METALLOREGULATION IN-VITRO OF THE AEROBACTIN PROMOTER OF ESCHERICHIA-COLI BY THE FUR (FERRIC UPTAKE REGULATION) PROTEIN

The mechanism of transcriptional repression of the aerobactin operon o f Escherichia coli by the Fe2+-responsive Fur (ferric uptake regulatio n) protein has been investigated. In the presence of a divalent metal, such as Mn2+, the Fur protein sequentially occupies two defined sites at the aerobactin promoter region, followed by a looser occupation of upstream DNA sequences. However, binding to the primary target site s uffices for the entire repression effect. Comparison of transcription patterns generated with run-off experiments in the presence and absenc e of heparin showed that access of the RNA polymerase to the principal -35/-10 hexamers of the promoter region was fully prevented by Fur-Mn 2+ bound to its primary site. Similarly, promoter-bound RNA polymerase could not be competed out from the DNA even in the presence of a larg e Fur-Mn2+ excess, although the repressor could immediately bind its t arget sequence at the region as soon as RNA polymerase moved away from the promoter during transcription. The high affinities of either prot ein for the promoter produce, in practice, a first-come, first-served effect that helps the system to respond instantly to changes in the ir on status of the cells.