Sulfate as a Synergistic Anion Facilitating Iron Binding by the Bacterial Transferrin FbpA:  The Origins and Effects of Anion Promiscuity

Heymann, Jared J.; Weaver, Katherine; Mietzner, Timothy A.; Crumbliss, Alvin L.

doi:10.1021/ja0709268

Cited by 28 publications

(37 citation statements)

References 45 publications

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“…The phosphate concentration dependence of the rate in Figure 4 may be attributed to the relative shift in equilibrium reaction 10 with increasing phosphate concentration. This assertion is supported by a plot of % total protein complexed as apo-FbpA-PO 4 as a function of [H x PO 4 y- ] (Figure 4, insert) (19). …”

Section: Resultsmentioning

confidence: 81%

Role of Citrate and Phosphate Anions in the Mechanism of Iron(III) Sequestration by Ferric Binding Protein: Kinetic Studies of the Formation of the Holoprotein of Wild-Type FbpA and Its Engineered Mutants

et al. 2010

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Ferric binding protein A (FbpA) plays a central role in the iron acquisition processes of pathogenic Neisseria gonorrhoeae, Neisseria meningitidis, and Haemophilus influenza. FbpA functions as an iron shuttle within the periplasmic space of these Gram-negative human pathogens. Iron is picked up by FbpA at the periplasmic aspect of the outer membrane with concomitant acquisition of a synergistic anion. Here we report the kinetics and mechanisms involved with iron(III) loading into iron-free FbpA using iron(III) citrate as an iron source in the presence of excess citrate or phosphate (physiologically available anions) at pH 6.5. In the presence of excess phosphate, iron(III) citrate loads into apo-FbpA in three kinetically distinguishable steps, while in the presence of excess citrate only two steps are discernible. A stable intermediate containing iron(III) citrate bound FbpA is observed in each case. The observation of an additional kinetic step and moderate increase in apparent rate constants suggests an active role for phosphate in the iron insertion process. To further elucidate a mechanism for iron-loading, we report on the sequestration kinetics of iron(III) citrate in the presence of phosphate with binding site mutant apo-FbpAs, H9E, E57D, E57Q, Q58A, Y195F, and Y196H. Tyrosine mutations drastically alter the kinetics, and reduce iron sequestration ability. H9E, E57D, and E57Q have near native iron sequestration behavior; however iron binding rates are altered enabling assignment of sequential side chain interactions. Additionally, this investigation elaborates on the function of FbpA as a carrier for iron chelates as well as “naked” or free iron as originally proposed.

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Section: Resultsmentioning

confidence: 81%

Role of Citrate and Phosphate Anions in the Mechanism of Iron(III) Sequestration by Ferric Binding Protein: Kinetic Studies of the Formation of the Holoprotein of Wild-Type FbpA and Its Engineered Mutants

et al. 2010

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“…These transporters are conventionally called Fe 3+ ABC transporters, but the precise form of inorganic Fe(III) transported is debated (e.g. Zhu et al ., 2003; Heymann et al ., 2007). Although only a few Fe 3+ ABC transporters have been characterized, orthologue detection via OrthoMCL produced coherent groups containing the well‐characterized Fe 3+ transporters for each Fe 3+ ABC transport component, allowing identification of Fe 3+ ABC components in marine prokaryotes.…”

Section: Discussionmentioning

confidence: 99%

Iron transporters in marine prokaryotic genomes and metagenomes

Hopkinson

Barbeau

2011

Environmental Microbiology

101

123

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In the pelagic environment, iron is a scarce but essential micronutrient. The iron acquisition capabilities of selected marine bacteria have been investigated, but the recent proliferation of marine prokaryotic genomes and metagenomes offers a more comprehensive picture of microbial iron uptake pathways in the ocean. Searching these data sets, we were able to identify uptake mechanisms for Fe(3+), Fe(2+) and iron chelates (e.g. siderophore and haem iron complexes). Transport of iron chelates is accomplished by TonB-dependent transporters (TBDTs). After clustering the TBDTs from marine prokaryotic genomes, we identified TBDT clusters for the transport of hydroxamate and catecholate siderophore iron complexes and haem using gene neighbourhood analysis and co-clustering of TBDTs of known function. The genomes also contained two classes of siderophore biosynthesis genes: NRPS (non-ribosomal peptide synthase) genes and NIS (NRPS Independent Siderophore) genes. The most common iron transporters, in both the genomes and metagenomes, were Fe(3+) ABC transporters. Iron uptake-related TBDTs and siderophore biosynthesis genes were less common in pelagic marine metagenomes relative to the genomic data set, in part because Pelagibacter ubique and Prochlorococcus species, which almost entirely lacked these Fe uptake systems, dominate the metagenomes. Our results are largely consistent with current knowledge of iron speciation in the ocean, but suggest that in certain niches the ability to acquire siderophores and/or haem iron chelates is beneficial.

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“…The proton independent stability constant for the tris catecholato siderophores are remarkably large with Fe(enterobactin) 3- at 1049,78 and Fe(bacillibactin) 3- at 10 47.6 77. In each case, Fe(III) is present in the high-spin electronic configuration.…”

Section: Siderophoresmentioning

confidence: 99%