Kynurenine aminotransferase II (KAT‑II), also known as alpha ‑Aminoadipate Aminotransferase (AADAT), catalyzes the PLP‑dependent transamination of aminoadipate to alpha‑oxoadipate in the catabolism of lysine in the liver and also is the primary brain enzyme catalyzing the transamination of kynurenine to kynurenic acid (KYNA) (1). KYNA is an endogenous antagonist of the N‑methyl‑D‑aspartate (NMDA) receptors with weaker effects on kainite and alpha‑amino‑3‑hydroxy‑5-methyl‑4‑isoxazole (AMPA) receptors, the other two ionotropic glutamate receptors (2, 3). KYNA also acts upon alpha‑7 nicotinic acetylcholine receptors ( alpha 7 nAChRs) and importantly may suppress the pre-synaptic release of glutamate to confer neuroprotective effects against NMDA‑receptor mediated over‑stimulation. Also, KYNA is an endogenous ligand of the orphaned G protein-coupled receptor 35 (GPR35), found primarily in immune cells, and may induce inositol phosphate production and Ca2+ mobilizaiton (4). Elevated levels of KYNA have been implicated in Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, epilepsy, schizophrenia and cognitive impairment (3, 5).
Recombinant Human alpha-Aminoadipate Aminotransferase, CF
R&D Systems | Catalog # 7927-AT
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Key Product Details
- R&D Systems E. coli-derived Recombinant Human alpha-Aminoadipate Aminotransferase (7927-AT)
- 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
E. coli
Accession Number
Applications
Enzyme Activity
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Product Specifications
Source
E. coli-derived human alpha-Aminoadipate Aminotransferase protein
Met1-Leu425, with an N-terminal Gly-Arg-Ala-His
Met1-Leu425, with an N-terminal Gly-Arg-Ala-His
Purity
>90%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
N-terminal Sequence Analysis
Gly
Predicted Molecular Mass
48 kDa
SDS-PAGE
40-42 kDa, reducing conditions
Activity
Measured by its ability to form Kynurenic Acid from Kynurenine and alpha-Ketoglutarate.
The specific activity is >170 pmol/min/μg, as measured under the described conditions.
The specific activity is >170 pmol/min/μg, as measured under the described conditions.
Formulation, Preparation, and Storage
7927-AT
| 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: alpha-Aminoadipate Aminotransferase
References
- Han, Q. et al. (2008) Biosci. Rep. 28:205.
- Passera, E. et al. (2011) FEBS J. 278:1882.
- Vecsei, L. et al. (2013) Nat. Rev. Drug Discov. 12:64.
- Wang, J. et al. (2006) J. Biol. Chem. 281:22021.
- Rossi, F. et al. (2008) J. Biol. Chem. 283:3559.
Long Name
alpha-Aminoadipate Aminotransferase
Alternate Names
AADAT, alphaAminoadipate Aminotransferase, KAT2
Gene Symbol
AADAT
UniProt
Additional alpha-Aminoadipate Aminotransferase Products
Product Documents for Recombinant Human alpha-Aminoadipate Aminotransferase, 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 alpha-Aminoadipate Aminotransferase, CF
Coomassie is a registered trademark of Imperial Chemical Industries Ltd.
For research use only
Related Research Areas
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Protocols
View specific protocols for Recombinant Human alpha-Aminoadipate Aminotransferase, CF (7927-AT):
Materials
- Assay Buffer: 20 mM HEPES, 50 mM NaCl, pH 8.0
- Recombinant Human alpha ‑Aminoadipate Aminotransferase (rhAADAT) (Catalog # 7927-AT)
- L-Kynurenine (Sigma, Catalog # K8625), 40 mM stock in 80 mM HEPES, 8 mM HCl, pH 8.0
- alpha -Ketoglutarate (Sigma, Catalog # K2010), 1 M stock in diH2O
- Pyridoxal 5’-phosphate (Sigma, Catalog # P9255), 100 mM stock in 1 M HEPES, pH 8.0
- KYNA Development Mixture: 2 M ZnCl2, 2 M NaOAc in diH2O
- Standard: Kynurenic acid (Catalog # 0223), 50 mM stock in 100 mM NaOH
- F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
- Fluorescent Plate Reader (Model: Gemini EM by Molecular Devices) or equivalent
- Dilute 50 mM Kynurenic acid standard to 400 µM in Assay Buffer. This is the first point of the standard curve.
- Perform six additional one-half serial dilutions in Assay Buffer. The standard curve has a range of 156‑10,000 pmol per well.
- Dilute rhAADAT to 5 µg/mL in Assay Buffer.
- Prepare Substrate Component Solution containing 50 mM HEPES, 20 mM alpha -ketoglutarate, 80 µM Pyridoxal 5’-phosphate, pH 8.0.
- Immediately before use, prepare Substrate Mixture by combining 25 µL of Kynurenine with 50 µL of Substrate Component Solution, per well assayed.
- Load 25 µL of each standard curve dilution into empty wells of a black well plate.
- Load 25 µL of dilute rhAADAT into empty wells. Include a Blank containing 25 µL of Assay Buffer. This will serve as both enzyme and standard curve blank.
- Add 75 µL of Substrate Mixture to all wells used.
- Cover the plate with a plate sealer and incubate at room temperature for 30 minutes.
- Add 100 µL of KYNA Development Mixture to all wells used.
- Read plate at excitation and emission wavelengths of 338 and 405 nm, respectively, in endpoint mode.
- Calculate specific activity:
|
Specific Activity (pmol/min/µg) = |
Adjusted Kynurenic acid produced* (pmol) |
| Incubation time (min) x amount of enzyme (µg) |
*Derived from the Kynurenic acid curve using linear fitting and adjusted for Blank.
Per Reaction:
- rhAADAT: 0.125 µg
- alpha -ketoglutarate: 10 mM
- Pyridoxal 5’-phosphate: 40 µM
- Kynurenine: 10 mM