@article {3070, title = {An Chassis for Production of Electrically Conductive Protein Nanowires.}, journal = {ACS Synth Biol}, volume = {9}, year = {2020}, month = {2020 Mar 20}, pages = {647-654}, abstract = {

pilin-based electrically conductive protein nanowires (e-PNs) are a revolutionary electronic material. They offer novel options for electronic sensing applications and have the remarkable ability to harvest electrical energy from atmospheric humidity. However, technical constraints limit mass cultivation and genetic manipulation of . Therefore, we designed a strain of to express e-PNs by introducing a plasmid that contained an inducible operon with genes for type IV pili biogenesis machinery and a synthetic gene designed to yield a peptide monomer that could be assembled into e-PNs. The e-PNs expressed in and harvested with a simple filtration method had the same diameter (3 nm) and conductance as e-PNs expressed in . These results, coupled with the robustness of for mass cultivation and the extensive toolbox for genetic manipulation, greatly expand the opportunities for large-scale fabrication of novel e-PNs.

}, keywords = {Electric Conductivity, Escherichia coli, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Graphite, Microorganisms, Genetically-Modified, Microscopy, Atomic Force, Nanowires, Operon, Protein Engineering}, issn = {2161-5063}, doi = {10.1021/acssynbio.9b00506}, author = {Ueki, Toshiyuki and Walker, David J F and Woodard, Trevor L and Nevin, Kelly P and Nonnenmann, Stephen S and Lovley, Derek R} } @article {3081, title = {Conductive Composite Materials Fabricated from Microbially Produced Protein Nanowires.}, journal = {Small}, volume = {14}, year = {2018}, month = {2018 Nov}, pages = {e1802624}, abstract = {

Protein-based electronic materials have numerous potential advantages with respect to sustainability and biocompatibility over electronic materials that are synthesized using harsh chemical processes and/or which contain toxic components. The microorganism Geobacter sulfurreducens synthesizes electrically conductive protein nanowires (e-PNs) with high aspect ratios (3 nm {\texttimes} 10-30 {\textmu}m) from renewable organic feedstocks. Here, the integration of G. Sulfurreducens e-PNs into poly(vinyl alcohol) (PVA) as a host polymer matrix is described. The resultant e-PN/PVA composites exhibit conductivities comparable to PVA-based composites containing synthetic nanowires. The relationship between e-PN density and conductivity of the resultant composites is consistent with percolation theory. These e-PNs confer conductivity to the composites even under extreme conditions, with the highest conductivities achieved from materials prepared at pH 1.5 and temperatures greater than 100 {\textdegree}C. These results demonstrate that e-PNs represent viable and sustainable nanowire compositions for the fabrication of electrically conductive composite materials.

}, keywords = {Geobacter, Nanocomposites, Nanowires, Polymers}, issn = {1613-6829}, doi = {10.1002/smll.201802624}, author = {Sun, Yun-Lu and Tang, Hai-Yan and Ribbe, Alexander and Duzhko, Volodimyr and Woodard, Trevor L and Ward, Joy E and Bai, Ying and Nevin, Kelly P and Nonnenmann, Stephen S and Russell, Thomas and Emrick, Todd and Lovley, Derek R} } @article {3083, title = {Construction of a Strain With Exceptional Growth on Cathodes.}, journal = {Front Microbiol}, volume = {9}, year = {2018}, month = {2018}, pages = {1512}, abstract = {

Insoluble extracellular electron donors are important sources of energy for anaerobic respiration in biogeochemical cycling and in diverse practical applications. The previous lack of a genetically tractable model microorganism that could be grown to high densities under anaerobic conditions in pure culture with an insoluble extracellular electron donor has stymied efforts to better understand this form of respiration. We report here on the design of a strain of , designated strain ACL, which grows as thick (ca. 35 μm) confluent biofilms on graphite cathodes poised at -500 mV ( Ag/AgCl) with fumarate as the electron acceptor. Sustained maximum current consumption rates were >0.8 A/m, which is >10-fold higher than the current consumption of the wild-type strain. The improved function on the cathode was achieved by introducing genes for an ATP-dependent citrate lyase, completing the complement of enzymes needed for a reverse TCA cycle for the synthesis of biosynthetic precursors from carbon dioxide. Strain ACL provides an important model organism for elucidating the mechanisms for effective anaerobic growth with an insoluble extracellular electron donor and may offer unique possibilities as a chassis for the introduction of synthetic metabolic pathways for the production of commodities with electrons derived from electrodes.

}, issn = {1664-302X}, doi = {10.3389/fmicb.2018.01512}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Aklujkar, Muktak A and Holmes, Dawn E and Lovley, Derek R} } @article {3117, title = {Carbon cloth stimulates direct interspecies electron transfer in syntrophic co-cultures.}, journal = {Bioresour Technol}, volume = {173}, year = {2014}, month = {2014 Dec}, pages = {82-86}, abstract = {

This study investigated the possibility that the electrical conductivity of carbon cloth accelerates direct interspecies electron transfer (DIET) in co-cultures. Carbon cloth accelerated metabolism of DIET co-cultures (Geobacter metallireducens-Geobacter sulfurreducens and G.metallireducens-Methanosarcina barkeri) but did not promote metabolism of co-cultures performing interspecies H2 transfer (Desulfovibrio vulgaris-G.sulfurreducens). On the other hand, DIET co-cultures were not stimulated by poorly conductive cotton cloth. Mutant strains lacking electrically conductive pili, or pili-associated cytochromes participated in DIET only in the presence of carbon cloth. In co-cultures promoted by carbon cloth, cells were primarily associated with the cloth although the syntrophic partners were too far apart for cell-to-cell biological electrical connections to be feasible. Carbon cloth seemingly mediated interspecies electron transfer between the distant syntrophic partners. These results suggest that the ability of carbon cloth to accelerate DIET should be considered in anaerobic digester designs that incorporate carbon cloth.

}, keywords = {Carbon, Cell Communication, Coculture Techniques, Electric Conductivity, Electron Transport, Geobacter, Materials Testing, Membranes, Artificial, Microbial Consortia, Oxidation-Reduction, Symbiosis}, issn = {1873-2976}, doi = {10.1016/j.biortech.2014.09.009}, author = {Chen, Shanshan and Rotaru, Amelia-Elena and Liu, Fanghua and Philips, Jo and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {3115, title = {Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii.}, journal = {mBio}, volume = {5}, year = {2014}, month = {2014 Oct 21}, pages = {e01636-14}, abstract = {

Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70\% of the carbon and electron flow was diverted to the production of butyrate with H2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. Importance: The development of a microbial chassis for efficient conversion of carbon dioxide directly to desired organic products would greatly advance the environmentally sustainable production of biofuels and other commodities. Clostridium ljungdahlii is an effective catalyst for microbial electrosynthesis, a technology in which electricity generated with renewable technologies, such as solar or wind, powers the conversion of carbon dioxide and water to organic products. Other electron donors for C. ljungdahlii include carbon monoxide, which can be derived from industrial waste gases or the conversion of recalcitrant biomass to syngas, as well as hydrogen, another syngas component. The finding that carbon and electron flow in C. ljungdahlii can be diverted from the production of acetate to butyrate synthesis is an important step toward the goal of renewable commodity production from carbon dioxide with this organism.

}, keywords = {Acetyl Coenzyme A, Butyrates, Carbon Dioxide, Clostridium, Metabolic Engineering, Metabolic Flux Analysis, Metabolic Networks and Pathways, Recombinant Proteins}, issn = {2150-7511}, doi = {10.1128/mBio.01636-14}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Lovley, Derek R} } @article {700, title = {Characterizing regulation of metabolism in Geobacter sulfurreducens through genome-wide expression data and sequence analysis.}, journal = {OMICS}, volume = {12}, year = {2008}, month = {2008 Mar}, pages = {33-59}, abstract = {Geobacteraceae are a family of metal reducing bacteria with important applications in bioremediation and electricity generation. G. sulfurreducens is a representative of Geobacteraceae that has been extensively studied with the goal of extending the understanding of this family of organisms for optimizing their practical applications. Here, we have analyzed gene expression data from 10 experiments involving environmental and genetic perturbations and have identified putative transcription factor binding sites (TFBS) involved in regulating key aspects of metabolism. Specifically, we considered data from both a subset of 10 microarray experiments (7 of 10) and all 10 experiments. The expression data from these two sets were independently clustered, and the upstream regions of genes and operons from the clusters in both sets were used to identify TFBS using the AlignACE program. This analysis resulted in the identification of motifs upstream of several genes involved in central metabolism, sulfate assimilation, and energy metabolism, as well as genes potentially encoding acetate permease. Further, similar TFBS were identified from the analysis of both sets, suggesting that these TFBS are significant in the regulation of metabolism in G. sulfurreducens. In addition, we have utilized microarray data to derive condition specific constraints on the capacity of key enzymes in central metabolism. We have incorporated these constraints into the metabolic model of G. sulfurreducens and simulated Fe(II)-limited growth. The resulting prediction was consistent with data, suggesting that regulatory constraints are important for simulating growth phenotypes in nonoptimal environments.}, keywords = {Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Models, Genetic, Oligonucleotide Array Sequence Analysis, Sequence Analysis, DNA, Transcription, Genetic}, issn = {1536-2310}, doi = {10.1089/omi.2007.0043}, author = {Mahadevan, Radhakrishnan and Yan, Bin and Postier, Brad and Nevin, Kelly P and Woodard, Trevor L and O{\textquoteright}Neil, Regina and Coppi, Maddalena V and Meth{\'e}, Barbara A and Krushkal, Julia} }