Abstract
BackgroundKynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA). KYNA is a neuroactive compound and functions as an antagonist of alpha7-nicotinic acetylcholine receptors and is the only known endogenous antagonist of N-methyl-D-aspartate receptors. Four KAT enzymes, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase, have been reported in mammalian brains. Because of the substrate overlap of the four KAT enzymes, it is difficult to assay the specific activity of each KAT in animal brains.ResultsThis study concerns the functional expression and comparative characterization of KAT I, II, III, and IV from mice. At the applied test conditions, equimolar tryptophan with kynurenine significantly inhibited only mouse KAT I and IV, equimolar methionine inhibited only mouse KAT III and equimolar aspartate inhibited only mouse KAT IV. The activity of mouse KAT II was not significantly inhibited by any proteinogenic amino acids at equimolar concentrations. pH optima, temperature preferences of four KATs were also tested in this study. Midpoint temperatures of the protein melting, half life values at 65°C, and pKa values of mouse KAT I, II, III, and IV were 69.8, 65.9, 64.8 and 66.5°C; 69.7, 27.4, 3.9 and 6.5 min; pH 7.6, 5.7, 8.7 and 6.9, respectively.ConclusionThe characteristics reported here could be used to develop specific assay methods for each of the four murine KATs. These specific assays could be used to identify which KAT is affected in mouse models for research and to develop small molecule drugs for prevention and treatment of KAT-involved human diseases.
Highlights
Kynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA)
In addition to its roles as an excitatory amino acid and α7-nicotinic acetylcholine antagonist, KYNA is involved in the control of the cardiovascular function by acting at the rostral ventrolateral medulla of the central nervous system (CNS)[11]
Freezing and thawing did not alter the activity of the isolated enzymes. mouse KAT (mKAT) I showed maximum activity at 60 to 70°C (Fig. 1a), mKAT II showed maximum activity at 50°C (Fig. 1a), both mKAT III and mKAT IV (Fig. 1b) showed maximum activity at 60°C
Summary
Kynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate subtype of glutamate receptors[1,2,3,4] It is an antagonist of the α7-nicotinic acetylcholine receptor[58]. Abnormal concentration of KYNA in cerebrospinal fluid/brain tissue has been observed in patients with mental and neurological disorders, including the Huntington's disease, Alzheimer's disease, schizophrenia, multiple sclerosis and others (for a review see [10]). These data suggest that KYNA, acting as an endogenous modulator of glutamatergic and cholinergic neurotransmission, may be functionally significant in the development and progression of these diseases. Hypertensive rat, the most widely used animal model for studying genetic hypertension, is associated with abnormally low KYNA levels in the area of CNS which controls physiological blood pressure[12,13]
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