INTRAMOLECULAR SIGNAL-TRANSDUCTION WITHIN THE FIXJ TRANSCRIPTIONAL ACTIVATOR - IN-VITRO EVIDENCE FOR THE INHIBITORY EFFECT OF THE PHOSPHORYLATABLE REGULATORY DOMAIN

FixJ is a phosphorylatable 'response regulator' controlling the transc ription of the key nitrogen fixation genes nifA and fixK in Rhizobium meliloti. Sequence and genetic analyses indicated that FixJ comprises an N-terminal phosphorylatable regulatory domain, FixJN, and a C-termi nal transcriptional activator domain, FixJC. We have now overexpressed and purified the FixJC protein and show that it is fully active in an in vitro transcription system with purified RNA polymerase. FixJC app eared to act synergistically with RNA polymerase at the nifA promoter. Furthermore FixJC was more active in vitro than the full-length depho sphorylated FixJ protein. Therefore activity of FixJC is inhibited by FixJN within the FixJ protein. This inhibition is relieved by phosphor ylation of FixJN. Such a negative mode of intramolecular signal transd uction may be generalizable to other response regulators.