The Mechanism of Action of 5′-Adenylic Acid-activated Threonine Dehydrase

The effect of AMP on the kinetic parameters, Vmax, Km, and Hill coefficient for l-threonine, as well as the effect of enzyme concentration on the Ka and Hill coefficient for AMP were determined for the biodegradative l-threonine dehydrase of Escherichia coli. These values were affected by the order of addition of assay components and the manner in which the enzyme was diluted. For instance, dilutions in the absence of AMP followed by assays with graded levels of AMP resulted in a degree of inactivation which was dependent upon the concentrations of AMP and enzyme; on the other hand, no inactivation occurred when the dehydrase was diluted and assayed in 5 mm AMP. A correction procedure is described which nullifies this spurious effect. The kinetic parameters determined following correction were Km for threonine = 70 mm without and 3.1 with 5 mm AMP; Vmax = 489 µmoles min-1 mg of protein-1 at 28° in the absence of AMP and 478 µmoles min-1 mg of protein-1 in the presence of 5 mm AMP. The Hill coefficients for l-threonine were 1.2 to 1.6 without and 1.0 with 5 mm AMP. The Ka value and Hill coefficient for AMP was dependent on dehydrase concentration.