Evidence for Compartmentation of Tryptophan in Cultured Plant Tissues: Free Tryptophan Levels and Inhibition of Anthranilate Synthetase

Abstract

Abstract1. Anthranilate synthetase activity in crude extracts from tissue cultures of Daucus carota L. (carrot), Nicotiana tabacum L. (tobacco; cv. Wisconsin 38 and xanthi), Glycine max Merr. (soybean) and Oryza sativa L. (rice) was completely inhibited by l‐tryptophan (5 to 50 μM). Mutant carrot and tobacco lines, capable of growth in the presence of 5‐methyltryptophan, required 500 to more than 1000 μM tryptophan for complete inhibition of enzyme activity, respectively.2. Except for the mutant tobacco line, the concentrations of free tryptophan in all tissue cultures tested were greater than the levels necessary to completely inhibit the respective anthranilate synthetase activities in vitro. These findings would indicate that much of the free tryptophan is compartmentalized away from the regulatory enzyme, anthranilate synthetase. This could implicate compartmentalization of the inhibitor as a biosynthetic control mechanism.3. During the growth of normal and mutant carrot tissues the anthranilate synthetase enzyme must be at least 7.8 and 10.8% active, respectively, in order to accumulate the amount of tryptophan found in the tissues.4. Of the substrates and cofactors required for anthranilate synthetase activity in vitro, Mg2+ and glutamine were present at near optimal levels in the carrot and tobacco tissues, but chorismate was found to be significantly below the optimal concentrations.