 460 nm with excitation at 380 nm. Perform assay rapidly. The specific activity is > 200 pmol/min/µg in the presence of 0.1 µg of recombinant human KLK5.
- Assay buffer: 50 mM Tris, 0.005% (w/v) Brij-35, pH 8.0
- Recombinant human KLK5
- Fluorogenic peptide substrate: Boc-Val-Pro-Arg-AMC
- F16 Black Maxisorp Plate (Nunc, Cat. No. 475515)
Activity Assay Procedures
- Dilute the substrate in assay buffer at 200 µM. Wrap the substrate container in aluminum foil.
- In assay buffer, prepare 50 µL of serial dilutions of human KLK5. Start at a concentration of 16 µg/mL and dilute 1 to 2 six times after.
- Add 50 µL of 200 µM substrate to wells with 50 µL of prepared dilutions.
- Include a substrate black with 50 µL of assay buffer and 50 µL of 200 µM substrate without any human KLK5.
- Read at excitation and emission wavelengths of 380 nm and 460 nm, respectively, in kinetic mode for 5 minutes.
- Calculate specific activity using the following equation:
Specific Activity (pmol/min/µg)= ((Adjusted Vmax* (RFU/min) x Conversion Factor** (pmol/RFU))
amount of enzyme (µg)
Final assay conditions per well:
hKLK5: 0.8, 0.4, 0.2, 0.1, 0.05, 0.025, 0.0125 µg
Substrate: 100 µM
BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are verified in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at firstname.lastname@example.org.
Highly expressed in endocrine or hormone-responsive tissues, ovary, testis, breast, and skin (epidermis). Secreted by T cells and neutrophils.
- Involved in skin desquamation, skin physiology. Upregulated by estrogens and progestins. Inhibited by serpins alpha(2)-antiplasmin and antithrombin. Inhibited by LEKTI fragments.
- DPP4 (CD26)
- Substrates: plasminogen, kininogen, fibrinogen, vitronectin, laminin, and collagens type I, II, III, IV
- Human KLK5 activity is measured by its ability to cleave fluorogenic peptide substrate Boc-Val-Pro-Arg-AMC.
- Cell Type
- Neutrophils, T cells, Th17
- Biology Area
- Cancer Biomarkers, Immuno-Oncology
- Molecular Family
- Enzymes and Regulators, Proteases
- Antigen References
- Brattsand M & Egelrud T. 1999. J Biol Chem. 274:30033-40.
- Komatsu N, et al. 2003. J Invest Dermatol. 121:542-9.
- Yousef GM, et al. 2004. Br J Cancer. 90:167-72.
- Michael IP, et al. 2005. J Biol Chem. 280:14628-35.
- Lizama AJ, et al. 2015. Innate Immun. 6:575-86.
- Nargis T, et al. 2017. Mol Metab. 6:1529-1539.
- Gene ID
- 25818 View all products for this Gene ID
- View information about KLK5 on UniProt.org
- Does specific activity of a recombinant protein vary between lots?
Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.
- Have your recombinants been tested for stability?
Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.
- How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?
- Use formula Specific activity (Units/mg) = 10e6/ ED50 (ng/mL)
- How does the activity of your recombinant proteins compare to competitors?
We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!
- What is the specific activity or ED50 of my recombinant protein?
The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.