Cell-secreted Flavins Bound to Membrane Cytochromes Dictate Electron Transfer Reactions to Surfaces with Diverse Charge and pH

Okamoto, Akimitsu; Kalathil, Shafeer; Deng, Xiao; Hashimoto, Kazuhito; Nakamura, Ryuhei; Nealson, Kenneth H.

doi:10.1038/srep05628

Cited by 148 publications

(123 citation statements)

References 57 publications

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“…To utilize the KIE to examine whether proton transport determines the rate of EET by OM Cyts c in in vivo measurements, the rate of EET by the OM Cyts c must limit or reflect the catalytic current ( I c ) production. For this reason, the experiments were conducted in an electrochemical system containing sufficient lactate as an electron donor and under pH and temperature conditions that support oxidative lactate metabolism, thereby limiting the I c to EET by OM Cyts c , as confirmed in previous reports 4c,4d, 9. The measurements were conducted following the electrochemical cultivation of MR‐1 as a monolayer biofilm on an ITO electrode, poised at +0.4 V [vs. the standard hydrogen electrode (SHE)].…”

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Proton Transport in the Outer‐Membrane Flavocytochrome Complex Limits the Rate of Extracellular Electron Transport

et al. 2017

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The microbial transfer of electrons to extracellularly located solid compounds, termed extracellular electron transport (EET), is critical for microbial electrode catalysis. Although the components of the EET pathway in the outer membrane (OM) have been identified, the role of electron/cation coupling in EET kinetics is poorly understood. We studied the dynamics of proton transport associated with EET in an OM flavocytochrome complex in Shewanella oneidensis MR‐1. Using a whole‐cell electrochemical assay, a significant kinetic isotope effect (KIE) was observed following the addition of deuterated water (D2O). The removal of a flavin cofactor or key components of the OM flavocytochrome complex significantly increased the KIE in the presence of D2O to values that were significantly larger than those reported for proton channels and ATP synthase, thus indicating that proton transport by OM flavocytochrome complexes limits the rate of EET.

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Proton Transport in the Outer‐Membrane Flavocytochrome Complex Limits the Rate of Extracellular Electron Transport

et al. 2017

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“…Reduction of insoluble acceptors occurs through a series of electron transfer proteins and molecules that span the inner membrane, periplasm, and outer membrane and transfer electrons from the quinone pool to the cell exterior. Multiple mechanisms for extracellular electron transfer (EET) have been studied in S. oneidensis, including direct contact, secretion of soluble electron shuttles, such as flavins, and the formation of structures termed bacterial nanowires that transfer electrons micrometers away from the cell body (1)(2)(3)(4). Each of these EET mechanisms requires outer membrane cytochromes, such as MtrC and OmcA (4,5).…”

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Regulation of Gene Expression in Shewanella oneidensis MR-1 during Electron Acceptor Limitation and Bacterial Nanowire Formation

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et al. 2016

Appl Environ Microbiol

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In limiting oxygen as an electron acceptor, the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 rapidly forms nanowires, extensions of its outer membrane containing the cytochromes MtrC and OmcA needed for extracellular electron transfer. RNA sequencing (RNA-Seq) analysis was employed to determine differential gene expression over time from triplicate chemostat cultures that were limited for oxygen. We identified 465 genes with decreased expression and 677 genes with increased expression. The coordinated increased expression of heme biosynthesis, cytochrome maturation, and transport pathways indicates that S. oneidensis MR-1 increases cytochrome production, including the transcription of genes encoding MtrA, MtrC, and OmcA, and transports these decaheme cytochromes across the cytoplasmic membrane during electron acceptor limitation and nanowire formation. In contrast, the expression of the mtrA and mtrC homologs mtrF and mtrD either remains unaffected or decreases under these conditions. The ompW gene, encoding a small outer membrane porin, has 40-fold higher expression during oxygen limitation, and it is proposed that OmpW plays a role in cation transport to maintain electrical neutrality during electron transfer. The genes encoding the anaerobic respiration regulator cyclic AMP receptor protein (CRP) and the extracytoplasmic function sigma factor RpoE are among the transcription factor genes with increased expression. RpoE might function by signaling the initial response to oxygen limitation. Our results show that RpoE activates transcription from promoters upstream of mtrC and omcA. The transcriptome and mutant analyses of S. oneidensis MR-1 nanowire production are consistent with independent regulatory mechanisms for extending the outer membrane into tubular structures and for ensuring the electron transfer function of the nanowires. IMPORTANCEShewanella oneidensis MR-1 has the capacity to transfer electrons to its external surface using extensions of the outer membrane called bacterial nanowires. These bacterial nanowires link the cell's respiratory chain to external surfaces, including oxidized metals important in bioremediation, and explain why S. oneidensis can be utilized as a component of microbial fuel cells, a form of renewable energy. In this work, we use differential gene expression analysis to focus on which genes function to produce the nanowires and promote extracellular electron transfer during oxygen limitation. Among the genes that are expressed at high levels are those encoding cytochrome proteins necessary for electron transfer. Shewanella coordinates the increased expression of regulators, metabolic pathways, and transport pathways to ensure that cytochromes efficiently transfer electrons along the nanowires.

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“…O 2 concentration was less than 0.1 ppm during the measurement in our experimental setup. 11 The lactate concentration in the medium was quantified using an ion chromatograph system (Shimadzu, HIC-20Asuper) as previously described.…”

Section: Electrochemical Measurementmentioning

confidence: 99%

Nanoscale Secondary Ion Mass Spectrometry Analysis of Individual Bacterial Cells Reveals Feedback from Extracellular Electron Transport to Upstream Reactions

2017

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We analyzed the metabolic activity of Shewanella oneidensis MR-1 respiring on an indium tin-doped oxide electrode with nanoscale secondary ion mass spectrometry (NanoSIMS) for quantification of the anabolism of 15 N-labeled NH 4 Cl. Although acceleration of extracellular electron transfer (EET) did not enhance the average metabolic activity, the distribution of 15 N intake among individual cells was drastically bipolarized upon EET rate enhancement, suggesting not only positive but also negative effects on cellular activity. The present data highlight the importance of controlling the downregulation of metabolic activity caused by the rate acceleration of EET to improve the efficiency of microbial electrode catalysis.

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Cell-secreted Flavins Bound to Membrane Cytochromes Dictate Electron Transfer Reactions to Surfaces with Diverse Charge and pH

Cited by 148 publications

References 57 publications

Proton Transport in the Outer‐Membrane Flavocytochrome Complex Limits the Rate of Extracellular Electron Transport

Proton Transport in the Outer‐Membrane Flavocytochrome Complex Limits the Rate of Extracellular Electron Transport

Regulation of Gene Expression in Shewanella oneidensis MR-1 during Electron Acceptor Limitation and Bacterial Nanowire Formation

Nanoscale Secondary Ion Mass Spectrometry Analysis of Individual Bacterial Cells Reveals Feedback from Extracellular Electron Transport to Upstream Reactions

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