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Enzymatic method for determination of branched-chain amino acid aminotransferase activity
P Schadewaldt1, W Hummel, U Wendel
1Institut für Physiologische Chemie II, Heinrich-Heine-Universität Düsseldorf, Federal Republic of Germany.
Abstract:
A spectrophotometric assay for the determination of branched-chain L-amino acid aminotransferase activity is described. It is based on the transamination of L-leucine in the presence of 2-oxoglutarate yielding 4-methyl-2-oxopentanoate. The rate of formation of the branched-chain 2-oxo acid is specifically monitored in a coupled enzymatic reaction using NAD(+)-dependent D-2-hydroxyisocaproate dehydrogenase from Lactobacillus casei ssp. pseudoplantarum as coupling enzyme by measuring the decrease in NADH absorbance at 334 nm. Optimized assay conditions are provided as evaluated for the (iso)enzyme from rat heart.