Recombinant Human Kallikrein 7 Protein, CF

(6 citations)
(1 Review)
  
  • Purity
    >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
  • 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, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002). The specific activity  is >150 pmol/min/µg, as measured under the described conditions. See Activity Assay Protocol on www.RnDSystems.com.
  • Source
    Mouse myeloma cell line, NS0-derived Glu23-His252, with a C-terminal 10-His tag
  • Accession #
  • N-terminal Sequence
    Analysis
    Glu23
  • Structure / Form
    Pro form
  • Predicted Molecular Mass
    26 kDa
  • SDS-PAGE
    29 kDa and 32 kDa, reducing conditions
2624-SE
 
Formulation Supplied as a 0.2 μm filtered solution in HEPES and NaCl.
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.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.
Assay Procedure
Materials
  • Activation Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
  • Assay Buffer: 50 mM Tris, 150 mM NaCl, pH 8.5
  • Recombinant Human Kallikrein 7 (rhKLK7) (Catalog # 2624-SE)
  • Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
  • EDTA (Sigma, Catalog # E-4884)
  • Substrate: MCA-Arg-Pro Lys-Pro-Val-Glu-Nval-Trp-Arg-Lys(Dnp)-NH2 (Catalog # ES002)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhKLK7 to 200 μg/mL in Activation Buffer.
  2. Dilute Thermolysin to 20 μg/mL in Activation Buffer.
  3. Combine equal volumes of 200 μg/mL rhKLK7 and 20 μg/mL Thermolysin.
  4. Incubate at 37 °C for 2 hours.
  5. Stop reaction with an equal volume 100 mM EDTA diluted in Assay Buffer (Cf = 50 mM).
  6. Dilute activated rhKLK7 to 4 ng/μL in Assay Buffer.
  7. Dilute Substrate to 20 μM in Assay Buffer.
  8. Load 50 μL of 4 ng/μL rhKLK7 in a plate, and start the reaction by adding 50 μL of 20 μM Substrate to each well. Include a Substrate Blank containing 50 μL Assay Buffer and 50 μL of 20 μM Substrate.
  9. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
  10. 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 MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).

Per Well:
  • rhKLK7: 0.2 μg
  • Substrate: 10 μM
Background: Kallikrein 7
Human tissue Kallikrein 7 (hK7), also known as stratum corneum chymotryptic enzyme (SCCE), is a member of the human tissue kallikrein family. Full-length hK7 consists of 253 amino acids, with a signal peptide (residues 1‑22), short pro peptide (residues 23‑29) and mature chain (residues 30‑252) (1). Predominantly expressed in the skin, a major physiological function of hK7 is to regulate the desquamation process through proteolysis of the intercellular adhesive structures between corneocytes (2). Thus, it is related to some inflammatory skin diseases, such as psoriasis and chronic itchy dermatitis (3, 4). Studies have shown that one potential physiological activator for hK7 is hK5, another member of the human tissue Kallikrein family. Along with hK14, these three kallikreins form a proteolytic cascade in the stratum corneum (5). The purified, secreted recombinant human K7 corresponds to the pro form. When activated by thermolysin, it displays enzymatic activity towards a fluorogenic synthetic peptide described in the Activity Assay Protocol. This activity can be inhibited by recombinant human Serpin A1, A3, A4, and A5 (Catalog # 1268-PI, 1295-PI1669-PI, and 1266-PI).
  • References:
    1. Hansson, L. et al. (1994) J. Bio. Chem. 269:19420.
    2. Caubet, C. et al. (2004) J. Invest. Dermatol. 122:1235.
    3. Ekholm, E. and Egelrud, T. (1999) Arch. Dermatol. Res. 291:195.
    4. Hansson, L. et al. (2002) J. Invest. Dermatol. 118:444.
    5. Brattsand, M. et al. (2004) J. Invest. Dermatol. 124:198.
  • Entrez Gene IDs:
    5650 (Human); 23993 (Mouse)
  • Alternate Names:
    EC 3.4.21; EC 3.4.21.117; hK7; hSCCE; kallikrein 7 (chymotryptic, stratum corneum); Kallikrein 7; kallikrein-related peptidase 7; KLK7; protease, serine, 6; PRSS6; SCCEkallikrein-7; Serine protease 6; signal protein; Stratum corneum chymotryptic enzyme
Related Research Areas
Citations:

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.

6 Citations: Showing 1 - 6
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Applications
Sample Type
  1. Mesotrypsin and caspase-14 participate in prosaposin processing: potential relevance to epidermal permeability barrier formation.
    Authors: Yamamoto-Tanaka M, Motoyama A, Miyai M, Matsunaga Y, Matsuda J, Tsuboi R, Hibino T
    J Biol Chem, 2014;289(29):20026-38.
    Species: Human
    Sample Type: Whole Cells
    Application: Bioassay
  2. Proteolytic activation cascade of the Netherton syndrome-defective protein, LEKTI, in the epidermis: implications for skin homeostasis.
    Authors: Fortugno P, Bresciani A, Paolini C, Pazzagli C, El Hachem M, D'Alessio M, Zambruno G
    J Invest Dermatol, 2011;131(11):2223-32.
    Species: Human
    Sample Type: Substrate
    Application: EnzAct
  3. Matriptase initiates activation of epidermal pro-kallikrein and disease onset in a mouse model of Netherton syndrome.
    Authors: Sales KU, Masedunskas A, Bey AL
    Nat. Genet., 2010;42(8):676-83.
    Species: Human
    Sample Type: Recombinant Protein
    Application: Bioassay
  4. Kallikrein expression and cathelicidin processing are independently controlled in keratinocytes by calcium, vitamin D(3), and retinoic acid.
    Authors: Morizane S, Yamasaki K, Kabigting FD, Gallo RL
    J. Invest. Dermatol., 2010;130(5):1297-306.
    Species: N/A
    Sample Type: N/A
    Application: ELISA Standard
  5. Isolation of SPINK6 in human skin: selective inhibitor of kallikrein-related peptidases.
    Authors: Meyer-Hoffert U, Wu Z, Kantyka T, Fischer J, Latendorf T, Hansmann B, Bartels J, He Y, Glaser R, Schroder JM
    J. Biol. Chem., 2010;285(42):32174-81.
    Species: Human
    Sample Type: Peptide
    Application: Enzyme Assay
  6. Specific inhibition of tissue kallikrein 1 with a human monoclonal antibody reveals a potential role in airway diseases.
    Authors: Sexton DJ, Chen T, Martik D, Kuzmic P, Kuang G, Chen J, Nixon AE, Zuraw BL, Forteza RM, Abraham WM, Wood CR
    Biochem. J., 2009;422(2):383-92.
    Species: Human
    Sample Type: Antibody
    Application: Surface Plasmon Resonance
Fluorogenic Peptide Substrates
Description Application Cat# Citations Images  

Mca-RPKPVE-Nval-WRK(Dnp)-NH2 Fluorogenic MMP Substrate

BA, BA2, Bind ES002 8
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Natural Enzymes
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Bacterial Thermolysin Protein, CF

EnzAct 3097-ZN 7
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Primary Antibodies
Description Application Cat# Citations Images  

His Tag Antibody

WB, Simple Western, Flow, AP, CyTOF-ready MAB050 26  
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His Tag HRP-conjugated Antibody

WB, Simple Western MAB050H 3  
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His Tag APC-conjugated Antibody

ICFlow IC050A  
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His Tag Biotinylated Antibody

WB BAM050 4  
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His Tag PE-conjugated Antibody

ICFlow IC050P 2  
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His Tag Antibody

WB, Flow, AP MAB050R  
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His Tag Alexa Fluor® 488-conjugated Antibody

ICFlow IC050G  
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His Tag PerCP-conjugated Antibody

ICFlow IC050C  
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His Tag Alexa Fluor® 700-conjugated Antibody

ICFlow IC050N  
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Protein Purification Kits
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Histidine-tagged Protein Purification Resin

IP999 1
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ApplicationEnzymatic activity in vitro

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