Ferric-enterobactin

The importance of iron for a bacteria-like E. coli can be illustrated by fact that 14 genes alone are required for enterobactin-mediated iron uptake, including those for its synthesis, export, transport of the ferric-enterobactin back into the cell and iron release (Figure 3.16). In total, E. coli has at least 8 uptake systems for iron, encoded by some 50 genes. 

Neilands JB, Ericson TJ, Rastetter WH (1981) Stereospecifity of the Ferric Enterobactin Receptor of Escherichia coli K-12. J Biol Chem 256 3831... 

Dean, C.R. Poole, K. Expression of the ferric enterobactin receptor (PfeA) of Pseudomonas aeruginosa involvement of a two-component regulatory system. Mol. Microbiol., 8, 1095-1103 (1993)... 

In the LI CAM series lipophilicity, hence ability to cross cell membranes, was introduced by substitution of alkyl groups on the terminal nitrogens262. Rastetter et al.256 have also produced a linear polycatecholamide. However, unlike the LICAM series their siderophore is a chiral analogue, synthesized from L-asparagine, for which a formation constant of io46-5 1-2 has been calculated for its ferric complex. In contrast the formation constant for ferric enterobactinate is 1052 263. ... 

Figure 3. Ferric enterobactin, the ferric complex of cyclotri-2,3-dihydroxy-N-benzoyl-i,-serine (14)...

The kinetic lability of ferric siderophores requires that transport experiments be performed with molecules bearing separate radioactive labels in the metal and ligand moieties. As coordination compounds the siderophores are thermodynamically stable and kinetically labile. The formation constants are typically 1030. In the case of ferrichrome the exchange half time at pH 6.3 and 37° is about 10 min (57). Published work (58, 59) with doubly labeled ferric schizokinen in Bacillus mega-terium and ferric aerobactin in A. aerogenes as well as a study of ferric enterobactin in E. coli (60) in each instance suggests a synchronous uptake mechanism for iron and ligand. 

Recently Frost and Rosenberg (56) demonstrated that strains of K-12 blocked in aromatic biosynthesis and also tonB" could fabricate enterobactin from precursors such as 2,3-dihydroxybenzoic acid. The siderophore synthesized endogenously from 2,3-dihydroxybenzoic acid was pictured as carrying iron from the periplasmic space across the cytoplasmic membrane. Evidently ferric enterobactin could not traverse the outer membrane in these strains. 

The existence of ferrichrome and ferric enterobactin receptors in the outer membrane of enteric bacteria confirms the discovery, first reported for vitamin Bi2 (34), for a genuine transport role for this segment of the cell envelope. The properties of the four analogous systems known at the present time are shown in Table IV. 

As seen in Table 7, colicins V and la require the tonB gene, but the receptors for these agents have not been correlated with any specific siderophore or other nutrient substance. The binding of ferric enterobactin has been defined as the biochemical function of the coli-cin B receptor (53,). Iron supply to the cell interferes with adsorption of colicin la, thus suggesting this receptor is designed for a siderophore, the specific nature of which is still unkown (73). 

The synthesis of the colicin la receptor is clearly derepressed at low iron (73, 96), but a specific siderophore has not been assigned to this large polypeptide constituent, which is programmed by the cir gene at 43 min on the chromosome map. FeuB is the specific locus for the colicin B-ferric enterobactin receptor (66). 

A transport-defective mutant of E. coli is advocated for the convenient preparation of ferric enterobactin (100). In S. typhirrmrium, as well as in E. coli, several proteins of the outer membrane are regulated by the iron content of the medium (101). 

It can be seen from molecular models that two diastereoisomers are possible for the ferric enterobactin complex, A-cis and A-cis. These are not mirror images because of the optical activity of the ligand. The similarity of the roles played by the ferrichromes and enterobactin lent additional speculative interest to the preferred absolute configuration of the iron complex (20). The structural studies of the tris catechol complexes (vide infra) and the spectroscopic properties of the chromic... 

Figure 5. A schematic of the A-cis isomer of chromic and ferric enterobactin. The metal lies at the center of a distorted octahedron formed by the oxygen atoms of tne three catechol dianions.

The largest /f-bar re Is have been observed with the monomeric iron transporter proteins FhuA and FepA. The structure of FhuA was established independently by two groups (Locher et al., 1998 Ferguson et al., 1998). It is known with and without a ligated siderophore. The structure of the ferric enterobactin receptor FepA is homologous to that of FhuA showing identical topology and a similar transport mechanism (Buchanan etal., 1999). In both cases there are more than 700 residues assembled in two domains an N-terminal 150-residue domain is located inside a C-terminal 22-stranded (6-barrel with a shear number S = 24. 

Fig. 10. A plot of the absorbance at 450 nm of ferric enterobactin solutions in 50 % aqueous methanol as the pH is changed. The data are from Figure 10 of Ref. 148. The vertical axis is absorbance (AobJ. The horizontal axis is the function (Ao — A0J/[H+] and has been multiplied by 104. A linear relationship implies a single-proton stoichiometry in the reaction. A least-squares fit (the line shown) gives a corelation coefficient of 0.978, a slope of 1.18 x 10-5 (the inverse of the protonation constant) and an intercept of. 425 (Aw)...

Parabactin A, derived from hydrolysis of parabactin (the oxazoline group) exhibits a physiological reducibility. Therefore, oxazoline ring cleavage 148) may be a part of metabolic iron removal in Paracoccus denitrificans. A similar process was initially proposed and broadly accepted for the utilization of enterobactin 144). An esterase was found in the cell extracts of E. coli which cleaved the ester backbone of ferric enterobactin 14s> and desferri-enterobactin 146). In addition, it was demonstrated that... 

Fig. 16. Mossbauer spectra of ferric enterobactin as a function of pH at 4.2 °K in a magnetic field with the Happ = 60 mT parallel to the y ray source. All samples were in frozen solution form. The vertical bars indicate 1 % absorption. With decreasing pH a fast relaxing Fe(IIl) high-spin species arises at about 6 = 0.5 mm/s...

A locally low pH region in the cellular uptake pathway of the ferric enterobactin complex with a resultant shift in the redox potential which would make it available via reduction. 

Fig. 18. Cyclic voltammograms of (top) ferrichrome A (pH 8) (middle) ferriox-amine B (pH 8) (bottom) ferric enterobactin (pH 10.5). All are in 1 M KC1, 0.05 M sodium borate/0.05 M sodium phosphate buffer. All cyclic voltammograms were at hanging mercury drop electrode with 100 mV/sec scan rate...

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