Dataset Information

Interactions of proteins with biogenic iron oxyhydroxides and a new culturing technique to increase biomass yields of neutrophilic, iron-oxidizing bacteria.

ABSTRACT: Neutrophilic, bacterial iron-oxidation remains one of the least understood energy-generating biological reactions to date. One of the reasons it remains under-studied is because there are inherent problems with working with iron-oxidizing bacteria (FeOB), including low biomass yields and interference from the iron oxides in the samples. In an effort to circumvent the problem of low biomass, a new large batch culturing technique was developed. Protein interactions with biogenic iron oxides were investigated confirming that such interactions are strong. Therefore, a protein extraction method is described to minimize binding of proteins to biogenic iron oxides. The combination of these two methods results in protein yields that are appropriate for activity assays in gels and for proteomic profiling.

SUBMITTER: Barco RA

PROVIDER: S-EPMC4038746 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Interactions of proteins with biogenic iron oxyhydroxides and a new culturing technique to increase biomass yields of neutrophilic, iron-oxidizing bacteria.

Barco Roman A RA Edwards Katrina J KJ

Frontiers in microbiology 20140530

Neutrophilic, bacterial iron-oxidation remains one of the least understood energy-generating biological reactions to date. One of the reasons it remains under-studied is because there are inherent problems with working with iron-oxidizing bacteria (FeOB), including low biomass yields and interference from the iron oxides in the samples. In an effort to circumvent the problem of low biomass, a new large batch culturing technique was developed. Protein interactions with biogenic iron oxides were i ...[more]

PMID: 24910632

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Project description:Neutrophilic microaerophilic iron-oxidizing bacteria (FeOB) are thought to play a significant role in cycling of carbon, iron and associated elements in both freshwater and marine iron-rich environments. However, the roles of the neutrophilic microaerophilic FeOB are still poorly understood due largely to the difficulty of cultivation and lack of functional gene markers. Here, we analyze the genomes of two freshwater neutrophilic microaerophilic stalk-forming FeOB, Ferriphaselus amnicola OYT1 and Ferriphaselus strain R-1. Phylogenetic analyses confirm that these are distinct species within Betaproteobacteria; we describe strain R-1 and propose the name F. globulitus. We compare the genomes to those of two freshwater Betaproteobacterial and three marine Zetaproteobacterial FeOB isolates in order to look for mechanisms common to all FeOB, or just stalk-forming FeOB. The OYT1 and R-1 genomes both contain homologs to cyc2, which encodes a protein that has been shown to oxidize Fe in the acidophilic FeOB, Acidithiobacillus ferrooxidans. This c-type cytochrome common to all seven microaerophilic FeOB isolates, strengthening the case for its common utility in the Fe oxidation pathway. In contrast, the OYT1 and R-1 genomes lack mto genes found in other freshwater FeOB. OYT1 and R-1 both have genes that suggest they can oxidize sulfur species. Both have the genes necessary to fix carbon by the Calvin-Benson-Basshom pathway, while only OYT1 has the genes necessary to fix nitrogen. The stalk-forming FeOB share xag genes that may help form the polysaccharide structure of stalks. Both OYT1 and R-1 make a novel biomineralization structure, short rod-shaped Fe oxyhydroxides much smaller than their stalks; these oxides are constantly shed, and may be a vector for C, P, and metal transport to downstream environments. Our results show that while different FeOB are adapted to particular niches, freshwater and marine FeOB likely share common mechanisms for Fe oxidation electron transport and biomineralization pathways.

Project description:Iron (Fe) oxidation is one of Earth's major biogeochemical processes, key to weathering, soil formation, water quality, and corrosion. However, our understanding of microbial contribution is limited by incomplete knowledge of microbial iron oxidation mechanisms, particularly in neutrophilic iron oxidizers. The genomes of many diverse iron oxidizers encode a homolog to an outer membrane cytochrome (Cyc2) shown to oxidize iron in two acidophiles. Phylogenetic analyses show Cyc2 sequences from neutrophiles cluster together, suggesting a common function, though this function has not been verified in these organisms. Therefore, we investigated the iron oxidase function of heterologously expressed Cyc2 from a neutrophilic iron oxidizer Mariprofundus ferrooxydans PV-1. Cyc2PV-1 is capable of oxidizing iron, and its redox potential is 208 ± 20 mV, consistent with the ability to accept electrons from Fe2+ at neutral pH. These results support the hypothesis that Cyc2 functions as an iron oxidase in neutrophilic iron-oxidizing organisms. The results of sequence analysis and modeling reveal that the entire Cyc2 family shares a unique fused cytochrome-porin structure, with a defining consensus motif in the cytochrome region. On the basis of results from structural analyses, we predict that the monoheme cytochrome Cyc2 specifically oxidizes dissolved Fe2+, in contrast to multiheme iron oxidases, which may oxidize solid Fe(II). With our results, there is now functional validation for diverse representatives of Cyc2 sequences. We present a comprehensive Cyc2 phylogenetic tree and offer a roadmap for identifying cyc2/Cyc2 homologs and interpreting their function. The occurrence of cyc2 in many genomes beyond known iron oxidizers presents the possibility that microbial iron oxidation may be a widespread metabolism. IMPORTANCE Iron is practically ubiquitous across Earth's environments, central to both life and geochemical processes, which depend heavily on the redox state of iron. Although iron oxidation, or "rusting," can occur abiotically at near-neutral pH, we find neutrophilic iron-oxidizing bacteria (FeOB) are widespread, including in aquifers, sediments, hydrothermal vents, pipes, and water treatment systems. FeOB produce highly reactive Fe(III) oxyhydroxides that bind a variety of nutrients and toxins; thus, these microbes are likely a controlling force in iron and other biogeochemical cycles. There has been mounting evidence that Cyc2 functions as an iron oxidase in neutrophiles, but definitive proof of its function has long eluded us. This work provides conclusive biochemical evidence of iron oxidation by Cyc2 from neutrophiles. Cyc2 is common to a wide variety of iron oxidizers, including acidophilic and phototrophic iron oxidizers, suggesting that this fused cytochrome-porin structure is especially well adapted for iron oxidation.

Dataset Information

Interactions of proteins with biogenic iron oxyhydroxides and a new culturing technique to increase biomass yields of neutrophilic, iron-oxidizing bacteria.

Publications

Interactions of proteins with biogenic iron oxyhydroxides and a new culturing technique to increase biomass yields of neutrophilic, iron-oxidizing bacteria.

Similar Datasets

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