Dataset Information

Use of SWATH mass spectrometry for quantitative proteomic investigation of Shewanella oneidensis MR-1 biofilms grown on graphite cloth electrodes

ABSTRACT: . In this study we show successful use of SWATH-MS for quantitative proteomic analysis of a microbial electrochemically active biofilm. Shewanella oneidensis MR-1 was grown on carbon cloth electrodes under continuous anodic electrochemical polarizations in a bioelectrochemical system. Using lactate as the electron donor, anodes serving as terminal microbial electron acceptors were operated at three different electrode potentials (+0.71V, +0.21V & -0.19V vs. SHE) and the development of catalytic activity was monitored by measuring the current traces over time. Once maximum current was reached (usually within 21-29 hours) the electrochemical systems were shut off and biofilm proteins were extracted from the electrodes for proteomic assessment.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Shewanella Oneidensis (strain Mr-1)

TISSUE(S): Biofilm

SUBMITTER: Christy Grobbler

LAB HEAD: Dr. Philip L. Bond

PROVIDER: PXD001472 | Pride | 2016-08-10

REPOSITORIES: Pride

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Use of SWATH mass spectrometry for quantitative proteomic investigation of Shewanella oneidensis MR-1 biofilms grown on graphite cloth electrodes.

Grobbler Christy C Virdis Bernardino B Nouwens Amanda A Harnisch Falk F Rabaey Korneel K Bond Philip L PL

Systematic and applied microbiology 20141129 2

Quantitative proteomics from low biomass, biofilm samples is not well documented. In this study we show successful use of SWATH-MS for quantitative proteomic analysis of a microbial electrochemically active biofilm. Shewanella oneidensis MR-1 was grown on carbon cloth electrodes under continuous anodic electrochemical polarizations in a bioelectrochemical system (BES). Using lactate as the electron donor, anodes serving as terminal microbial electron acceptors were operated at three different el ...[more]

PMID: 25523930

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Project description:The formation of electroactive biofilms is a crucial process for the generation of bioelectricity and bioremediation. G. sulfurreducens is a dissimilatory metal-reducing microorganism that can couple oxidation of organic matter with extracellular electron transfer to different insoluble electron acceptors. It has the capability to form biofilms in insoluble metal oxides and electroconductive biofilms in electrodes in bioelectrochemical systems. The formation of electroactive biofilms in this microorganism is a process that has been studied from a physiological, genetic, physical, and electrochemical approach. In G. sulfurreducens, we found that the transcriptional regulator GSU1771 participates in the gene expression of essential genes involved in electron transfer and biofilm formation. Strains deficient in GSU1771 increases Fe(III) reduction, produces more c-type cytochromes and exopolysaccharides. Furthermore, the biofilms produced are thicker and more electroactive than wild-type. In this work, we investigate the global gene expression profile performing RNA-seq comparing Δgsu1771 mutant biofilm grown in non-conductive support (glass) and respiring-graphite electrode. RNA-seq analysis of Δgsu1771 biofilm grown in glass support revealed a total of 467 (167 upregulated and 300 downregulated) differentially expressed genes versus the wild-type biofilm. Meanwhile, in Δgsu1771 biofilm developed in respiring-electrode graphite, we detect 119 (79 upregulated and 40 downregulated) differentially expressed genes with respect to wild-type biofilm. Moreover, transcriptional changes of 67 (56 with the same regulation and in 11 counterregulation) genes were shared in Δgsu1771 biofilm developed in glass and graphite electrodes. We locate upregulated in Δgsu1771 biofilms potential target genes, involved in exopolysaccharide synthesis (gsu1961-63, gsu1959, gsu1972-73, gsu1976-77). We confirmed the upregulation of gsu1979, gsu0972, gsu0783, pgcA, omcM, aroG, panC gnfK, gsu2507, and the downregulation of asnA, ato-1, gsu0810, pilA, csrA, ppcD, and gsu3356 genes by RT-qPCR. DNA-protein binding assay shows direct binding of the GSU1771 regulator to the promoter region of pgcA, pulF, relA, and gsu3356. Also, heme-staining and western blotting revealed an increase of c-type cytochromes in Δgsu1771 biofilms such as OmcS and OmcZ. In general, our data shows that GSU1771 is a global regulator involved in controlling the extracellular electron transfer and exopolysaccharide synthesis, processes required for electroconductive biofilm development.

Project description:G. sulfurreducens can generate electricity from the oxidation of organic compounds. This is because it can take electrons from organic compounds and ship them out to the outer surface of the cell where they can then be deposited on various insoluble electron acceptors including electrodes. Cells attatched to the surface of an electrode oxidize acetate and and deposit the electrons derived from acetate onto the surface of the electrode after which they can travel through an electrical circuit, producing a current. Microbial fuel cells powered by acetate oxidation by Geobacter species are called Geobatteries. In this experiment we compared gene expression in a biofilm of the wild type strain growing on the surface of an electrode within a current-producing Geobattery to gene expression in a wild type biofilm that is not producing current, but is growing on the surface of an electrode. In both cases, the cells were growing in a flow-through two chambered H-cell Geobattery setup. This consists of two glass chambers, an anoxic anode chamber containing G. sulfurreducens, a graphite electrode, a reference electrode and growth medium and an oxic chamber containing the counter electrode. The two chambers are connected by a cation selective membrane and a wire connected to a potentionstat. A potentiostat is an instrument which maintains the redox potential of the anode at a fixed value relative to a reference electrode. Media continuously flowed through the anoxic anode chamber at a dilution rate of 0.15. In the experimental condition, the Geobattery was operational. The circuit was closed and G. sulfurreducens attached to the electrode generated current as it oxidized acetate. The redox potential at the anode was poised at 300 mV by the potentiostat. In the control condition, everything was the same, except that the medium in the anode chamber contained fumarate as electron acceptor, and the anode was not hooked up to the potentiostat i.e. the circuit was open. This prevented the anode from serving as an electron acceptor. Nevertheless a thick G. sulfurreducens biofilm grew on the surface of the electrode. The control and experimental geobatteries were harvested when current in the operational/experimental Geobatteries reached 10 mA. Keywords: two condition comparison

Dataset Information

Use of SWATH mass spectrometry for quantitative proteomic investigation of Shewanella oneidensis MR-1 biofilms grown on graphite cloth electrodes

Publications

Use of SWATH mass spectrometry for quantitative proteomic investigation of Shewanella oneidensis MR-1 biofilms grown on graphite cloth electrodes.

Similar Datasets

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