Three binding sites for AraC protein are required for autoregulation of araC in Escherichia coli

Three binding sites for AraC protein were shown to be required for the autoregulation of araC: araI1, araO1, and araO2. Selective inactivation of AraC-binding sites on the DNA demonstrated that araO1 and araO2 are required in vivo to produce repression of araC in the presence of arabinose, whereas araI1 and araO2 are required in its absence. We found that the low-affinity site araO2 is essential for araC autoregulation; araO1 and araI1 provide high-affinity AraC-binding sites, which allow cooperative binding at araO2. Profound effects on the araBAD promoter and the araC promoter are produced by ligand-induced changes in AraC occupancy of functional sites on the DNA. We suggest that AraC exerts its multiplicity of controls through two alternative states of cooperative interactions with DNA and we illustrate this with a model. This model presents our interpretations of activation and repression of the araBAD operon and the autoregulation of the araC gene.