Structural basis of promiscuous inhibition of Listeria virulence activator PrfA by oligopeptides

The facultative pathogen Listeria monocytogenes uses a master regulator, PrfA, to tightly control the fitness-costly expression of its virulence factors. We found that PrfA activity is repressed via competitive occupancy of the binding site for the PrfA-activating cofactor glutathione by exogenous nutritional oligopeptides. The inhibitory peptides show different sequence and physicochemical properties, but how such wide variety of oligopeptides can bind PrfA was unclear. Using crystal structure analysis of PrfA complexed with inhibitory tri- and tetrapeptides, we show here that the binding promiscuity is due to the ability of PrfA {beta}5 in the glutathione-binding tunnel to establish parallel or antiparallel {beta}-sheet-like interactions with the peptide backbone. Spacious tunnel pockets provide additional flexibility for unspecific peptide accommodation while providing selectivity for hydrophobic residues. Hydrophobic contributions from two adjacent peptide residues appears to be critical for PrfA inhibitory binding. In contrast to glutathione, peptide binding prevents the conformational change required for PrfA activation and formation of the DNA-binding helix-turn-helix motifs, effectively inhibiting virulence expression.