Purification and properties of 3-deoxy-D-arabionheptulosonic acid-7-phosphate synthetase (trp) from Escherichia coli.

Abstract

The 3-deoxy-d-arabinoheptulosonic acid-7-phosphate synthetase which is subject to regulation by tryptophan has been partially purified from a strain of Escherichia coli K-12, in which this is the only functional form of this enzyme activity, and from a similar strain possessing a feedback-resistant form of the enzyme. Maximal observed inhibition by tryptophan of the feedback-sensitive enzyme was 56%. There was no evidence for cooperativity in the saturation of the enzyme with tryptophan or E4P. The molecular weights of the feedback-sensitive and feedback-resistant forms of the enzyme were the same (52,000), and no change was detected in the molecular weight of the feedback-sensitve enzyme in the presence of tryptophan. The effect of tryptophan analogues was tested to determine the nature of the tryptophan binding site. Treatment with ethylenediaminetetraacetic acid removed 80% of the activity of the feedback-sensitive enzyme. This activity was restored upon the addition of Co(2+) or Mn(2+). Neither treatment with ethylenediaminetetraacetic acid nor addition of Co(2+) or Mn(2+) affected the activity of the feedback-resistant enzyme.