Structural Basis of a Novel Heme Binding Bacterial One-Component Switch

One-component systems (OCSs) integrate sensory and effector functions within a single protein, enabling rapid gene expression changes in response to environmental cues. Here, we characterized a novel putative OCS protein, FG214, from Fimbriimonas ginsengisoli, which drew our attention as a potential redox or O2-regulated helix-turn-helix (HTH)-Per-ARNT-Sim (PAS) transcription factor. Data supporting this included our observation of the FG214 PAS domain binding a hexacoordinate heme b in oxidized conditions and undergoing a slate of redox and ligand-dependent conformational changes, transitioning from a monomer to a homodimer. Spectroscopic and structural data revealed that oxidation stabilizes the likely HTH-PAS intramolecular domain interface, while reduction of the heme iron dissociates the HTH, freeing previously-sequestered homodimerization surfaces. Similar effects were seen by addition of a small molecule ferric heme ligand, as directly visualized with a 1.47 [A] crystal structure of an imidazole-bound truncated construct. Using in vitro DNA-binding assays, we identified an artificial promoter sequence and demonstrated ligand-enhanced protein-DNA binding. Finally, we performed proof of concept experiments exploring the ability of FG214 to homodimerize in vivo, setting the stage for a redox or gas sensitive biosensor. Together, these findings define FG214 as a novel heme-binding PAS DNA binding protein and potential transcription factor, complementing known heme-PAS two-component signaling switches. Significance StatementStudying sensory protein structure-function relationships is paramount to fully understanding cellular adaptation to environmental cues. Here, we have discovered a heme-regulated DNA binding switch, FG214, that undergoes a monomeric to dimeric transition upon redox-triggered changes. The nature of this transition is reminiscent of other proteins with similar domain architecture, although those have been observed to instead sense changes in the presence of other distinct stimuli. Here we not only describe the activation mechanisms of FG214, but also provide support for its ability to be used as a regulatory gene expression tool.