Ferrichrome structure

Figure 3.3 Comparison of the FepA and FhuA crystal structures. A portion of the 13-barrel (in violet is removed to show the globular cork domain (in yellow) that inserts from the periplasm into the channel of the 11-barrel. FhuA is loaded with ferrichrome (iron is shown as a green ball) (Ferguson et ah, 1998 Locher et ah, 1998). The FepA crystal structure does not reveal Fe3+-enterobactin, but the FepA structure shown might be partially occupied by enterobactin (Buchanan et ah, 1999). 

Figure 3.5 Structures of FhuA ligands as determined by X-ray analysis of co-crystals with FhuA. Albomycin adopts an extended and a compact conformation in the FhuA crystal, and rifamycin CGP 4832 binds to the same FhuA site as ferrichrome and albomycin although it assumes a different conformation.

Figure 3.2 Chemical structures of selected siderophores to demonstrate the four major structural classes and the different solutions of microorganisms to scavenge iron. See for comparison the conformations of the Fe3+-complexes of ferrichrome and albomycin shown in Figure 3.5.

Mainly the outer membrane ferrichrome receptor and transporter FhuA will be discussed because most structural and functional studies have been performed with this protein. In fact, FhuA was the first outer membrane protein identified (called TonA), with known functions as a phage and colicin receptor, that are related to iron transport (for a historical account, see Braun and Hantke 1977). 

The crystal structure of FhuA, with and without bound ferrichrome, has been determined (Ferguson et ah, 1998 Locher et al, 1998). FhuA consists of 22 antiparallel transmembrane 3-strands extending from residue 161 to residue 723, which form a (3-barrel (Figure 3.3, Plate 4). The -barrel strands are interconnected by large loops at the cell surface and small turns in the periplasm. Such a 3-barrel structure is the... 

Figure 3.6 Comparison of the chemical structures of rifamycin (Rifampicin ) and rifamycin CGP 4832 the latter is transported by FhuA. Note the entirely different chemical structures (Figure 3.2) and conformations (Figure 3.5) of the ferrichrome and albomycin FhuA transport substrates.

Another factor that relates complex stability and siderophore architecture is the chelate effect. The chelate effect is represented by an increase in complex stability for a multidentate ligand when compared to complexes with homologous donor atoms of lower denticity. The effect can be observed when comparing the stability of complexes of mono-hydroxamate ligands to their tris-hydroxamate analogs, such as ferrichrome (6) or desferrioxamine B (4). However, the increase in stability alone is not sufficient to explain the preponderance of hexadentate siderophores over tetradentate or bidentate siderophores in nature, and the chelate effect is not observed to a great extent in some siderophore structures (10,22,50,51). 

The FhuA receptor of E. coli transports the hydroxamate-type siderophore ferrichrome (see Figure 9), the structural similar antibiotic albomycin and the antibiotic rifamycin CGP 4832. Likewise, FepA is the receptor for the catechol-type siderophore enterobactin. As monomeric proteins, both receptors consist of a hollow, elliptical-shaped, channel-like 22-stranded, antiparallel (3-barrel, which is formed by the large C-terminal domain. A number of strands extend far beyond the lipid bilayer into the extracellular space. The strands are connected sequentially using short turns on the periplasmic side, and long loops on the extracellular side of the barrel. 

Figure 8 (Plate 7). Structure of the Escherichia coli FhuA protein serving as receptor for ferrichrome and the antibiotic albomycin. (a) side view (b) side aspect with partly removed barrel to allow the view on the cork domain (c) top view. A single lipo-polysaccharide molecule is tightly associated with the transmembrane region of FhuA (reproduced by permission of W. Welte and A. Brosig)...

Figure 9. Structure of the siderophore ferrichrome (and derivatives) produced by certain fungal species...

As mentioned above, transport of siderophores across the cytoplasmic membrane is less specific than the translocation through the outer membrane. In E. coli three different outer membrane proteins (among them FepA the receptor for enterobactin produced by most E. coli strains) recognise siderophores of the catechol type (enterobactin and structurally related compounds), while only one ABC system is needed for the passage into the cytosol. Likewise, OM receptors FhuA, FhuE, and Iut are needed to transport a number of different ferric hydroxamates, whereas the FhuBCD proteins accept a variety of hydroxamate type ligands such as albomycin, ferrichrome, coprogen, aerobactin, shizokinen, rhodotorulic acid, and ferrioxamine B [165,171], For the vast majority of systems, the substrate specificity has not been elucidated, but it can be assumed that many siderophore ABC permeases might be able to transport several different but structurally related substrates. 

Locher, K. P., Rees, B., Koebnik, R., Mitschler, A., Moulinier, L., Rosenbusch, J. P. and Moras, D. (1998). Transmembrane signaling across the ligand-gated FhuA receptor crystal structures of free and ferrichrome-bound states reveal allosteric changes, Cell, 95, 771-778. 

Figure 7.3 Structures of the siderophores enterobactin and ferrichrome. (From Andrews et al., 2003. Reproduced with permission from Blackwell Publishing Ltd.)...

The members of the ferrichrome group are cyclohexapeptides with the general structure [-(lV -acyl-lV -hydroxy-L-Om)3-A-B-Gly-] where A and B can be Gly, Ala, or Ser (Table 1) the various acyl groups are depicted in Fig. 5. Exceptions are tetraglycylferrichrome, a cycloheptapeptide with four Gly units in sequence and three acetyl residues in the Om part (ferrichrome with an additional Gly) 82), and des(diserylglycyl)ferrirhodin, a linear tripeptide containing only the three Om units... 

Ferrichrome (as do also at least the members of the group for which structural data are available ((366) and references noted in Table 1) shows A-, synthetic euauf/o-ferrichrome based on o-Om A-configuration (253). Uptake studies performed with Ustilago sphaerogena (103) using and [ C]-ferrichrome under... 

Relationships between the structure of the siderophores and the iron transport were investigated for the fungus Neurospora crassa (160, 160a). Apparently two different receptors exist for ferrichromes and for coprogenes. For the recognition and the binding to the cell surface the iron configuration and the nature of the acyl chains is of importance. However, the transport system seems to be the same for both siderophore types dependent on the peptide part of the molecules. 

Llinas L, Neilands JB (1976) The Structure of Two Alanine Containing Ferrichromes Sequence Determination by Proton Magnetic Resonance. Biophys Struct Mechanism 2 105... 

Zalkin A, Forrester JD, Templeton DH (1966) Ferrichrome A Tetrahydrate. Determination of Crystal and Molecular Structure. J Am Chem Soc 88 1810... 

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. ... 

TABLE 1. Natural ferrichromes and their structural variations ... 

Substitution of the terminal amine with ligating groups, such as catecholate, hydrox-amate or diketonate, lead to the formation of A-cis chiral complexes. In these structures the chiral information content is located in the amino acid bridges, instead of the macrocyclic peptide ring structure used in ferrichrome. It should be emphasized... 

The closely related structures show completely different microbial uptake characteristics. The 3D structures described above show distinct different orientation of the backbone amide (tangental in type 1 versus radial in type 2), which can be explained by the interactions that take place between the FhuA receptor and the ferrichrome siderophore -As mentioned, the second coordination sphere of natural ferrichrome in FhuA receptor is very sensitive to the distance and orientation between a proton donor and the proton acceptor, therefore the orientation of the amide groups in the biomimetic siderophore plays a crucial role in receptor recognition. 

In a later work, the iron(III) complex stability and structure of 75 was compared to other NTA-based analogs 76, 77, 78 and 79 °. Complex stability was in the order 75 > 79 > 76 > 77 = 78. CD spectra revealed that the enantiomeric pair 77 and 78 gave A-and A-configurations, respectively, by reference to the assignment for ferrichrome Both 75 and its diastereomer 76 showed preference for the A-cis configuration. 

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