Testican 1, encoded by the SPOCK1 gene, is a proteoglycan first identified in human seminal plasma (1). The cDNAs from the human testis and mouse brain indicate 95% identity between the deduced amino acid sequences from the two species, predicting a conserved function (2, 3). Human Testican 1 is able to inhibit attachment of Neuro-2a cells and their ability to form neurite extensions (4). At R&D Systems, recombinant human (rh) Testican 1 has also been shown to be able to enhance neurite outgrowth of E18 rat embryonic hippocampal neurons. Testican 1 contains Ca2+-binding domain and the C-terminal acidic domain with putative glycosaminoglycan attachment sites (5). In addition, it contains three potential inhibitory domains targeted toward three different classes of proteases, metallo, cysteine and serine proteases. The N-terminal region, which is unique to testicans, is responsible for the inhibition of testican 1 towards MMP-14 (MT1-MMP, a metalloprotease) activation of MMP-2 (6). The thyropin domain may be responsible for the inhibition of testican 1 towards cathepsin L, a cysteine protease (5). The follistatin-like domain with a six cysteine Kazal-like motif may inhibit serine proteases. The purified rhTestican 1 is capable of inhibiting rhMMP-14 and rhCathepsin L (Catalog # http://www.rndsystems.com/product_results.aspx?k=918-MP">918-MP and http://www.rndsystems.com/product_results.aspx?k=952-CY">952-CY) in assays using the fluorogenic peptide substrates (Catalog # http://www.rndsystems.com/product_results.aspx?k=ES001">ES001 and http://www.rndsystems.com/product_results.aspx?k=ES008">ES008).
Recombinant Human Testican 1/SPOCK1 Protein, CF
R&D Systems | Catalog # 2327-PI
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Key Product Details
- R&D Systems NS0-derived Recombinant Human Testican 1/SPOCK1 Protein (2327-PI)
- Quality control testing to verify active proteins with lot specific assays by in-house scientists
- All R&D Systems proteins are covered with a 100% guarantee
Source
NS0
Accession Number
Applications
Inhibition Activity
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Product Specifications
Source
Mouse myeloma cell line, NS0-derived human Testican 1/SPOCK1 protein
Arg22-Trp439, with a C-terminal 6-His tag
Arg22-Trp439, with a C-terminal 6-His tag
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by silver stain.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
Arg22
Predicted Molecular Mass
48 kDa
SDS-PAGE
50-56 kDa, reducing conditions
Activity
Measured by its ability to inhibit active Cathepsin L cleavage of a fluorogenic peptide substrate Z-LR-AMC (Catalog # ES008).
The IC50 value is <10 nM, as measured under the described conditions.
The IC50 value is <10 nM, as measured under the described conditions.
Formulation, Preparation, and Storage
2327-PI
| Formulation | Lyophilized from a 0.2 μm filtered solution in PBS. |
| Reconstitution | Reconstitute at 100 μg/mL in sterile PBS. |
| Shipping | The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Calculators
Background: Testican 1/SPOCK1
References
- Bonnet, F. et al. (1992) Biochem. J. 288:565.
- Alliel, P.M. et al. (1993) Eur. J. Biochem. 214:347.
- Bonnet, F. et al. (1996) J. Biol. Chem. 271:565.
- Marr, H.S. and C.-J.S Edgell (2003) Matrix Biol. 22:259.
- Bocock, J.P. et al. (2003) Eur. J. Biochem. 270:4008.
- Nakada, M. et al. (2001) Cancer Res. 61:8896.
Alternate Names
SPOCK1, TIC1, TICN1
Entrez Gene IDs
6695 (Human)
Gene Symbol
SPOCK1
UniProt
Additional Testican 1/SPOCK1 Products
Product Documents for Recombinant Human Testican 1/SPOCK1 Protein, CF
Certificate of Analysis
To download a Certificate of Analysis, please enter a lot or batch number in the search box below.
Note: Certificate of Analysis not available for kit components.
Product Specific Notices for Recombinant Human Testican 1/SPOCK1 Protein, CF
For research use only
Related Research Areas
Citations for Recombinant Human Testican 1/SPOCK1 Protein, CF
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Protocols
View specific protocols for Recombinant Human Testican 1/SPOCK1 Protein, CF (2327-PI):
Materials
- Assay Buffer: 50 mM MES, 5 mM DTT, pH 6.0
- Recombinant Human Testican 1/SPOCK1 (rhTestican 1) (Catalog # 2327-PI)
- Recombinant Human Cathepsin L (rhCathepsin L) (Catalog # 952-CY)
- Substrate: Z-Leu-Arg-AMC (Catalog # ES008)
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhCathepsin L to 1 µg/mL in Assay Buffer.
- Incubate diluted rhCathepsin L on ice for 15 minutes.
- Dilute incubated rhCathepsin L to 0.25 µg/mL in Assay Buffer.
- Prepare a curve of rhTestican 1 (MW: 47,800 Da) in Assay Buffer. Make the following serial dilutions: 500, 250, 125, 83.3, 55.6, 37, 24.7, and 12.3 nM.
- Combine equal volumes of diluted rhCathepsin L and rhTestican 1 at each concentration of the curve. Include two controls containing equal volumes of Assay buffer and diluted rhCathepsin L without any rhTestican 1.
Incubate mixtures at 37 ˚C for 15 minutes. - Dilute the reaction mixtures 1/5 in Assay Buffer.
- Dilute Substrate to 20 µM in Assay Buffer.
- Load 50 µL of the incubated mixtures in a plate, and start the reaction by adding 50 µL of 20 µM Substrate.
- Read at excitation and emission wavelengths of 380 nm and 460 nm (top read), respectively, in kinetic mode for 5 minutes.
- Derive the 50% inhibition concentration (IC50) value for rhTestican 1 by plotting RFU/min (or specific activity) vs. concentration with 4‑PL fitting.
- The specific activity for rhCathepsin L at each point may be determined using the following formula (if needed):
|
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 # A-9891).
Per Well:
- rhCathepsin L: 0.00125 µg
- rhTestican 1 curve: 25, 12.5, 6.25, 4.165, 2.78, 1.85, 1.235, and 0.615 nM
- Substrate: 10 µM
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