Characterization of rat brain kynurenine aminotransferases I and II.
The endogenous neuroprotectant kynurenic acid (KYNA) is produced by irreversible transamination of L-kynurenine (KYN). In the brain, two distinct kynurenine aminotransferases (KAT I and KAT II) are responsible for the formation of KYNA. The present experiments were designed to examine the respective roles of the two KATs in the normal rat brain. To this end, the two enzymes were partially purified, and their characteristics were examined. KAT I (identical with glutamine transaminase K) had an optimal pH of 9.5, preferred pyruvate as a cosubstrate and was potently inhibited by glutamine. KAT II (identical with L-alpha-aminoadipate transaminase) had a neutral optimal pH, showed no preference for pyruvate, and was essentially insensitive to inhibition by glutamine. KAT II was selectively inhibited by quisqualic acid (IC50: 520 microM). The endogenous substrate 3-hydroxykynurenine had an approximately 10-fold preference for KAT II. The distinct properties of the two enzymes made it possible to measure brain KAT I and KAT II in parallel by using dialyzed tissue homogenate (to remove interfering endogenous amino acids). Under these conditions, both enzymes presented essentially the same apparent Km values as the partially purified enzymes. In lesioned, neurondepleted brain tissue and in brain regions other than the cerebellum, KYNA derived primarily from KAT II at physiologic pH. In summary, the present study describes a simple methodology for the simultaneous determination of the two KYNA-producing enzymes in small rat brain tissue samples and provides baseline values for future work in experimentally challenged animals.
Guidetti
P
P
Maryland Psychiatric Research Center, University of Maryland School of Medicine, Baltimore 21228, USA.
Okuno
E
E
Schwarcz
R
R
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002852:Chromatography, Ion Exchange; D003342:Corpus Striatum / Q000187:drug effects / Q000201:enzymology; D007839:Functional Laterality; D005968:Glutamate Decarboxylase / Q000378:metabolism*; D005973:Glutamine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007051:Ibotenic Acid / Q000633:toxicity; D007700:Kinetics; D008190:Lyases / Q000302:isolation & purification / Q000378:metabolism*; D008297:Male; D009928:Organ Specificity; D051381:Rats; D017207:Rats, Sprague-Dawley; D013379:Substrate Specificity; D000637:Transaminases / Q000302:isolation & purification / Q000378:metabolism*
D005973:Glutamine; D007051:Ibotenic Acid; D000637:Transaminases; C427605:glutamine - phenylpyruvate transaminase; C021481:kynurenine-oxoglutarate transaminase; D008190:Lyases; D005968:Glutamate Decarboxylase