Mycobactin and the competition for iron between Mycobacterium neoaurum and M. vaccae

Two closely related species of mycobacteria, Mycobacterium vaccae and M. neoaurum, were grown under conditions of iron-deficiency (0.02-0.05 microgram Fe ml-1) and iron-sufficiency (2-4 micrograms Fe ml-1) in a simple glycerol/asparagine medium. The strain of M. vaccae used was a nonmycobactin producer whereas M. neoaurum synthesized between 6-8% of its cell biomass as the lipid-soluble siderophore when grown under iron-limitation. The role of mycobactin for iron-acquisition was examined using both pure and mixed cultures, with cell viability determined following growth at various iron concentrations. M. neoaurum, the mycobactin producer, outgrew M. vaccae when iron was readily available. When grown under conditions where iron was limiting, M. neoaurum showed a decline in viable cell number compared with its competitor, highlighting its increased requirement for the metal. Some recovery was observed following mycobactin biosynthesis, this being greatly enhanced by the addition of an iron supplement to the growing cells. Mycobactin biosynthesis allowed M. neoaurum to rapidly acquire any additional iron presented to the bacteria when growing under iron-limitation. However, M. vaccae did not synthesize the lipid-soluble siderophore with its iron-requirement satisfied by production of extracellular exochelin.     

Isolation and characterization of carboxymycobactins as the second extracellular siderophores in Mycobacterium smegmatis

The structure of a chloroform-extractable siderophore from the supernatant of a nonpathogenic mycobacteria, Mycobacterium smegmatis, has been determined and it closely resembles the structure of mycobactin, the intracellular siderophore found in all mycobacteria. The difference in structures is that the extracellular siderophore has a family of short carboxylic chains attached to the mycobactin nucleus instead of a long alkyl chain and hence the name 'carboxymycobactin' is proposed to distinguish it from mycobactin itself as well as from the major siderophore that is produced - exochelin MS. © Rapid Science 1998.     

Mutational and phylogenetic analyses of the mycobacterial mbt gene cluster

The mycobactin siderophore system is present in many Mycobacterium species, including M. tuberculosis and other clinically relevant mycobacteria. This siderophore system is believed to be utilized by both pathogenic and nonpathogenic mycobacteria for iron acquisition in both in vivo and ex vivo iron-limiting environments, respectively. Several M. tuberculosis genes located in a so-called mbt gene cluster have been predicted to be required for the biosynthesis of the core scaffold of mycobactin based on sequence analysis. A systematic and controlled mutational analysis probing the hypothesized essential nature of each of these genes for mycobactin production has been lacking. The degree of conservation of mbt gene cluster orthologs remains to be investigated as well. In this study, we sought to conclusively establish whether each of nine mbt genes was required for mycobactin production and to examine the conservation of gene clusters orthologous to the M. tuberculosis mbt gene cluster in other bacteria. We report a systematic mutational analysis of the mbt gene cluster ortholog found in Mycobacterium smegmatis. This mutational analysis demonstrates that eight of the nine mbt genes investigated are essential for mycobactin production. Our genome mining and phylogenetic analyses reveal the presence of orthologous mbt gene clusters in several bacterial species. These gene clusters display significant organizational differences originating from an intricate evolutionary path that might have included horizontal gene transfers. Altogether, the findings reported herein advance our understanding of the genetic requirements for the biosynthesis of an important mycobacterial secondary metabolite with relevance to virulence.     

Mycobactins in the classification and identification of armadillo-derived mycobacteria

Abstract Seven strains of armadillo-derived mycobacteria (ADMs) were encouraged to produce the lipid-soluble siderophore mycobactin when grown under conditions of iron limitation. These compounds have recently been shown to be excellent chemotaxonomic markers for the mycobacteria being both species-specific and highly conserved. Characterization of the mycobactins was carried out by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). Examination of the mycobactins isolated from the ADM strains showed them to be heterogeneous. Four strains synthesized mycobactins closely resembling those of the M. avium-intracellularescrofulaceum (MAIS) complex, whereas the remaining three strains formed mycobactins which differed in structure to those of any other mycobacterium previously examined.     

Directing the Biosynthesis of Putrebactin or Desferrioxamine B in Shewanella putrefaciens through the Upstream Inhibition of Ornithine Decarboxylase

To manage iron acquisition in an oxic environment, Shewanella putrefaciens produces the macrocyclic dihydroxamic acid putrebactin (PB) as its native siderophore. In this work, we have established the siderophore profile of S. putrefaciens in cultures augmented with the native PB precursor putrescine and in putrescine-depleted cultures. Compared to base medium, PB increased by two-fold in cultures of S. putrefaciens with 10?mM NaCl and 20?mM exogenous putrescine. In cultures augmented with 1,4-diaminobutan-2-one (DAB), PB decreased with only 0.02-fold PB detectable at 10?mM DAB. As an ornithine decarboxylase (ODC) inhibitor, DAB depleted levels of endogenous putrescine which attenuated downstream PB assembly. Under putrescine-depleted conditions, S. putrefaciens produced as its replacement siderophore the cadaverine-based desferrioxamine B (DFO-B), as characterised by ESI-MS of the Fe(III) -loaded form (m/z(obs) 614.13; m/z(calc) 614.27). A third siderophore, independent of DAB, was observed in low levels. LC/MS Analysis of the Fe(III) -loaded extract gave m/z(obs) 440.93, which, formulated as a 1?:?1 Fe(III) complex with a macrocyclic dihydroxamic acid, comprising one putrescine- and one cadaverine-based precursor (m/z(calc) 440.14). These results show that the production of native PB or non-native DFO-B by S. putrefaciens can be directed though upstream inhibition of ODC. This approach could be used to increase the molecular diversity of siderophores produced by S. putrefaciens and to map alternative diamine-dependent metabolites.     

Factors that influence siderophoremediated iron bioavailability: catalysis of interligand iron (III) transfer from ferrioxamine B to EDTA by hydroxamic acids

Deferriferrioxamine B (H3DFB) is a linear trihydroxamic acid siderophore with molecular formula NH2(CH2)5[N(OH)C(O)(CH2)2C(O)NH(CH2)5]2N(OH)C(O)CH3 that forms a kinetically and thermodynamically stable complex with iron(III), ferrioxamine B. Under the conditions of our study (pH = 4.30, 25 degrees C), ferrioxamine B, Fe(HDFB)+, is hexacoordinated and the terminal amine group is protonated. Addition of simple hydroxamic acids, R1C(O)N(OH)R2 (R1 = CH3, R2 = H; R1 = C6H5, R2 = H; R1 = R2 = CH3), to an aqueous solution of ferrioxamine B at pH = 4.30, 25.0 degrees C, I = 2.0, results in the formation of ternary complexes Fe(H2DFB)A+ and Fe(H3DFB)A2+, and tris complexes FeA3, where A- represents the bidendate hydroxamate anion R1C(O)N(O)R2-. The addition of a molar excess of ethylenediaminetetraacetic acid (EDTA) to an aqueous solution of ferrioxamine B at pH 4.30 results in a slow exchange of iron(III) to eventually completely form Fe(EDTA)- and H4DFB+. The addition of a hydroxamic acid, HA, catalyzes the rate of this iron exchange reaction: (formula; see text) A four parallel path mechanism is proposed for reaction (1) in which catalysis occurs via transient formation of the ternary and tris complexes Fe(H2DFB) A+, Fe(H3DFB)A2+, and FeA3. Rate and equilibrium constants for the various reaction paths to products were obtained and the influence of hydroxamic acid structure on catalytic efficiency is discussed. The importance of a low energy pathway for iron dissociation from a siderophore complex in influencing microbial iron bio-availability is discussed. The system represented by reaction (1) is proposed as a possible model for in vivo catalyzed release of iron from its siderophore complex at the cell wall or interior, where EDTA represents the intracellular storage depot or membrane-bound carrier and HA represents a low molecular weight hydroxamate-based metabolite capable of catalyzing interligand iron exchange.     

The structure of MbtI from Mycobacterium tuberculosis, the first enzyme in the biosynthesis of the siderophore mycobactin, reveals it to be a salicylate synthase

The ability to acquire iron from the extracellular environment is a key determinant of pathogenicity in mycobacteria. Mycobacterium tuberculosis acquires iron exclusively via the siderophore mycobactin T, the biosynthesis of which depends on the production of salicylate from chorismate. Salicylate production in other bacteria is either a two-step process involving an isochorismate synthase (chorismate isomerase) and a pyruvate lyase, as observed for Pseudomonas aeruginosa, or a single-step conversion catalyzed by a salicylate synthase, as with Yersinia enterocolitica. Here we present the structure of the enzyme MbtI (Rv2386c) from M. tuberculosis, solved by multiwavelength anomalous diffraction at a resolution of 1.8 A, and biochemical evidence that it is the salicylate synthase necessary for mycobactin biosynthesis. The enzyme is critically dependent on Mg2+ for activity and produces salicylate via an isochorismate intermediate. MbtI is structurally similar to salicylate synthase (Irp9) from Y. enterocolitica and the large subunit of anthranilate synthase (TrpE) and shares the overall architecture of other chorismate-utilizing enzymes, such as the related aminodeoxychorismate synthase PabB. Like Irp9, but unlike TrpE or PabB, MbtI is neither regulated by nor structurally stabilized by bound tryptophan. The structure of MbtI is the starting point for the design of inhibitors of siderophore biosynthesis, which may make useful lead compounds for the production of new antituberculosis drugs, given the strong dependence of pathogenesis on iron acquisition in M. tuberculosis.     

The mechanism and specificity of iron transport in Rhodotorula pilimanae probed by synthetic analogs of rhodotorulic acid

The yeast Rhodotorula pilimanae produces the dihydroxamate siderophore rhodotorulic acid (RA) in prodigious amounts when starved for iron. Synthetic dihydroxamate analogs of RA have been prepared in which the diketopiperazine ring of RA is replaced by a simple chain of n methylene groups. It is found that R. pilimanae is able to accumulate iron using these achiral complexes, as well as from simple monohydroxamate analogs, at rates comparable to those of RA. While the Fe2RA3 complex does not enter the cell, there is a receptor system whose geometric requirements for siderophore recognition have been probed using analogs. In contrast to mono- or dihydroxamate ligands, the trihydroxamate siderophores such as ferrioxamine B are completely ineffective at delivering iron to R. pilimanae. This is ascribed to the greater stability of these complexes, which blocks release of the Fe(III) in a ligand exchange process that is required for uptake. To explore whether this ligand exchange involves redox catalysis, Ga(III) was substituted for Fe(III). The gallium was taken up at rates near those of iron and were also energy-dependent, as determined by metabolic inhibition with KCN.     

The iron-binding properties of aminochelin, the mono(catecholamide) siderophore of Azotobacter vinelandii

Azotobacter vinelandii produces siderophores with different metal-binding properties, depending on the concentration of Fe(III) and molybdate in the growth medium. The three protonation constants of the mono(catecholamide) siderophore aminochelin were determined by simultaneous spectrophotometric and potentiometric titrations as log K(1)=12.1, log K(2)=10.22 and log K(3)=7.04. Based on the two catechol protonation constants, log K(1) and log K(3), the overall stability constant of the aminochelin iron 3:1 complex was found to be log beta(3)=41.3, resulting in a pFe(3+) value of 17.6 at pH 7.45. In order to further investigate the properties of the siderophore, the solubilization of Fe(III) hydroxide by a 8x10(-4) M solution of aminochelin at pH 7 and 25 degrees C was followed spectrophotometrically in the absence and in the presence of molybdate. It was observed that the addition of molybdate resulted in a significant delay in the solubilization.     

Metabolic utilization of 57Fe-labeled coprogen in Neurospora crassa. An in vivo M?ssbauer study

M?ssbauer spectra of whole cells of Neurospora crassa arg-5 ota aga (a siderophore-free mutant) show that the siderophore coprogen is accumulated inside the cell as an entity. 57Fe from 57Fe-labeled coprogen is slowly removed from the complex (45% in 27 h). The rate of removal depends on the degree of iron starvation of the cells. The distribution of 55Fe from [55Fe]coprogen in vacuoles, membranes, and cytoplasm has been also determined. From this it is clear that coprogen is accumulated in the cytoplasm. In addition to its role as a siderophore, coprogen serves as an iron-storage compound. No holoferritins could be detected. We therefore conclude that this type of iron-storage protein is lacking in N. crassa. Metabolized iron was found predominantly to exist as an envelope of Fe(II) high-spin (delta = 1.2-1.3 mm s-1; delta EQ = 3.0-3.1 mm s-1 at 4.2 K) and fast-relaxing Fe(III) high-spin species (delta approximately equal to 0.25 mm s-1 and 0.45 mm s-1; delta EQ approximately equal to 0.6 mm s-1 and 0.55 mm s-1, respectively, at 4.2 K). An assignment of these major iron metabolites is difficult. The M?ssbauer data of the Fe(II) species do not fit those reported for heme, cytochromes and ferredoxins. We therefore assume that this iron metabolite represents a novel internal iron compound. One of the Fe(III) species becomes the dominant component of the cell spectra after 65 h of metabolization and might correspond to an iron-storage compound with iron oxide cores similar to bacterioferritin. After 27 h of growth in mycelia supplied with 57Fe-labeled coprogen, the siderophore ferricrocin was observed in the cell spectra. This is unexpected, since N. crassa arg-5 ota aga is unable to synthesize ornithine. We assume that ferricrocin is synthesized by the use of coprogen degradation products.     

Iron as a candidate in virulence and pathogenesis in mycobacteria and other microorganisms

Iron acquisition is one of the important virulence characteristics of a pathogen. Microorganisms elaborate different ways of acquiring iron. Siderophore-mediated iron acquisition is common in many microorganisms, while some like Neisseriae directly acquire iron from the host proteins. Microorganisms also elaborate systems for iron acquisition from ferric citrate and xenosiderophores. Such modes of uptake are also seen in mycobacteria. The siderophores produced by mycobacteria are well characterised. Iron-regulated envelope proteins (IREPs) are expressed in both saprophytic and pathogenic mycobacteria. In many bacterial systems, like that of Corynebacterium diphtheriae, low iron conditions favour the expression of virulence factors. The siderophore mycobactin has been recently implicated as a virulence factor in Mycobacterium tuberculosis. The role of the IdeR repressor in controlling the expression of the iron acquisition machinery was studied with the generation of the IdeR mutant. Development of other such mutants will facilitate the study of these mechanisms in greater detail and help to develop new drugs for combating tuberculosis and leprosy. This revised version was published online in November 2006 with corrections to the Cover Date.     

Exchange of iron by gallium in siderophores

Siderophores are iron transport compounds produced by numerous microorganisms and which strongly chelate Fe(III), but not Fe(II). Other trivalent metals, such as Al(III), Cr(III), or Ga(III), are not capable of significantly displacing iron from siderophores. However, I demonstrate here that Ga(III) can effectively displace iron under reducing conditions. With ascorbate as reductant and ferrozine as Fe(II) trapping agent, the kinetics of reductive displacement of iron by Ga(III) were followed spectroscopically by the increase of absorbance at 562 nm due to formation of the Fe(II)-ferrozine complex. No significant reduction of siderophore occurred in the absence of Ga(III). With excess Ga(III), the displacement was quantitative and very rapid. The rate of metal exchange was pseudo first order with respect to Ga(III) concentration and highly pH dependent, suggesting that siderophore ligands are displaced from the iron in a concerted mechanism by Ga(III) and protonation to expose the Fe(III) to reduction by ascorbate. Reaction rates were dependent upon the structure of the siderophore, being greatest for ferric rhodotorulic acid and slowest for ferrichrome A at pH 5.4. The pH profile for ferric rhodotorulic acid was unusual in that it showed a maximum at pH 6.5, while all other siderophores examined showed an increase in rate as pH was lowered from 7.0. The physiological significance of this reaction to the clinical use of gallium is discussed.     

Participation of fad and mbt genes in synthesis of mycobactin in Mycobacterium smegmatis

Colonies of Mycobacterium smegmatis LR222 on iron-limiting (0.1 micro M Fe) minimal medium agar fluoresce under UV light due to the accumulation in the cells of the deferri form of the siderophore mycobactin. Two mutants with little or no fluorescence, designated LUN8 and LUN9, were isolated by screening colonies of transposon (Tn611)-mutagenized M. smegmatis. Ferrimycobactin prepared from iron-restricted cells of the wild type had an R(f) of 0.62 on high-performance thin-layer chromatography (HPTLC) and a characteristic visible absorption spectrum with a peak near 450 nm. Similar extracts from LUN8 cells contained a small amount of ferrimycobactin with an R(f) of 0.58 on HPTLC and an absorption spectrum with the peak shifted to a wavelength lower than that of the wild-type ferrimycobactin. Nuclear magnetic resonance spectroscopy studies suggested that the LUN8 mycobactin may have an altered fatty acid side chain. Mutant strain LUN9 produced no detectable mycobactin. Neither mutant strain produced measurable amounts of excreted mycobactin, although both excreted exochelin (the mycobacterial peptido-hydroxamate siderophore), and both mutants were more sensitive than the wild-type strain to growth inhibition by the iron chelator ethylenediamine-di(o-hydroxyphenylacetic acid). The transposon insertion sites were identified, and sequence analyses of the cloned flanking chromosome regions showed that the mutated gene in LUN9 was an orthologue of the Mycobacterium tuberculosis mycobactin biosynthetic gene mbtE. The mutated gene in LUN8 had homology with M. tuberculosis fadD33 (Rv1345), a gene that may encode an acyl-coenzyme A synthase and which previously was not known to participate in synthesis of mycobactin.     

A simple and reliable EDDA method for mycobactin production in mycobacteria: Optimal conditions and use in mycobacterial speciation

Ethylenediamino-di-(0-hydroxyphenylacetic acid) (EDDA) was found to be a suitable iron chelator for investigating mycobactin production in mycobacteria. The optimal conditions of EDDA concentration and time of exposure were established and were used to develop a twostep method for mycobactin production. Applied to representative strains of selected species, the method was found to yield reliable results useful for identification and speciation of mycobacteria.     

Genomic insights into the iron uptake mechanisms of the biomining microorganism Acidithiobacillus ferrooxidans

Commercial bioleaching of copper and the biooxidation of gold is a cost-effective and environmentally friendly process for metal recovery. A partial genome sequence of the acidophilic, bioleaching bacterium Acidithiobacillus ferrooxidans is available from two public sources. This information has been used to build preliminary models that describe how this microorganism confronts unusually high iron loads in the extremely acidic conditions (pH 2) found in natural environments and in bioleaching operations. A. ferrooxidans contains candidate genes for iron uptake, sensing, storage, and regulation of iron homeostasis. Predicted proteins exhibit significant amino acid similarity with known proteins from neutrophilic organisms, including conservation of functional motifs, permitting their identification by bioinformatics tools and allowing the recognition of common themes in iron transport across distantly related species. However, significant differences in amino acid sequence were detected in pertinent domains that suggest ways in which the periplasmic and outer membrane proteins of A. ferrooxidans maintain structural integrity and relevant protein-protein contacts at low pH. Unexpectedly, the microorganism also contains candidate genes, organized in operon-like structures that potentially encode at least 11 siderophore systems for the uptake of Fe(III), although it does not exhibit genes that could encode the biosynthesis of the siderophores themselves. The presence of multiple Fe(III) uptake systems suggests that A. ferrooxidans can inhabit aerobic environments where iron is scarce and where siderophore producers are present. It may also help to explain why it cannot tolerate high Fe(III) concentrations in bioleaching operations where it is out-competed by Leptospirillum species.     

The kinetics and mechanisms of reactions of iron(III) with caffeic acid, chlorogenic acid, sinapic acid, ferulic acid and naringin

The kinetics and mechanisms of the reactions of iron(III) with the hydroxy cinnamic acid based ligands caffeic, chlorogenic, sinapic and ferulic acids and the flavonoid naringin have been investigated in aqueous solution. The mechanisms for caffeic and chlorogenic acid are generally consistent with the formation of a 1:1 complex that subsequently decays through an electron transfer reaction. On reaction with iron(III), ferulic and sinapic acids undergo an electron transfer without the prior formation of any complex. There was no evidence of electron transfer occurring in the complex formed when iron(III) is reacted with naringin. Rate constants for k1 (formation) and k(-1) (dissociation) have been evaluated for the complex formation reactions of [Fe(H2O)6(OH)]2+ with caffeic acid, chlorogenic acid and naringin. Analysis of the kinetic data yielded stability constants, equilibrium constants for protonation of the iron(III) chlorogenic acid complex initially formed, together with the rate constants for complex decomposition through intramolecular electron transfers and in the case of caffeic acid and chlorogenic acid, rate constants for the iron(III) assisted decomposition of the initial complex formed. Some of the suggested mechanisms and calculated rate constants are validated by calculations carried out using global analysis of time dependent spectra.     

Specificity of Staphyloferrin B Recognition by the SirA Receptor from Staphylococcus aureus

Many organisms use sophisticated systems to acquire growth-limiting iron. Iron limitation is especially apparent in bacterial pathogens of mammalian hosts where free iron concentrations are physiologically negligible. A common strategy is to secrete low molecular weight iron chelators, termed siderophores, and express high affinity receptors for the siderophore-iron complex. Staphylococcus aureus, a widespread pathogen, produces two siderophores, staphyloferrin A (SA) and staphyloferrin B (SB). We have determined the crystal structure of the staphyloferrin B receptor, SirA, at high resolution in both the apo and Fe(III)-SB (FeSB)-bound forms. SirA, a member of the class III binding protein family of metal receptors, has N- and C-terminal domains, each composed of mainly a β-stranded core and α-helical periphery. The domains are bridged by a single α-helix and together form the FeSB binding site. SB coordinates Fe(III) through five oxygen atoms and one nitrogen atom in distorted octahedral geometry. SirA undergoes conformational change upon siderophore binding, largely securing two loops from the C-terminal domain to enclose FeSB with a low nanomolar dissociation constant. The staphyloferrin A receptor, HtsA, homologous to SirA, also encloses its cognate siderophore (FeSA); however, the largest conformational rearrangements involve a different region of the C-terminal domain. FeSB is uniquely situated in the binding pocket of SirA with few of the contacting residues being conserved with those of HtsA interacting with FeSA. Although both SirA and HtsA bind siderophores from the same α-hydroxycarboxylate class, the unique structural features of each receptor provides an explanation for their distinct specificity.     

Characterization of the aqueous iron(III) chelation chemistry of a potential Trojan Horse antimicrobial agent: chelate structure, stability and pH dependent speciation

The aqueous solution equilibria of a β-lactam antimicrobial agent containing a 3-hydroxy, 4-pyridinone group (L (PF)) binding to Fe(III) in aqueous solution has been characterized through spectrophotometric and potentiometric titrations. The metal-free ligand has four observable protonation constants, pK(a1)?=?2.6, pK(a2)?=?3.43, pK(a3)?=?6.43, and pK(a4)?=?9.62. L (PF) forms a 3:1 ligand:Fe(III) complex in aqueous solution through coordinate-covalent bond formation exclusively involving the bidentate hydroxypyridinone moiety. This 3:1 L (PF):Fe complex was found to have a stability constant of log β(130)?=?33.46. A speciation diagram for the L (PF) system demonstrates that in the region of physiological pH the tris-(L (PF))Fe(III) complex, Fe(L(PF)) (3) (6-) , predominates. This complex exhibits two irreversible reduction waves in solution at -30?mV versus NHE, corresponding to a ligand-based reduction, and at -385?mV versus NHE, corresponding to an irreversible Fe(3+)/Fe(2+) reduction of the Fe(L(PF)) (3) (6-) complex.     

Spin, oxidation, and ligand states of p-nitrothiophenolatoiron(III) complex of protoporphyrin-IX-dimethylester in the presence of 1-MeIm: magnetic circular dichroism and 1H nuclear magnetic resonance spectroscopic studies

Complex formation of 5-coordinated iron(III) heme containing thiolate anion (p-nitrothiophenol) with imidazole (1-methylimidazole) showed very interesting features depending on the nature of the solvent and the ratio of the ligand to heme. The complexes formed under different conditions were not only low spin iron(III) complexes with a thiolate anion and an imidazole or with two imidazoles, but also reduced (iron(II] complexes with a thiolate and an imidazole or with two imidazoles. Absorption, magnetic circular dichroism, and 1H NMR spectroscopies could identify the complex formed when they were used concurrently. The dependence of polarity of the solvents used on the resultant chemical species was ascribed to the stability of Fe(III) or Fe(II) complex in the different solvents. The iron(III) complex with a thiolate anion and an imidazole was found to be reduced automatically to the iron(II) complex with a thiolate and an imidazole which exchanged ligand to the iron(II) bisimidazoles in the presence of excess imidazole. This study showed that the ligands of heme are easily exchanged and that the heme iron(III) is automatically reduced in several conditions. Possible significance with respect to biological systems containing a sulfur ligand is discussed.     

Genomic insights into the iron uptake mechanisms of the biomining microorganism Acidithiobacillus ferrooxidans

Commercial bioleaching of copper and the biooxidation of gold is a cost-effective and environmentally friendly process for metal recovery. A partial genome sequence of the acidophilic, bioleaching bacterium Acidithiobacillus ferrooxidans is available from two public sources. This information has been used to build preliminary models that describe how this microorganism confronts unusually high iron loads in the extremely acidic conditions (pH 2) found in natural environments and in bioleaching operations. A. ferrooxidans contains candidate genes for iron uptake, sensing, storage, and regulation of iron homeostasis. Predicted proteins exhibit significant amino acid similarity with known proteins from neutrophilic organisms, including conservation of functional motifs, permitting their identification by bioinformatics tools and allowing the recognition of common themes in iron transport across distantly related species. However, significant differences in amino acid sequence were detected in pertinent domains that suggest ways in which the periplasmic and outer membrane proteins of A. ferrooxidans maintain structural integrity and relevant protein-protein contacts at low pH. Unexpectedly, the microorganism also contains candidate genes, organized in operon-like structures that potentially encode at least 11 siderophore systems for the uptake of Fe(III), although it does not exhibit genes that could encode the biosynthesis of the siderophores themselves. The presence of multiple Fe(III) uptake systems suggests that A. ferrooxidans can inhabit aerobic environments where iron is scarce and where siderophore producers are present. It may also help to explain why it cannot tolerate high Fe(III) concentrations in bioleaching operations where it is out-competed by Leptospirillum species.