Regulation of glutamine synthetase: III. Cumulative feedback inhibition of glutamine synthetase from Escherichia coli

Abstract

The glutamine Synthetase of Escherichia coli has been examined for its susceptibility to inhibition by a large number of nitrogen-containing compounds of biological importance. Eight of the compounds tested caused inhibition of the enzyme; namely, alanine, glycine, histidine, tryptophan, CTP, AMP, carbamyl phosphate, and glucosamine-6-phosphate. At high concentrations each compound causes only partial inhibition. When mixtures of the inhibitors are present at nearly saturating concentrations their effects are cumulative. That is, the total residual enzyme activity in the presence of several inhibitors is equal to the product of the fractional activities observed when each of the inhibitors is tested alone. Thus, whereas each inhibitor is able to cause only partial inhibition, collectively they can cause almost complete inhibition of the enzyme. The results suggest that the enzyme possesses separate binding sites for each of the eight inhibitors and that there is no significant interaction between these binding sites; i.e., each inhibitor is independent in its action. This suggestion is further supported by kinetic studies showing that three of the inhibitors are noncompetitive with respect to any substrate, three are partially competitive with respect to glutamate, and two are partially competitive with respect to ammonia. Moreover, the inhibitory responses to different inhibitors are differentially effected by Mg ++ and Mn ++, one or the other of which is required for glutamine synthetase activity. Finally, the occurrence of separate binding sites for each inhibitor is indicated by the selective sensitization or desensitization of the enzyme to inhibition by particular inhibitors as a result of structural modification induced by aging, or treatment with urea, or acetone.