Catabolite gene activator protein and integration host factor act in concert to regulate tdc operon expression in Escherichia coli

Abstract

Anaerobic expression of the tdcABC operon of Escherichia coli requires cyclic AMP and the catabolite gene activator protein (CAP). Purified CAP binds to a 30-bp sequence in the tdc promoter between positions -55 and -26, and a mutant CAP site with base substitutions at positions -48, -47, and -45 failed to bind CAP and also drastically reduced the beta-galactosidase expression from a tdcB'-'lacZ fusion plasmid. Recently, we showed that efficient expression of the tdc operon also requires a functional integration host factor (IHF) and an IHF-binding site in the tdc promoter between positions -118 and -88. The levels of beta-galactosidase activity from the tdcB'-'lacZ fusion plasmids were also reduced in an IHF-deficient strain with the wild-type or mutant plasmid CAP sequence. In vitro footprinting experiments revealed that CAP and IHF occupy their specific binding sites on tdc DNA when they are present separately or together. These regulatory proteins also induced significant bending of the tdc promoter DNA. Our results suggest that CAP and IHF act in concert as positive transcription factors for tdc operon expression in vivo.