Kinetic properties of phosphoribosyladenosine triphosphate synthetase. Inhibition by aggregation at high enzyme concentrations

1. 1. The specific activity of phosphoribosyladenosine triphosphate: pyrophosphate phosphoribosyltransferase (phosphoribosyladenosine triphosphate synthetase) from Escherichia coli decreases with increasing concentration of the enzyme, indicating aggregation to less active forms. 2. 2. The decrease in specific activity at high enzyme concentrations is more pronounced at 35° than at 15°, and for this reason a very low activation energy for the synthetase reaction is found at high enzyme concentrations. 3. 3. 2.5 mM AMP inhibits the synthetase reaction under standard assay conditions and low enzyme concentrations while little inhibition is seen at high enzyme concentrations. Even at high enzyme levels, 2.5 mM AMP changes the response of the enzyme to histidine from a cooperative to a more hyperbolic type. 4. 4. At high enzyme concentrations, inhibition by the product, phosphoribosyladenosine triphosphate, is temperature dependent and most pronounced at low temperatures. 5. 5. The apparently sigmoidal response of synthetase activity to magnesium concentration may be explained by the binding of Mg2+ to ATP. After correction for this binding, half saturation of the enzyme with Mg2+ is observed at 0.13 mM.