Anthranilate Synthetase

PARTIAL PURIFICATION AND SOME KINETIC STUDIES ON THE ENZYME FROM ESCHERICHIA COLI

From the ¹ From the Department of Microbiology, Western Reserve University, School of Medicine, Cleveland, Ohio 44106

Anthranilate synthetase was purified by ammonium sulfate precipitation and gel filtration from extracts of an episome-bearing Escherichia coli mutant grown under conditions of tryptophan pathway derepression. This purification represented an 18-fold increase in specific activity over the activity of the derepressed mutant extract. Starch gel electrophoresis and ultracentrifugation revealed only slight contamination of the anthranilate synthetase protein. An s_20,w⁰ 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.

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				K Bertrand, L Korn, F Lee, T Platt, C. Squires, C Squires, and C Yanofsky New features of the regulation of the tryptophan operon Science, July 4, 1975; 189(4196): 22 - 26. [PDF]