In this thesis, we discuss three topics concerning extracellular electron transfer in the Dissimilatory Metal Reducing Bacterium (DMRB) Shewanella oneidensis MR-1. One proposed strategy to accomplish extracellular charge transfer in Shewanella involves forming a conductive pathway to electrodes by incorporating redox components on outer cell membranes and along extracellular appendages known as bacterial nanowires within biofilms. In the first part of this dissertation, to describe extracellular charge transfer in microbial redox chains, we employed a model based on incoherent hopping between sites in the chain and an interfacial treatment of electrochemical interactions with the surrounding electrodes. Based on this model, we calculated the current-voltage (I-V) characteristics and found the results to be in good agreement with I-V measurements across and along individual microbial nanowires produced by the bacterium S. oneidensis MR-1. Based on our analysis, we propose that multistep hopping in redox chains constitutes a viable strategy for extracellular charge transfer in microbial biofilms.
In the second part, we report the first in vivo observations of the formation and respiratory impact of nanowires in the model metal-reducing microbe S. oneidensis MR-1. Live fluorescence measurements, immunolabeling, and quantitative gene expression analysis point to S. oneidensis MR-1 nanowires as extensions of the outer membrane and periplasm that include the multiheme cytochromes responsible for EET, rather than pilin-based structures as previously thought. These membrane extensions are associated with outer membrane vesicles, structures ubiquitous in Gram-negative bacteria, and are consistent with bacterial nanowires that mediate long-range EET by our proposed multistep redox hopping mechanism. Redox-functionalized membrane and vesicular extensions may represent a general microbial strategy for electron transport and energy distribution.
In addition, to elucidate the membranous nature of Shewanella nanowires, we imaged these filaments using Transmission Electron Microscopy. The TEM images reported in this thesis also provide the most accurate estimates of bacterial nanowire dimensions to date. Future TEM and cryo-TEM imaging can establish the specific alignment and configuration of outer membrane cytochromes that facilitate electron transport along bacterial nanowires.
In the third part of this thesis, we focus on the molecular conductance of MtrF, the first decaheme outer membrane cytochrome with a solved crystal structure. Decaheme outer membrane cytochromes of Shewanella play a crucial role in all the suggested pathways of extracellular electron transfer. An understanding of the electron transfer properties in MtrF will therefore impact all aspects of extracellular electron transfer research. In this thesis, using purified MtrF, we form monolayers of the protein on atomically flat gold substrates and address the dry monolayer with a Scanning Tunneling Microscope (STM) tip. This technique can be used in the future to examine the conductivity of individual MtrF molecules within the monolayer in the form of I-V curves. This methodology will allow experimental comparison with recently developed simulations of MtrF conductance.
|Advisor:||El-Naggar, Mohamed Y.|
|Commitee:||Finkel, Steven E., Haas, Stephan, Haselwandter, Christoph, Kresin, Vitaly|
|School:||University of Southern California|
|School Location:||United States -- California|
|Source:||DAI-B 76/10(E), Dissertation Abstracts International|
|Keywords:||Bacterial nanowire, Extracellular electron transfer, Membrane cytochrome, Membrane extension, Microbial fuel cell, Shewanella|
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