Spelling suggestions: "subject:"shewanella oneidensis"" "subject:"schewanella oneidensis""
1 |
Electron transfer in multiheme cytochromes of Shewanella oneidensis MR-1: CymA and the dissimilatory metal reduction pathwaySherwood, Mackenzie A. Firer January 2012 (has links)
Thesis (Ph.D.)--Boston University / PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you. / Shewanella oneidensis is a facultative, gram-negative microbe that, in the absence of oxygen, can use a wide variety of terminal electron acceptors including iron, manganese, uranium, nitrite, nitrate, sulfate, fumarate, and DMSO. The anaerobic versatility is believed to be the result of a highly branched electron transfer pathway involving many redox-active proteins. Shewanella is capable of dissimilatory metal reduction (DMR) of insoluble iron and manganese oxides, in which electrons are transferred from the cell's interior to its exterior. Several multiheme c -type cytochromes comprise a pathway for this electron transfer. These cytochromes, specifically the tetraheme protein, CymA, and the decaheme protein, MtrA, are the primary focus of this thesis.
The current model of electron transfer indicates that electrons originate in the cytoplasmic membrane from the menaquinol pool, and are transferred into the periplasm by CymA. From here the pathway branches and electrons are transferred into several potential periplasmic targets, including MtrA. MtrA may then transfer electrons directly or indirectly to MtrC and OmcA, which have been shown to reduce exogenous electron acceptors such as iron oxides. Recently, it has been suggested that MtrA and MtrC dock with 13-barrel protein, MtrB and transfer electrons through the porin sheath.
Here, the DMR pathway has been studied with respect to four aims: (1) purification and characterization of the multiheme cytochromes through the use of non-catalytic protein film voltammetry (PFV), (2) structural analysis of MtrA by small angle X-ray scattering (SAXS), (3) investigation of protein-protein interactions via catalytic PFV and anaerobic affinity chromatography, and (4) exploration of heme cofactor function within the tetraheme cytochrome, CymA and MtrA by characterizing heme knockout mutants of the two proteins. We demonstrate that these proteins interact to form an electron transfer pathway from the cytoplasm to terminal electron acceptors on the outside of the cell through a "wire" of heme cofactors. Additionally, the data support the model that MtrA can span a large portion of the peri plasmic space to act as an intermediary by accepting electrons from CymA and subsequently docking with MtrB to transfer electrons to MtrC. / 2031-01-02
|
2 |
Bacterial iron and manganese reduction driven by organic sulfur electron shuttlesCooper, Rebecca Elizabeth 27 May 2016 (has links)
Dissimilatory metal-reducing bacteria (DMRB) play an important role in the biogeochemical cycling of metals. DMRB are unique in that they possess the ability to couple metal reduction with their metabolism. Microbial Fe(III) respiration is a central component of a variety of environmentally important processes, including the biogeochemical cycling of iron and carbon in redox stratified water and sediments, the bioremediation of radionuclide-contaminated water, the degradation of toxic hazardous pollutants, and the generation of electricity in microbial fuel cells. Despite this environmental and evolutionary importance, the molecular mechanism of microbial Fe(III) respiration is poorly understood. Current models of the molecular mechanism of microbial metal respiration are based on direct enzymatic, Fe(III) solubilization, and electron shuttling pathways. Fe(III) oxides are solid at circumneutral pH and therefore unable to come into direct contact with the microbial inner membrane, these bacteria must utilize an alternative strategy for iron reduction. Reduced organic compounds such as thiols are prominent in natural environments where DMRB are found. These thiol compounds are redox reactive and are capable of abiotically reducing Fe(III) oxides at high rates
S. oneidensis wild-type and ΔluxS anaerobic biofilm formation phenotypes were examined under a variety of electron donor-electron acceptor pairs, including lactate or formate as the electron donor and fumarate, thiosulfate, or Fe(III) oxide-coated silica surfaces as the terminal electron acceptor. The rates of biofilm formation under the aforementioned growth conditions as well as in the presence of exogenous thiol compounds indicate that ∆luxS formed biofilms at rates only 5-10% of the wild-type strain and ∆luxS biofilm formation rates were restored to wild-type levels by addition of a variety of exogenous compounds including cysteine, glutathione, homocysteine, methionine, serine, and homoserine. Cell adsorption isotherm analyses results indicate that wild-type is can attach to the surface of hematite particles attachment , but ΔluxS is unable to attach the hematite surfaces. These results indicate that biofilm formation is not required for Fe(III) oxide reduction by S. oneidensis
∆luxS anaerobic biofilm formation rates were restored to wild-type levels by addition of exogenous auntoinducer-2 (AI-2), a by-product of homocysteine production in the Activated Methyl Cycle. This discovery led to subsequent experiments performed to detect the production and utilization of AI-2 by wild-type and ∆luxS strains under aerobic and anaerobic conditions. AI-2 production experiments showed wild-type, but not ΔluxS, was capable of producing AI-2. The addition of exogenous S. oneidensis and Vibrio harveyi-produced AI-2 to wild-type and ∆luxS resulted in the swift depletion of AI-2 from the media. These results provide evidence that S. oneidensis can produce AI-2 and subsequently utilize its’ own AI-2 as well as AI-2 produced by other bacteria as a carbon and electron source in the absence of preferred carbon sources.
S. oneidensis produces and secretes a suite of extracellular thiols under anaerobic Fe(III)-reducing and Mn(III) and Mn(IV)-reducing conditions including cysteine, homocysteine, glutathione, and cyteamine. Exogenous thiols produced by S. oneidensis are intermediates of the Activated Methyl Cycle (AMC) and Transulfurylation Pathway (TSP). Reduced and oxidized thiols were detected, indicating that the thiols are in a constant state of flux between the reduced and oxidized forms and that the concentration of reduced thiols to its’ oxidized counterpart is indicative of the state of metal reduction by the microorganisms. Respiratory phenotypes Based on Fe(III) and Mn(IV) respiratory phenotypes observed in the AMC and TSP pathway mutants (∆luxS, ∆metB, ∆metC and ∆metY) we can infer that cysteine, glutathione, and cysteamine contribute to metal reduction by serving as efficient electron shuttling molecules, while homocysteine is critical for maintenance of the AMC, propagation of thiol biosynthesis, and maintenance of cellular metabolism via the AMC intermediate SAM. Furthermore, these findings suggest that all metal-reducing bacteria require thiol formation to reduce solid metal oxides. Direct contact mechanism is not the dominant means through electrons are transferred and metals are reduced, instead electron shuttles are the maid reduction mechanism.
|
3 |
Novel octaheme cytochrome c tetrathionate reductase (OTR) from Shewanella oneidensis MR-1Wu, Fei January 2010 (has links)
Octa-heme cytochrome c tetrathionate reductase (OTR) from Shewanella oneidensis MR-1 is a periplasmic protein and shows several extraordinary structural features around its active-site heme. OTR has been found able to catalyse the in vitro reduction of tetrathionate, nitrite, hydroxylamine and hydrogen peroxide. However the physiological function of this novel protein remains unknown. The subject of this thesis is the in vitro catalytic mechanism and the in vivo function of OTR. As OTR displays great similarity with bacterial penta-heme cytochrome c nitrite reductase (NrfA) in several aspects, it has been proposed that OTR might be physiologically involved in the metabolism of nitrite or other nitrogenous compounds. However kinetics assays and phenotypes studies carried out in this project suggest this is not the case. In vitro kinetic assays of the reduction of nitrite and hydroxylamine catalysed by OTR showed no significant difference in enzyme activities among the wild-type OTR and its mutant forms which have one active site residue replaced by alanine, namely OTR K153A, C64A, N61A and D150A. And the nitrite reductase activity of OTR (kcat/Km = 1.0×105 M-1•s-1) are much lower than that of NrfA (kcat/Km = ~108 M-1•s-1). These results indicate that OTR is not specifically adapted to reduce nitrite and it cannot compete for nitrite against NrfA in vivo. No phenotype difference was identified between the wild-type and the Δotr strain of Shewanella oneidensis MR-1 when nitrite or nitrate served as the sole electron acceptor. OTR appears not to be involved in the respiration or detoxification of nitrite, which is consistent with previous transcriptional and phenotype reports that involve OTR or its homologues. The in vitro tetrathionate reduction activity of OTR was unable to be reproduced in this project for unknown reasons. Although transcriptomic data from the literature suggest that OTR may be related to the metabolism of sulphur-containing compounds, kinetic and phenotype studies reveal that OTR does not directly participate in the respiration of thiosulfate, sulfite, tetrathionate, polysulfide or elemental sulphur. Cysteine 64 is a highly-conserved amino acid residue of OTR close to the active site and its side-chain sulphur atom is covalently bonded by either an oxygen or a sulphur atom as observed in the crystal structure. Such a modification is potentially important to the function of OTR. ESI mass spectroscopy results show that in native OTR the modified form is around 48 Da heavier than the unmodified form, and the MALDITOF peptide mass spectra show that the modified form could be converted into the unmodified form by reducing agent DTT. These results suggest that the modification could be a cysteine persulfide attaching an extra oxygen atom in the form of water or hydroxide anion.
|
4 |
Shewanella oneidensis MR-1 cell-to-cell signaling and its influences on biogeochemical processesLearman, Deric Ronald 26 June 2008 (has links)
The goal of this project is to decipher the quorum sensing (cell-to-cell signaling) abilities of Shewanella oneidensis MR-1, a Gram-negative bacterium well known for its ability to use geologic substrates, such as Fe and Mn oxides, for respiratory purposes. Overall our results show that S. oneidensis cannot utilize either an acyl-homoserine lactone (AHL) or AI-2 quorum sensing signal, despite previous work that indicated that it produced an AHL that would enhance it ability to growth in certain anaerobic environments. Using a variety of quorum sensing signal sensors, no evidence could be found that S. oneidensis has a typical AHL signal. An in silco analysis of the genome also produced little evidence that S. oneidensis has the genes to accept or relay an AHL signal. S. oneidensis can produce a luminescence response in the AI-2 reporter strain, Vibrio harveyi MM32. This luminescence response is abolished upon deletion of luxS, the gene responsible for catalyzing AI-2. Deletion of luxS also affected biofilm formation. Within 16 hours of growth in a biofilm flow-through reactor, the luxS mutant had an inhibited ability to initiate biofilm formation. After 48 hours of growth, the mutant's biofilm had developed similarly to wild-type. The addition of synthetic AI-2 did not restore the mutant's ability to initiation biofilm formation, which led to the conclusion that AI-2 is not likely used as a quorum sensing signal in S. oneidensis for this phenotype. Because of the involvement of LuxS in the activated methyl cycle (AMC) in other organisms, growth on various sulfur sources was examined. A mutation in luxS produced a reduced ability to growth with methionine as the sole sulfur source. Methionine is a key metabolite used in the AMC to produce a methyl source in the cell and homocysteine. This data suggests that LuxS is important in metabolizing methionine and the AMC in S. oneidensis. / Ph. D.
|
5 |
Conception de biofilms bactériens artificiels électroactifs en vue d’optimiser les réactions de transferts extracellulaires d’électrons / Conception of an artificial electroactive biofilm in order to promote electron transfer reactionsPinck, Stéphane 24 November 2017 (has links)
Nous avons cherché dans ce travail à élaborer un biofilm artificiel électroactif dans le but de promouvoir les réactions de transfert extracellulaire d’électrons (EET) en reconstituant artificiellement un biofilm en présence de matériaux exogènes. Un matériau composite auto-assemblé constitué de cellules bactériennes (Shewanella oneidensis), de nanotubes de carbone et de cytochromes c exogènes (issue de cellules de cœur de bœuf) a été tout d’abord proposé. Le processus d’auto-assemblage a été étudié par diffusion de lumière dynamique, microscopie électronique à balayage et spectroscopie Raman. Ces analyses ont mis en évidence l’importance du cytochrome exogène dans l’assemblage et l’organisation du matériau. La viabilité bactérienne a été étudiée et l’activité métabolique a été caractérisée par électrochimie. Les courants à l’anode étaient 10 et 4 fois plus importants avec ce biofilm artificiel (0,027 A m-2) qu’avec les électrodes modifiées par les bactéries seules (0,003 A m-2) ou associées au cytochrome c (0,007 A m-2). Le biofilm artificiel a été testé en substituant S. oneidensis par Pseudomonas fluorescens, produisant un courant d’oxydation lors de l’ajout de 1,5 mM de glucose. Le cytochrome c possède, outre son rôle structurant, une activité de navette à électrons. Son potentiel redox, 254 mV (vs NHE), était adapté à l’oxydation du formiate mais inadapté à la réduction du fumarate. Pour cette raison, il a été substitué par d’autres cytochromes (c3DvH, c7Da, c553DvH, c3DdN ou c3Dg) possédant des potentiels redox plus bas, de 20 mV à -400 mV. Ces cytochromes variaient aussi au niveau de leur charge à pH neutre, permettant de valider l’importance des forces électrostatiques dans l’assemblage du biocomposite. Les résultats optimaux obtenus avec c3DvH et c7Da ont montré l’importance du potentiel redox des éléments exogènes pour l’EET. Nous avons ensuite remplacé le cytochrome c par la protamine. Cette protéine non électroactive a permis l’assemblage du biocomposite tout en maintenant les transferts directs d’électrons entre les bactéries et les différents nanomatériaux testés. Les optimisations ont permis d’atteindre des courants cathodiques de plus de 12 A m-2 en présence de 50 mM de fumarate. Les expériences de stabilité ont montré la présence d’un courant biotique de 1,75 A m-2 après 24 h de réduction de 50 mM de fumarate / The aim of this PhD work was to design an artificial electroactive biofilm in order to optimize extracellular electron transfers (EET) by artificially reconstituting the biofilm in the presence of exogenous materials. A biocomposite material was proposed from the self-assembly of the bacteria Shewanella oneidensis with carbon nanotubes and cytochrome c (extract from bovine heart). The self-assembly was first studied by diffusion light scattering, scanning electron microscopy and Raman spectroscopy. These analyzes showed the importance of the cytochrome c in the assembly and organization of the biocomposite. Bacterial viability was studied and metabolic activity was characterized with the help of electrochemistry. The current at the anode was 10 and 4 times higher with the artificial biofilm (0.027 A m2) than with film composed with bacteria alone (0.003 A m2) or associated with cytochrome c (0.007 A m2). Artificial biofilm was also tested with Pseudomonas fluorescens instead of S. oneidensis, producing an oxidative current upon the addition of 1.5 mM glucose. That indicates cytochrome c has, in addition to its structuring role, an electron shuttle activity. Its redox potential, +254 mV (vs. NHE), was adapted to the oxidation of formate but was unsuitable for the reduction of fumarate. For this reason, it has been substituted by other cytochromes, c3DvH, c7Da, c553DvH, c3DdN, and c3Dg, possessing lower redox potentials, in the range of 20 mV to -400 mV. These cytochromes also varied at the level of their charge at neutral pH and allowed to validate the importance of the electrostatic forces in the assembly of the biocomposite. The optimal results obtained with c3DvH and c7Da showed the importance of the redox potential of the exogenous elements for the EET. We then replaced the cytochrome c with protamine. This non-electroactive protein allowed the assembly of the biocomposite by promoting direct electrons transfer between the bacteria and the different nanomaterials tested. The optimizations made it possible to reach cathodic currents of more than 12 A m2 in the presence of 50 mM of fumarate. The stability experiments showed the presence of a biotic current of 1.75 A m2 after 24 h of reduction of 50 mM of fumarate
|
6 |
Dissimilatory iron reduction: insights from the interaction between Shewanella oneidensis MR-1 and ferric iron (oxy)(hydr)oxide mineral surfacesZhang, Mengni 17 November 2010 (has links)
Dissimilatory iron reduction (DIR) is significant to the biogeochemical cycling of iron, carbon and other elements, and may be applied to bioremediation of organic pollutants, toxic metals, and radionuclides; however, the mechanism(s) of DIR and factors controlling its kinetics are still unclear. To provide insights into these questions, the interaction between a common dissimilatory iron reducing bacterium (DIRB)was studied, Shewanella oneidensis MR-1, and ferric iron (Fe(III)) (oxy)(hydr)oxide mineral surfaces. Firstly, atomic force microscopy was used to study how S. oneidensis MR-1 dissolved Fe(III) (oxy)(hydr)oxides and compared it to two other cases where Fe(III) (oxy)(hydr)oxides were either dissolved by a chemical reductant or by a mutant with an electron shuttling compound. Without the electron shuttling compound, the mutant is unable to respire on Fe(III) (oxy)(hydr)oxides, but with the electron shuttling compound, it can. It was found that the cells of S. oneidensis MR-1 formed microcolonies on mineral surfaces and dissolved the minerals in a non-uniform way which was consistent with the shape of microcolonies, whereas Fe(III) (oxy)(hydr)oxides were uniformly dissolved in both of the other cases. Secondly, confocal microscopy was used to study the adhesion behavior of S. oneidensis MR-1 cells on Fe(III) (oxy)(hydr)oxide surfaces across a broad range of bulk cell densities. While the cells were evenly distributed under low bulk cell densities, microcolonies were observed at high bulk cell densities. This adhesion behavior was modeled by a new, two-step adhesion isotherm which fit better than a simple Langmuir or Freundlich isotherm. The results of these studies suggest that DIR is in-part transport limited and the surface cell density may control DIR.
|
7 |
Chromium (VI) Reduction by <i>Shewanella oneidensis</i> MR-1 in Elevated Chromium Concentrations Exhibited in Corrosion Resistant Coatings.Miller, Robert B., II 05 June 2014 (has links)
No description available.
|
8 |
Bioreduction of Hematite Nanoparticles by Shewanella oneidensis MR-1Bose, Saumyaditya 09 January 2007 (has links)
A dissertation is presented on the bioreduction of hematite (α-Fe2O3) nanoparticles. The study shows that an alternative extracellular electron transfer mechanism other than the classical 'direct-contact' mechanism may be simultaneously employed by Shewanella oneidensis MR-1 during solid-phase metal reduction. This conclusion is supported by analysis of the bioreduction kinetics of hematite nanoparticles coupled with microscopic investigations of cell-mineral interactions. The reduction kinetics of metal-oxide nanoparticles were examined to determine how S. oneidensis utilizes these environmentally-relevant solid-phase electron acceptors. Nanoparticles involved in geochemical reactions show different properties relative to larger particles of the same phase, and their reactivity is predicted to change as a function of size. To demonstrate these size-dependent effects, the surface area normalized reduction rates of hematite nanoparticles by S. oneidensis MR-1 with lactate as the sole electron donor were measured. As evident from whole cell TEM analysis, the mode of nanoparticle adhesion to cells is different between the more aggregated, pseudo-hexagonal to irregular shaped 11 nm, 12 nm, 99 nm and the less aggregated 30 nm and 43 nm rhombohedral particles. The 11 nm, 12 nm and 99 nm particles show less cell contact and coverage than the 30 nm and 43 nm particles but still show significant rates of reduction. This leads to the provisional speculation that S. oneidensis MR-1 employs a pathway of indirect electron transfer in conjunction with the direct-contact pathway, and the relative importance of the mechanism employed depends upon aggregation level and the shape of the particles or crystal faces exposed. In accord with the proposed increase in electronic band-gap for hematite nanoparticles, the smallest particles (11 nm) exhibit one order of magnitude decrease in reduction when compared with larger (99 nm) particles, and the 12 nm rates fall in between these two. This effect may also be due to the passivation of the mineral and cell surfaces by Fe(II), or decreasing solubility due to decrease in size. / Ph. D.
|
9 |
Assessing the Microbial Consequences of Remediation: Surrogate Microbial Screening and Native Metabolic Signatures in Tc(VII) Contaminated SedimentsBailey, Kathryn Lafaye 01 January 2012 (has links)
The chemical and physical processes controlling contaminant fate and transport in the vadose zone limit the options for application of many remedial technologies. Foam delivery technology (FDT) has been developed as a potential solution to overcome these limitations for remediating subsurface and deep vadose zone environments using reactive amendments. Although there are many advantages to utilizing FDT for treatment in the deep vadose zone, little information is available on how the addition of these surfactants and remedial amendments affect the indigenous microbial communities in the deep vadose zone as well as the impact of biological transformations of surfactant-based foams on remediation efforts. The purpose of this study was to develop a rapid method for assessment of microbial communities in contaminated subsurface environments. This research was divided into two phases: (1) assess the toxicity of proposed FDT components on a single bacterial species, Shewanella oneidensis MR-1; and (2) determine the effects of these components on a microbial community from the vadose zone.
In Phase I, S. oneidensis MR-1 was exposed to proposed FDT components to assess potential growth inhibition or stimulation caused by these chemicals. S. oneidensis MR-1 cultures were exposed to the surfactants sodium laureth sulfate (SLES), sodium dodecyl sulfate (SDS), cocamidopropyl betaine (CAPB), and NINOL 40-CO, and the remedial amendment, calcium polysulfide (CPS). Results from this phase revealed that the relative acute toxicity order for these compounds was SDS>>CPS>>NINOL40-CO>SLES≥CAPB. High concentrations of SDS were toxic to the growth of S. oneidensis MR-1 but low concentrations were stimulatory. This benchtop system provided a capability to assess adverse microbial-remediation responses and contributed to the development of in situ remedial chemistries before they are deployed in the field.
For Phase II, sediments from the BC Cribs and Trenches (BCCT) area of the Hanford Site, WA, were characterized before and after exposure to potential FDT components. First, the phylogenetic and metabolic diversity of sediment from the BCCT was assessed by sequencing the microbial community and measuring the metabolic activity. The sediment was also incubated with various concentrations of SDS, CAPB, and CPS. Phylogenetic analysis detected phylotypes from the Alpha-, Beta-, Delta-, and Gammaproteobacteria, and Actinobacteria. Unlike the S. oneidensis MR-1 studies conducted in Phase I, the surfactants and CPS stimulated the metabolic activity of the native microbial communities. The observed stimulation could be caused by sorption of the chemicals to the sediment particles, or utilization of the surfactants by the microbial communities. These findings emphasize the importance of monitoring microbial activity at remediation sites in order to determine short and long term efficacy of the treatment, compliance with regulatory mandates, and act as an early warning indicator of unintended changes to the subsurface.
|
10 |
INVESTIGATING MICROBIOLOGICALLY INFLUENCED CORROSION USING THE ZERO-RESISTANCE AMMETRY TECHNIQUE IN A SPLIT CELL FORMATMiller, Robert B., II January 2019 (has links)
No description available.
|
Page generated in 0.067 seconds