Two alternatively spliced forms of HAI-2 have been found in human tissues (1). HAI-2A, the full-length molecule and also known as placental bikunin, is a major form expressed in human tissues. Encoded by the SPINT2 gene, HAI-2A consists of two Kunitz domains, and a C-terminal transmembrane domain (1‑5). Both Kunitz domains can function as inhibitors independent of each other. In addition to HGF activator and trysin, HAI-2A strongly inhibits plasmin, tissue and plasma kallikreins, and factor XIa. In comparison, HAI-2A is a weaker inhibitor of factor VIIa-tissue factor, factors IXa, Xa, and XIIa. Recombinant HAI-2A prolonged the clotting time in an activated partial thromboplastin time assay.
Recombinant Human HAI-2A Protein, CF
R&D Systems | Catalog # 1106-PI
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Key Product Details
- R&D Systems NS0-derived Recombinant Human HAI-2A Protein (1106-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 HAI-2A protein
Met1-Lys197, with a C-terminal 10-His tag
Met1-Lys197, with a C-terminal 10-His tag
Purity
>95%, 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
Ala28
Predicted Molecular Mass
21 kDa
SDS-PAGE
27 kDa and 32 kDa, reducing conditions
Activity
Measured by its ability to inhibit trypsin cleavage of a fluorogenic peptide substrate, Mca-RPKPVE-Nval-WRK(Dnp)-NH2 (Catalog # ES002).
The IC50 value is <4.6 nM, as measured under the described conditions.
The IC50 value is <4.6 nM, as measured under the described conditions.
Formulation, Preparation, and Storage
1106-PI
| Formulation | Lyophilized from a 0.2 μm filtered solution in Tris, NaCl and CaCl2. |
| Reconstitution | Reconstitute at 100 μg/mL in sterile, deionized water.
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| 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: HAI-2A
References
- Kataoka, H. et al. (2002) Biochem. Biophys. Res. Comm. 290:1096.
- Kawaguchi, T. et al. (1997) J. Biol. Chem. 272:27558.
- Marlor. C.W. et al. (1997) J. Biol. Chem. 272:12202.
- Muller-Pillasch, F. et al. (1998) Biochim. Biophys. Acta 1395:88.
- Delaria, K.A. et al. (1997) J. Biol. Chem. 272:12209.
Long Name
HGF Activator Inhibitor Type 2A
Alternate Names
HAI2A, Placental Bikunin, SPINT2
Entrez Gene IDs
10653 (Human)
Gene Symbol
SPINT2
UniProt
Additional HAI-2A Products
Product Documents for Recombinant Human HAI-2A 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 HAI-2A Protein, CF
For research use only
Related Research Areas
Citations for Recombinant Human HAI-2A Protein, CF
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Protocols
View specific protocols for Recombinant Human HAI-2A Protein, CF (1106-PI):
Materials
- Assay Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5 (TCNB)
- Recombinant Human HAI-2A (rhHAI-2A) (Catalog # 1106-PI)
- Trypsin (Sigma, Catalog # T-1426)
- Substrate: MCA-Arg-Pro-Lys-Pro-Val-Glu-NVAL-Trp-Arg-Lys(DNP)-NH2 (Catalog # ES002) ), 2 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute Trypsin to 0.25 µg/mL with Assay Buffer.
- Prepare a curve of rhHAI-2A (MW: 20539 Da) in Assay Buffer. Make the following serial dilutions: 1000, 100, 33.3, 16.7, 8.33, 4.17, 2.08, and 0.417 nM.
- Mix equal volumes of the rhHAI-2A curve dilutions and the diluted Trypsin. Include a control (in duplicate) containing Assay Buffer and the diluted Trypsin without any rhHAI-2A.
- Incubate reactions for 30 minutes at room temperature.
- After incubation, dilute the mixtures by 5 fold in Assay Buffer.
- Dilute Substrate to 20 µM in Assay Buffer.
- Load 50 µL of the diluted incubated mixtures into a plate, and start the reaction by adding 50 µL of 20 µM Substrate.
- Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in kinetic mode for 5 minutes.
- Derive the 50% inhibition concentration (IC50) by plotting RFU/min (or specific activity) vs. concentration with 4-PL fitting.
- The specific activity for Trypsin 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 MCA-Pro-Leu-OH (Bachem, Catalog # M-1975)
Per Well:
- Trypsin: 0.00125 µg
- rhHAI-2A curve: 50, 5, 1.67, 0.83, 0.417, 0.209, 0.104, and 0.0209 nM
- Substrate: 10 µM
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