Siderophore receptors

J. Morris, D. F. Donnelly, E. O Neill, F. McConnell, and F. O Gara, F. Nucleotide sequence analysis and potential environmental distribution of a ferric pseudobactin receptor gene of Pseudomonas sp. strain Ml 14. Mol. Gen. Genet. 242 9 (1994). J. M. Raaijmakers, W. Bitter, H. L. M. Punte, P. A. H. M. Bakker, P. J. Weisbeek, and B. Schippers, Siderophore receptor PupA as a marker to monitor wild-type Pseudomonas piitida WCS358 in natural environments. Appl. Environ. Microbiol. 60 1184 (1994). 

We will deal here only with proteins of class (ii) that contain diiron centres and reserve the discussion of proteins containing polyiron oxo aggregates (ferritins) until Chapter 6. As for the proteins of class (iii), their discussion is deferred until Chapter 3 concerning siderophore receptors and Chapter 5 concerning the transferrins and their receptors respectively. 

Once the siderophore-iron complexes are inside the bacteria, the iron is released and utilized for vital cell functions. The iron-free hydroxamate siderophores are commonly re-excreted to bring in an additional iron load (Enterobactin is at least partially degraded by a cytoplasmic esterase This cycle is repeated until specific intracellular ferric uptake regulation proteins (Fur proteins) bind iron, and signal that the intracellular iron level is satisfactory, at -which point ne-w siderophore and siderophore-receptor biosynthesis are halted and the iron-uptake process stops. This intricate feedback mechanism allows a meticulous control over iron(III) uptake and accumulation against an unfavorable concentration gradient so as to maintain the intracellular iron(III) level within the required narrow window. Several excellent reviews concerning siderophore-iron transport mechanisms have been recently published i.3,i6, is,40,45,60-62 ... 

Since siderophore receptors, being membrane-bound, are hard to crystallize, there are to date few structures in the Protein Data Bank (PDB). One such structure is the E. call FhuA entries IbyS, with and without ferrichrome, Ifcp and 2fcp , respectively. ... 

However, this approach still produces racemic mixtures that preclude assessing chiral recognition events taking place between siderophore mimics and their bacterial outer-membrane receptors. Since many siderophore receptors were shown to exhibit high enan-tioselectivity " " , it was anticipated that pure enantiomeric siderophore analogs would allow one to identify the preferred directionality of the helical twist about the iron for the specihc receptors. 

A series of analogs with probes of varying size and polarity was prepared to determine the effect on siderophore receptor transport. These include 7-nitrobenz-2-oxa-l,3-diazole (NBD) (176-179), fluorescein-5-isothiocyanate methyl ether methyl ester (diMe-FITC)... 

Figure 13. Schematic of the organization and properties of siderophore receptors in Escherichia coli K-12.

While most workers report the outer membrane siderophore receptors to have molecular weights in the 75-95K range, some variation in the magnitude of these numbers may be attributed to the preparative and analytical methods as well as to the particular standards used. Since enterobactin will rapidly remove iron from ferrichrome, the transport of the latter must perforce be studied in mutants lacking the former. However, such mutants often display multiple lesions. Additionally, isogenic strains have seldom been used and variations in media and cultural conditions will further confound attempts to compare results reported from different laboratories. 

In times of iron deficiency, many bacteria and fungi release low molecular weight chelators called siderophores (see Iron Transport Siderophores). These molecules bind ferric iron tightly and the ferric-siderophore complexes are then transported into the cell by a system of uptake proteins. The first stage in the uptake process involves an outer membrane receptor specific to each siderophore. One of the best characterized of these receptors is FhuA, the ferrichrome uptake receptor of E. coli, and we will describe this in detail. However, though other ferric-siderophore complexes are taken up by cells, and their iron released by systems similar to those of ferrichrome, their mechanisms may vary from those of ferrichrome in some respects. FepA and FecA" are two of the outer membrane ferric-siderophore receptors that have recently been structurally characterized. 

In the past few years, crystal structmes have been published of three of the seven outer-membrane-siderophore receptors from E.coli, namely of FepA,FhuA, and FecA. FepA and FhuA represent similar monomeric transmembrane proteins that are composed of 22 antiparallel B-strands of approximately 70 A height (Figme 11). The right-handed twist of the /3-strands produces an ellipticalshaped barrel with a diameter of 35 x 47 A constituting a transmembrane pore. Targe extracellular loops extend... 

Figure 11 Crystal structure of the OM-siderophore receptor FhuA with the bound siderophore ferrichrome (emphasized by a space filling atom presentation). The plot is based on PDB file IQFF, taken from the NCBl database using the program PYMOL. Resolution of the structure is 2.70 A, and the space group is P6i. The upper side of the figure corresponds to the extracellular side of the outer membrane. See Section 5.2 for a detailed discussion...

D. van der Hekn and R. Chakraborty, Structures of Siderophore Receptors, in Microbial Transport Systems , ed. G. Winkelmann, Wiley-VCH, Weinbeim, 2001, p. 261. 

Relatively few membrane transport proteins have been structurally characterized. Some of the best understood examples to date are the lactose permease and glycerol-3-phosphate transporter and the Ca + P-type ATPase (which is a primary ion pump). Other structurally well-characterized transport proteins include the bacterial porins and siderophore receptor proteins. In addition structures have been determined for several ion channels and additional bacterial transporters that are either directly relevant to or models for proteins important in drug transport. The following web sites maintained by Hartmut Michel and Stephen White respectively, contain exceptionally useful listings of these and other solved membrane protein structures and are frequently updated ... 

All outer-membrane transporters (OMTs) involved in iron uptake are made up of a 22-stranded p—barrel, which is occluded by an independently folded mixed a P globular cork domain of around 160 amino acid residues. This is illustrated for a vitamin B12 receptor and the ferric siderophore receptors for citrate, enterobactin, ferrichrome, pyochelin, and pyoverdin from E. coli and P. aeruginosa in Fig. 7.6. The ferric siderophore sits on top of the cork domain, as can be seen in Fig. 7.7 for FecA. The binding of the ligand induces a conformational... 

A vaccine targeting siderophore receptor and porin proteins (Epitopix, LLC, Wilmar, MN) recently received a conditional license for use in cattle in fhe United States. (Epitopix, LLC, 2009). The vaccine tended to reduce... 

Fox, J. T., Thomson, D. U., Drouillard, J. S., Thornton, A. B., Burkhardt, D. T., Emery, D. A., and Nagaraja, T. G. (2009a). Efficacy of Escherichia coli 0157 H7 siderophore receptor/ porin proteins-based vaccine in feedlot cattle naturally shedding E. coli 0157. Foodborne Pathog. Dis. 6, 893-899. 

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