PROMOTER AND OPERATOR DETERMINANTS FOR FUR-MEDIATED IRON REGULATION IN THE BIDIRECTIONAL FEPA-FES CONTROL REGION OF THE ESCHERICHIA-COLI ENTEROBACTIN GENE SYSTEM

The fepA-entD and fes-entF operons in the enterobactin synthesis and t ransport system are divergently transcribed from overlapping promoters , and both are inhibited by the Fur repressor protein under iron-reple te conditions, A plasmid harboring divergent fepA'-'phoA and fes-entF' -'lacZ fusions, both under the control of this bidirectional regulator y region, was constructed for the purpose of monitoring changes in exp ression of the two operons simultaneously. Deletion analysis, site dir ected mutagenesis, and primer extension were employed to define both a single promoter governing the expression of fes-entF and two tandemly arranged promoters giving rise to the opposing fepA-entD transcript. A single Fur-binding site that coordinately regulates the expression o f all transcripts emanating from this control region was identified by in vitro protection from DNase I nicking. The substitution of one bas e pair in the Fur recognition sequence relieved Fur repression but did not change the in vitro affinity of Fur for its binding site. Additio nal mutations in a limited region outside of the promoter determinants for either transcript inhibited expression of both fes and fepA. Thes e observations suggest a mechanism of Fur-mediated regulation in this compact control region that may involve other regulatory components.