Biosynthesis of selenate reductase in Salmonella enterica: Critical roles for the signal peptide and DmsD

Abstract

Salmonella enterica serovar Typhimurium is a Gram-negative bacterium with a flexible respiratory capability. Under anaerobic conditions, S. enterica can utilize a range of terminal electron acceptors, including selenate, to sustain respiratory electron transport. The S. enterica selenate reductase is a membrane-bound enzyme encoded by the ynfEFGH-dmsD operon. The active enzyme is predicted to comprise at least three subunits where YnfE is a molybdenum-containing catalytic subunit. The YnfE protein is synthesized with an N-terminal twin-arginine signal peptide and biosynthesis of the enzyme is coordinated by a signal peptide binding chaperone called DmsD. In this work, the interaction between S. enterica DmsD and the YnfE signal peptide has been studied by chemical crosslinking. These experiments were complemented by genetic approaches, which identified the DmsD binding epitope within the YnfE signal peptide. YnfE signal peptide residues L24 and A28 were shown to be important for assembly of an active selenate reductase. Conversely, a random genetic screen identified the DmsD V16 residue as being important for signal peptide recognition and selenate reductase assembly.