The Solution Structure, Binding Properties, and Dynamics of the Bacterial Siderophore-binding Protein FepB

Chu, Byron C. H.; Otten, Renee; Krewulak, Karla D.; Mulder, Frans A. A.; Vogel, Hans J.

doi:10.1074/jbc.m114.564021

Cited by 29 publications

(31 citation statements)

References 91 publications

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“…For instance, in bacterial importer MalFGK2, the transporter rests in an inward-open conformation and switches to outward-open upon MBP and ATP binding (27). For YbtPQ, although no specific SBP has been identified, one possibility is that SBPs from other siderophore uptake systems may be used here, for example, FepB of the enterobactin uptake system (36) and FhuD of the hydroxamate siderophore uptake system (37). Once SBP delivers the substrate to the transporter, YbtPQ's conformational transition happens, which will effectively stimulate ATP binding and hydrolysis.…”

Section: Discussionmentioning

confidence: 99%

Pathogenic siderophore ABC importer YbtPQ adopts a surprising fold of exporter

Wang

Zheng

2020

Sci. Adv.

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To fight for essential metal ions, human pathogens secrete virulence-associated siderophores and retake the metal-chelated siderophores through a subfamily of adenosine triphosphate (ATP)–binding cassette (ABC) importer, whose molecular mechanisms are completely unknown. We have determined multiple structures of the yersiniabactin importer YbtPQ from uropathogenic Escherichia coli (UPEC) at inward-open conformation in both apo and substrate-bound states by cryo–electron microscopy. YbtPQ does not adopt any known fold of ABC importers but surprisingly adopts the fold of type IV ABC exporters. To our knowledge, it is the first time an exporter fold of ABC importer has been reported. We have also observed two unique features in YbtPQ: unwinding of a transmembrane helix in YbtP upon substrate release and tightly associated nucleotide-binding domains without bound nucleotides. Together, our study suggests that siderophore ABC importers have a distinct transport mechanism and should be classified as a separate subfamily of ABC importers.

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Section: Discussionmentioning

confidence: 99%

Pathogenic siderophore ABC importer YbtPQ adopts a surprising fold of exporter

Wang

Zheng

2020

Sci. Adv.

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“…For Gram-positive bacteria, these include EntS-type major facilitator superfamily (MFS) exporter and import through a FepCDG-like multifunctional ABC transporter system associated with a FepB-like transporter through the ligand exchange mechanism. 33,34,35 Imported ferric iron bound to siderophore must be released to be made available to cellular machinery. One strategy for iron release is through hydrolytic destruction of the siderophore by esterases of the α,β-hydrolase family of enzymes, represented by the E. coli fes, iroD and iroE gene products and is a common strategy for macrolactone-based siderophores.…”

Section: Introductionmentioning

confidence: 99%

Structure and Mechanism of the Siderophore-Interacting Protein from the Fuscachelin Gene Cluster of Thermobifida fusca

Chen

Bruner

2015

Biochemistry

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Microbial iron acquisition is a complex process and frequently a key and necessary step for survival. Among the several paths for iron assimilation, small molecule siderophore-mediated transport is a commonly employed strategy of many microorganisms. The chemistry and biology of the extraordinary tight and specific binding of siderophores to metal is also exploited in therapeutic treatments for microbial virulence and metal toxicity. The intracellular fate of iron acquired via the siderophore pathway is one of the least understood steps in the complex process at the molecular level. A common route to cellular incorporation is the single-electron reduction of ferric to ferrous iron catalyzed by specific and/or nonspecific reducing agents. The biosynthetic gene clusters for siderophores often contain representatives of one or two families of redox-active enzymes: the flavin-containing "siderophore-interacting protein" and iron-sulfur ferric siderophore reductases. Here we present the structure and characterization of the siderophore-interacting protein, FscN, from the fuscachelin siderophore gene cluster of Thermobifida fusca. The structure shows a flavoreductase fold with a noncovalently bound FAD cofactor along with an unexpected metal bound adjacent to the flavin site. We demonstrated that FscN is redox-active and measured the binding and reduction of ferric fuscachelin. This work provides a structural basis for the activity of a siderophore-interacting protein and further insight into the complex and important process of iron acquisition and utilization.

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“…The equilibrium between open and closed conformations of E. coli periplasmic ligand binding proteins has previously been investigated by solution NMR spectroscopy for the maltose binding protein (Tang et al 2007), the glutamine binding protein (Bermejo et al 2010), the histidine binding protein (Chu et al 2013), the siderophore binding protein (Chu et al 2014), and the ribose and glucose/galactose binding proteins (Ortega et al 2012 In the case of DEBP, the PCSs confirm that the glutamate-bound protein assumes the closed structure observed in the crystal structure of the S. flexneri homologue. In contrast to the DEER data, which indicated that the closed conformation is predominantly populated even in the absence of glutamate (Abdelkader et al 2015), the changes in chemical shifts as well as PCSs indicate that the transition to the substrate-free protein involves a movement of the domains as rigid entities relative to each other.…”

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confidence: 82%

Pulse EPR-enabled interpretation of scarce pseudocontact shifts induced by lanthanide binding tags

et al. 2015

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Pseudocontact shifts (PCS) induced by tags loaded with paramagnetic lanthanide ions provide powerful long-range structure information, provided the location of the metal ion relative to the target protein is known. Usually, the metal position is determined by fitting the magnetic susceptibility anisotropy (Δχ) tensor to the 3D structure of the protein in an 8-parameter fit, which requires a large set of PCSs to be reliable. In an alternative approach, we used multiple Gd(3+)-Gd(3+) distances measured by double electron-electron resonance (DEER) experiments to define the metal position, allowing Δχ-tensor determinations from more robust 5-parameter fits that can be performed with a relatively sparse set of PCSs. Using this approach with the 32 kDa E. coli aspartate/glutamate binding protein (DEBP), we demonstrate a structural transition between substrate-bound and substrate-free DEBP, supported by PCSs generated by C3-Tm(3+) and C3-Tb(3+) tags attached to a genetically encoded p-azidophenylalanine residue. The significance of small PCSs was magnified by considering the difference between the chemical shifts measured with Tb(3+) and Tm(3+) rather than involving a diamagnetic reference. The integrative sparse data approach developed in this work makes poorly soluble proteins of limited stability amenable to structural studies in solution, without having to rely on cysteine mutations for tag attachment.

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The Solution Structure, Binding Properties, and Dynamics of the Bacterial Siderophore-binding Protein FepB

Cited by 29 publications

References 91 publications

Pathogenic siderophore ABC importer YbtPQ adopts a surprising fold of exporter

Pathogenic siderophore ABC importer YbtPQ adopts a surprising fold of exporter

Structure and Mechanism of the Siderophore-Interacting Protein from the Fuscachelin Gene Cluster of Thermobifida fusca

Pulse EPR-enabled interpretation of scarce pseudocontact shifts induced by lanthanide binding tags

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