COW BRAIN GLUTAMINASE - PARTIAL-PURIFICATION AND MECHANISM OF ACTION

We have purified glutaminase 65-fold from cow brain; the final specific activity is 24 μmol/min/mg. The enzyme is stable between pH 7.5 and 9.0 and has maximal activity at pH 8.8. It requires Pi for activity. The dependence of activity on Pi concentration is sigmoidal; 50 mm Pi gives half-maximal velocity at pH 8.8. At 0.2 m Pi, pH 8.8, the dependence of activity on glutamine concentration is hyperbolic; the observed KGln was 30 mm. Increasing Pi concentrations increase the apparent Vm and decrease the apparent KGln. NH4+ does not inhibit at concentrations up to 0.1 m. Glutamic acid inhibits competitively with respect to glutamine; at 0.2 m Pi pH 8.8, KGln was 30 mm and KGlu was 19 mm. The results are consistent with a model in which NH4+ is released irreversibly from the enzyme-substrate complex and is the first product released. The activity of glutaminase appears to be independent of the nature of the buffer with which it is equilibrated before being assayed. © 1979.