SPECIFIC BINDING OF DNAA PROTEIN TO A DNAA BOX IN THE GUAB GENE OF ESCHERICHIA-COLI K12

Expression of the guaBA operon of Escherichia coli is regulated by the DNA replication-initiating protein, DnaA. Two DnaA boxes, which are p otential binding sites for DnaA, are present in the gun operon. One bo x (with 8/9 match to the DnaA box consensus sequence) is at the gua pr omoter; the other box, which has a consensus sequence, is on the non-t ranscribed strand within the guaB coding region approximately 200 bp d ownstream of the initiation codon. The binding in vitro of purified Dn aA protein to these boxes was investigated by filter retention and gel retardation analysis, and by deoxyribonuclease I footprinting, using restriction fragments of gun operon DNA. DnaA protein was shown to bin d specifically only to the fragment carrying the consensus sequence Dn aA box, and to protect this box from deoxyribonuclease I. Transcriptio n termination resulting from the binding of DnaA to this box within th e guaB gene explains repression by DnaA of the gun operon in vivo.