BINDING IN THE DNAA OPERON PROMOTER REGION

The region between the rpmH and dnaA genes contains five promoters tha t divergently express the ribosomal protein L34 and the proteins of th e dnaA operon, including DnaA, the beta clamp of DNA polymerase III ho loenzyme, and RecF. The DNA-binding protein Fis was shown by the band shift assay to bind near the rpmHp2 and dnaAp2 promoters and by DNase I footprinting to bind to a single site in the dnaAp2 promoter overlap ping the -35 and spacer sequences. There were no observable difference s in Fis affinity or the angle of bending induced by Fis between methy lated and unmethylated DNA fragments containing the Fis binding site i n the dnaAp2 promoter. Fis directly or indirectly represses the expres sion of DnaA protein and the beta clamp of DNA polymerase III. A fis n ull mutant containing a dnaA-lacZ in-frame fusion had twofold greater beta-galactosidase activity than a fis wild-type strain, and induced e xpression of Fis eliminated the increase in activity of the fusion pro tein. A two- to threefold increase in the levels of DnaA and beta clam p proteins was found in a ps null mutant by immunoblot gel analysis.