THE 2 OPPOSING ACTIVITIES OF ADENYLYL TRANSFERASE RESIDE IN DISTINCT HOMOLOGOUS DOMAINS, WITH INTRAMOLECULAR SIGNAL-TRANSDUCTION

Adenylyl transferase (ATase) is the bifunctional effector enzyme in th e nitrogen assimilation cascade that controls the activity of glutamin e synthetase (GS) in Escherichia coli. This study addresses the questi on of whether the two antagonistic activities of ATase (adenylylation and deadenylylation) occur at the same or at different active sites. T he 945 amino acid residue ATase has been truncated in two ways, so as to produce two homologous polypeptides corresponding to amino acids 1- 423 (AT-N) and 425-945 (AT-C). We demonstrate that ATase has two activ e sites; AT-N carries a deadenylylation activity and AT-C carries an a denylylation activity. Glutamine activates the adenylylation reaction of the AT-C domain, whereas alpha-ketoglutarate activates the deadenyl ylation reaction catalysed by the AT-N domain. With respect to the reg ulation by the nitrogen status monitor PII, however, the adenylylation domain appears to be dependent on the deadenylylation domain: the dea denylylation activity of AT-N depends on PII-UMP and is inhibited by P II. The adenylylation activity of AT-C is independent of PII (or PII-U MP), whereas in the intact enzyme PII is required for this activity. T he implications of this intramolecular signal transduction for the pre vention of futile cycling are discussed.