Reaction of muscimol with 4-aminobutyrate aminotransferase

The reaction of muscimol as amino donor substrate for GABA transaminase (GABA-T) has been studied using enzyme purified from rabbit brain. Enzyme activity was assayed by measuring the glutamate produced using glutamate dehydrogenase. Kinetic parameters determined at 37 degrees C were for GABA, Km (app) = 1.92 +/- 0.24 mM, specific activity = 7.33 +/- 0.27 mumol/min/mg (kcat = 13.7s-1), and for muscimol, Km (app) = 1.27 +/- 0.15 mM, specific activity = 0.101 +/- 0.009 mumol/min/mg (kcat = 0.19s-1). Addition of muscimol to the enzyme caused the spectral changes associated with conversion of the pyridoxaldimine form to the pyridoxamine form, and the first-order rate constant for the reaction showed a dependence on muscimol concentration that followed saturation kinetics, with a K = 1.1 +/- 0.18 mM and kmax = 0.065 +/- 0.004 s-1 (19 degrees C). The rate of spectral change observed on addition of muscimol to ornithine transaminase was extremely slow--at least an order of magnitude slower than that seen with GABA-T.