scholarly journals Single molecule tracking of bacterial cell surface cytochromes reveals dynamics that impact long-distance electron transport

2021 ◽  
Author(s):  
Grace W Chong ◽  
Sahand Pirbadian ◽  
Yunke Zhao ◽  
Lori A Zacharoff ◽  
Fabien Pinaud ◽  
...  
Keyword(s):  
Electron Transport ◽  
Quantum Dot ◽  
Single Molecule ◽  
Shewanella Oneidensis ◽  
Single Particle Tracking ◽  
Extracellular Electron Transfer ◽  
Long Distance ◽  
Electron Hopping ◽  
Diffusive Dynamics

Using a series of multiheme cytochromes, the metal-reducing bacterium Shewanella oneidensis MR-1 can perform extracellular electron transfer (EET) to respire redox-active surfaces, including minerals and electrodes outside the cell. While the role of multiheme cytochromes in transporting electrons across the cell wall is well established, these cytochromes were also recently found to facilitate long-distance (micrometer-scale) redox conduction along outer membranes and across multiple cells bridging electrodes. Recent studies proposed that long-distance conduction arises from the interplay of electron hopping and cytochrome diffusion, which allows collisions and electron exchange between cytochromes along membranes. However, the diffusive dynamics of the multiheme cytochromes have never been observed or quantified in vivo, making it difficult to assess their hypothesized contribution to the collision-exchange mechanism. Here we use quantum dot labeling, total internal reflection fluorescence microscopy, and single-particle tracking to quantify the lateral diffusive dynamics of the outer membrane-associated decaheme cytochromes MtrC and OmcA, two key components of EET in S. oneidensis. We observe confined diffusion behavior for both quantum dot-labeled MtrC and OmcA along cell surfaces (diffusion coefficients DMtrC = 0.0192 ± 0.0018 μm2/s, DOmcA = 0.0125 ± 0.0024 μm2/s) and the membrane extensions thought to function as bacterial nanowires. We find that these dynamics can trace a path for electron transport via overlap of cytochrome trajectories, consistent with the long-distance conduction mechanism. The measured dynamics inform kinetic Monte Carlo simulations that combine direct electron hopping and redox molecule diffusion, revealing significant electron transport rates along cells and membrane nanowires.

scholarly journals Spin-Dependent Electron Transport through Bacterial Cell Surface Multiheme Electron Conduits

2019 ◽  
Author(s):  
Suryakant Mishra ◽  
Sahand Pirbadian ◽  
Amit Kumar Mondal ◽  
Moh El-Naggar ◽  
Ron Naaman
Keyword(s):  
Electron Transport ◽  
Cell Surface ◽  
Bacterial Cell ◽  
Shewanella Oneidensis ◽  
Extracellular Electron Transfer ◽  
Long Distance ◽  
Force Microscopy ◽  
Bacterial Cell Surface ◽  
Redox Active ◽  
Gain Energy

Multiheme cytochromes, located on the bacterial cell surface, function as long-distance (> 10 nm) electron conduits linking intracellular reactions to external surfaces. This extracellular electron transfer process, which allows microorganisms to gain energy by respiring solid redox-active minerals, also facilitates the wiring of cells to electrodes. While recent studies suggested that a chiral induced spin selectivity effect is linked to efficient electron transmission through biomolecules, this phenomenon has not been investigated in the extracellular electron conduits. Using magnetic conductive probe atomic force microscopy, Hall voltage measurements, and spin-dependent electrochemistry of the decaheme cytochromes MtrF and OmcA from the metal-reducing bacterium <i>Shewanella oneidensis</i> MR-1, we show that electron transport through these extracellular conduits is spin-selective. Our study has implications for understanding how spin-dependent interactions and magnetic fields may control electron transport across biotic-abiotic interfaces in both natural and biotechnological systems.

scholarly journals Spin-Dependent Electron Transport through Bacterial Cell Surface Multiheme Electron Conduits

2019 ◽  
Author(s):  
Suryakant Mishra ◽  
Sahand Pirbadian ◽  
Amit Kumar Mondal ◽  
Moh El-Naggar ◽  
Ron Naaman
Keyword(s):  
Electron Transport ◽  
Cell Surface ◽  
Bacterial Cell ◽  
Shewanella Oneidensis ◽  
Extracellular Electron Transfer ◽  
Long Distance ◽  
Force Microscopy ◽  
Bacterial Cell Surface ◽  
Redox Active ◽  
Gain Energy

Multiheme cytochromes, located on the bacterial cell surface, function as long-distance (> 10 nm) electron conduits linking intracellular reactions to external surfaces. This extracellular electron transfer process, which allows microorganisms to gain energy by respiring solid redox-active minerals, also facilitates the wiring of cells to electrodes. While recent studies suggested that a chiral induced spin selectivity effect is linked to efficient electron transmission through biomolecules, this phenomenon has not been investigated in the extracellular electron conduits. Using magnetic conductive probe atomic force microscopy, Hall voltage measurements, and spin-dependent electrochemistry of the decaheme cytochromes MtrF and OmcA from the metal-reducing bacterium <i>Shewanella oneidensis</i> MR-1, we show that electron transport through these extracellular conduits is spin-selective. Our study has implications for understanding how spin-dependent interactions and magnetic fields may control electron transport across biotic-abiotic interfaces in both natural and biotechnological systems.

scholarly journals Syncrip/hnRNP Q is required for activity-induced Msp300/Nesprin-1 expression and new synapse formation

2020 ◽  
Vol 219 (3) ◽  
Author(s):  
Joshua Titlow ◽  
Francesca Robertson ◽  
Aino Järvelin ◽  
David Ish-Horowicz ◽  
Carlas Smith ◽  
...  
Keyword(s):  
Single Molecule ◽  
Posttranscriptional Regulation ◽  
Rna Binding ◽  
Synapse Formation ◽  
Long Distance ◽  
Key Factor ◽  
Larval Neuromuscular Junction ◽  
Rnp Granules ◽  
Activity Dependent

Memory and learning involve activity-driven expression of proteins and cytoskeletal reorganization at new synapses, requiring posttranscriptional regulation of localized mRNA a long distance from corresponding nuclei. A key factor expressed early in synapse formation is Msp300/Nesprin-1, which organizes actin filaments around the new synapse. How Msp300 expression is regulated during synaptic plasticity is poorly understood. Here, we show that activity-dependent accumulation of Msp300 in the postsynaptic compartment of the Drosophila larval neuromuscular junction is regulated by the conserved RNA binding protein Syncrip/hnRNP Q. Syncrip (Syp) binds to msp300 transcripts and is essential for plasticity. Single-molecule imaging shows that msp300 is associated with Syp in vivo and forms ribosome-rich granules that contain the translation factor eIF4E. Elevated neural activity alters the dynamics of Syp and the number of msp300:Syp:eIF4E RNP granules at the synapse, suggesting that these particles facilitate translation. These results introduce Syp as an important early acting activity-dependent regulator of a plasticity gene that is strongly associated with human ataxias.

scholarly journals In vivo assembly and single-molecule characterization of the transcription machinery from Shewanella oneidensis MR-1

2009 ◽  
Vol 65 (1) ◽  
pp. 66-76 ◽  
Author(s):  
Natalie R. Gassman ◽  
Sam On Ho ◽  
You Korlann ◽  
Janet Chiang ◽  
Yim Wu ◽  
...  
Keyword(s):  
Single Molecule ◽  
Shewanella Oneidensis ◽  
Transcription Machinery

scholarly journals Surface-induced formation and redox-dependent staining of outer membrane extensions inShewanella oneidensisMR-1

2019 ◽  
Author(s):  
Grace W. Chong ◽  
Sahand Pirbadian ◽  
Mohamed Y. El-Naggar
Keyword(s):  
Outer Membrane ◽  
Shewanella Oneidensis ◽  
Medium Composition ◽  
Extracellular Electron Transfer ◽  
Active Components ◽  
Transmission Electron ◽  
Redox Active ◽  
Redox Centers

AbstractThe metal-reducing bacteriumShewanella oneidensisMR-1 produces extensions of its outer membrane (OM) and periplasm that contain cytochromes responsible for extracellular electron transfer (EET) to external redox-active surfaces, including minerals and electrodes. While the role of multi-heme cytochromes in transporting electrons across the cell wall is well established, their distribution alongS. oneidensisOM extensions is also thought to allow lateral electron transport along these filaments. These proposed bacterial nanowires, which can be several times the cell length, would thereby extend EET to more distant electron acceptors. However, it is still unclear why these extensions form, and to what extent they contribute to respiration in living cells. Here, we investigate physical contributors to their formation usingin vivofluorescence microscopy. While previous studies focused on the display ofS. oneidensisouter membrane extensions (OMEs) as a response to oxygen limitation, we find that cell-to-surface contact is sufficient to trigger the production of OMEs, including some that reach >100 µm in length, irrespective of medium composition, agitation, or aeration. To visualize the extent of heme redox centers along OMEs, and help distinguish these structures from other extracellular filaments, we also performed histochemical redox-dependent staining with transmission electron microscopy on wild type and cytochrome-deficient strains. We demonstrate that redox-active components are limited to OMEs and not present on other extracellular appendages, such as pili and flagella. We also observed that the loss of 8 functional periplasmic and outer membrane cytochromes significantly decreased both the frequency and intensity of redox-dependent staining found widespread on OMEs. These results will improve our understanding of the environmental conditions that influence the formation ofS. oneidensisOMEs, as well as the distribution and functionality of EET components along extracellular appendages.

scholarly journals A Novel Complex: A Quantum Dot Conjugated to an Active T7RNA Polymerase

2013 ◽  
Vol 2013 ◽  
pp. 1-9 ◽  
Author(s):  
Mette Eriksen ◽  
Peter Horvath ◽  
Michael A. Sørensen ◽  
Szabolcs Semsey ◽  
Lene B. Oddershede ◽  
...  
Keyword(s):  
Quantum Dot ◽  
Rna Polymerase ◽  
Single Molecule ◽  
Rna Polymerases ◽  
Fluorescent Signal ◽  
Single Molecule Level ◽  
Single Molecule Studies ◽  
Novel Complex ◽  
Luminescent Nanoparticle

To perform single-molecule studies of the T7RNA polymerase, it is crucial to visualize an individual T7RNA polymerase, for example, through a fluorescent signal. We present a novel complex combining two different molecular functions, an active T7RNA polymerase and a highly luminescent nanoparticle, a quantum dot. The complex has the advantage of both constituents: the complex can traffic along DNA and simultaneously be visualized, both at the ensemble and at the single-molecule level. The labeling was mediated through anin vivobiotinylation of a His-tagged T7RNA polymerase and subsequent binding of a streptavidin-coated quantum dot. Our technique allows for easy purification of the quantum dot labeled T7RNA polymerases from the reactants. Also, the conjugation does not alter the functionality of the polymerase; it retains the ability to bind and transcribe.

scholarly journals In vivo single-molecule imaging of syntaxin1A reveals polyphosphoinositide- and activity-dependent trapping in presynaptic nanoclusters

2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Adekunle T. Bademosi ◽  
Elsa Lauwers ◽  
Pranesh Padmanabhan ◽  
Lorenzo Odierna ◽  
Ye Jin Chai ◽  
...  
Keyword(s):  
Single Molecule ◽  
Neurotransmitter Release ◽  
Motor Nerve ◽  
Living Organism ◽  
Neurosecretory Cells ◽  
Single Particle Tracking ◽  
Snare Complex ◽  
Motor Nerve Terminal ◽  
Activity Dependent

Abstract Syntaxin1A is organized in nanoclusters that are critical for the docking and priming of secretory vesicles from neurosecretory cells. Whether and how these nanoclusters are affected by neurotransmitter release in nerve terminals from a living organism is unknown. Here we imaged photoconvertible syntaxin1A-mEos2 in the motor nerve terminal of Drosophila larvae by single-particle tracking photoactivation localization microscopy. Opto- and thermo-genetic neuronal stimulation increased syntaxin1A-mEos2 mobility, and reduced the size and molecular density of nanoclusters, suggesting an activity-dependent release of syntaxin1A from the confinement of nanoclusters. Syntaxin1A mobility was increased by mutating its polyphosphoinositide-binding site or preventing SNARE complex assembly via co-expression of tetanus toxin light chain. In contrast, syntaxin1A mobility was reduced by preventing SNARE complex disassembly. Our data demonstrate that polyphosphoinositide favours syntaxin1A trapping, and show that SNARE complex disassembly leads to syntaxin1A dissociation from nanoclusters. Lateral diffusion and trapping of syntaxin1A in nanoclusters therefore dynamically regulate neurotransmitter release.

scholarly journals Shewanella oneidensis as a living electrode for controlled radical polymerization

2018 ◽  
Vol 115 (18) ◽  
pp. 4559-4564 ◽  
Author(s):  
Gang Fan ◽  
Christopher M. Dundas ◽  
Austin J. Graham ◽  
Nathaniel A. Lynd ◽  
Benjamin K. Keitz
Keyword(s):  
Electron Transfer ◽  
Metabolic Engineering ◽  
Electron Transport ◽  
Radical Polymerization ◽  
Copper Catalyst ◽  
Shewanella Oneidensis ◽  
Extracellular Electron Transfer ◽  
Metal Catalyst ◽  
General Strategy ◽  
Significant Activity

Metabolic engineering has facilitated the production of pharmaceuticals, fuels, and soft materials but is generally limited to optimizing well-defined metabolic pathways. We hypothesized that the reaction space available to metabolic engineering could be expanded by coupling extracellular electron transfer to the performance of an exogenous redox-active metal catalyst. Here we demonstrate that the electroactive bacterium Shewanella oneidensis can control the activity of a copper catalyst in atom-transfer radical polymerization (ATRP) via extracellular electron transfer. Using S. oneidensis, we achieved precise control over the molecular weight and polydispersity of a bioorthogonal polymer while similar organisms, such as Escherichia coli, showed no significant activity. We found that catalyst performance was a strong function of bacterial metabolism and specific electron transport proteins, both of which offer potential biological targets for future applications. Overall, our results suggest that manipulating extracellular electron transport pathways may be a general strategy for incorporating organometallic catalysis into the repertoire of metabolically controlled transformations.

scholarly journals Syncrip/hnRNPQ is required for activity-induced Msp300/Nesprin-1 expression and new synapse formation

2019 ◽  
Author(s):  
Josh Titlow ◽  
Francesca Robertson ◽  
Aino Järvelin ◽  
David Ish-Horowicz ◽  
Carlas Smith ◽  
...  
Keyword(s):  
Synaptic Plasticity ◽  
Single Molecule ◽  
Rna Binding ◽  
Synapse Formation ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Long Distance ◽  
Key Factor ◽  
Activity Dependent

AbstractMemory and learning involve activity-driven expression of proteins and cytoskeletal reorganisation at new synapses, often requiring post-transcriptional regulation a long distance from corresponding nuclei. A key factor expressed early in synapse formation is Msp300/Nesprin-1, which organises actin filaments around the new synapse. How Msp300 expression is regulated during synaptic plasticity is not yet known. Here, we show that the local translation of msp300 is promoted during activity-dependent plasticity by the conserved RNA binding protein Syncrip/hnRNP Q, which binds to msp300 transcripts and is essential for plasticity. Single molecule imaging shows that Syncrip is associated in vivo with msp300 mRNA in ribosome-rich particles. Elevated neural activity alters the dynamics of Syncrip RNP granules at the synapse, suggesting a change in particle composition or binding that facilitates translation. These results introduce Syncrip as an important early-acting activity-dependent translational regulator of a plasticity gene that is strongly associated with human ataxias.Syncrip regulates synaptic plasticity via msp300Titlow et al. find that Syncrip (hnRNPQ RNA binding protein) acts directly on msp300 to modulate activity-dependent synaptic plasticity. In vivo biophysical experiments reveal activity-dependent changes in RNP complex sizes compatible with an increase in translation at the synapse.

scholarly journals Establishing Live-Cell Single-Molecule Localization Microscopy Imaging and Single-Particle Tracking in the Archaeon Haloferax volcanii

2020 ◽  
Vol 11 ◽  
Author(s):  
Bartosz Turkowyd ◽  
Sandra Schreiber ◽  
Julia Wörtz ◽  
Ella Shtifman Segal ◽  
Moshe Mevarech ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fluorescent Proteins ◽  
Single Particle Tracking ◽  
Haloferax Volcanii ◽  
Microscopy Imaging ◽  
Localization Microscopy ◽  
Cellular Dna ◽  
Eukaryotic Organisms ◽  
Single Molecule Localization Microscopy

In recent years, fluorescence microscopy techniques for the localization and tracking of single molecules in living cells have become well-established and are indispensable tools for the investigation of cellular biology and in vivo biochemistry of many bacterial and eukaryotic organisms. Nevertheless, these techniques are still not established for imaging archaea. Their establishment as a standard tool for the study of archaea will be a decisive milestone for the exploration of this branch of life and its unique biology. Here, we have developed a reliable protocol for the study of the archaeon Haloferax volcanii. We have generated an autofluorescence-free H. volcanii strain, evaluated several fluorescent proteins for their suitability to serve as single-molecule fluorescence markers and codon-optimized them to work under optimal H. volcanii cultivation conditions. We found that two of them, Dendra2Hfx and PAmCherry1Hfx, provide state-of-the-art single-molecule imaging. Our strategy is quantitative and allows dual-color imaging of two targets in the same field of view (FOV) as well as DNA co-staining. We present the first single-molecule localization microscopy (SMLM) images of the subcellular organization and dynamics of two crucial intracellular proteins in living H. volcanii cells, FtsZ1, which shows complex structures in the cell division ring, and RNA polymerase, which localizes around the periphery of the cellular DNA.This work should provide incentive to develop SMLM strategies for other archaeal organisms in the near future.

Sign in / Sign up

Export Citation Format

Share Document