Characterization of the Functional Activities of the Subunits of 3-Deoxy-d-arabinoheptulosonate 7-phosphate Synthetase-Chorismate Mutase from Bacillus subtilis 168

Abstract

Abstract The interrelationships between the 3-deoxy-d-arabinoheptulosonate 7-phosphate (DAHP) synthetase-chorismate mutase enzyme and shikimate kinase were studied. Attempts were made to distinguish between the various catalytic and regulatory activities of this ternary enzyme complex by a variety of techniques. Limited trypsin treatment specifically cleaved the active region of chorismate mutase and the region required for the activation of shikimate kinase activity while most of the catalytic activity of DAHP synthetase remained intact. The loss of catalytic activity of chorismate mutase paralleled the loss of feedback control of DAHP synthetase and substrate binding studies revealed only one site for prephenate binding on each polypeptide chain. Anti-serum to DAHP synthetase-chorismate mutase specifically inhibited chorimate mutase and shikimate kinase activity but did not inhibit DAHP synthetase activity. These data indicate that DAHP synthetase activity does not depend on a catalytically functional chorismate mutase activity. The data further suggest that the chorismate mutase active site is identical with the feedback inhibition site for DAHP synthetase, thus facilitating feedback control of the aromatic amino acid biosynthetic pathway. The distribution of the catalytic and regulatory activities of this enzyme on one polypeptide chain is discussed.