Interactions of the periplasmic binding protein CeuE with Fe(III) n-LICAM(4-) siderophore analogues of varied linker length

Bacteria use siderophores to mediate the transport of essential Fe(III) into the cell. In Campylobacter jejuni the periplasmic binding protein CeuE, an integral part of the Fe(III) transport system, has adapted to bind tetradentate siderophores using a His and a Tyr side chain to complete the Fe(III) coordination. A series of tetradentate siderophore mimics was synthesized in which the length of the linker between the two iron-binding catecholamide units was increased from four carbon atoms (4-LICAM(4-)) to five, six and eight (5-, 6-, 8-LICAM(4-), respectively). Co-crystal structures with CeuE showed that the inter-planar angles between the iron-binding catecholamide units in the 5-, 6- and 8-LICAM(4-) structures are very similar (111°, 110° and 110°) and allow for an optimum fit into the binding pocket of CeuE, the inter-planar angle in the structure of 4-LICAM(4-) is significantly smaller (97°) due to restrictions imposed by the shorter linker. Accordingly, the protein-binding affinity was found to be slightly higher for 5- compared to 4-LICAM(4-) but decreases for 6- and 8-LICAM(4-). The optimum linker length of five matches that present in natural siderophores such as enterobactin and azotochelin. Site-directed mutagenesis was used to investigate the relative importance of the Fe(III)-coordinating residues H227 and Y288

Iron traffics in circulation bound to a siderocalin (Ngal)–catechol complex

The lipocalins are secreted proteins that bind small organic molecules. Scn-Ngal [known as Neutrophil Gelatinase Associated Lipocalin, Siderocalin, Lipocalin 2] sequesters bacterial iron chelators, called siderophores, and consequently blocks bacterial growth. However, Scn-Ngal is also prominently expressed in aseptic diseases, implying that it binds additional ligands and serves additional functions. Using chemical screens, crystallography, and fluorescence methods, we report that Scn-Ngal binds iron together with a small metabolic product called catechol. The formation of the complex blocked the reactivity of iron and permitted its transport once introduced into circulation in vivo. Scn-Ngal then recycled its iron in endosomes by a pH sensitive mechanism. Since catechols derive from bacterial and mammalian metabolism of dietary compounds, the Scn-Ngal:catechol:iron complex represents an unforeseen microbial-host interaction, which mimics Scn-Ngal:siderophore interactions, but instead traffics iron in aseptic tissues. These results identify an endogenous siderophore, which may link the disparate roles of Scn-Ngal in different diseases