Cloning and functional expression of a soluble form of kynurenine/alpha-aminoadipate aminotransferase from rat kidney.
Several aminotransferases with kynurenine aminotransferase (KAT) activity are able to convert L-kynurenine into kynurenic acid, a putative endogenous modulator of glutamatergic neurotransmission. In the rat, one of the described KAT isoforms has been found to correspond to glutamine transaminase K. In addition, rat kidney alpha-aminoadipate aminotransferase (AadAT) also shows KAT activity. In this report, we describe the isolation of a cDNA clone encoding the soluble form of this aminotransferase isoenzyme from rat (KAT/AadAT). Degenerate oligonucleotides were designed from the amino acid sequences of rat kidney KAT/AadAT tryptic peptides for use as primers for reverse transcription-polymerase chain reaction of rat kidney RNA. The resulting polymerase chain reaction fragment was used to screen a rat kidney cDNA library and to isolate a cDNA clone encoding KAT/AadAT. Analysis of the combined DNA sequences indicated the presence of a single 1275-base pair open reading frame coding for a soluble protein of 425 amino acid residues. KAT/AadAT appears to be structurally homologous to aspartate aminotransferase in the pyridoxal 5'-phosphate binding domain. RNA blot analysis of rat tissues, including brain, revealed a single species of KAT/AadAT mRNA of approximately 2.1 kilobases. HEK-293 cells transfected with the KAT/AadAT cDNA exhibited both KAT and AadAT activities with enzymatic properties similar to those reported for the rat native protein.
D051307:2-Aminoadipate Transaminase; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000302:isolation & purification; D007668:Kidney / Q000201:enzymology*; D008190:Lyases; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis; D000637:Transaminases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D011994:Recombinant Proteins; D000637:Transaminases; D051307:2-Aminoadipate Transaminase; C427605:glutamine - phenylpyruvate transaminase; C021481:kynurenine-oxoglutarate transaminase; D008190:Lyases