Studies on plant aspartate transcarbamylase: regulatory properties of the enzyme from mung bean (Phaseolus aureus) seedlings

Abstract

Aspartate transcarbamylase (EC 2·1·3·2) purified from mung bean seedlings was used as a model to understand the mechanism of allosteric regulation. The enzyme exhibited homotropic interactions with carbamyl phosphate. Preincubation of the enzyme with aspartate abolished the sigmoidicity of the carbamyl phosphate saturation curve. UMP was the most potent inhibitor of the reaction and was noncompetitive with respect to aspartate. The sigmoidicity of carbamyl phosphate saturation curves increased with increase in UMP concentration. These results were analysed by an iterative least squares procedure. There was no change in V_max values with increase in the UMP concentration, although the K_0.5 values (concentration of carbamyl phosphate required to reach half maximal velocity) increased. This implied that the effect of UMP was on the binding of carbamyl phosphate only and not on the catalytic function of the enzyme. The allosteric properties of the enzyme could be explained in terms of K system of the symmetry model. The values of the allosteric constantsn, L and c calculated for mung bean enzyme, making use of the Monod equation accounted for all the observed properties. The enzyme appeared to be a tetramer (n=4) and in the absence of ligands was predominantly in the T form (L_o=2.25). Carbamyl phosphate bound preferentially to theR form (c=10⁻³), while UMP bound preferentially to the T form and hence these two ligands exhibited the typical heterotropic interactions as expected of antagonistic ligands.