2-Methylcitrate-dependent activation of the propionate catabolic operon (prpBCDE) of Salmonella enterica by the PrpR protein

Abstract

The function of the PrpR protein ofSalmonella entericaserovar Typhimurium LT2 was studiedin vitroandin vivo. The PrpR protein is a sensor of 2-methylcitrate (2-MC), an intermediate of the 2-methylcitric acid cycle used by this bacterium to convert propionate to pyruvate. PrpR was unresponsive to citrate (a close structural analogue of 2-MC) and to propionate, suggesting that 2-MC, not propionate, is the metabolite that signals the presence of propionate in the environment toS. enterica.prpRalleles encoding mutant proteins with various levels of 2-MC-independent activity were isolated. All lesions causing constitutive PrpR activity were mapped to the N-terminal domain of the protein. Removal of the entire sensing domain resulted in a protein (PrpRc) with the highest 2-MC-independent activity. Residue A162 is critical to 2-MC sensing, since the mutant PrpR protein PrpRA162Twas as active as the PrpRcprotein in the absence of 2-MC. DNA footprinting studies identified the site in the region betweenprpRand theprpBCDEoperon to which the PrpR protein binds. Analysis of the binding-site sequence revealed two sites with dyad symmetry. Results from DNase I footprinting assays suggested that the PrpR protein may have higher affinity for the site proximal to the PprpBCDEpromoter.