Siderophore-dependent ferrichelatases.

Abstract

Iron is a crucial secondary metabolite for bacterial proliferation, but its bioavailability under infection conditions is limited by the low solubility of ferric ion and the host's ability to sequester iron by protein chelation. In these iron limiting conditions, bacteria produce and secrete low molecular weight ferric ion chelators, siderophores, to scavenge host iron. Iron bound siderophores are recognized by surface displayed receptors and internalized by active transport preceding the liberation of the iron payload by reduction or cleavage of the siderophore. The traditional paradigms surrounding the interactions between siderophores and their corresponding receptors have relied on canonical protein-ligand binding models that do not accurately reflect the conditions experienced by siderophore binding proteins (SBPs). Research by the Raymond group suggested that a ligand displacement model does not fully describe the role of SBPs in siderophore transport where the ferric ion can be shuttled between siderophore molecules during the transport process. This work inspired further research by the Wencewicz group, which demonstrated that the Staphylococcus aureusSBP FhuD2 can catalyze the transfer of iron from the biological iron source holo-transferrin to a SBP bound iron-free siderophore. The discovery of this ferrichelatase activity represents a novel mechanism of receptor mediated active transport which raises the question: is ferrichelatase activity a unique feature of FhuD2 or a previously unappreciated hallmark of SBPs? This chapter highlights a series of protocols for the general functional characterization of SBPs and methodologies to assay ferrichelatase activity with the hopes of providing the tools to answer this question.