@article {1559, title = {Identification of 4-Hydroxycumyl Alcohol As the Major MnO2-Mediated Bisphenol A Transformation Product and Evaluation of Its Environmental Fate.}, journal = {Environ Sci Technol}, volume = {49}, year = {2015}, month = {2015 May 19}, pages = {6214-21}, abstract = {

Bisphenol A (BPA), an environmental contaminant with weak estrogenic activity, resists microbial degradation under anoxic conditions but is susceptible to abiotic transformation by manganese dioxide (MnO2). BPA degradation followed pseudo-first-order kinetics with a rate constant of 0.96 ({\textpm}0.03) min(-1) in the presence of 2 mM MnO2 (0.017\% w/w) at pH 7.2. 4-hydroxycumyl alcohol (HCA) was the major transformation product, and, on a molar basis, up to 64\% of the initial amount of BPA was recovered as HCA. MnO2 was also reactive toward HCA, albeit at 5-fold lower rates, and CO2 evolution (i.e., mineralization) occurred. In microcosms established with freshwater sediment, HCA was rapidly biodegraded under oxic, but not anoxic conditions. With a measured octanol-water partition coefficient (Log K(ow)) of 0.76 and an aqueous solubility of 2.65 g L(-1), HCA is more mobile in saturated media than BPA (Log K(ow) = 2.76; aqueous solubility = 0.31 g L(-1)), and therefore more likely to encounter oxic zones and undergo aerobic biodegradation. These findings corroborate that BPA is not inert under anoxic conditions and suggest that MnO2-mediated coupled abiotic-biotic processes may be relevant for controlling the fate and longevity of BPA in sediments and aquifers.

}, issn = {1520-5851}, doi = {10.1021/acs.est.5b00372}, author = {Im, Jeongdae and Prevatte, Carson W and Campagna, Shawn R and L{\"o}ffler, Frank E} } @article {1558, title = {Response to Comment on "Environmental Fate of the Next Generation Refrigerant 2,3,3,3-Tetrafluoropropene (HFO-1234yf)".}, journal = {Environ Sci Technol}, volume = {49}, year = {2015}, month = {2015 Jul 7}, pages = {8265-6}, issn = {1520-5851}, doi = {10.1021/acs.est.5b01970}, author = {Im, Jeongdae and Walshe-Langford, Gillian E and Moon, Ji-Won and L{\"o}ffler, Frank E} } @article {1438, title = {Variation in diazotrophic community structure in forest soils reflects land use history.}, journal = {Soil Biology and Biochemistry}, volume = {80}, year = {2015}, month = {2014 Sep}, pages = {1-8}, doi = {10.1016/j.soilbio.2014.09.017}, url = {http://www.sciencedirect.com/science/article/pii/S0038071714003253}, author = {N{\"u}sslein, K and Izquierdo, J} } @article {1561, title = {4-methylphenol produced in freshwater sediment microcosms is not a bisphenol A metabolite.}, journal = {Chemosphere}, volume = {117}, year = {2014}, month = {2014 Dec}, pages = {521-6}, abstract = {

4-Methylphenol (4-MP), a putative bisphenol A (BPA) degradation intermediate, was detected at concentrations reaching 2.1 mg L(-1) in anoxic microcosms containing 10 mg L(-1) BPA and 5 g of freshwater sediment material collected from four geographically distinct locations and amended with nitrate, nitrite, ferric iron, or bicarbonate as electron acceptors. 4-MP accumulation was transient, and 4-MP degradation was observed under all redox conditions tested. 4-MP was not detected in microcosms not amended with BPA. Unexpectedly, incubations with (13)C-labeled BPA failed to produce (13)C-labeled 4-MP suggesting that 4-MP was not derived from BPA. The detection of 4-MP in live microcosms amended with lactate, but not containing BPA corroborated that BPA was not the source of 4-MP. These findings demonstrate that the transient formation of 4-MP as a possible BPA degradation intermediate must be interpreted cautiously, as microbial activity in streambed microcosms may generate 4-MP from sediment-associated organic material.

}, keywords = {Anaerobiosis, Benzhydryl Compounds, Biodegradation, Environmental, Chromatography, High Pressure Liquid, Cresols, Environmental Monitoring, Fresh Water, Gas Chromatography-Mass Spectrometry, Geologic Sediments, Phenols, Water Pollutants, Chemical}, issn = {1879-1298}, doi = {10.1016/j.chemosphere.2014.09.008}, author = {Im, Jeongdae and Prevatte, Carson W and Lee, Hong Geun and Campagna, Shawn R and L{\"o}ffler, Frank E} } @article {1560, title = {Environmental fate of the next generation refrigerant 2,3,3,3-tetrafluoropropene (HFO-1234yf).}, journal = {Environ Sci Technol}, volume = {48}, year = {2014}, month = {2014 Nov 18}, pages = {13181-7}, abstract = {

The hydrofluoroolefin 2,3,3,3-tetrafluoropropene (HFO-1234yf) has been introduced to replace 1,1,1,2-tetrafluoroethane (HFC-134a) as refrigerant in mobile, including vehicle, air conditioning systems because of its lower global warming potential. HFO-1234yf is volatile at ambient temperatures; however, high production volumes and widespread handling are expected to release this fluorocarbon into terrestrial and aquatic environments, including groundwater. Laboratory experiments explored HFO-1234yf degradation by (i) microbial processes under oxic and anoxic conditions, (ii) abiotic processes mediated by reactive mineral phases and zerovalent iron (Fe(0), ZVI), and (iii) cobalamin-catalyzed biomimetic transformation. These investigations demonstrated that HFO-1234yf was recalcitrant to microbial (co)metabolism and no transformation was observed in incubations with ZVI, makinawite (FeS), sulfate green rust (GR(SO4)), magnetite (Fe(3)O(4)), and manganese oxide (MnO2). Sequential reductive defluorination of HFO-1234yf to 3,3,3-trifluoropropene and 3,3-dichloropropene with concomitant stoichiometric release of fluoride occurred in incubations with reduced cobalamins (e.g., vitamin B12) indicating that biomolecules can transform HFO-1234yf at circumneutral pH and at ambient temperature. Taken together, these findings suggest that HFO-1234yf recalcitrance in aquifers should be expected; however, HFO-1234yf is not inert and a biomolecule may mediate reductive transformation in low redox environments, albeit at low rates.

}, issn = {1520-5851}, doi = {10.1021/es5032147}, author = {Im, Jeongdae and Walshe-Langford, Gillian E and Moon, Ji-Won and L{\"o}ffler, Frank E} } @article {1562, title = {Regulation of ethanol-related behavior and ethanol metabolism by the Corazonin neurons and Corazonin receptor in Drosophila melanogaster.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e87062}, abstract = {

Impaired ethanol metabolism can lead to various alcohol-related health problems. Key enzymes in ethanol metabolism are alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH); however, neuroendocrine pathways that regulate the activities of these enzymes are largely unexplored. Here we identified a neuroendocrine system involving Corazonin (Crz) neuropeptide and its receptor (CrzR) as important physiological regulators of ethanol metabolism in Drosophila. Crz-cell deficient (Crz-CD) flies displayed significantly delayed recovery from ethanol-induced sedation that we refer to as hangover-like phenotype. Newly generated mutant lacking Crz Receptor (CrzR(01) ) and CrzR-knockdown flies showed even more severe hangover-like phenotype, which is causally associated with fast accumulation of acetaldehyde in the CrzR(01) mutant following ethanol exposure. Higher levels of acetaldehyde are likely due to 30\% reduced ALDH activity in the mutants. Moreover, increased ADH activity was found in the CrzR(01) mutant, but not in the Crz-CD flies. Quantitative RT-PCR revealed transcriptional upregulation of Adh gene in the CrzR(01) . Transgenic inhibition of cyclic AMP-dependent protein kinase (PKA) also results in significantly increased ADH activity and Adh mRNA levels, indicating PKA-dependent transcriptional regulation of Adh by CrzR. Furthermore, inhibition of PKA or cAMP response element binding protein (CREB) in CrzR cells leads to comparable hangover-like phenotype to the CrzR(01) mutant. These findings suggest that CrzR-associated signaling pathway is critical for ethanol detoxification via Crz-dependent regulation of ALDH activity and Crz-independent transcriptional regulation of ADH. Our study provides new insights into the neuroendocrine-associated ethanol-related behavior and metabolism.

}, keywords = {Acetaldehyde, Alcohol Dehydrogenase, Aldehyde Dehydrogenase, Alleles, Animals, Cyclic AMP-Dependent Protein Kinases, Drosophila melanogaster, Drosophila Proteins, Ethanol, Genes, Reporter, Male, Mutation, Neurons, Neuropeptides, Phenotype, Receptors, Neuropeptide, RNA, Messenger, Transcription, Genetic}, issn = {1932-6203}, doi = {10.1371/journal.pone.0087062}, author = {Sha, Kai and Choi, Seung-Hoon and Im, Jeongdae and Lee, Gyunghee G and Loeffler, Frank and Park, Jae H} } @article {1415, title = {Evidence supporting dissimilatory and assimilatory lignin degradation in Enterobacter lignolyticus SCF1.}, journal = {Front Microbiol}, volume = {4}, year = {2013}, month = {2013}, pages = {280}, abstract = {

Lignocellulosic biofuels are promising as sustainable alternative fuels, but lignin inhibits access of enzymes to cellulose, and by-products of lignin degradation can be toxic to cells. The fast growth, high efficiency and specificity of enzymes employed in the anaerobic litter deconstruction carried out by tropical soil bacteria make these organisms useful templates for improving biofuel production. The facultative anaerobe Enterobacter lignolyticus SCF1 was initially cultivated from Cloud Forest soils in the Luquillo Experimental Forest in Puerto Rico, based on anaerobic growth on lignin as sole carbon source. The source of the isolate was tropical forest soils that decompose litter rapidly with low and fluctuating redox potentials, where bacteria using oxygen-independent enzymes likely play an important role in decomposition. We have used transcriptomics and proteomics to examine the observed increased growth of SCF1 grown on media amended with lignin compared to unamended growth. Proteomics suggested accelerated xylose uptake and metabolism under lignin-amended growth, with up-regulation of proteins involved in lignin degradation via the 4-hydroxyphenylacetate degradation pathway, catalase/peroxidase enzymes, and the glutathione biosynthesis and glutathione S-transferase (GST) proteins. We also observed increased production of NADH-quinone oxidoreductase, other electron transport chain proteins, and ATP synthase and ATP-binding cassette (ABC) transporters. This suggested the use of lignin as terminal electron acceptor. We detected significant lignin degradation over time by absorbance, and also used metabolomics to demonstrate moderately significant decreased xylose concentrations as well as increased metabolic products acetate and formate in stationary phase in lignin-amended compared to unamended growth conditions. Our data show the advantages of a multi-omics approach toward providing insights as to how lignin may be used in nature by microorganisms coping with poor carbon availability.

}, issn = {1664-302X}, doi = {10.3389/fmicb.2013.00280}, author = {Deangelis, Kristen M and Sharma, Deepak and Varney, Rebecca and Simmons, Blake and Isern, Nancy G and Markilllie, LM and Nicora, Carrie and Norbeck, Angela D and Taylor, Ronald C and Aldrich, Joshua T and Robinson, Errol W} } @article {1564, title = {A field trial of nutrient stimulation of methanotrophs to reduce greenhouse gas emissions from landfill cover soils.}, journal = {J Air Waste Manag Assoc}, volume = {63}, year = {2013}, month = {2013 Mar}, pages = {300-9}, abstract = {

UNLABELLED: Landfills are among the major sources of anthropogenic methane (CH4) estimated to reach 40 x 10(9) kg per year worldwide by 2015 (IPCC, 2007). A 2 1/2-year field experiment was conducted at a closed landfill in western Michigan where methanotrophs, methane-consuming bacteria, were stimulated by nutrient addition to the soil without significantly increasing biogenic nitrous oxide (N2O) production. The effects of the nitrogen amendments (KNO3 and NH4Cl), phenylacetylene (a selective inhibitor of nitrifying bacteria that contribute to N2O production), and a canopy (to reduce direct water infiltration) on the vertical soil gas profiles of CH4, CO2, and O2 were measured in the top meter of the soil. Methane and nitrous oxide fluxes were calculated from the corresponding soil gas concentration gradients with respect to depth and a Millington-Quirk diffusivity coefficient in soil derived empirically from soil porosity, water content, and diffusivity coefficients in air from the literature. Methane flux estimates were as high as 218.4 g m(-2) day(-1) in the fall and 12.8 g/m(-2) day(-1) in the summer. During the spring and summer CH4 fluxes were reduced by more than half by adding KNO3 and NH4Cl into the soil as compared to control plots, while N2O fluxes increased substantially. The concurrent addition of phenylacetylene to the amendment decreased peak N2O production by half and the rate of peak methane oxidation by about one-third. The seasonal average methane and N2O flux data were extrapolated to estimate the reduction of CH4 and N2O fluxes into the atmosphere by nitrogen and inhibitor addition to the cover soils. The results suggest that such additions coupled with soil moisture management may provide a potential strategy to significantly reduce greenhouse gas emissions from landfills.

IMPLICATIONS: The results of a 2 1/2-year study of effects of nutrient stimulation on methane oxidation in landfill cover soils demonstrates that nutrient addition does decrease methane emissions. The work further underscores the control which soil moisture exerts on methane oxidation. Water management is critical to the success of methane oxidation strategies.

}, keywords = {Biodegradation, Environmental, Gases, Greenhouse Effect, Methane, Oxidation-Reduction, Soil, Soil Microbiology, Waste Management}, issn = {1096-2247}, author = {Lizik, William and Im, Jeongdae and Semrau, Jeremy D and Barcelona, Michael J} } @article {1565, title = {Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity.}, journal = {Philos Trans R Soc Lond B Biol Sci}, volume = {368}, year = {2013}, month = {2013 Apr 19}, pages = {20120320}, abstract = {

Dehalococcoides mccartyi strains are corrinoid-auxotrophic Bacteria and axenic cultures that require vitamin B12 (CN-Cbl) to conserve energy via organohalide respiration. Cultures of D. mccartyi strains BAV1, GT and FL2 grown with limiting amounts of 1 {\textmu}g l(-1) CN-Cbl quickly depleted CN-Cbl, and reductive dechlorination of polychlorinated ethenes was incomplete leading to vinyl chloride (VC) accumulation. In contrast, the same cultures amended with 25 {\textmu}g l(-1) CN-Cbl exhibited up to 2.3-fold higher dechlorination rates, 2.8-9.1-fold increased growth yields, and completely consumed growth-supporting chlorinated ethenes. To explore whether known cobamide-producing microbes supply Dehalococcoides with the required corrinoid cofactor, co-culture experiments were performed with the methanogen Methanosarcina barkeri strain Fusaro and two acetogens, Sporomusa ovata and Sporomusa sp. strain KB-1, as Dehalococcoides partner populations. During growth with H2/CO2, M. barkeri axenic cultures produced 4.2 {\textpm} 0.1 {\textmu}g l(-1) extracellular cobamide (factor III), whereas the Sporomusa cultures produced phenolyl- and p-cresolyl-cobamides. Neither factor III nor the phenolic cobamides supported Dehalococcoides reductive dechlorination activity suggesting that M. barkeri and the Sporomusa sp. cannot fulfil Dehalococcoides{\textquoteright} nutritional requirements. Dehalococcoides dechlorination activity and growth occurred in M. barkeri and Sporomusa sp. co-cultures amended with 10 {\textmu}M 5{\textquoteright},6{\textquoteright}-dimethylbenzimidazole (DMB), indicating that a cobalamin is a preferred corrinoid cofactor of strains BAV1, GT and FL2 when grown with chlorinated ethenes as electron acceptors. Even though the methanogen and acetogen populations tested did not produce cobalamin, the addition of DMB enabled guided biosynthesis and generated a cobalamin that supported Dehalococcoides{\textquoteright} activity and growth. Guided cobalamin biosynthesis may offer opportunities to sustain and enhance Dehalococcoides activity in contaminated subsurface environments.

}, keywords = {Bacteriological Techniques, Benzimidazoles, Biodegradation, Environmental, Chloroflexi, Chromatography, High Pressure Liquid, Coculture Techniques, Culture Media, Dichloroethylenes, Halogenation, Oxidation-Reduction, Trichloroethylene, Vitamin B 12}, issn = {1471-2970}, doi = {10.1098/rstb.2012.0320}, author = {Yan, Jun and Im, Jeongdae and Yang, Yi and L{\"o}ffler, Frank E} } @article {1563, title = {Interference of ferric ions with ferrous iron quantification using the ferrozine assay.}, journal = {J Microbiol Methods}, volume = {95}, year = {2013}, month = {2013 Dec}, pages = {366-7}, abstract = {

The ferrozine assay is a widely used colorimetric method for determining soluble iron concentrations. We provide evidence for a heretofore unrecognized interference of ferric ions (Fe(3+)) on ferrous iron (Fe(2+)) measurements performed in the dark. Fe(3+) concentrations affected the absorbance measurements, which linearly increased with incubation time.

}, keywords = {Chemistry Techniques, Analytical, Colorimetry, Darkness, Diagnostic Errors, Ferric Compounds, Ferrous Compounds, Ferrozine, Ions, Iron, Time Factors}, issn = {1872-8359}, doi = {10.1016/j.mimet.2013.10.005}, author = {Im, Jeongdae and Lee, Jaejin and L{\"o}ffler, Frank E} } @article {3137, title = {U(VI) reduction by diverse outer surface c-type cytochromes of Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Oct}, pages = {6369-74}, abstract = {

Early studies with Geobacter sulfurreducens suggested that outer-surface c-type cytochromes might play a role in U(VI) reduction, but it has recently been suggested that there is substantial U(VI) reduction at the surface of the electrically conductive pili known as microbial nanowires. This phenomenon was further investigated. A strain of G. sulfurreducens, known as Aro-5, which produces pili with substantially reduced conductivity reduced U(VI) nearly as well as the wild type, as did a strain in which the gene for PilA, the structural pilin protein, was deleted. In order to reduce rates of U(VI) reduction to levels less than 20\% of the wild-type rates, it was necessary to delete the genes for the five most abundant outer surface c-type cytochromes of G. sulfurreducens. X-ray absorption near-edge structure spectroscopy demonstrated that whereas 83\% {\textpm} 10\% of the uranium associated with wild-type cells correspond to U(IV) after 4 h of incubation, with the quintuple mutant, 89\% {\textpm} 10\% of uranium was U(VI). Transmission electron microscopy and X-ray energy dispersion spectroscopy revealed that wild-type cells did not precipitate uranium along pili as previously reported, but U(IV) was precipitated at the outer cell surface. These findings are consistent with those of previous studies, which have suggested that G. sulfurreducens requires outer-surface c-type cytochromes but not pili for the reduction of soluble extracellular electron acceptors.

}, keywords = {Cytochromes, Fimbriae, Bacterial, Gene Deletion, Geobacter, Microscopy, Electron, Transmission, Oxidation-Reduction, Uranium, X-Ray Absorption Spectroscopy}, issn = {1098-5336}, doi = {10.1128/AEM.02551-13}, author = {Orellana, Roberto and Leavitt, Janet J and Comolli, Luis R and Csencsits, Roseann and Janot, Noemie and Flanagan, Kelly A and Gray, Arianna S and Leang, Ching and Izallalen, Mounir and Mester, T{\"u}nde and Lovley, Derek R} } @article {412, title = {Identification of multicomponent histidine-aspartate phosphorelay system controlling flagellar and motility gene expression in Geobacter species.}, journal = {J Biol Chem}, volume = {287}, year = {2012}, month = {2012 Mar 30}, pages = {10958-66}, abstract = {Geobacter species play an important role in the natural biogeochemical cycles of aquatic sediments and subsurface environments as well as in subsurface bioremediation by oxidizing organic compounds with the reduction of insoluble Fe(III) oxides. Flagellum-based motility is considered to be critical for Geobacter species to locate fresh sources of Fe(III) oxides. Functional and comparative genomic approaches, coupled with genetic and biochemical methods, identified key regulators for flagellar gene expression in Geobacter species. A master transcriptional regulator, designated FgrM, is a member of the enhancer-binding protein family. The fgrM gene in the most studied strain of Geobacter species, Geobacter sulfurreducens strain DL-1, is truncated by a transposase gene, preventing flagellar biosynthesis. Integrating a functional FgrM homolog restored flagellar biosynthesis and motility in G. sulfurreducens DL-1 and enhanced the ability to reduce insoluble Fe(III) oxide. Interrupting the fgrM gene in G. sulfurreducens strain KN400, which is motile, removed the capacity for flagellar production and inhibited Fe(III) oxide reduction. FgrM, which is also a response regulator of the two-component His-Asp phosphorelay system, was phosphorylated by histidine kinase GHK4, which was essential for flagellar production and motility. GHK4, which is a hybrid kinase with a receiver domain at the N terminus, was phosphorylated by another histidine kinase, GHK3. Therefore, the multicomponent His-Asp phosphorelay system appears to control flagellar gene expression in Geobacter species.}, issn = {1083-351X}, doi = {10.1074/jbc.M112.345041}, author = {Ueki, Toshiyuki and Leang, Ching and Inoue, Kengo and Lovley, Derek R} } @article {416, title = {Laboratory evolution of Geobacter sulfurreducens for enhanced growth on lactate via a single-base-pair substitution in a transcriptional regulator.}, journal = {ISME J}, volume = {6}, year = {2012}, month = {2012 May}, pages = {975-83}, abstract = {The addition of organic compounds to groundwater in order to promote bioremediation may represent a new selective pressure on subsurface microorganisms. The ability of Geobacter sulfurreducens, which serves as a model for the Geobacter species that are important in various types of anaerobic groundwater bioremediation, to adapt for rapid metabolism of lactate, a common bioremediation amendment, was evaluated. Serial transfer of five parallel cultures in a medium with lactate as the sole electron donor yielded five strains that could metabolize lactate faster than the wild-type strain. Genome sequencing revealed that all five strains had non-synonymous single-nucleotide polymorphisms in the same gene, GSU0514, a putative transcriptional regulator. Introducing the single-base-pair mutation from one of the five strains into the wild-type strain conferred rapid growth on lactate. This strain and the five adaptively evolved strains had four to eight-fold higher transcript abundance than wild-type cells for genes for the two subunits of succinyl-CoA synthase, an enzyme required for growth on lactate. DNA-binding assays demonstrated that the protein encoded by GSU0514 bound to the putative promoter of the succinyl-CoA synthase operon. The binding sequence was not apparent elsewhere in the genome. These results demonstrate that a single-base-pair mutation in a transcriptional regulator can have a significant impact on the capacity for substrate utilization and suggest that adaptive evolution should be considered as a potential response of microorganisms to environmental change(s) imposed during bioremediation.}, issn = {1751-7370}, doi = {10.1038/ismej.2011.166}, author = {Summers, Zarath M and Ueki, Toshiyuki and Ismail, Wael and Haveman, Shelley A and Lovley, Derek R} } @article {427, title = {Biochemical characterization of purified OmcS, a c-type cytochrome required for insoluble Fe(III) reduction in Geobacter sulfurreducens.}, journal = {Biochim Biophys Acta}, volume = {1807}, year = {2011}, month = {2011 Apr}, pages = {404-12}, abstract = {Previous studies with Geobacter sulfurreducens have demonstrated that OmcS, an abundant c-type cytochrome that is only loosely bound to the outer surface, plays an important role in electron transfer to Fe(III) oxides as well as other extracellular electron acceptors. In order to further investigate the function of OmcS, it was purified from a strain that overproduces the protein. Purified OmcS had a molecular mass of 47015 Da, and six low-spin bis-histidinyl hexacoordinated heme groups. Its midpoint redox potential was -212 mV. A thermal stability analysis showed that the cooperative melting of purified OmcS occurs in the range of 65-82 {\textdegree}C. Far UV circular dichroism spectroscopy indicated that the secondary structure of purified OmcS consists of about 10\% α-helix and abundant disordered structures. Dithionite-reduced OmcS was able to transfer electrons to a variety of substrates of environmental importance including insoluble Fe(III) oxide, Mn(IV) oxide and humic substances. Stopped flow analysis revealed that the reaction rate of OmcS oxidation has a hyperbolic dependence on the concentration of the studied substrates. A ten-fold faster reaction rate with anthraquinone-2,6-disulfonate (AQDS) (25.2 s$^{-}${\textonesuperior}) was observed as compared to that with Fe(III) citrate (2.9 s$^{-}${\textonesuperior}). The results, coupled with previous localization and gene deletion studies, suggest that OmcS is well-suited to play an important role in extracellular electron transfer.}, keywords = {Circular Dichroism, Cytochrome c Group, Geobacter, Heme, Iron, Kinetics, Molecular Weight, Oxidation-Reduction, Solubility}, issn = {0006-3002}, doi = {10.1016/j.bbabio.2011.01.003}, author = {Qian, Xinlei and Mester, T{\"u}nde and Morgado, Leonor and Arakawa, Tsutomu and Sharma, Manju L and Inoue, Kengo and Joseph, Crisjoe and Salgueiro, Carlos A and Maroney, Michael J and Lovley, Derek R} } @article {1567, title = {Characterization of a novel facultative Methylocystis species capable of growth on methane, acetate and ethanol.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {174-81}, abstract = {

A non-motile strain of Methylocystis, strain SB2, isolated from a spring bog in southeast Michigan, had a curved rod morphology with a typical type II intracytoplasmic membrane system. This organism expressed the membrane-bound or particulate methane monooxygenase (pMMO) as well as a chalkophore with high affinity for copper and did not express the cytoplasmic or soluble methane monooxygenase (sMMO). Strain SB2 was found to grow within the pH range of 6-9, with optimal growth at 6.8. Growth was observed at temperatures ranging between 10{\textdegree}C and 30{\textdegree}C, with no growth at 37{\textdegree}C. The DNA G+C content was 62.9 mol\%. Predominant fatty acids were 18:1ω7c (72.7\%) and 18:1ω9c (24\%) when grown on methane. Phylogenetic comparisons based on both pmoA and 16S rRNA sequences indicated that this organism belonged to the Methylocystis genus, and was closely related to Methylocystis rosea SV97(T) and Methylocystis echinoides IMET10491(T) (98\% 16S rRNA gene sequence similarity to both strains). DNA : DNA hybridizations indicated that strain SB2 had 70\% similarity with M. rosea SV97(T) . Unlike M. rosea SV97(T) , strain SB2 was able to utilize not only methane for growth, but also ethanol and acetate. Furthermore, the predominant fatty acids in strain SB2 were different from those found in M. rosea SV97(T) , i.e. 54.2\% and 39.7\% of fatty acids are 18:1ω8 and 18:1ω7 in M. rosea SV97(T) , while 18:1ω8 is completely absent in strain SB2.

}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00204.x}, author = {Im, Jeongdae and Lee, Sung-Woo and Yoon, Sukhwan and Dispirito, Alan A and Semrau, Jeremy D} } @article {371, title = {Complete genome sequence of "Enterobacter lignolyticus" SCF1.}, journal = {Stand Genomic Sci}, volume = {5}, year = {2011}, month = {2011 Oct 15}, pages = {69-85}, abstract = {In an effort to discover anaerobic bacteria capable of lignin degradation, we isolated "Enterobacter lignolyticus" SCF1 on minimal media with alkali lignin as the sole source of carbon. This organism was isolated anaerobically from tropical forest soils collected from the Short Cloud Forest site in the El Yunque National Forest in Puerto Rico, USA, part of the Luquillo Long-Term Ecological Research Station. At this site, the soils experience strong fluctuations in redox potential and are net methane producers. Because of its ability to grow on lignin anaerobically, we sequenced the genome. The genome of "E. lignolyticus" SCF1 is 4.81 Mbp with no detected plasmids, and includes a relatively small arsenal of lignocellulolytic carbohydrate active enzymes. Lignin degradation was observed in culture, and the genome revealed two putative laccases, a putative peroxidase, and a complete 4-hydroxyphenylacetate degradation pathway encoded in a single gene cluster.}, issn = {1944-3277}, doi = {10.4056/sigs.2104875}, author = {Deangelis, Kristen M and D{\textquoteright}haeseleer, Patrik and Chivian, Dylan and Fortney, Julian L and Khudyakov, Jane and Simmons, Blake and Woo, Hannah and Arkin, Adam P and Davenport, Karen Walston and Goodwin, Lynne and Chen, Amy and Ivanova, Natalia and Kyrpides, Nikos C and Mavromatis, Konstantinos and Woyke, Tanja and Hazen, Terry C} } @article {1566, title = {Constitutive expression of pMMO by Methylocystis strain SB2 when grown on multi-carbon substrates: implications for biodegradation of chlorinated ethenes.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {182-8}, abstract = {

The particulate methane monooxygenase (pMMO) in Methylocystis strain SB2 was found to be constitutively expressed in the absence of methane when the strain was grown on either acetate or ethanol. Real-time quantitative polymerase chain reaction (PCR) and reverse transcription-PCR showed that the expression of pmoA decreased by one to two orders of magnitude when grown on acetate as compared with growth of strain SB2 on methane. The capability of strain SB2 to degrade a mixture of chlorinated ethenes in the absence of methane was examined to verify the presence and activity of pMMO under acetate-growth conditions as well determine the effectiveness of such conditions for bioremediation. It was found that when strain SB2 was grown on acetate and exposed to 40 {\textmu}M each of trichloroethylene (TCE), trans-dichloroethylene (t-DCE) and vinyl chloride (VC), approximately 30\% of VC and t-DCE was degraded but no appreciable TCE removal was measured after 216 h of incubation. The ability to degrade VC and t-DCE was lost when acetylene was added, confirming that pMMO was responsible for the degradation of these chlorinated ethenes by Methylocystis strain SB2 when the strain was grown on acetate.

}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00205.x}, author = {Yoon, Sukhwan and Im, Jeongdae and Bandow, Nathan and Dispirito, Alan A and Semrau, Jeremy D} } @article {1569, title = {Field application of nitrogen and phenylacetylene to mitigate greenhouse gas emissions from landfill cover soils: effects on microbial community structure.}, journal = {Appl Microbiol Biotechnol}, volume = {89}, year = {2011}, month = {2011 Jan}, pages = {189-200}, abstract = {

Landfills are large sources of CH(4), but a considerable amount of CH(4) can be removed in situ by methanotrophs if their activity can be stimulated through the addition of nitrogen. Nitrogen can, however, lead to increased N(2)O production. To examine the effects of nitrogen and a selective inhibitor on CH(4) oxidation and N(2)O production in situ, 0.5 M of NH(4)Cl and 0.25 M of KNO(3), with and without 0.01\% (w/v) phenylacetylene, were applied to test plots at a landfill in Kalamazoo, MI from 2007 November to 2009 July. Nitrogen amendments stimulated N(2)O production but had no effect on CH(4) oxidation. The addition of phenylacetylene stimulated CH(4) oxidation while reducing N(2)O production. Methanotrophs possessing particulate methane monooxygenase and archaeal ammonia-oxidizers (AOAs) were abundant. The addition of nitrogen reduced methanotrophic diversity, particularly for type I methanotrophs. The simultaneous addition of phenylacetylene increased methanotrophic diversity and the presence of type I methanotrophs. Clone libraries of the archaeal amoA gene showed that the addition of nitrogen increased AOAs affiliated with Crenarchaeal group 1.1b, while they decreased with the simultaneous addition of phenylacetylene. These results suggest that the addition of phenylacetylene with nitrogen reduces N(2)O production by selectively inhibiting AOAs and/or type II methanotrophs.

}, keywords = {Acetylene, Archaea, Archaeal Proteins, Bacteria, Bacterial Proteins, Gases, Greenhouse Effect, Methane, Molecular Sequence Data, Nitrogen, Refuse Disposal, Soil, Soil Microbiology}, issn = {1432-0614}, doi = {10.1007/s00253-010-2811-0}, author = {Im, Jeongdae and Lee, Sung-Woo and Bodrossy, Levente and Barcelona, Michael J and Semrau, Jeremy D} } @article {438, title = {Genome-scale dynamic modeling of the competition between Rhodoferax and Geobacter in anoxic subsurface environments.}, journal = {ISME J}, volume = {5}, year = {2011}, month = {2011 Feb}, pages = {305-16}, abstract = {The advent of rapid complete genome sequencing, and the potential to capture this information in genome-scale metabolic models, provide the possibility of comprehensively modeling microbial community interactions. For example, Rhodoferax and Geobacter species are acetate-oxidizing Fe(III)-reducers that compete in anoxic subsurface environments and this competition may have an influence on the in situ bioremediation of uranium-contaminated groundwater. Therefore, genome-scale models of Geobacter sulfurreducens and Rhodoferax ferrireducens were used to evaluate how Geobacter and Rhodoferax species might compete under diverse conditions found in a uranium-contaminated aquifer in Rifle, CO. The model predicted that at the low rates of acetate flux expected under natural conditions at the site, Rhodoferax will outcompete Geobacter as long as sufficient ammonium is available. The model also predicted that when high concentrations of acetate are added during in situ bioremediation, Geobacter species would predominate, consistent with field-scale observations. This can be attributed to the higher expected growth yields of Rhodoferax and the ability of Geobacter to fix nitrogen. The modeling predicted relative proportions of Geobacter and Rhodoferax in geochemically distinct zones of the Rifle site that were comparable to those that were previously documented with molecular techniques. The model also predicted that under nitrogen fixation, higher carbon and electron fluxes would be diverted toward respiration rather than biomass formation in Geobacter, providing a potential explanation for enhanced in situ U(VI) reduction in low-ammonium zones. These results show that genome-scale modeling can be a useful tool for predicting microbial interactions in subsurface environments and shows promise for designing bioremediation strategies.}, keywords = {Acetates, Anaerobiosis, Biodegradation, Environmental, Biomass, Comamonadaceae, Genome, Genome, Bacterial, Geobacter, Models, Biological, Nitrogen Fixation, Quaternary Ammonium Compounds, RNA, Ribosomal, 16S, Uranium, Water Microbiology, Water Pollutants, Radioactive}, issn = {1751-7370}, doi = {10.1038/ismej.2010.117}, author = {Zhuang, Kai and Izallalen, Mounir and Mouser, Paula and Richter, Hanno and Risso, Carla and Mahadevan, Radhakrishnan and Lovley, Derek R} } @article {1568, title = {Pollutant degradation by a Methylocystis strain SB2 grown on ethanol: bioremediation via facultative methanotrophy.}, journal = {FEMS Microbiol Lett}, volume = {318}, year = {2011}, month = {2011 May}, pages = {137-42}, abstract = {

A facultative methanotroph, Methylocystis strain SB2, was examined for its ability to degrade chlorinated hydrocarbons when grown on methane or ethanol. Strain SB2 grown on methane degraded vinyl chloride (VC), trans-dichloroethylene (t-DCE), trichloroethylene (TCE), 1,1,1-trichloroethane (1,1,1-TCA), and chloroform (CF), but not dichloromethane (DCM). Growth on methane was reduced in the presence of any chlorinated hydrocarbon. Strain SB2 grown on ethanol degraded VC, t-DCE, and TCE, and 1,1,1-TCA, but not DCM or CF. With the exception of 1,1,1-TCA, the growth of strain SB2 on ethanol was not affected by any individual chlorinated hydrocarbon. No degradation of any chlorinated hydrocarbon was observed when acetylene was added to ethanol-grown cultures, indicating that this degradation was due to particulate methane monooxygenase (pMMO) activity. When mixtures of chlorinated alkanes or alkenes were added to cultures growing on methane or ethanol, chlorinated alkene degradation occurred, but chlorinated alkanes were not, and growth was reduced on both methane and ethanol. Collectively, these data indicate that competitive inhibition of pMMO activity limits methanotrophic growth and pollutant degradation. Facultative methanotrophy may thus be useful to extend the utility of methanotrophs for bioremediation as the use of alternative growth substrates allows for pMMO activity to be focused on pollutant degradation.

}, keywords = {Autotrophic Processes, Biodegradation, Environmental, Environmental Pollutants, Ethanol, Hydrocarbons, Chlorinated, Methane, Methylocystaceae}, issn = {1574-6968}, doi = {10.1111/j.1574-6968.2011.02249.x}, author = {Im, Jeongdae and Semrau, Jeremy D} } @article {719, title = {Taxonomical developments in the family Polyomaviridae.}, journal = {Arch Virol}, volume = {156}, year = {2011}, month = {2011 Sep}, pages = {1627-34}, abstract = {The Polyomaviridae Study Group of the International Committee on Taxonomy of Viruses (ICTV) has recommended several taxonomical revisions, as follows: The family Polyomaviridae, which is currently constituted as a single genus (Polyomavirus), will be comprised of three genera: two containing mammalian viruses and one containing avian viruses. The two mammalian genera will be designated Orthopolyomavirus and Wukipolyomavirus, and the avian genus will be named Avipolyomavirus. These genera will be created by the redistribution of species from the current single genus (Polyomavirus) and by the inclusion of several new species. In addition, the names of several species will be changed to reflect current usage.}, keywords = {Phylogeny, Polyomaviridae, Terminology as Topic}, issn = {1432-8798}, doi = {10.1007/s00705-011-1008-x}, author = {Johne, Reimar and Buck, Christopher B and Allander, Tobias and Atwood, Walter J and Garcea, Robert L and Imperiale, Michael J and Major, Eugene O and Ramqvist, Torbjorn and Norkin, Leonard C} } @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 {690, title = {Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle.}, journal = {J Exp Med}, volume = {206}, year = {2009}, month = {2009 Dec 21}, pages = {2879-88}, abstract = {Tuberculosis remains a fatal disease caused by Mycobacterium tuberculosis, which contains various unique components that affect the host immune system. Trehalose-6,6{\textquoteright}-dimycolate (TDM; also called cord factor) is a mycobacterial cell wall glycolipid that is the most studied immunostimulatory component of M. tuberculosis. Despite five decades of research on TDM, its host receptor has not been clearly identified. Here, we demonstrate that macrophage inducible C-type lectin (Mincle) is an essential receptor for TDM. Heat-killed mycobacteria activated Mincle-expressing cells, but the activity was lost upon delipidation of the bacteria; analysis of the lipid extracts identified TDM as a Mincle ligand. TDM activated macrophages to produce inflammatory cytokines and nitric oxide, which are completely suppressed in Mincle-deficient macrophages. In vivo TDM administration induced a robust elevation of inflammatory cytokines in sera and characteristic lung inflammation, such as granuloma formation. However, no TDM-induced lung granuloma was formed in Mincle-deficient mice. Whole mycobacteria were able to activate macrophages even in MyD88-deficient background, but the activation was significantly diminished in Mincle/MyD88 double-deficient macrophages. These results demonstrate that Mincle is an essential receptor for the mycobacterial glycolipid, TDM.}, keywords = {Animals, Cord Factors, Granuloma, Lectins, C-Type, Ligands, Lung Diseases, Macrophage Activation, Membrane Proteins, Mice, Mice, Inbred C57BL, Myeloid Differentiation Factor 88, Receptors, IgG}, issn = {1540-9538}, doi = {10.1084/jem.20091750}, author = {Ishikawa, Eri and Ishikawa, Tetsuaki and Morita, Yasu S and Toyonaga, Kenji and Yamada, Hisakata and Takeuchi, Osamu and Kinoshita, Taroh and Akira, Shizuo and Yoshikai, Yasunobu and Yamasaki, Sho} } @article {1570, title = {Effect of nutrient and selective inhibitor amendments on methane oxidation, nitrous oxide production, and key gene presence and expression in landfill cover soils: characterization of the role of methanotrophs, nitrifiers, and denitrifiers.}, journal = {Appl Microbiol Biotechnol}, volume = {85}, year = {2009}, month = {2009 Nov}, pages = {389-403}, abstract = {

Methane and nitrous oxide are both potent greenhouse gasses, with global warming potentials approximately 25 and 298 times that of carbon dioxide. A matrix of soil microcosms was constructed with landfill cover soils collected from the King Highway Landfill in Kalamazoo, Michigan and exposed to geochemical parameters known to affect methane consumption by methanotrophs while also examining their impact on biogenic nitrous oxide production. It was found that relatively dry soils (5\% moisture content) along with 15 mg NH (4) (+) (kg soil)(-1) and 0.1 mg phenylacetylene(kg soil)(-1) provided the greatest stimulation of methane oxidation while minimizing nitrous oxide production. Microarray analyses of pmoA showed that the methanotrophic community structure was dominated by Type II organisms, but Type I genera were more evident with the addition of ammonia. When phenylacetylene was added in conjunction with ammonia, the methanotrophic community structure was more similar to that observed in the presence of no amendments. PCR analyses showed the presence of amoA from both ammonia-oxidizing bacteria and archaea, and that the presence of key genes associated with these cells was reduced with the addition of phenylacetylene. Messenger RNA analyses found transcripts of pmoA, but not of mmoX, nirK, norB, or amoA from either ammonia-oxidizing bacteria or archaea. Pure culture analyses showed that methanotrophs could produce significant amounts of nitrous oxide, particularly when expressing the particulate methane monooxygenase (pMMO). Collectively, these data suggest that methanotrophs expressing pMMO played a role in nitrous oxide production in these microcosms.

}, keywords = {Archaea, Bacteria, Base Sequence, DNA, DNA Primers, DNA, Archaeal, DNA, Bacterial, Global Warming, Greenhouse Effect, Inorganic Chemicals, Methane, Nitrites, Nitrogen, Nitrogen Oxides, Nitrous Oxide, Oxidation-Reduction, Polymerase Chain Reaction, Reverse Transcriptase Polymerase Chain Reaction, RNA, Bacterial, RNA, Messenger, Soil, Water Pollutants, Chemical}, issn = {1432-0614}, doi = {10.1007/s00253-009-2238-7}, author = {Lee, Sung-Woo and Im, Jeongdae and Dispirito, Alan A and Bodrossy, Levente and Barcelona, Michael J and Semrau, Jeremy D} } @article {456, title = {Genome-scale comparison and constraint-based metabolic reconstruction of the facultative anaerobic Fe(III)-reducer Rhodoferax ferrireducens.}, journal = {BMC Genomics}, volume = {10}, year = {2009}, month = {2009}, pages = {447}, abstract = {BACKGROUND: Rhodoferax ferrireducens is a metabolically versatile, Fe(III)-reducing, subsurface microorganism that is likely to play an important role in the carbon and metal cycles in the subsurface. It also has the unique ability to convert sugars to electricity, oxidizing the sugars to carbon dioxide with quantitative electron transfer to graphite electrodes in microbial fuel cells. In order to expand our limited knowledge about R. ferrireducens, the complete genome sequence of this organism was further annotated and then the physiology of R. ferrireducens was investigated with a constraint-based, genome-scale in silico metabolic model and laboratory studies. RESULTS: The iterative modeling and experimental approach unveiled exciting, previously unknown physiological features, including an expanded range of substrates that support growth, such as cellobiose and citrate, and provided additional insights into important features such as the stoichiometry of the electron transport chain and the ability to grow via fumarate dismutation. Further analysis explained why R. ferrireducens is unable to grow via photosynthesis or fermentation of sugars like other members of this genus and uncovered novel genes for benzoate metabolism. The genome also revealed that R. ferrireducens is well-adapted for growth in the subsurface because it appears to be capable of dealing with a number of environmental insults, including heavy metals, aromatic compounds, nutrient limitation and oxidative stress. CONCLUSION: This study demonstrates that combining genome-scale modeling with the annotation of a new genome sequence can guide experimental studies and accelerate the understanding of the physiology of under-studied yet environmentally relevant microorganisms.}, keywords = {Comamonadaceae, Comparative Genomic Hybridization, DNA, Bacterial, Ferric Compounds, Genome, Bacterial, Genomics, Models, Biological, Oxidation-Reduction, Sequence Analysis, DNA}, issn = {1471-2164}, doi = {10.1186/1471-2164-10-447}, author = {Risso, Carla and Sun, Jun and Zhuang, Kai and Mahadevan, Radhakrishnan and DeBoy, Robert and Ismail, Wael and Shrivastava, Susmita and Huot, Heather and Kothari, Sagar and Daugherty, Sean and Bui, Olivia and Schilling, Christophe H and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {476, title = {Geobacter sulfurreducens strain engineered for increased rates of respiration.}, journal = {Metab Eng}, volume = {10}, year = {2008}, month = {2008 Sep}, pages = {267-75}, abstract = {Geobacter species are among the most effective microorganisms known for the bioremediation of radioactive and toxic metals in contaminated subsurface environments and for converting organic compounds to electricity in microbial fuel cells. However, faster rates of electron transfer could aid in optimizing these processes. Therefore, the Optknock strain design methodology was applied in an iterative manner to the constraint-based, in silico model of Geobacter sulfurreducens to identify gene deletions predicted to increase respiration rates. The common factor in the Optknock predictions was that each resulted in a predicted increase in the cellular ATP demand, either by creating ATP-consuming futile cycles or decreasing the availability of reducing equivalents and inorganic phosphate for ATP biosynthesis. The in silico model predicted that increasing the ATP demand would result in higher fluxes of acetate through the TCA cycle and higher rates of NADPH oxidation coupled with decreases in flux in reactions that funnel acetate toward biosynthetic pathways. A strain of G. sulfurreducens was constructed in which the hydrolytic, F(1) portion of the membrane-bound F(0)F(1) (H(+))-ATP synthase complex was expressed when IPTG was added to the medium. Induction of the ATP drain decreased the ATP content of the cell by more than half. The cells with the ATP drain had higher rates of respiration, slower growth rates, and a lower cell yield. Genome-wide analysis of gene transcript levels indicated that when the higher rate of respiration was induced transcript levels were higher for genes involved in energy metabolism, especially in those encoding TCA cycle enzymes, subunits of the NADH dehydrogenase, and proteins involved in electron acceptor reduction. This was accompanied by lower transcript levels for genes encoding proteins involved in amino acid biosynthesis, cell growth, and motility. Several changes in gene expression that involve processes not included in the in silico model were also detected, including increased expression of a number of redox-active proteins, such as c-type cytochromes and a putative multicopper outer-surface protein. The results demonstrate that it is possible to genetically engineer increased respiration rates in G. sulfurreducens in accordance with predictions from in silico metabolic modeling. To our knowledge, this is the first report of metabolic engineering to increase the respiratory rate of a microorganism.}, keywords = {Adenosine Triphosphate, Bacterial Proteins, Biodegradation, Environmental, Citric Acid Cycle, Electron Transport, Geobacter, Metals, Models, Biological, NADH Dehydrogenase, NADP, Oxygen Consumption, Phosphates, Proton-Translocating ATPases, Radioactive Pollutants}, issn = {1096-7184}, doi = {10.1016/j.ymben.2008.06.005}, author = {Izallalen, Mounir and Mahadevan, Radhakrishnan and Burgard, Anthony and Postier, Bradley and DiDonato, Raymond and Sun, Jun and Schilling, Christopher H and Lovley, Derek R} } @article {812, title = {RecA-mediated SOS induction requires an extended filament conformation but no ATP hydrolysis.}, journal = {Mol Microbiol}, volume = {69}, year = {2008}, month = {2008 Sep}, pages = {1165-79}, abstract = {The Escherichia coli SOS response to DNA damage is modulated by the RecA protein, a recombinase that forms an extended filament on single-stranded DNA and hydrolyzes ATP. The RecA K72R (recA2201) mutation eliminates the ATPase activity of RecA protein. The mutation also limits the capacity of RecA to form long filaments in the presence of ATP. Strains with this mutation do not undergo SOS induction in vivo. We have combined the K72R variant of RecA with another mutation, RecA E38K (recA730). In vitro, the double mutant RecA E38K/K72R (recA730,2201) mimics the K72R mutant protein in that it has no ATPase activity. The double mutant protein will form long extended filaments on ssDNA and facilitate LexA cleavage almost as well as wild-type, and do so in the presence of ATP. Unlike recA K72R, the recA E38K/K72R double mutant promotes SOS induction in vivo after UV treatment. Thus, SOS induction does not require ATP hydrolysis by the RecA protein, but does require formation of extended RecA filaments. The RecA E38K/K72R protein represents an improved reagent for studies of the function of ATP hydrolysis by RecA in vivo and in vitro.}, keywords = {Adenosine Triphosphate, Amino Acid Substitution, Bacterial Proteins, DNA, Bacterial, DNA, Single-Stranded, Escherichia coli, Escherichia coli Proteins, Hydrolysis, Rec A Recombinases, Serine Endopeptidases, SOS Response (Genetics), Ultraviolet Rays}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06341.x}, author = {Gruenig, Marielle C and Renzette, Nicholas and Long, Edward and Chitteni-Pattu, Sindhu and Inman, Ross B and Cox, Michael M and Sandler, Steven J} } @article {748, title = {Distribution of extensive nifH gene diversity across physical soil microenvironments.}, journal = {Microb Ecol}, volume = {51}, year = {2006}, month = {2006 May}, pages = {441-52}, abstract = {The diversity of nitrogen-fixing bacteria is well described for aquatic environments; however, terrestrial analyses remain mostly biased to rhizobial plant-microbe associations. We maximized the level of resolution for this study through the use of nucleotide sequence information extracted from a series of soil microenvironments, ranging from macroaggregates at 2000 microm to the clay fraction at < 75 microm in diameter. In addition, we attempted to create an overview of the distribution of terrestrial nitrogen fixers across such microenvironments by combining culture-independent techniques with a suite of natural soil environments from uniquely different origins. Soil diazotroph diversity was analyzed phylogenetically for 600 terrestrial nifH sequences from 12 midsized clone libraries based on microenvironments of three separate soils across a global scale. Statistical analyses of nifH gene clone libraries were used to estimate coverage, establish degrees of sequence overlap, and compare cluster distributions. These analyses revealed an extensive diversity in a tropical (19 phylotypes) and an arctic soil (17 phylotypes), and moderate diversity in a temperate soil (11 phylotypes). Within each soil, comparisons across aggregate size fractions delineated nifH gene cluster shifts within populations and degrees of sequence overlap that ranged from significantly different (arctic, tropical) to significantly similar (temperate). We suggest that this is due to population separation across aggregates of different size classes, which results from differences in the temporal stability of aggregates as niches for microbial communities. This study not only provides new knowledge of the arrangement of diazotrophic communities at the soil microscale, but it also contributes to the underrepresented knowledge of soil nifH sequences in the public databases.}, keywords = {Bacteria, Base Sequence, DNA Primers, Genes, Bacterial, Multigene Family, Oxidoreductases, Phylogeny, Polymerase Chain Reaction, Soil Microbiology}, issn = {0095-3628}, doi = {10.1007/s00248-006-9044-x}, author = {Izquierdo, Javier A and N{\"u}sslein, Klaus} } @article {533, title = {Characterization of citrate synthase from Geobacter sulfurreducens and evidence for a family of citrate synthases similar to those of eukaryotes throughout the Geobacteraceae.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Jul}, pages = {3858-65}, abstract = {Members of the family Geobacteraceae are commonly the predominant Fe(III)-reducing microorganisms in sedimentary environments, as well as on the surface of energy-harvesting electrodes, and are able to effectively couple the oxidation of acetate to the reduction of external electron acceptors. Citrate synthase activity of these organisms is of interest due to its key role in acetate metabolism. Prior sequencing of the genome of Geobacter sulfurreducens revealed a putative citrate synthase sequence related to the citrate synthases of eukaryotes. All citrate synthase activity in G. sulfurreducens could be resolved to a single 49-kDa protein via affinity chromatography. The enzyme was successfully expressed at high levels in Escherichia coli with similar properties as the native enzyme, and kinetic parameters were comparable to related citrate synthases (kcat= 8.3 s(-1); Km= 14.1 and 4.3 microM for acetyl coenzyme A and oxaloacetate, respectively). The enzyme was dimeric and was slightly inhibited by ATP (Ki= 1.9 mM for acetyl coenzyme A), which is a known inhibitor for many eukaryotic, dimeric citrate synthases. NADH, an allosteric inhibitor of prokaryotic hexameric citrate synthases, did not affect enzyme activity. Unlike most prokaryotic dimeric citrate synthases, the enzyme did not have any methylcitrate synthase activity. A unique feature of the enzyme, in contrast to citrate synthases from both eukaryotes and prokaryotes, was a lack of stimulation by K+ ions. Similar citrate synthase sequences were detected in a diversity of other Geobacteraceae members. This first characterization of a eukaryotic-like citrate synthase from a prokaryote provides new insight into acetate metabolism in Geobacteraceae members and suggests a molecular target for tracking the presence and activity of these organisms in the environment.}, keywords = {Amino Acid Sequence, Citrate (si)-Synthase, Culture Media, Deltaproteobacteria, DNA, Bacterial, Eukaryotic Cells, Geobacter, Kinetics, Molecular Sequence Data, Phylogeny, Sequence Analysis, DNA}, issn = {0099-2240}, doi = {10.1128/AEM.71.7.3858-3865.2005}, author = {Bond, Daniel R and Mester, T{\"u}nde and Nesb{\o}, Camilla L and Izquierdo-Lopez, Andrea V and Collart, Frank L and Lovley, Derek R} } @article {752, title = {Tuning the hemolytic and antibacterial activities of amphiphilic polynorbornene derivatives.}, journal = {J Am Chem Soc}, volume = {126}, year = {2004}, month = {2004 Dec 8}, pages = {15870-5}, abstract = {Amphiphilic cationic polynorbornene derivatives, soluble in water, were prepared from modular norbornene monomers, with a wide range of molecular weights (M(n) = 1600-137 500 g/mol) and narrow polydispersities (PDI = 1.1-1.3). The antibacterial activity determined by growth inhibition assays and the hemolytic activity against human red blood cells were measured and compared to determine the selectivity of the polymers for bacterial over mammalian cells. The effects of monomer repeat unit hydrophobicity and polymer molecular weight on antibacterial and hemolytic activities were determined. The hydrophobicity of the repeat unit was observed to have dramatic effects on antibacterial and hemolytic activities. Lipid membrane disruption activities of the polymers was confirmed by measuring polymer-induced dye leakage from large unilamellar vesicles. By tuning the overall hydrophobicity of the polymer through random copolymerizations of modular norbornene derivatives, highly selective, nonhemolytic antibacterial activities were obtained. For appropriate monomer composition, selectivity against bacteria versus human red blood cells was determined to be over 100.}, keywords = {Anti-Bacterial Agents, Bacillus subtilis, Cell Membrane, Cholesterol, Erythrocytes, Escherichia coli, Hemolysis, Humans, Liposomes, Microbial Sensitivity Tests, Molecular Weight, Norbornanes, Polymers, Structure-Activity Relationship}, issn = {0002-7863}, doi = {10.1021/ja045664d}, author = {Ilker, M Firat and N{\"u}sslein, Klaus and Tew, Gregory N and Coughlin, E Bryan} } @article {590, title = {Isolation, characterization and gene sequence analysis of a membrane-associated 89 kDa Fe(III) reducing cytochrome c from Geobacter sulfurreducens.}, journal = {Biochem J}, volume = {359}, year = {2001}, month = {2001 Oct 1}, pages = {147-52}, abstract = {Geobacter sulfurreducens is capable of anaerobic respiration with Fe(III) as a terminal electron acceptor via a membrane-bound Fe(III) reductase activity associated with a large molecular mass cytochrome c. This cytochrome was purified by detergent extraction of the membrane fraction, Q-Sepharose ion-exchange chromatography, preparative electrophoresis, and MonoQ ion-exchange chromatography. Spectrophotometric analysis of the purified cytochrome reveals a c-type haem, with no evidence of haem a, haem b or sirohaem. The cytochrome has an M(r) of 89000 as determined by denaturing PAGE, and has an isoelectric point of 5.2 as determined by analytical isoelectric focusing. Dithionite-reduced cytochrome can donate electrons to Fe(III)-nitrilotriacetic acid and synthetic ferrihydrite, thus demonstrating that the cytochrome has redox and thermodynamic properties required for reduction of Fe(III). Analysis using cyclic voltammetry confirmed that the reduced cytochrome can catalytically transfer electrons to ferrihydrite, further demonstrating its ability to be an electron transport mediator in anaerobic Fe(III) respiration. Sequence analysis of a cloned chromosomal DNA fragment revealed a 2307 bp open reading frame (ferA) encoding a 768 amino acid protein corresponding to the 89 kDa cytochrome. The deduced amino acid sequence (FerA) translated from the open reading frame contained 12 putative haem-binding motifs, as well as a hydrophobic N-terminal membrane anchor sequence, a lipid-attachment site and an ATP/GTP-binding site. FerA displayed 20\% or less identity with amino acid sequences of other known cytochromes, although it does share some features with characterized polyhaem cytochromes c.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Chromatography, Ion Exchange, Cloning, Molecular, Cytochrome c Group, Deltaproteobacteria, DNA Primers, Electron Transport, Electrophoresis, Polyacrylamide Gel, Ferric Compounds, Gene Expression Regulation, Bacterial, Molecular Sequence Data, Molecular Weight, Nitrilotriacetic Acid, Oxidation-Reduction, Polymerase Chain Reaction, Sequence Homology, Amino Acid}, issn = {0264-6021}, author = {Magnuson, T S and Isoyama, N and Hodges-Myerson, A L and Davidson, G and Maroney, M J and Geesey, G G and Lovley, D R} }