Anthranilate synthetase. Partial purification and some kinetic studies on the enzyme from Escherichia coli.

Anthranilate synthetase was purified by ammonium sulfate precipitation and gel filtration from extracts of an episomebearing Escherichia coli mutant grown under conditions of tryptophan pathway derepression. This purification represented an l&fold increase in specific activity over the activity of the derepressedmutant extract. Starch gel electrophoresis and ultracentrifugation revealed only slight contamination of the anthranilate synthetase protein. An a&~,~ of 10.7 S was obtained for the native enzyme. Kinetic studies with anthranilate synthetase have been initiated. The reaction catalyzed by this enzyme requires two substrates, chorismate and L-glutamine. The reaction mechanism was found to be sequential, and the presence of one substrate on the active site of the enzyme does not affect the binding of the second substrate. In the presence of the feedback inhibitor, L-tryptophan, L-glutamine utilization is noncompetitively inhibited. In addition, L-tryptophan is a partially competitive inhibitor of chorismate utilization.