Vascular adhesion protein-1 (VAP-1) is a copper amine oxidase with a topaquinone cofactor. VAP-1 is a Type II integral membrane protein, but a soluble form of the enzyme is present in human serum, and its level increases in diabetes and some inflammatory liver diseases (1, 2). VAP-1 catalyzes the oxidative deamination of small primary amines such as methylamine, benzylamine, and aminoacetone in a reaction that produces an aldehyde, ammonia, and H2O2 (3). The enzyme is sensitive to inhibition by semicarbazide. VAP-1 expression is highest in the endothelium of lung, heart, and intestine, but low in tissues such as brain, spleen, kidney, and liver (4). VAP-1 vascular expression is regulated at sites of inflammation through its release from intracellular granules in which the protein is stored (5). The adhesive function of VAP-1 has been demonstrated in studies showing that the protein is important for the adherence of certain lymphocyte subtypes to inflamed endothelial tissues (6). VAP-1 mediated adhesion is involved in the process of leukocyte extravasation, an important feature of inflammatory responses. The role of VAP-1 amine oxidase activity in this process is not fully defined, but it appears to be carbohydrate-dependent (7). VAP-1 is considered to be a therapeutic target for diabetes, oxidative stress, and inflammatory diseases (8).
Recombinant Human VAP-1 Protein, CF
R&D Systems | Catalog # 3957-AO
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Key Product Details
- R&D Systems Sf 21 (baculovirus)-derived Recombinant Human VAP-1 Protein (3957-AO)
- 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
Sf 21 (baculovirus)
Accession Number
Structure / Form
Disulfide-linked homodimer
Applications
Enzyme Activity
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Product Specifications
Source
Spodoptera frugiperda, Sf 21 (baculovirus)-derived human VAP-1/AOC3 protein
Gly27-Asn763, with an N-terminal Met and 10-His tag, followed by a Gly-Gly-Gly-Ser-Gly-Gly-Gly-Ser-Gly-Gly-Gly-Ser-Ile-Glu-Gly-Arg linker
Gly27-Asn763, with an N-terminal Met and 10-His tag, followed by a Gly-Gly-Gly-Ser-Gly-Gly-Gly-Ser-Gly-Gly-Gly-Ser-Ile-Glu-Gly-Arg linker
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.
N-terminal Sequence Analysis
Met
Predicted Molecular Mass
85 kDa
SDS-PAGE
80-90 kDa, reducing conditions
Activity
Measured by its ability to produce hydrogen peroxide during the oxidation of benzylamine.
The specific activity is >8 pmol/min/μg, as measured under the described conditions.
The specific activity is >8 pmol/min/μg, as measured under the described conditions.
Formulation, Preparation, and Storage
3957-AO
| 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.
|
Background: VAP-1/AOC3
References
- Kurkijärvi, R. et al. (1998) J. Immunol. 161:1549.
- Gearing, A.J.H. and W. Newman (1993) Immunol. Today 14:506.
- Lizcano, J.M. et al. (1998) Biochem. J. 331:69.
- Smith, D.J. et al. (1998) J. Exp. Med. 188:17.
- Jaakkala K. et al. (2000) Am. J. Pathol. 157:463.
- Salmi, M. and J. Jalkanen (2001) Trends Immunol. 22:211.
- Salmi, M. and J. Jalkanen (1996) J. Exp. Med. 183:569.
- Dunkel, P. et al. (2008) Curr. Med. Chem. 15:1827.
Long Name
Vascular Adhesion Protein-1
Alternate Names
AOC3, HPAO, SSAO, VAP1
Gene Symbol
AOC3
UniProt
Additional VAP-1/AOC3 Products
Product Documents for Recombinant Human VAP-1 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 VAP-1 Protein, CF
For research use only
Related Research Areas
Citations for Recombinant Human VAP-1 Protein, CF
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Protocols
View specific protocols for Recombinant Human VAP-1 Protein, CF (3957-AO):
Materials
- Assay Buffer: 50 mM HEPES, pH 7.5
- Recombinant Human VAP‑1/AOC3 (rhVAP‑1) (Catalog # 3957-AO)
- Coupling Enzyme: Horseradish Peroxidase (HRP) (250 - 330 U/mg) (Sigma, Catalog # P8375), 250 units/mL stock in 0.1 M Sodium Phosphate, pH 8.0
- Substrate Component 1: Benzylamine (Sigma, Catalog # B5136), 100 mM stock in deionized water
- Substrate Component 2: Amplex® Ultra Red (AUR) (Molecular Probes, Catalog # A36006), 10 mM stock in DMSO
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
- Dilute rhVAP-1 to 20 ng/µL in Assay Buffer.
- Prepare the Substrate mixture 2 mM Benzylamine, 2 units/mL HRP and 100 µM AUR in Assay Buffer.
- In a plate, load 50 µL of 20 ng/µL rhVAP-1 and start the reaction by adding 50 µL of the Substrate mixture (step 2). Include a Substrate Blank containing 50 µL of the Assay Buffer and 50 µL of the Substrate mixture.
- Read at excitation and emission wavelengths of 544 nm and 590 nm (top read), respectively in kinetic mode for 5 minutes. Note: A cutoff must be set at a wavelength of 570 nm.
- 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 a fluorescent standard prepared by incubating 50 µM AUR, 1 unit/mL HRP, 1 mM Benzylamine, and a curve of
Hydrogen Peroxide (Sigma, Catalog # H1009) in Assay Buffer. Use this oxidized AUR curve to determine the conversion factor.
- rhVAP-1: 1.0 µg
- Benzylamine: 1 mM
- HRP: 1 unit/mL
- AUR: 50 µM
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