@article {418, title = {Tunable metallic-like conductivity in microbial nanowire networks.}, journal = {Nat Nanotechnol}, volume = {6}, year = {2011}, month = {2011 Sep}, pages = {573-9}, abstract = {Electronic nanostructures made from natural amino acids are attractive because of their relatively low cost, facile processing and absence of toxicity. However, most materials derived from natural amino acids are electronically insulating. Here, we report metallic-like conductivity in films of the bacterium Geobacter sulfurreducens and also in pilin nanofilaments (known as microbial nanowires) extracted from these bacteria. These materials have electronic conductivities of \~{}5~mS~cm(-1), which are comparable to those of synthetic metallic nanostructures. They can also conduct over distances on the centimetre scale, which is thousands of times the size of a bacterium. Moreover, the conductivity of the biofilm can be tuned by regulating gene expression, and also by varying the gate voltage in a transistor configuration. The conductivity of the nanofilaments has a temperature dependence similar to that of a disordered metal, and the conductivity could be increased by processing.}, keywords = {Electric Conductivity, Geobacter, Nanowires, Transistors, Electronic}, issn = {1748-3395}, doi = {10.1038/nnano.2011.119}, author = {Malvankar, Nikhil S and Vargas, Madeline and Nevin, Kelly P and Franks, Ashley E and Leang, Ching and Kim, Byoung-Chan and Inoue, Kengo and Mester, T{\"u}nde and Covalla, Sean F and Johnson, Jessica P and Rotello, Vincent M and Tuominen, Mark T and Lovley, Derek R} } @article {445, title = {Purification and characterization of OmcZ, an outer-surface, octaheme c-type cytochrome essential for optimal current production by Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {76}, year = {2010}, month = {2010 Jun}, pages = {3999-4007}, abstract = {Previous studies have demonstrated that Geobacter sulfurreducens requires the c-type cytochrome OmcZ, which is present in large (OmcZ(L); 50-kDa) and small (OmcZ(S); 30-kDa) forms, for optimal current production in microbial fuel cells. This protein was further characterized to aid in understanding its role in current production. Subcellular-localization studies suggested that OmcZ(S) was the predominant extracellular form of OmcZ. N- and C-terminal amino acid sequence analysis of purified OmcZ(S) and molecular weight measurements indicated that OmcZ(S) is a cleaved product of OmcZ(L) retaining all 8 hemes, including 1 heme with the unusual c-type heme-binding motif CX(14)CH. The purified OmcZ(S) was remarkably thermally stable (thermal-denaturing temperature, 94.2 degrees C). Redox titration analysis revealed that the midpoint reduction potential of OmcZ(S) is approximately -220 mV (versus the standard hydrogen electrode [SHE]) with nonequivalent heme groups that cover a large reduction potential range (-420 to -60 mV). OmcZ(S) transferred electrons in vitro to a diversity of potential extracellular electron acceptors, such as Fe(III) citrate, U(VI), Cr(VI), Au(III), Mn(IV) oxide, and the humic substance analogue anthraquinone-2,6-disulfonate, but not Fe(III) oxide. The biochemical properties and extracellular localization of OmcZ suggest that it is well suited for promoting electron transfer in current-producing biofilms of G. sulfurreducens.}, keywords = {Binding Sites, Bioelectric Energy Sources, Cytochromes c, Electricity, Electron Transport, Geobacter, Heme, Hot Temperature, Molecular Sequence Data, Molecular Weight, Oxidation-Reduction, Protein Binding, Protein Stability, Sequence Alignment, Sequence Analysis, Protein}, issn = {1098-5336}, doi = {10.1128/AEM.00027-10}, author = {Inoue, Kengo and Qian, Xinlei and Morgado, Leonor and Kim, Byoung-Chan and Mester, T{\"u}nde and Izallalen, Mounir and Salgueiro, Carlos A and Lovley, Derek R} } @article {447, title = {Role of Geobacter sulfurreducens outer surface c-type cytochromes in reduction of soil humic acid and anthraquinone-2,6-disulfonate.}, journal = {Appl Environ Microbiol}, volume = {76}, year = {2010}, month = {2010 Apr}, pages = {2371-5}, abstract = {Deleting individual genes for outer surface c-type cytochromes in Geobacter sulfurreducens partially inhibited the reduction of humic substances and anthraquinone-2,6,-disulfonate. Complete inhibition was obtained only when five of these genes were simultaneously deleted, suggesting that diverse outer surface cytochromes can contribute to the reduction of humic substances and other extracellular quinones.}, keywords = {Anthraquinones, Bacterial Proteins, Cytochromes, Gene Deletion, Genes, Bacterial, Geobacter, Humic Substances, Oxidation-Reduction, Soil, Soil Microbiology}, issn = {1098-5336}, doi = {10.1128/AEM.02250-09}, author = {Voordeckers, James W and Kim, Byoung-Chan and Izallalen, Mounir and Lovley, Derek R} } @article {462, title = {Anode biofilm transcriptomics reveals outer surface components essential for high density current production in Geobacter sulfurreducens fuel cells.}, journal = {PLoS One}, volume = {4}, year = {2009}, month = {2009}, pages = {e5628}, abstract = {The mechanisms by which Geobacter sulfurreducens transfers electrons through relatively thick (>50 microm) biofilms to electrodes acting as a sole electron acceptor were investigated. Biofilms of Geobacter sulfurreducens were grown either in flow-through systems with graphite anodes as the electron acceptor or on the same graphite surface, but with fumarate as the sole electron acceptor. Fumarate-grown biofilms were not immediately capable of significant current production, suggesting substantial physiological differences from current-producing biofilms. Microarray analysis revealed 13 genes in current-harvesting biofilms that had significantly higher transcript levels. The greatest increases were for pilA, the gene immediately downstream of pilA, and the genes for two outer c-type membrane cytochromes, OmcB and OmcZ. Down-regulated genes included the genes for the outer-membrane c-type cytochromes, OmcS and OmcT. Results of quantitative RT-PCR of gene transcript levels during biofilm growth were consistent with microarray results. OmcZ and the outer-surface c-type cytochrome, OmcE, were more abundant and OmcS was less abundant in current-harvesting cells. Strains in which pilA, the gene immediately downstream from pilA, omcB, omcS, omcE, or omcZ was deleted demonstrated that only deletion of pilA or omcZ severely inhibited current production and biofilm formation in current-harvesting mode. In contrast, these gene deletions had no impact on biofilm formation on graphite surfaces when fumarate served as the electron acceptor. These results suggest that biofilms grown harvesting current are specifically poised for electron transfer to electrodes and that, in addition to pili, OmcZ is a key component in electron transfer through differentiated G. sulfurreducens biofilms to electrodes.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Bioelectric Energy Sources, Biofilms, Cytochromes, Electrodes, Electron Transport, Fumarates, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genetic Complementation Test, Geobacter, Microscopy, Confocal, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, RNA, Messenger, Up-Regulation}, issn = {1932-6203}, doi = {10.1371/journal.pone.0005628}, author = {Nevin, Kelly P and Kim, Byoung-Chan and Glaven, Richard H and Johnson, Jessica P and Woodard, Trevor L and Meth{\'e}, Barbara A and Didonato, Raymond J and Covalla, Sean F and Franks, Ashley E and Liu, Anna and Lovley, Derek R} } @article {490, title = {PilR, a transcriptional regulator for pilin and other genes required for Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Mol Microbiol Biotechnol}, volume = {16}, year = {2009}, month = {2009}, pages = {146-58}, abstract = {Growth using Fe(III) as a terminal electron acceptor is a critical physiological process in Geobacter sulfurreducens. However, the mechanisms of electron transfer during Fe(III) reduction are only now being understood. It has been demonstrated that the pili in G. sulfurreducens function as microbial nanowires conducting electrons onto Fe(III) oxides. A number of c-type cytochromes have also been shown to play important roles in Fe(III) reduction. However, the regulatory networks controlling the expression of the genes involved in such processes are not well known. Here we report that the expression of pilA, which encodes the pilistructural protein, is directly regulated by a two-component regulatory system in which PilR functions as an RpoN-dependent enhancer binding protein. Surprisingly, a deletion of the pilR gene affected not only insoluble Fe(III) reduction, which requires pili, but also soluble Fe(III) reduction, which, in contrast, does not require pili. Gene expression profiling using whole-genome DNA microarray and quantitative RT-PCR analyses obtained with a PilR-deficient mutant revealed that the expression of pilA and other pilin-related genes are downregulated, while many c-type cytochromes involved in Fe(III) reduction were differentially regulated. This is the first instance of an enhancer binding protein implicated in regulating genes involved in Fe(III) respiratory functions.}, keywords = {Bacterial Proteins, Ferric Compounds, Fimbriae Proteins, Gene Expression Regulation, Bacterial, Genes, Regulator, Geobacter, Oxidation-Reduction, Transcription, Genetic}, issn = {1660-2412}, doi = {10.1159/000115849}, author = {Ju{\'a}rez, Katy and Kim, Byoung-Chan and Nevin, Kelly and Olvera, Leticia and Reguera, Gemma and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {460, title = {Selection of a variant of Geobacter sulfurreducens with enhanced capacity for current production in microbial fuel cells.}, journal = {Biosens Bioelectron}, volume = {24}, year = {2009}, month = {2009 Aug 15}, pages = {3498-503}, abstract = {Geobacter sulfurreducens produces current densities in microbial fuel cells that are among the highest known for pure cultures. The possibility of adapting this organism to produce even higher current densities was evaluated. A system in which a graphite anode was poised at -400 mV (versus Ag/AgCl) was inoculated with the wild-type strain of G. sulfurreducens, strain DL-1. An isolate, designated strain KN400, was recovered from the biofilm after 5 months of growth on the electrode. KN400 was much more effective in current production than strain DL-1. This was apparent with anodes poised at -400 mV, as well as in systems run in true fuel cell mode. KN400 had current (7.6A/m(2)) and power (3.9 W/m(2)) densities that respectively were substantially higher than those of DL1 (1.4A/m(2) and 0.5 W/m(2)). On a per cell basis KN400 was more effective in current production than DL1, requiring thinner biofilms to make equivalent current. The enhanced capacity for current production in KN400 was associated with a greater abundance of electrically conductive microbial nanowires than DL1 and lower internal resistance (0.015 versus 0.130 Omega/m(2)) and mass transfer limitation in KN400 fuel cells. KN400 produced flagella, whereas DL1 does not. Surprisingly, KN400 had much less outer-surface c-type cytochromes than DL1. KN400 also had a greater propensity to form biofilms on glass or graphite than DL1, even when growing with the soluble electron acceptor, fumarate. These results demonstrate that it is possible to enhance the ability of microorganisms to electrochemically interact with electrodes with the appropriate selective pressure and that improved current production is associated with clear differences in the properties of the outer surface of the cell that may provide insights into the mechanisms for microbe-electrode interactions.}, keywords = {Bioelectric Energy Sources, Electrochemistry, Equipment Design, Equipment Failure Analysis, Geobacter, Species Specificity}, issn = {1873-4235}, doi = {10.1016/j.bios.2009.05.004}, author = {Yi, Hana and Nevin, Kelly P and Kim, Byoung-Chan and Franks, Ashely E and Klimes, Anna and Tender, Leonard M and Lovley, Derek R} } @article {480, title = {Insights into genes involved in electricity generation in Geobacter sulfurreducens via whole genome microarray analysis of the OmcF-deficient mutant.}, journal = {Bioelectrochemistry}, volume = {73}, year = {2008}, month = {2008 Jun}, pages = {70-5}, abstract = {Geobacter sulfurreducens effectively produces electricity in microbial fuel cells by oxidizing acetate with an electrode serving as the sole electron acceptor. Deletion of the gene encoding OmcF, a monoheme outer membrane c-type cytochrome, substantially decreased current production. Previous studies demonstrated that inhibition of Fe(III) reduction in the OmcF-deficient mutant could be attributed to poor transcription of the gene for OmcB, an outer membrane c-type cytochrome that is required for Fe(III) reduction. However, a mutant in which omcB was deleted produced electricity as well as wild type. Microarray analysis of the OmcF-deficient mutant versus the wild type revealed that many of the genes with the greatest decreases in transcript levels were genes whose expression was previously reported to be upregulated in cells grown with an electrode as the sole electron acceptor. These included genes with putative functions related to metal efflux and/or type I secretion and two hypothetical proteins. The outer membrane cytochromes, OmcS and OmcE, which previous studies have demonstrated are required for optimal current generation, were not detected on the outer surface of the OmcF-deficient mutant even though the omcS and omcE genes were still transcribed, suggesting that the putative secretion system could be involved in the export of outer membrane proteins necessary for electron transfer to the fuel cell anode. These results suggest that the requirement for OmcF for optimal current production is not because OmcF is directly involved in extracellular electron transfer but because OmcF is required for the appropriate transcription of other genes either directly or indirectly involved in electricity production.}, keywords = {Bacterial Outer Membrane Proteins, Cytochromes c, Down-Regulation, Electricity, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Mutation, Oligonucleotide Array Sequence Analysis, Transcription, Genetic}, issn = {1567-5394}, doi = {10.1016/j.bioelechem.2008.04.023}, author = {Kim, Byoung-Chan and Postier, Bradley L and Didonato, Raymond J and Chaudhuri, Swades K and Nevin, Kelly P and Lovley, Derek R} } @article {478, title = {Investigation of direct vs. indirect involvement of the c-type cytochrome MacA in Fe(III) reduction by Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {286}, year = {2008}, month = {2008 Sep}, pages = {39-44}, abstract = {The electron transfer pathway to Fe(III) reduction in Geobacter sulfurreducens has been hypothesized to consist of a series of c-type cytochromes. Previous genetic studies suggested that the inner membrane-associated, c-type cytochrome, MacA, was a component of the electron transfer chain leading to Fe(III) reduction in the dissimilatory Fe(III)-reducer, G. sulfurreducens. However, investigation of the expression of OmcB, an outer-membrane c-type cytochrome demonstrated previously to be critical for optimal Fe(III) reduction, revealed that both omcB transcript and protein levels were dramatically reduced in the MacA-deficient mutant. Expression of the omcB gene in trans enabled the MacA-deficient mutant to reduce Fe(III) at a rate that was proportional to the level of omcB expression. These results suggest that MacA is not directly involved in electron transfer to Fe(III) and further confirm the importance of OmcB in Fe(III) reduction by G. sulfurreducens.}, keywords = {ATP-Binding Cassette Transporters, Bacterial Proteins, Cytochromes c, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Oxidation-Reduction}, issn = {0378-1097}, doi = {10.1111/j.1574-6968.2008.01252.x}, author = {Kim, Byoung-Chan and Lovley, Derek R} } @article {505, title = {Importance of c-Type cytochromes for U(VI) reduction by Geobacter sulfurreducens.}, journal = {BMC Microbiol}, volume = {7}, year = {2007}, month = {2007}, pages = {16}, abstract = {BACKGROUND: In order to study the mechanism of U(VI) reduction, the effect of deleting c-type cytochrome genes on the capacity of Geobacter sulfurreducens to reduce U(VI) with acetate serving as the electron donor was investigated. RESULTS: The ability of several c-type cytochrome deficient mutants to reduce U(VI) was lower than that of the wild type strain. Elimination of two confirmed outer membrane cytochromes and two putative outer membrane cytochromes significantly decreased (ca. 50-60\%) the ability of G. sulfurreducens to reduce U(VI). Involvement in U(VI) reduction did not appear to be a general property of outer membrane cytochromes, as elimination of two other confirmed outer membrane cytochromes, OmcB and OmcC, had very little impact on U(VI) reduction. Among the periplasmic cytochromes, only MacA, proposed to transfer electrons from the inner membrane to the periplasm, appeared to play a significant role in U(VI) reduction. A subpopulation of both wild type and U(VI) reduction-impaired cells, 24-30\%, accumulated amorphous uranium in the periplasm. Comparison of uranium-accumulating cells demonstrated a similar amount of periplasmic uranium accumulation in U(VI) reduction-impaired and wild type G. sulfurreducens. Assessment of the ability of the various suspensions to reduce Fe(III) revealed no correlation between the impact of cytochrome deletion on U(VI) reduction and reduction of Fe(III) hydroxide and chelated Fe(III). CONCLUSION: This study indicates that c-type cytochromes are involved in U(VI) reduction by Geobacter sulfurreducens. The data provide new evidence for extracellular uranium reduction by G. sulfurreducens but do not rule out the possibility of periplasmic uranium reduction. Occurrence of U(VI) reduction at the cell surface is supported by the significant impact of elimination of outer membrane cytochromes on U(VI) reduction and the lack of correlation between periplasmic uranium accumulation and the capacity for uranium reduction. Periplasmic uranium accumulation may reflect the ability of uranium to penetrate the outer membrane rather than the occurrence of enzymatic U(VI) reduction. Elimination of cytochromes rarely had a similar impact on both Fe(III) and U(VI) reduction, suggesting that there are differences in the routes of electron transfer to U(VI) and Fe(III). Further studies are required to clarify the pathways leading to U(VI) reduction in G. sulfurreducens.}, keywords = {Biodegradation, Environmental, Cytochrome c Group, Ferric Compounds, Geobacter, Microscopy, Electron, Transmission, Mutation, Oxidation-Reduction, Periplasm, Uranium}, issn = {1471-2180}, doi = {10.1186/1471-2180-7-16}, author = {Shelobolina, Evgenya S and Coppi, Maddalena V and Korenevsky, Anton A and DiDonato, Laurie N and Sullivan, Sara A and Konishi, Hiromi and Xu, Huifang and Leang, Ching and Butler, Jessica E and Kim, Byoung-Chan and Lovley, Derek R} } @article {521, title = {Two putative c-type multiheme cytochromes required for the expression of OmcB, an outer membrane protein essential for optimal Fe(III) reduction in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Apr}, pages = {3138-42}, abstract = {Deletion of two homologous Geobacter sulfurreducens c-type cytochrome genes, omcG and omcH, decreased the rate of Fe(III) reduction and decreased the level of an outer membrane cytochrome critical for Fe(III) reduction, OmcB, without affecting its transcription. Expression of either gene restored Fe(III) reduction and OmcB expression, suggesting functional similarity.}, keywords = {Bacterial Outer Membrane Proteins, Bacterial Proteins, Blotting, Northern, Blotting, Western, Cytochromes c, Ferric Compounds, Gene Deletion, Gene Expression, Genes, Bacterial, Geobacter, Oxidation-Reduction, RNA, Bacterial, RNA, Messenger}, issn = {0021-9193}, doi = {10.1128/JB.188.8.3138-3142.2006}, author = {Kim, Byoung-Chan and Qian, Xinlei and Leang, Ching and Coppi, Maddalena V and Lovley, Derek R} } @article {536, title = {OmcF, a putative c-Type monoheme outer membrane cytochrome required for the expression of other outer membrane cytochromes in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {187}, year = {2005}, month = {2005 Jul}, pages = {4505-13}, abstract = {Outer membrane cytochromes are often proposed as likely agents for electron transfer to extracellular electron acceptors, such as Fe(III). The omcF gene in the dissimilatory Fe(III)-reducing microorganism Geobacter sulfurreducens is predicted to code for a small outer membrane monoheme c-type cytochrome. An OmcF-deficient strain was constructed, and its ability to reduce and grow on Fe(III) citrate was found to be impaired. Following a prolonged lag phase (150 h), the OmcF-deficient strain developed the ability to grow in Fe(III) citrate medium with doubling times and yields that were ca. 145\% and 70\% of those of the wild type, respectively. Comparison of the c-type cytochrome contents of outer membrane-enriched fractions prepared from wild-type and OmcF-deficient cultures confirmed the outer membrane association of OmcF and revealed multiple changes in the cytochrome content of the OmcF-deficient strain. These changes included loss of expression of two previously characterized outer membrane cytochromes, OmcB and OmcC, and overexpression of a third previously characterized outer membrane cytochrome, OmcS, during growth on Fe(III) citrate. The omcB and omcC transcripts could not be detected in the OmcF-deficient mutant by either reverse transcriptase PCR or Northern blot analyses. Expression of the omcF gene in trans restored both the capacity of the OmcF-deficient mutant to reduce Fe(III) and wild-type levels of omcB and omcC mRNA and protein. Thus, elimination of OmcF may impair Fe(III) reduction by influencing expression of OmcB, which has previously been demonstrated to play a critical role in Fe(III) reduction.}, keywords = {Amino Acid Sequence, Bacterial Outer Membrane Proteins, Cytochromes c, Ferric Compounds, Gene Deletion, Gene Expression Regulation, Bacterial, Geobacter, Molecular Sequence Data, Oxidation-Reduction, Sequence Alignment}, issn = {0021-9193}, doi = {10.1128/JB.187.13.4505-4513.2005}, author = {Kim, Byoung-Chan and Leang, Ching and Ding, Yan-Huai R and Glaven, Richard H and Coppi, Maddalena V and Lovley, Derek R} }