The structure of Vibrio cholerae FeoC reveals conservation of the helix-turn-helix motif but not the cluster-binding domain
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2022-02-26
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Attribution-NonCommercial-NoDerivs 3.0 United States
Attribution-NonCommercial-NoDerivs 3.0 United States
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Abstract
Most pathogenic bacteria require ferrous iron (Fe²⁺) in order to sustain infection within hosts. The ferrous iron transport (Feo) system is the most highly-conserved prokaryotic transporter of Fe²⁺, but its mechanism remains to be fully characterized. Most Feo systems are composed of two proteins: FeoA, a soluble SH3-like accessory protein and FeoB, a membrane protein that translocates Fe²⁺ across a lipid bilayer. Some bacterial feo operons encode FeoC, a third soluble, winged-helix protein that remains enigmatic in function. We previously demonstrated that select FeoC proteins bind O₂-sensitive [4Fe-4S] clusters via Cys residues, leading to the proposal that some FeoCs could sense O₂ to regulate Fe²⁺ transport. However, not all FeoCs conserve these Cys residues, and FeoC from the causative agent of cholera (Vibrio cholerae) notably lacks any Cys residues, precluding cluster binding. In this work, we determined the NMR structure of VcFeoC, which is monomeric and conserves the helix-turn-helix domain seen in other FeoCs. In contrast, however, the structure of VcFeoC reveals a truncated winged β-sheet in which the cluster-binding domain is notably absent. To test the interactions of VcFeoC with VcFeoB, we used NMR to demonstrate that these proteins interact in a 1:1 stoichiometry with a Kᵈ of ca. 25 μM. Finally, using homology modeling, we predicted the structure of VcNFeoB and used docking to identify an interaction site with VcFeoC, which is confirmed by NMR spectroscopy. These findings provide the first atomic-level structure of VcFeoC and contribute to a better understanding of its role vis-à-vis FeoB.