Dataset Information

Vibrio cholerae FeoB contains a dual nucleotide-specific NTPase domain essential for ferrous iron uptake.

ABSTRACT: The Feo ferrous iron transporter is widely distributed among bacteria and archaea, but its mechanism of transport has not been fully elucidated. In Vibrio cholerae, the transport system requires three proteins: the small cytosolic proteins FeoA and FeoC and a large cytoplasmic-membrane-associated protein FeoB, which has an N-terminal G-protein domain. We show that, in contrast to Escherichia coli FeoB, which is solely a GTPase, the V. cholerae and Helicobacter pylori FeoB proteins have both GTPase and ATPase activity. In V. cholerae, mutation of the G4 motif, responsible for hydrogen bonding with the guanine base, abolished the GTPase activity but not ATPase activity. The ATPase activity of the G4 motif mutants was sufficient for Feo function in the absence of GTPase. We show that the serine and asparagine residues in the G5 motif likely play a role in the ATPase activity, and substitution of these residues with those found in the corresponding positions in E. coli FeoB resulted in similar nucleotide hydrolysis activity in the E. coli protein. These results add significantly to our understanding of the NTPase domain of FeoB and its role in Feo function.

SUBMITTER: Shin M

PROVIDER: S-EPMC6410867 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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Vibrio cholerae FeoB contains a dual nucleotide-specific NTPase domain essential for ferrous iron uptake.

Shin Minhye M Mey Alexandra R AR Payne Shelley M SM

Proceedings of the National Academy of Sciences of the United States of America 20190213 10

The Feo ferrous iron transporter is widely distributed among bacteria and archaea, but its mechanism of transport has not been fully elucidated. In Vibrio cholerae, the transport system requires three proteins: the small cytosolic proteins FeoA and FeoC and a large cytoplasmic-membrane-associated protein FeoB, which has an N-terminal G-protein domain. We show that, in contrast to Escherichia coli FeoB, which is solely a GTPase, the V. cholerae and Helicobacter pylori ...[more]

PMID: 30760591

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Project description:Vibrio cholerae is the causative agent of the severe diarrheal disease cholera. V. cholerae thrives within the human host, where it replicates to high numbers, but it also persists within the aquatic environments of ocean and brackish water. To survive within these nutritionally diverse environments, V. cholerae must encode the necessary tools to acquire the essential nutrient iron in all forms it may encounter. A prior study of systems involved in iron transport in V. cholerae revealed the existence of vciB, which, while unable to directly transport iron, stimulates the transport of iron through ferrous (Fe2+) iron transport systems. We demonstrate here a role for VciB in V. cholerae in which VciB stimulates the reduction of Fe3+ to Fe2+, which can be subsequently transported into the cell with the ferrous iron transporter Feo. Iron reduction is independent of functional iron transport but is associated with the electron transport chain. Comparative analysis of VciB orthologs suggests a similar role for other proteins in the VciB family. Our data indicate that VciB is a dimer located in the inner membrane with three transmembrane segments and a large periplasmic loop. Directed mutagenesis of the protein reveals two highly conserved histidine residues required for function. Taken together, our results support a model whereby VciB reduces ferric iron using energy from the electron transport chain.IMPORTANCEVibrio cholerae is a prolific human pathogen and environmental organism. The acquisition of essential nutrients such as iron is critical for replication, and V. cholerae encodes a number of mechanisms to use iron from diverse environments. Here, we describe the V. cholerae protein VciB that increases the reduction of oxidized ferric iron (Fe3+) to the ferrous form (Fe2+), thus promoting iron acquisition through ferrous iron transporters. Analysis of VciB orthologs in Burkholderia and Aeromonas spp. suggest that they have a similar activity, allowing a functional assignment for this previously uncharacterized protein family. This study builds upon our understanding of proteins known to mediate iron reduction in bacteria.

Project description:The Na+ ion-translocating NADH:quinone oxidoreductase (NQR) from Vibrio cholerae is a membrane-bound respiratory enzyme which harbors flavins and Fe-S clusters as redox centers. The NQR is the main producer of the sodium motive force (SMF) and drives energy-dissipating processes such as flagellar rotation, substrate uptake, ATP synthesis, and cation-proton antiport. The NQR requires for its maturation, in addition to the six structural genes nqrABCDEF, a flavin attachment gene, apbE, and the nqrM gene, presumably encoding a Fe delivery protein. We here describe growth studies and quantitative real-time PCR for the V. cholerae O395N1 wild-type (wt) strain and its mutant ?nqr and ?ubiC strains, impaired in respiration. In a comparative proteome analysis, FeoB, the membrane subunit of the uptake system for Fe2+ (Feo), was increased in V. cholerae ?nqr In this study, the upregulation was confirmed on the mRNA level and resulted in improved growth rates of V. cholerae ?nqr with Fe2+ as an iron source. We studied the expression of feoB on other respiratory enzyme deletion mutants such as the ?ubiC mutant to determine whether iron transport is specific to the absence of NQR resulting from impaired respiration. We show that the nqr operon comprises, in addition to the structural nqrABCDEF genes, the downstream apbE and nqrM genes on the same operon and demonstrate induction of the nqr operon by iron in V. cholerae wt. In contrast, expression of the nqrM gene in V. cholerae ?nqr is repressed by iron. The lack of functional NQR has a strong impact on iron homeostasis in V. cholerae and demonstrates that central respiratory metabolism is interwoven with iron uptake and regulation.IMPORTANCE Investigating strategies of iron acquisition, storage, and delivery in Vibrio cholerae is a prerequisite to understand how this pathogen thrives in hostile, iron-limited environments such as the human host. In addition to highlighting the maturation of the respiratory complex NQR, this study points out the influence of NQR on iron metabolism, thereby making it a potential drug target for antibiotics.

Dataset Information

Vibrio cholerae FeoB contains a dual nucleotide-specific NTPase domain essential for ferrous iron uptake.

Publications

<i>Vibrio cholerae</i> FeoB contains a dual nucleotide-specific NTPase domain essential for ferrous iron uptake.

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