Recombinant Human Kallikrein 11 Protein, CF

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Product Details

Summary
Reactivity HuSpecies Glossary
Applications Enzyme Activity
Format
Carrier-Free

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Recombinant Human Kallikrein 11 Protein, CF Summary

Details of Functionality
Measured by its ability to cleave a colorimetric peptide substrate D-Val-Leu-Lys-ThioBenzyl ester (VLK-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 >200 pmol/min/µg, as measured under the described conditions.
Source
Mouse myeloma cell line, NS0-derived human Kallikrein 11 protein
Glu19-Asn250, with an N-terminal signal peptide and a C-terminal 10-His tag
Accession #
N-terminal Sequence
Glu19
Structure / Form
Pro form
Protein/Peptide Type
Recombinant Enzymes
Gene
KLK11
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Endotoxin Note
<1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

Theoretical MW
27 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 kDa, reducing conditions
Publications
Read Publications using
1595-SE in the following applications:

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 Tris, NaCl and CaCl2.
Purity
>95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
Assay Procedure
  • Activation Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, pH 7.5 (TCN)
  • Assay Buffer: 50 mM Tris, 1 M NaCl, 10 mM EDTA, pH 8.5
  • Recombinant Human Kallikrein 11 (rhKLK11) (Catalog # 1595-SE)
  • Bacterial Thermolysin (Thermolysin) (Catalog # 3097-ZN)
  • EDTA (Sigma, Catalog # E-4884)
  • Fluorogenic Peptide Substrate: Z-Lys-SBzl (Bachem, Catalog # M-1300), 10 mM stock in DMSO
  • 5,5’-dithio-bis (2-nitrobenzoic acid) (DTNB) (Sigma, Catalog # D-8130), 10 mM stock in DMSO
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute rhKLK11 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 rhKLK11 and 20 µg/mL Thermolysin.
  4. Incubate at 37 °C for 15 minutes.
  5. Stop Thermolysin activity by adding EDTA to a final concentration of 10 mM.
  6. Dilute incubated rhKLK11 to 2 ng/µL in Assay Buffer.
  7. Prepare Substrate Mixture containing 600 μM Substrate and 200 μM DTNB in Assay Buffer.
  8. Load into a clear plate 50 µL of the 2 ng/µL rhKLK11, and start the reaction by adding 50 µL of Substrate Mixture. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of Substrate Mixture.
  9. Read at a wavelength of 405 nm (bottom read) in kinetic mode for 5 minutes.
  10. 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 0.320 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rhKLK11: 0.1 µg
  • Substrate: 300 µM
  • DTNB: 100 µM

Notes

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

Alternate Names for Recombinant Human Kallikrein 11 Protein, CF

  • EC 3.4.21
  • EC 3.4.21.-
  • EC 3.4.21.4
  • Hippostasin
  • hK11
  • Kallikrein 11
  • kallikrein-related peptidase 11
  • KLK11
  • MGC33060
  • PRSS20kallikrein-11
  • Serine protease 20
  • TLSPhippostasin
  • Trypsin-like protease

Background

As a member of human tissue kallikrein family, Kallikrein 11, also known as hippostasin, trypsin-like serine protease and PRSS20, is encoded by the KLK11 gene (1). Two alternatively spliced forms exist, resulting in 250 (isoform 1) and 282 (isoform 2) amino acid sequences,respectively (2-5). Isoform 1 consists of a signal peptide (residues 1-18), a short pro peptide (residues 19-21) and the mature chain (residues 22-250). Isoform 2 is identical to isoform 1, except that a 32 amino acid segment is inserted in isoform 2 before residue 1 in isoform 1. Isoform 1 is predominantly expressed in brain whereas isoform 2 is preferentially expressed in prostate. KLK11 is a novel marker for ovarian and prostate cancer carcinomas (6-8). Recombinant human KLK11, after being activated by thermolysin, is active against a thioester substrate. This activity can be inhibited by AEBSF (Catalog # EI001), dichloroisocoumarin, and aprotinin. Recombinant human KLK11 produced by R&D Systems corresponds to isoform 1.

  1. Yousef, G.M. and E.P. Diamandis (2001) Endocrine Rev. 22:184.
  2. Yoshida, S. et al. (1998) Biochim. Biophys. Acta 1399:225. 
  3. Yousef, G.M. et al. (2000) Genomics 63:88.
  4. Mitsui, S. et al. (2000) Biochem. Biophys. Res. Commun. 272:205.
  5. Gan, L. et al. (2000) Gene 257:119.
  6. Diamandis, E.P. et al. (2002) Cancer Res. 62:295.
  7. Nakamura, T. et al. (2003) Urology 61:1042.
  8. Borgono, C.A. et al. (2003) Int. J. Cancer 106:605.

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Publications for Kallikrein 11 (1595-SE)(4)

We have publications tested in 2 confirmed species: Human, N/A.

We have publications tested in 3 applications: Bioassay, Enzyme Assay, Surface Plasmon Resonance.


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(1)
Enzyme Assay
(2)
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(1)
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Bioinformatics

Gene Symbol KLK11
Uniprot