@article {3051, title = {Direct microbial electron uptake as a mechanism for stainless steel corrosion in aerobic environments.}, journal = {Water Res}, volume = {219}, year = {2022}, month = {2022 Jul 01}, pages = {118553}, abstract = {

Shewanella oneidensis MR-1 is an attractive model microbe for elucidating the biofilm-metal interactions that contribute to the billions of dollars in corrosion damage to industrial applications each year. Multiple mechanisms for S. oneidensis-enhanced corrosion have been proposed, but none of these mechanisms have previously been rigorously investigated with methods that rule out alternative routes for electron transfer. We found that S. oneidensis grown under aerobic conditions formed thick biofilms (\~{}50~{\textmu}m) on stainless steel coupons, accelerating corrosion over sterile controls. H and flavins were ruled out as intermediary electron carriers because stainless steel did not reduce riboflavin and previous studies have demonstrated stainless does not generate H. Strain ∆mtrCBA, in which the genes for the most abundant porin-cytochrome conduit in S. oneidensis were deleted, corroded stainless steel substantially less than wild-type in aerobic cultures. Wild-type biofilms readily reduced nitrate with stainless steel as the sole electron donor under anaerobic conditions, but strain ∆mtrCBA did not. These results demonstrate that S. oneidensis can directly consume electrons from iron-containing metals and illustrate how direct metal-to-microbe electron transfer can be an important route for corrosion, even in aerobic environments.

}, keywords = {Biofilms, Corrosion, Electron Transport, Electrons, Metals, Oxidation-Reduction, Stainless Steel, Steel}, issn = {1879-2448}, doi = {10.1016/j.watres.2022.118553}, author = {Zhou, Enze and Li, Feng and Zhang, Dawei and Xu, Dake and Li, Zhong and Jia, Ru and Jin, Yuting and Song, Hao and Li, Huabing and Wang, Qiang and Wang, Jianjun and Li, Xiaogang and Gu, Tingyue and Homborg, Axel M and Mol, Johannes M C and Smith, Jessica A and Wang, Fuhui and Lovley, Derek R} } @article {3058, title = {Direct Observation of Electrically Conductive Pili Emanating from .}, journal = {mBio}, volume = {12}, year = {2021}, month = {2021 Aug 31}, pages = {e0220921}, abstract = {

Geobacter sulfurreducens is a model microbe for elucidating the mechanisms for extracellular electron transfer in several biogeochemical cycles, bioelectrochemical applications, and microbial metal corrosion. Multiple lines of evidence previously suggested that electrically conductive pili (e-pili) are an essential conduit for long-range extracellular electron transport in G. sulfurreducens. However, it has recently been reported that G. sulfurreducens does not express e-pili and that filaments comprised of multi-heme -type cytochromes are responsible for long-range electron transport. This possibility was directly investigated by examining cells, rather than filament preparations, with atomic force microscopy. Approximately 90\% of the filaments emanating from wild-type cells had a diameter (3 nm) and conductance consistent with previous reports of e-pili harvested from G. sulfurreducens or heterologously expressed in Escherichia coli from the G. sulfurreducens pilin gene. The remaining 10\% of filaments had a morphology consistent with filaments comprised of the -type cytochrome OmcS. A strain expressing a modified pilin gene designed to yield poorly conductive pili expressed 90\% filaments with a 3-nm diameter, but greatly reduced conductance, further indicating that the 3-nm diameter conductive filaments in the wild-type strain were e-pili. A strain in which genes for five of the most abundant outer-surface -type cytochromes, including OmcS, were deleted yielded only 3-nm-diameter filaments with the same conductance as in the wild type. These results demonstrate that e-pili are the most abundant conductive filaments expressed by G. sulfurreducens, consistent with previous functional studies demonstrating the need for e-pili for long-range extracellular electron transfer. Electroactive microbes have significant environmental impacts, as well as applications in bioenergy and bioremediation. The composition, function, and even existence of electrically conductive pili (e-pili) has been one of the most contentious areas of investigation in electromicrobiology, in part because e-pili offer a mechanism for long-range electron transport that does not involve the metal cofactors common in much of biological electron transport. This study demonstrates that e-pili are abundant filaments emanating from Geobacter sulfurreducens, which serves as a model for long-range extracellular electron transfer in direct interspecies electron transfer, dissimilatory metal reduction, microbe-electrode exchange, and corrosion caused by direct electron uptake from Fe(0). The methods described in this study provide a simple strategy for evaluating the distribution of conductive filaments throughout the microbial world with an approach that avoids artifactual production and/or enrichment of filaments that may not be physiologically relevant.

}, keywords = {Electric Conductivity, Electrons, Escherichia coli, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Microscopy, Atomic Force, Oxidation-Reduction}, issn = {2150-7511}, doi = {10.1128/mBio.02209-21}, author = {Liu, Xinying and Walker, David J F and Nonnenmann, Stephen S and Sun, Dezhi and Lovley, Derek R} } @article {3075, title = {Decorating the Outer Surface of Microbially Produced Protein Nanowires with Peptides.}, journal = {ACS Synth Biol}, volume = {8}, year = {2019}, month = {2019 Aug 16}, pages = {1809-1817}, abstract = {

The potential applications of electrically conductive protein nanowires (e-PNs) harvested from might be greatly expanded if the outer surface of the wires could be modified to confer novel sensing capabilities or to enhance binding to other materials. We developed a simple strategy for functionalizing e-PNs with surface-exposed peptides. The gene for the monomer that assembles into e-PNs was modified to add peptide tags at the carboxyl terminus of the monomer. Strains of were constructed that fabricated synthetic e-PNs with a six-histidine "His-tag" or both the His-tag and a nine-peptide "HA-tag" exposed on the outer surface. Addition of the peptide tags did not diminish e-PN conductivity. The abundance of HA-tag in e-PNs was controlled by placing expression of the gene for the synthetic monomer with the HA-tag under transcriptional regulation. These studies suggest broad possibilities for tailoring e-PN properties for diverse applications.

}, keywords = {Carboxy-Lyases, Ethylene Glycols, Molecular Structure, Nanowires, Oxygenases, Peptides, Phenylalanine Ammonia-Lyase, Plasmids, Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Styrenes}, issn = {2161-5063}, doi = {10.1021/acssynbio.9b00131}, author = {Ueki, Toshiyuki and Walker, David J F and Tremblay, Pier-Luc and Nevin, Kelly P and Ward, Joy E and Woodard, Trevor L and Nonnenmann, Stephen S and Lovley, Derek R} } @article {2219, title = {DivIVA concentrates mycobacterial cell envelope assembly for initiation and stabilization of polar growth.}, journal = {Cytoskeleton}, volume = {75}, year = {2018}, month = {11/2018}, chapter = {498}, doi = {10.1002/cm.21490}, author = {Melzer, ES and Sein, CE and Siegrist, MS and Chambers, JJ} } @article {3125, title = {Direct interspecies electron transfer between Geobacter metallireducens and Methanosarcina barkeri.}, journal = {Appl Environ Microbiol}, volume = {80}, year = {2014}, month = {2014 Aug}, pages = {4599-605}, abstract = {

Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri to participate in DIET was evaluated in coculture with Geobacter metallireducens. Cocultures formed aggregates that shared electrons via DIET during the stoichiometric conversion of ethanol to methane. Cocultures could not be initiated with a pilin-deficient G. metallireducens strain, suggesting that long-range electron transfer along pili was important for DIET. Amendments of granular activated carbon permitted the pilin-deficient G. metallireducens isolates to share electrons with M. barkeri, demonstrating that this conductive material could substitute for pili in promoting DIET. When M. barkeri was grown in coculture with the H2-producing Pelobacter carbinolicus, incapable of DIET, M. barkeri utilized H2 as an electron donor but metabolized little of the acetate that P.carbinolicus produced. This suggested that H2, but not electrons derived from DIET, inhibited acetate metabolism. P. carbinolicus-M. barkeri cocultures did not aggregate, demonstrating that, unlike DIET, close physical contact was not necessary for interspecies H2 transfer. M. barkeri is the second methanogen found to accept electrons via DIET and the first methanogen known to be capable of using either H2 or electrons derived from DIET for CO2 reduction. Furthermore, M. barkeri is genetically tractable,making it a model organism for elucidating mechanisms by which methanogens make biological electrical connections with other cells.

}, keywords = {Biological Transport, Electron Transport, Ethanol, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Hydrogen, Methane, Methanosarcina barkeri}, issn = {1098-5336}, doi = {10.1128/AEM.00895-14}, author = {Rotaru, Amelia-Elena and Shrestha, Pravin Malla and Liu, Fanghua and Markovaite, Beatrice and Chen, Shanshan and Nevin, Kelly P and Lovley, Derek R} } @article {3129, title = {The Dnmt2 RNA methyltransferase homolog of Geobacter sulfurreducens specifically methylates tRNA-Glu.}, journal = {Nucleic Acids Res}, volume = {42}, year = {2014}, month = {2014 Jun}, pages = {6487-96}, abstract = {

Dnmt2 enzymes are conserved in eukaryotes, where they methylate C38 of tRNA-Asp with high activity. Here, the activity of one of the very few prokaryotic Dnmt2 homologs from Geobacter species (GsDnmt2) was investigated. GsDnmt2 was observed to methylate tRNA-Asp from flies and mice. Unexpectedly, it had only a weak activity toward its matching Geobacter tRNA-Asp, but methylated Geobacter tRNA-Glu with good activity. In agreement with this result, we show that tRNA-Glu is methylated in Geobacter while the methylation is absent in tRNA-Asp. The activities of Dnmt2 enzymes from Homo sapiens, Drosophila melanogaster, Schizosaccharomyces pombe and Dictyostelium discoideum for methylation of the Geobacter tRNA-Asp and tRNA-Glu were determined showing that all these Dnmt2s preferentially methylate tRNA-Asp. Hence, the GsDnmt2 enzyme has a swapped transfer ribonucleic acid (tRNA) specificity. By comparing the different tRNAs, a characteristic sequence pattern was identified in the variable loop of all preferred tRNA substrates. An exchange of two nucleotides in the variable loop of murine tRNA-Asp converted it to the corresponding variable loop of tRNA-Glu and led to a strong reduction of GsDnmt2 activity. Interestingly, the same loss of activity was observed with human DNMT2, indicating that the variable loop functions as a specificity determinant in tRNA recognition of Dnmt2 enzymes.

}, keywords = {Animals, Bacterial Proteins, Geobacter, Humans, Methylation, Mice, Nucleic Acid Conformation, RNA, Transfer, Asp, RNA, Transfer, Glu, Substrate Specificity, tRNA Methyltransferases}, issn = {1362-4962}, doi = {10.1093/nar/gku256}, author = {Shanmugam, Raghuvaran and Aklujkar, Muktak and Sch{\"a}fer, Matthias and Reinhardt, Richard and Nickel, Olaf and Reuter, Gunter and Lovley, Derek R and Ehrenhofer-Murray, Ann and Nellen, Wolfgang and Ankri, Serge and Helm, Mark and Jurkowski, Tomasz P and Jeltsch, Albert} } @article {1445, title = {(D)-Amino acid chemical reporters reveal peptidoglycan dynamics of an intracellular pathogen.}, journal = {ACS Chem Biol}, volume = {8}, year = {2013}, month = {2013 Mar 15}, pages = {500-5}, abstract = {

Peptidoglycan (PG) is an essential component of the bacterial cell wall. Although experiments with organisms in vitro have yielded a wealth of information on PG synthesis and maturation, it is unclear how these studies translate to bacteria replicating within host cells. We report a chemical approach for probing PG in vivo via metabolic labeling and bioorthogonal chemistry. A wide variety of bacterial species incorporated azide and alkyne-functionalized d-alanine into their cell walls, which we visualized by covalent reaction with click chemistry probes. The d-alanine analogues were specifically incorporated into nascent PG of the intracellular pathogen Listeria monocytogenes both in vitro and during macrophage infection. Metabolic incorporation of d-alanine derivatives and click chemistry detection constitute a facile, modular platform that facilitates unprecedented spatial and temporal resolution of PG dynamics in vivo.

}, keywords = {Alanine, Click Chemistry, Listeria monocytogenes, Molecular Conformation, Molecular Dynamics Simulation, Peptidoglycan}, issn = {1554-8937}, doi = {10.1021/cb3004995}, author = {Siegrist, M Sloan and Whiteside, Sarah and Jewett, John C and Aditham, Arjun and Cava, Felipe and Bertozzi, Carolyn R} } @article {727, title = {Desiccation induces viable but Non-Culturable cells in Sinorhizobium meliloti 1021.}, journal = {AMB Express}, volume = {2}, year = {2012}, month = {2012}, pages = {6}, abstract = {ABSTRACT: Sinorhizobium meliloti is a microorganism commercially used in the production of e.g. Medicago sativa seed inocula. Many inocula are powder-based and production includes a drying step. Although S. meliloti survives drying well, the quality of the inocula is reduced during this process. In this study we determined survival during desiccation of the commercial strains 102F84 and 102F85 as well as the model strain USDA1021.The survival of S. meliloti 1021 was estimated during nine weeks at 22\% relative humidity. We found that after an initial rapid decline of colony forming units, the decline slowed to a steady 10-fold reduction in colony forming units every 22 days. In spite of the reduction in colony forming units, the fraction of the population identified as viable (42-54\%) based on the Baclight live/dead stain did not change significantly over time. This change in the ability of viable cells to form colonies shows (i) an underestimation of the survival of rhizobial cells using plating methods, and that (ii) in a part of the population desiccation induces a Viable But Non Culturable (VBNC)-like state, which has not been reported before. Resuscitation attempts did not lead to a higher recovery of colony forming units indicating the VBNC state is stable under the conditions tested. This observation has important consequences for the use of rhizobia. Finding methods to resuscitate this fraction may increase the quality of powder-based seed inocula.}, issn = {2191-0855}, doi = {10.1186/2191-0855-2-6}, author = {Vriezen, Jan Ac and de Bruijn, Frans J and N{\"u}sslein, Klaus R} } @article {410, title = {The design of long-term effective uranium bioremediation strategy using a community metabolic model.}, journal = {Biotechnol Bioeng}, volume = {109}, year = {2012}, month = {2012 Oct}, pages = {2475-83}, abstract = {Acetate amendment at uranium contaminated sites in Rifle, CO. leads to an initial bloom of Geobacter accompanied by the removal of U(VI) from the groundwater, followed by an increase of sulfate-reducing bacteria (SRBs) which are poor reducers of U(VI). One of the challenges associated with bioremediation is the decay in Geobacter abundance, which has been attributed to the depletion of bio-accessible Fe(III), motivating the investigation of simultaneous amendments of acetate and Fe(III) as an alternative bioremediation strategy. In order to understand the community metabolism of Geobacter and SRBs during artificial substrate amendment, we have created a genome-scale dynamic community model of Geobacter and SRBs using the previously described Dynamic Multi-species Metabolic Modeling framework. Optimization techniques are used to determine the optimal acetate and Fe(III) addition profile. Field-scale simulation of acetate addition accurately predicted the in situ data. The simulations suggest that batch amendment of Fe(III) along with continuous acetate addition is insufficient to promote long-term bioremediation, while continuous amendment of Fe(III) along with continuous acetate addition is sufficient to promote long-term bioremediation. By computationally minimizing the acetate and Fe(III) addition rates as well as the difference between the predicted and target uranium concentration, we showed that it is possible to maintain the uranium concentration below the environmental safety standard while minimizing the cost of chemical additions. These simulations show that simultaneous addition of acetate and Fe(III) has the potential to be an effective uranium bioremediation strategy. They also show that computational modeling of microbial community is an important tool to design effective strategies for practical applications in environmental biotechnology. Biotechnol. Bioeng. 2012; 109: 2475-2483. {\textcopyright} 2012 Wiley Periodicals, Inc.}, issn = {1097-0290}, doi = {10.1002/bit.24528}, author = {Zhuang, K and Ma, E and Lovley, Derek R and Mahadevan, Radhakrishnan} } @article {720, title = {Disassembly of simian virus 40 during passage through the endoplasmic reticulum and in the cytoplasm.}, journal = {J Virol}, volume = {86}, year = {2012}, month = {2012 Feb}, pages = {1555-62}, abstract = {The nonenveloped polyomavirus simian virus 40 (SV40) is taken up into cells by a caveola-mediated endocytic process that delivers the virus to the endoplasmic reticulum (ER). Within the ER lumen, the capsid undergoes partial disassembly, which exposes its internal capsid proteins VP2 and VP3 to immunostaining with antibodies. We demonstrate here that the SV40 genome does not become accessible to detection while the virus is in the ER. Instead, the genome becomes accessible two distinct detection procedures, one using anti-bromodeoxyuridine antibodies and the other using a 5-ethynyl-2-deoxyuridine-based chemical reaction, only after the emergence of partially disassembled SV40 particles in the cytoplasm. These cytoplasmic particles retain some of the SV40 capsid proteins, VP1, VP2, and VP3, in addition to the viral genome. Thus, SV40 particles undergo discrete disassembly steps during entry that are separated temporally and topologically. First, a partial disassembly of the particles occurs in the ER, which exposes internal capsid proteins VP2 and VP3. Then, in the cytoplasm, disassembly progresses further to also make the genomic DNA accessible to immune detection.}, keywords = {Animals, Cell Line, Cercopithecus aethiops, Cytoplasm, Endoplasmic Reticulum, Genome, Viral, Microscopy, Fluorescence, Simian virus 40, Virus Assembly}, issn = {1098-5514}, doi = {10.1128/JVI.05753-11}, author = {Kuksin, Dmitry and Norkin, Leonard C} } @article {721, title = {Disassociation of the SV40 Genome from Capsid Proteins Prior to Nuclear Entry.}, journal = {Virol J}, volume = {9}, year = {2012}, month = {2012 Aug 10}, pages = {158}, abstract = {ABSTRACT: BACKGROUND: Previously, we demonstrated that input SV40 particles undergo a partial disassembly in the endoplasmic reticulum, which exposes internal capsid proteins VP2 and VP3 to immunostaining. Then, in the cytoplasm, disassembly progresses further to also make the genomic DNA accessible to immune detection, as well as to detection by an ethynyl-2-deoxyuridine (EdU)-based chemical reaction. The cytoplasmic partially disassembled SV40 particles retain some of the SV40 capsid proteins, VP1, VP2, and VP3, in addition to the viral genome. FINDINGS: In the current study, we asked where in the cell the SV40 genome might disassociate from capsid components. We observed partially disassembled input SV40 particles around the nucleus and, beginning at 12 hours post-infection, 5-Bromo-2-deoxyuridine (BrdU)-labeled parental SV40 DNA in the nucleus, as detected using anti-BrdU antibodies. However, among the more than 1500 cells examined, we never detected input VP2/VP3 in the nucleus. Upon translocation of the BrdU-labeled SV40 genomes into nuclei, they were transcribed and, thus, are representative of productive infection CONCLUSIONS: Our findings imply that the SV40 genome disassociates from the capsid proteins before or at the point of entry into the nucleus, and then enters the nucleus devoid of VP2/3..}, issn = {1743-422X}, doi = {10.1186/1743-422X-9-158}, author = {Kuksin, Dmitry and Norkin, Leonard C} } @article {379, title = {Dynamic localization of Trypanosoma brucei mitochondrial DNA polymerase ID.}, journal = {Eukaryot Cell}, volume = {11}, year = {2012}, month = {2012 Jul}, pages = {844-55}, abstract = {Trypanosomes contain a unique form of mitochondrial DNA called kinetoplast DNA (kDNA) that is a catenated network composed of minicircles and maxicircles. Several proteins are essential for network replication, and most of these localize to the antipodal sites or the kinetoflagellar zone. Essential components for kDNA synthesis include three mitochondrial DNA polymerases TbPOLIB, TbPOLIC, and TbPOLID). In contrast to other kDNA replication proteins, TbPOLID was previously reported to localize throughout the mitochondrial matrix. This spatial distribution suggests that TbPOLID requires redistribution to engage in kDNA replication. Here, we characterize the subcellular distribution of TbPOLID with respect to the Trypanosoma brucei cell cycle using immunofluorescence microscopy. Our analyses demonstrate that in addition to the previously reported matrix localization, TbPOLID was detected as discrete foci near the kDNA. TbPOLID foci colocalized with replicating minicircles at antipodal sites in a specific subset of the cells during stages II and III of kDNA replication. Additionally, the TbPOLID foci were stable following the inhibition of protein synthesis, detergent extraction, and DNase treatment. Taken together, these data demonstrate that TbPOLID has a dynamic localization that allows it to be spatially and temporally available to perform its role in kDNA replication.}, issn = {1535-9786}, doi = {10.1128/EC.05291-11}, author = {Concepci{\'o}n-Acevedo, Jeniffer and Luo, Juemin and Klingbeil, Michele M} } @article {426, title = {Development of a biomarker for Geobacter activity and strain composition; proteogenomic analysis of the citrate synthase protein during bioremediation of U(VI).}, journal = {Microb Biotechnol}, volume = {4}, year = {2011}, month = {2011 Jan}, pages = {55-63}, abstract = {Monitoring the activity of target microorganisms during stimulated bioremediation is a key problem for the development of effective remediation strategies. At the US Department of Energy{\textquoteright}s Integrated Field Research Challenge (IFRC) site in Rifle, CO, the stimulation of Geobacter growth and activity via subsurface acetate addition leads to precipitation of U(VI) from groundwater as U(IV). Citrate synthase (gltA) is a key enzyme in Geobacter central metabolism that controls flux into the TCA cycle. Here, we utilize shotgun proteomic methods to demonstrate that the measurement of gltA peptides can be used to track Geobacter activity and strain evolution during in situ biostimulation. Abundances of conserved gltA peptides tracked Fe(III) reduction and changes in U(VI) concentrations during biostimulation, whereas changing patterns of unique peptide abundances between samples suggested sample-specific strain shifts within the Geobacter population. Abundances of unique peptides indicated potential differences at the strain level between Fe(III)-reducing populations stimulated during in situ biostimulation experiments conducted a year apart at the Rifle IFRC. These results offer a novel technique for the rapid screening of large numbers of proteomic samples for Geobacter species and will aid monitoring of subsurface bioremediation efforts that rely on metal reduction for desired outcomes.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Biodegradation, Environmental, Biological Markers, Citrate (si)-Synthase, Geobacter, Groundwater, Molecular Sequence Data, Phylogeny, Proteomics, Sequence Alignment, Uranium}, issn = {1751-7915}, doi = {10.1111/j.1751-7915.2010.00194.x}, author = {Wilkins, Michael J and Callister, Stephen J and Miletto, Marzia and Williams, Kenneth H and Nicora, Carrie D and Lovley, Derek R and Long, Philip E and Lipton, Mary S} } @article {429, title = {Direct coupling of a genome-scale microbial in silico model and a groundwater reactive transport model.}, journal = {J Contam Hydrol}, volume = {122}, year = {2011}, month = {2011 Mar 25}, pages = {96-103}, abstract = {The activity of microorganisms often plays an important role in dynamic natural attenuation or engineered bioremediation of subsurface contaminants, such as chlorinated solvents, metals, and radionuclides. To evaluate and/or design bioremediated systems, quantitative reactive transport models are needed. State-of-the-art reactive transport models often ignore the microbial effects or simulate the microbial effects with static growth yield and constant reaction rate parameters over simulated conditions, while in reality microorganisms can dynamically modify their functionality (such as utilization of alternative respiratory pathways) in response to spatial and temporal variations in environmental conditions. Constraint-based genome-scale microbial in silico models, using genomic data and multiple-pathway reaction networks, have been shown to be able to simulate transient metabolism of some well studied microorganisms and identify growth rate, substrate uptake rates, and byproduct rates under different growth conditions. These rates can be identified and used to replace specific microbially-mediated reaction rates in a reactive transport model using local geochemical conditions as constraints. We previously demonstrated the potential utility of integrating a constraint-based microbial metabolism model with a reactive transport simulator as applied to bioremediation of uranium in groundwater. However, that work relied on an indirect coupling approach that was effective for initial demonstration but may not be extensible to more complex problems that are of significant interest (e.g., communities of microbial species and multiple constraining variables). Here, we extend that work by presenting and demonstrating a method of directly integrating a reactive transport model (FORTRAN code) with constraint-based in silico models solved with IBM ILOG CPLEX linear optimizer base system (C library). The models were integrated with BABEL, a language interoperability tool. The modeling system is designed in such a way that constraint-based models targeting different microorganisms or competing organism communities can be easily plugged into the system. Constraint-based modeling is very costly given the size of a genome-scale reaction network. To save computation time, a binary tree is traversed to examine the concentration and solution pool generated during the simulation in order to decide whether the constraint-based model should be called. We also show preliminary results from the integrated model including a comparison of the direct and indirect coupling approaches and evaluated the ability of the approach to simulate field experiment.}, keywords = {Biodegradation, Environmental, Biological Transport, Colorado, Computer Simulation, Genome, Bacterial, Geobacter, Models, Biological, Uranium}, issn = {1873-6009}, doi = {10.1016/j.jconhyd.2010.11.007}, author = {Fang, Yilin and Scheibe, Timothy D and Mahadevan, Radhakrishnan and Garg, Srinath and Long, Philip E and Lovley, Derek R} } @article {443, title = {De Novo assembly of the complete genome of an enhanced electricity-producing variant of Geobacter sulfurreducens using only short reads.}, journal = {PLoS One}, volume = {5}, year = {2010}, month = {2010}, pages = {e10922}, abstract = {State-of-the-art DNA sequencing technologies are transforming the life sciences due to their ability to generate nucleotide sequence information with a speed and quantity that is unapproachable with traditional Sanger sequencing. Genome sequencing is a principal application of this technology, where the ultimate goal is the full and complete sequence of the organism of interest. Due to the nature of the raw data produced by these technologies, a full genomic sequence attained without the aid of Sanger sequencing has yet to be demonstrated.We have successfully developed a four-phase strategy for using only next-generation sequencing technologies (Illumina and 454) to assemble a complete microbial genome de novo. We applied this approach to completely assemble the 3.7 Mb genome of a rare Geobacter variant (KN400) that is capable of unprecedented current production at an electrode. Two key components of our strategy enabled us to achieve this result. First, we integrated the two data types early in the process to maximally leverage their complementary characteristics. And second, we used the output of different short read assembly programs in such a way so as to leverage the complementary nature of their different underlying algorithms or of their different implementations of the same underlying algorithm.The significance of our result is that it demonstrates a general approach for maximizing the efficiency and success of genome assembly projects as new sequencing technologies and new assembly algorithms are introduced. The general approach is a meta strategy, wherein sequencing data are integrated as early as possible and in particular ways and wherein multiple assembly algorithms are judiciously applied such that the deficiencies in one are complemented by another.}, keywords = {Algorithms, Electricity, Genome, Bacterial, Geobacter, Polymerase Chain Reaction}, issn = {1932-6203}, doi = {10.1371/journal.pone.0010922}, author = {Nagarajan, Harish and Butler, Jessica E and Klimes, Anna and Qiu, Yu and Zengler, Karsten and Ward, Joy and Young, Nelson D and Meth{\'e}, Barbara A and Palsson, Bernhard {\O} and Lovley, Derek R and Barrett, Christian L} } @article {31, title = {Derepression of INO1 transcription requires cooperation between the Ino2p-Ino4p heterodimer and Cbf1p and recruitment of the ISW2 chromatin-remodeling complex.}, journal = {Eukaryot Cell}, volume = {9}, year = {2010}, month = {2010 Dec}, pages = {1845-55}, abstract = {

The Saccharomyces cerevisiae INO1 gene encodes the structural enzyme inositol-3-phosphate synthase for the synthesis de novo of inositol and inositol-containing phospholipids. The transcription of INO1 is completely derepressed in the absence of inositol and choline (I(-) C(-)). Derepression requires the binding of the Ino2p-Ino4p basic helix-loop-helix (bHLH) heterodimer to the UAS(INO) promoter element. We report here the requirement of a third bHLH protein, centromere-binding factor 1 (Cbf1p), for the complete derepression of INO1 transcription. We found that Cbf1p regulates INO1 transcription by binding to sites distal to the INO1 promoter and encompassing the upstream SNA3 open reading frame (ORF) and promoter. The binding of Cbf1p requires Ino2p-Ino4p binding to the UAS(INO) sites in the INO1 promoter and vice versa, suggesting a cooperative mechanism. Furthermore, Cbf1p binding to the upstream sites was required for the binding of the ISW2 chromatin-remodeling complex to the Ino2p-Ino4p-binding sites on the INO1 promoter. Consistent with this, ISW2 was also required for the complete derepression of INO1 transcription.

}, keywords = {Adenosine Triphosphatases, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors, Basic Helix-Loop-Helix Transcription Factors, Chromatin, Chromatin Assembly and Disassembly, Dimerization, Gene Expression Regulation, Fungal, Myo-Inositol-1-Phosphate Synthase, Promoter Regions, Genetic, Protein Binding, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic}, issn = {1535-9786}, doi = {10.1128/EC.00144-10}, author = {Shetty, Ameet and Lopes, John M} } @article {431, title = {Direct exchange of electrons within aggregates of an evolved syntrophic coculture of anaerobic bacteria.}, journal = {Science}, volume = {330}, year = {2010}, month = {2010 Dec 3}, pages = {1413-5}, abstract = {Microbial consortia that cooperatively exchange electrons play a key role in the anaerobic processing of organic matter. Interspecies hydrogen transfer is a well-documented strategy for electron exchange in dispersed laboratory cultures, but cooperative partners in natural environments often form multispecies aggregates. We found that laboratory evolution of a coculture of Geobacter metallireducens and Geobacter sulfurreducens metabolizing ethanol favored the formation of aggregates that were electrically conductive. Sequencing aggregate DNA revealed selection for a mutation that enhances the production of a c-type cytochrome involved in extracellular electron transfer and accelerates the formation of aggregates. Aggregate formation was also much faster in mutants that were deficient in interspecies hydrogen transfer, further suggesting direct interspecies electron transfer.}, keywords = {Anaerobiosis, Bacterial Proteins, Biological Evolution, Culture Media, Cytochrome c Group, Electron Transport, Electrons, Ethanol, Fimbriae Proteins, Geobacter, Hydrogen, Microbial Consortia, Microbial Interactions, Mutation, Oxidation-Reduction, Selection, Genetic}, issn = {1095-9203}, doi = {10.1126/science.1196526}, author = {Summers, Zarath M and Fogarty, Heather E and Leang, Ching and Franks, Ashley E and Malvankar, Nikhil S 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 {473, title = {Diversity of promoter elements in a Geobacter sulfurreducens mutant adapted to disruption in electron transfer.}, journal = {Funct Integr Genomics}, volume = {9}, year = {2009}, month = {2009 Feb}, pages = {15-25}, abstract = {The delta-proteobacterium, Geobacter sulfurreducens, can obtain energy by coupling the oxidation of organic matter to the reduction of insoluble Fe(III) or the anode of a microbial fuel cell. Because Fe(III) oxide or the anode surface, in contrast to oxygen, nitrate, or sulfate, is not soluble nor can it be reduced readily, Geobacter species have developed mechanisms which allow electrons to be delivered across outer membrane to the cell surface. OmcB is an outer-membrane c-type cytochrome important for G. sulfurreducens Fe(III) respiration. In the absence of OmcB, cells lost the ability to reduce soluble or insoluble Fe(III). However, the omcB deletion mutant can slowly adapt to growth on soluble Fe(III) over prolonged incubation in the medium with acetate as the electron donor. We discuss available information about predicted or experimentally validated promoters and transcription regulatory sites identified upstream of operons with transcriptional expression significantly changed in the adapted omcB mutant. DNA sequences of upstream regions of coregulated operons in the adapted mutant are divergent, suggesting the presence of recognition sites for different transcriptional regulators and indicating that adaptation of the omcB mutant to growth on soluble Fe(III) has shifted the relevant expression networks involved to a more diverse molecular basis.}, keywords = {Adaptation, Physiological, Electron Transport, Genetic Variation, Geobacter, Mutation, Promoter Regions, Genetic}, issn = {1438-7948}, doi = {10.1007/s10142-008-0094-7}, author = {Krushkal, Julia and Leang, Ching and Barbe, Jose F and Qu, Yanhua and Yan, Bin and Puljic, Marko and Adkins, Ronald M and Lovley, Derek R} } @article {734, title = {"Doubly selective" antimicrobial polymers: how do they differentiate between bacteria?}, journal = {Chemistry}, volume = {15}, year = {2009}, month = {2009 Nov 2}, pages = {11710-4}, abstract = {We have investigated how doubly selective synthetic mimics of antimicrobial peptides (SMAMPs), which can differentiate not only between bacteria and mammalian cells, but also between Gram-negative and Gram-positive bacteria, make the latter distinction. By dye-leakage experiments on model vesicles and complementary experiments on bacteria, we were able to relate the Gram selectivity to structural differences of these bacteria types. We showed that the double membrane of E. coli rather than the difference in lipid composition between E. coli and S. aureus was responsible for Gram selectivity. The molecular-weight-dependent antimicrobial activity of the SMAMPs was shown to be a sieving effect: while the 3000 g mol(-1) SMAMP was able to penetrate the peptidoglycan layer of the Gram-positive S. aureus bacteria, the 50000 g mol(-1) SMAMP got stuck and consequently did not have antimicrobial activity.}, keywords = {Anti-Infective Agents, Biomimetic Materials, Escherichia coli, Microbial Sensitivity Tests, Peptidoglycan, Polymers, Staphylococcus aureus}, issn = {1521-3765}, doi = {10.1002/chem.200802558}, author = {Lienkamp, Karen and Kumar, Kushi-Nidhi and Som, Abhigyan and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {395, title = {A degenerate tripartite DNA-binding site required for activation of ComA-dependent quorum response gene expression in Bacillus subtilis.}, journal = {J Mol Biol}, volume = {381}, year = {2008}, month = {2008 Aug 29}, pages = {261-75}, abstract = {In Bacillus subtilis, the transcription factor ComA activates several biological processes in response to increasing population density. Extracellular peptide signaling is used to coordinate the activity of ComA with population density. At low culture densities, when the concentration of signaling peptides is lowest, ComA is largely inactive. At higher densities, when the concentration of signaling peptides is higher, ComA is active and activates the transcription of at least nine operons involved in the development of competence and in the production of degradative enzymes and antibiotics. We found that ComA binds a degenerate tripartite sequence consisting of three DNA-binding determinants or "recognition elements." Mutational analyses showed that all three recognition elements are required for transcription activation in vivo and for specific DNA binding by ComA in vitro. Degeneracy of the recognition elements in the ComA-binding site is an important regulatory feature for coordinating transcription with population density (i.e., promoters containing an optimized binding site have high activity at low culture density and are no longer regulated in the normal-density-dependent manner). We found that purified ComA forms a dimer in solution, and we propose a model for how two dimers of ComA bind to an odd number of DNA-binding determinants to activate transcription of target genes. This DNA-protein architecture for transcription activation appears to be conserved for ComA homologs in other Bacillus species.}, keywords = {Bacillus subtilis, Bacterial Proteins, Base Sequence, Binding Sites, Dimerization, DNA, Bacterial, DNA-Binding Proteins, Electrophoretic Mobility Shift Assay, Gene Expression Regulation, Bacterial, Peptide Synthases, Phosphoprotein Phosphatases, Promoter Regions, Genetic, Protein Binding, Quorum Sensing, Sequence Analysis, DNA, Transcription, Genetic, Transcriptional Activation}, issn = {1089-8638}, doi = {10.1016/j.jmb.2008.06.035}, author = {Griffith, Kevin L and Grossman, Alan D} } @article {498, title = {The degree of redundancy in metabolic genes is linked to mode of metabolism.}, journal = {Biophys J}, volume = {94}, year = {2008}, month = {2008 Feb 15}, pages = {1216-20}, abstract = {An understanding of the factors favoring the maintenance of duplicate genes in microbial genomes is essential for developing models of microbial evolution. A genome-scale flux-balance analysis of the metabolic network of Saccharomyces cerevisiae has suggested that gene duplications primarily provide increased enzyme dosage to enhance metabolic flux because the incidence of gene duplications in essential genes is no higher than that in nonessential genes. Here, we used genome-scale metabolic models to analyze the extent of genetic and biochemical redundancy in prokaryotes that are either specialists, with one major mode of energy generation, or generalists, which have multiple metabolic strategies for conservation of energy. Surprisingly, the results suggest that generalists, such as Escherichia coli and Bacillus subtilis, are similar to the eukaryotic generalist, S. cerevisiae, in having a low percentage (<10\%) of essential genes and few duplications of these essential genes, whereas metabolic specialists, such as Geobacter sulfurreducens and Methanosarcina barkeri, have a high percentage (>30\%) of essential genes and a high degree of genetic redundancy in these genes compared to nonessential genes. Furthermore, the specialist organisms appear to rely more on gene duplications rather than alternative-but-equivalent metabolic pathways to provide resilience to gene loss. Generalists rely more on alternative pathways. Thus, the concept that the role of gene duplications is to boost enzymatic flux rather than provide metabolic resilience may not be universal. Rather, the degree of gene duplication in microorganisms may be linked to mode of metabolism and environmental niche.}, keywords = {Bacterial Physiological Phenomena, Bacterial Proteins, Computer Simulation, Gene Expression Regulation, Bacterial, Genes, Duplicate, Models, Biological, Signal Transduction}, issn = {1542-0086}, doi = {10.1529/biophysj.107.118414}, author = {Mahadevan, R and Lovley, D R} } @article {810, title = {Differential requirements of two recA mutants for constitutive SOS expression in Escherichia coli K-12.}, journal = {PLoS One}, volume = {3}, year = {2008}, month = {2008}, pages = {e4100}, abstract = {BACKGROUND: Repairing DNA damage begins with its detection and is often followed by elicitation of a cellular response. In E. coli, RecA polymerizes on ssDNA produced after DNA damage and induces the SOS Response. The RecA-DNA filament is an allosteric effector of LexA auto-proteolysis. LexA is the repressor of the SOS Response. Not all RecA-DNA filaments, however, lead to an SOS Response. Certain recA mutants express the SOS Response (recA(C)) in the absence of external DNA damage in log phase cells. METHODOLOGY/PRINCIPAL FINDINGS: Genetic analysis of two recA(C) mutants was used to determine the mechanism of constitutive SOS (SOS(C)) expression in a population of log phase cells using fluorescence of single cells carrying an SOS reporter system (sulAp-gfp). SOS(C) expression in recA4142 mutants was dependent on its initial level of transcription, recBCD, recFOR, recX, dinI, xthA and the type of medium in which the cells were grown. SOS(C) expression in recA730 mutants was affected by none of the mutations or conditions tested above. CONCLUSIONS/SIGNIFICANCE: It is concluded that not all recA(C) alleles cause SOS(C) expression by the same mechanism. It is hypothesized that RecA4142 is loaded on to a double-strand end of DNA and that the RecA filament is stabilized by the presence of DinI and destabilized by RecX. RecFOR regulate the activity of RecX to destabilize the RecA filament. RecA730 causes SOS(C) expression by binding to ssDNA in a mechanism yet to be determined.}, keywords = {Escherichia coli K12, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Models, Biological, Mutation, Rec A Recombinases, SOS Response (Genetics)}, issn = {1932-6203}, doi = {10.1371/journal.pone.0004100}, author = {Long, Jarukit Edward and Renzette, Nicholas and Centore, Richard C and Sandler, Steven J} } @article {818, title = {DinI and RecX modulate RecA-DNA structures in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {63}, year = {2007}, month = {2007 Jan}, pages = {103-15}, abstract = {RecA plays a central role in recombination, DNA repair and SOS induction through forming a RecA-DNA helical filament. Biochemical observations show that at low ratios to RecA, DinI and RecX stabilize and destabilize RecA-DNA filaments, respectively, and that the C-terminal 17 residues of RecA are important for RecX function. RecA-DNA filament formation was assayed in vivo using RecA-GFP foci formation in log-phase and UV-irradiated cells. In log-phase cells, dinI mutants have fewer foci than wild type and that recX mutants have more foci than wild type. A recADelta17::gfp mutant had more foci like a recX mutant. dinI recX double mutants have the same number of foci as dinI mutants alone, suggesting that dinI is epistatic to recX. After UV treatment, the dinI, recX and dinI recX mutants differed in their ability to form foci. All three mutants had fewer foci than wild type. The dinI mutant{\textquoteright}s foci persisted longer than wild-type foci. Roles of DinI and RecX after UV treatment differed from those during log-phase growth and may reflect the different DNA substrates, population of proteins or amounts during the SOS response. These experiments give new insight into the roles of these proteins.}, keywords = {Bacterial Proteins, DNA, Bacterial, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Bacterial, Rec A Recombinases}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2006.05496.x}, author = {Renzette, Nicholas and Gumlaw, Nathan and Sandler, Steven J} } @article {749, title = {Desiccation responses and survival of Sinorhizobium meliloti USDA 1021 in relation to growth phase, temperature, chloride and sulfate availability.}, journal = {Lett Appl Microbiol}, volume = {42}, year = {2006}, month = {2006 Feb}, pages = {172-8}, abstract = {AIMS: To identify physical and physiological conditions that affect the survival of Sinorhizobium meliloti USDA 1021 during desiccation. METHODS AND RESULTS: An assay was developed to study desiccation response of S. meliloti USDA 1021 over a range of environmental conditions. We determined the survival during desiccation in relation to (i) matrices and media, (ii) growth phase, (iii) temperature, and (iv) chloride and sulfate availability. CONCLUSIONS: This study indicates that survival of S. meliloti USDA 1021 during desiccation is enhanced: (i) when cells were dried in the stationary phase, (ii) with increasing drying temperature at an optimum of 37 degrees C, and (iii) during an increase of chloride and sulfate, but not sodium or potassium availability. In addition, we resolved that the best matrix to test survival was nitrocellulose filters. SIGNIFICANCE AND IMPACT OF THE STUDY: The identification of physical and physiological factors that determine the survival during desiccation of S. meliloti USDA 1021 may aid in (i) the strategic development of improved seed inocula, (ii) the isolation, and (iii) the development of rhizobial strains with improved ability to survive desiccation. Furthermore, this work may provide insights into the survival of rhizobia under drought conditions.}, keywords = {Bacteriological Techniques, Chlorides, Desiccation, Models, Biological, Sinorhizobium meliloti, Sulfur Oxides, Temperature}, issn = {0266-8254}, doi = {10.1111/j.1472-765X.2005.01808.x}, author = {Vriezen, J A C and de Bruijn, F J and N{\"u}sslein, K} } @article {855, title = {Detection of Chlamydia in the peripheral blood cells of normal donors using in vitro culture, immunofluorescence microscopy and flow cytometry techniques.}, journal = {BMC Infect Dis}, volume = {6}, year = {2006}, month = {2006}, pages = {23}, abstract = {BACKGROUND: Chlamydia trachomatis (Ct) and Chlamydia pneumoniae (Cp) are medically significant infectious agents associated with various chronic human pathologies. Nevertheless, specific roles in disease progression or initiation are incompletely defined. Both pathogens infect established cell lines in vitro and polymerase chain reaction (PCR) has detected Chlamydia DNA in various clinical specimens as well as in normal donor peripheral blood monocytes (PBMC). However, Chlamydia infection of other blood cell types, quantification of Chlamydia infected cells in peripheral blood and transmission of this infection in vitro have not been examined. METHODS: Cp specific titers were assessed for sera from 459 normal human donor blood (NBD) samples. Isolated white blood cells (WBC) were assayed by in vitro culture to evaluate infection transmission of blood cell borne chlamydiae. Smears of fresh blood samples (FB) were dual immunostained for microscopic identification of Chlamydia-infected cell types and aliquots also assessed using Flow Cytometry (FC). RESULTS: ELISA demonstrated that 219 (47.7\%) of the NBD samples exhibit elevated anti-Cp antibody titers. Imunofluorescence microscopy of smears demonstrated 113 (24.6\%) of samples contained intracellular Chlamydia and monoclonals to specific CD markers showed that in vivo infection of neutrophil and eosinophil/basophil cells as well as monocytes occurs. In vitro culture established WBCs of 114 (24.8\%) of the NBD samples harbored infectious chlamydiae, clinically a potentially source of transmission, FC demonstrated both Chlamydia infected and uninfected cells can be readily identified and quantified. CONCLUSION: NBD can harbor infected neutrophils, eosinophil/basophils and monocytes. The chlamydiae are infectious in vitro, and both total, and cell type specific Chlamydia carriage is quantifiable by FC.}, keywords = {Adult, Blood Donors, Chlamydophila pneumoniae, Female, Flow Cytometry, Humans, Leukocytes, Male, Microscopy, Fluorescence}, issn = {1471-2334}, doi = {10.1186/1471-2334-6-23}, author = {Cirino, Frances and Webley, Wilmore C and West, Corrie and Croteau, Nancy L and Andrzejewski, Chester and Stuart, Elizabeth S} } @article {702, title = {Differential protein expression in the metal-reducing bacterium Geobacter sulfurreducens strain PCA grown with fumarate or ferric citrate.}, journal = {Proteomics}, volume = {6}, year = {2006}, month = {2006 Jan}, pages = {632-40}, abstract = {Geobacter sulfurreducens, generally considered to be a strict anaerobe, is a predominant microbe in subsurface environments, where it utilizes available metals as electron acceptors. To better understand the metabolic processes involved in the metal-reduction capability of this microbe, the proteins expressed by cells grown anaerobically with either fumarate or ferric citrate as electron acceptor were compared. Proteins were separated by 2-DE under denaturing or nondenaturing conditions, and proteins varying in abundance with a high level of statistical significance (p<0.0001) were identified by peptide mass analysis. Denaturing 2-DE revealed significant differences in the relative abundance of the membrane proteins OmpA and peptidoglycan-associated lipoprotein, several metabolic enzymes, and, in addition, superoxide dismutase and rubredoxin oxidoreductase. Nondenaturing 2-DE revealed elevated catalase in cells grown with ferric citrate. These results suggest that, in addition to adjustments in membrane transport and specific metabolic pathways in response to these two different electron acceptors, distinct differences exist in the oxidative environment within the cell when fumarate or soluble ferric citrate is provided as electron acceptor. Although an anaerobe, G. sulfurreducens appears to have alternate mechanisms for dealing with reactive oxygen species in response to increased intracellular soluble iron.}, keywords = {Anaerobiosis, Bacterial Proteins, Electrophoresis, Gel, Two-Dimensional, Ferric Compounds, Fumarates, Geobacter, Oxidation-Reduction, Proteomics, Reactive Oxygen Species, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization}, issn = {1615-9853}, doi = {10.1002/pmic.200500137}, author = {Khare, Tripti and Esteve-N{\'u}{\~n}ez, Abraham and Nevin, Kelly P and Zhu, Wenhong and Yates, John R and Lovley, Derek and Giometti, Carol S} } @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 {522, title = {DNA microarray and proteomic analyses of the RpoS regulon in Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Apr}, pages = {2792-800}, abstract = {The regulon of the sigma factor RpoS was defined in Geobacter sulfurreducens by using a combination of DNA microarray expression profiles and proteomics. An rpoS mutant was examined under steady-state conditions with acetate as an electron donor and fumarate as an electron acceptor and with additional transcriptional profiling using Fe(III) as an electron acceptor. Expression analysis revealed that RpoS acts as both a positive and negative regulator. Many of the RpoS-dependent genes determined play roles in energy metabolism, including the tricarboxylic acid cycle, signal transduction, transport, protein synthesis and degradation, and amino acid metabolism and transport. As expected, RpoS activated genes involved in oxidative stress resistance and adaptation to nutrient limitation. Transcription of the cytochrome c oxidase operon, necessary for G. sulfurreducens growth using oxygen as an electron acceptor, and expression of at least 13 c-type cytochromes, including one previously shown to participate in Fe(III) reduction (MacA), were RpoS dependent. Analysis of a subset of the rpoS mutant proteome indicated that 15 major protein species showed reproducible differences in abundance relative to those of the wild-type strain. Protein identification using mass spectrometry indicated that the expression of seven of these proteins correlated with the microarray data. Collectively, these results indicate that RpoS exerts global effects on G. sulfurreducens physiology and that RpoS is vital to G. sulfurreducens survival under conditions typically encountered in its native subsurface environments.}, keywords = {Adaptation, Physiological, Amino Acids, Bacterial Proteins, Biological Transport, Citric Acid Cycle, Cytochromes, Electrophoresis, Gel, Two-Dimensional, Gene Deletion, Gene Expression Regulation, Bacterial, Geobacter, Mass Spectrometry, Mutagenesis, Insertional, Oligonucleotide Array Sequence Analysis, Oxidative Stress, Protein Biosynthesis, Proteome, Regulon, Sigma Factor, Signal Transduction}, issn = {0021-9193}, doi = {10.1128/JB.188.8.2792-2800.2006}, author = {N{\'u}{\~n}ez, Cinthia and Esteve-N{\'u}{\~n}ez, Abraham and Giometti, Carol and Tollaksen, Sandra and Khare, Tripti and Lin, Winston and Lovley, Derek R and Meth{\'e}, Barbara A} } @article {538, title = {DNA microarray analysis of nitrogen fixation and Fe(III) reduction in Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 May}, pages = {2530-8}, abstract = {A DNA microarray representing the genome of Geobacter sulfurreducens was constructed for use in global gene expression profiling of cells under steady-state conditions with acetate as the electron donor and Fe(III) or fumarate as the electron acceptor. Reproducible differences in transcript levels were also observed in comparisons between cells grown with ammonia and those fixing atmospheric nitrogen. There was a high correlation between changes in transcript levels determined with microarray analyses and an evaluation of a subset of the genome with quantitative PCR. As expected, cells required to fix nitrogen had higher levels of transcripts of genes associated with nitrogen fixation, further demonstrating that the microarray approach could reliably detect important physiological changes. Cells grown with Fe(III) as the electron acceptor had higher levels of transcripts for omcB, a gene coding for an outer membrane c-type cytochrome that is essential for Fe(III) reduction. Several other c-type cytochrome genes also appeared to be up-regulated. An unexpected result was significantly higher levels of transcripts for genes which have a role in metal efflux, potentially suggesting the importance of maintaining metal homeostasis during release of soluble metals when reducing Fe(III). A substantial proportion (30\%) of significantly expressed genes during Fe(III) reduction were genes of unknown function or hypothetical proteins, suggesting differences in Fe(III) reduction physiology among microorganisms which perform this metabolic process.}, keywords = {Ferric Compounds, Fumarates, Geobacter, Microarray Analysis, Nitrogen Fixation, Oxidation-Reduction, Polymerase Chain Reaction}, issn = {0099-2240}, doi = {10.1128/AEM.71.5.2530-2538.2005}, author = {Meth{\'e}, Barbara A and Webster, Jennifer and Nevin, Kelly and Butler, Jessica and Lovley, Derek R} } @article {548, title = {Direct correlation between rates of anaerobic respiration and levels of mRNA for key respiratory genes in Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Sep}, pages = {5183-9}, abstract = {The predominance of Geobacter species in environments in which Fe(III) reduction is important has suggested that Fe(III) reduction rates might be estimated in Geobacter-dominated environments by assessing in situ activity with molecular techniques. To determine whether mRNA levels of key respiratory genes might be correlated with respiration rates in Geobacter sulfurreducens, studies were conducted with fumarate as the electron acceptor and acetate as the limiting electron donor in anaerobic continuous cultures. Levels of mRNA for a fumarate reductase gene, frdA, quantified by real-time reverse transcription-PCR were directly correlated with fumarate reduction rates. In similar studies with Fe(III) as the electron acceptor, mRNA levels for omcB, a gene for an outer membrane c-type cytochrome involved in Fe(III) reduction, were positively correlated with Fe(III) reduction rates. Levels of mRNA for frdA and omcB were also positively correlated with fumarate and Fe(III) reduction rates, respectively, when growth was limited by the availability of fumarate or Fe(III), but mRNA levels were higher than in acetate-limited cultures. Levels of mRNA for omcC, which encodes a c-type cytochrome highly similar to OmcB but not necessary for Fe(III) reduction, followed patterns different than those of omcB. This agrees with the previous finding that OmcC is not involved in Fe(III) reduction and suggests that changes in mRNA levels of omcB are related to its role in Fe(III) reduction. These results demonstrate that mRNA levels for respiratory genes might be used to estimate in situ Fe(III) reduction rates in Geobacter-dominated environments but suggest that information on environmental conditions and/or the metabolic state of Geobacter species is also required for accurate rate estimates.}, keywords = {Anaerobiosis, Base Sequence, DNA Primers, Geobacter, Kinetics, Oxygen Consumption, Polymerase Chain Reaction, RNA, Messenger}, issn = {0099-2240}, doi = {10.1128/AEM.70.9.5183-5189.2004}, author = {Chin, Kuk-Jeong and Esteve-N{\'u}{\~n}ez, Abraham and Leang, Ching and Lovley, Derek R} } @article {545, title = {Dissimilatory Fe(III) and Mn(IV) reduction.}, journal = {Adv Microb Physiol}, volume = {49}, year = {2004}, month = {2004}, pages = {219-86}, abstract = {Dissimilatory Fe(III) and Mn(IV) reduction has an important influence on the geochemistry of modern environments, and Fe(III)-reducing microorganisms, most notably those in the Geobacteraceae family, can play an important role in the bioremediation of subsurface environments contaminated with organic or metal contaminants. Microorganisms with the capacity to conserve energy from Fe(III) and Mn(IV) reduction are phylogenetically dispersed throughout the Bacteria and Archaea. The ability to oxidize hydrogen with the reduction of Fe(III) is a highly conserved characteristic of hyperthermophilic microorganisms and one Fe(III)-reducing Archaea grows at the highest temperature yet recorded for any organism. Fe(III)- and Mn(IV)-reducing microorganisms have the ability to oxidize a wide variety of organic compounds, often completely to carbon dioxide. Typical alternative electron acceptors for Fe(III) reducers include oxygen, nitrate, U(VI) and electrodes. Unlike other commonly considered electron acceptors, Fe(III) and Mn(IV) oxides, the most prevalent form of Fe(III) and Mn(IV) in most environments, are insoluble. Thus, Fe(III)- and Mn(IV)-reducing microorganisms face the dilemma of how to transfer electrons derived from central metabolism onto an insoluble, extracellular electron acceptor. Although microbiological and geochemical evidence suggests that Fe(III) reduction may have been the first form of microbial respiration, the capacity for Fe(III) reduction appears to have evolved several times as phylogenetically distinct Fe(III) reducers have different mechanisms for Fe(III) reduction. Geobacter species, which are representative of the family of Fe(III) reducers that predominate in a wide diversity of sedimentary environments, require direct contact with Fe(III) oxides in order to reduce them. In contrast, Shewanella and Geothrix species produce chelators that solubilize Fe(III) and release electron-shuttling compounds that transfer electrons from the cell surface to the surface of Fe(III) oxides not in direct contact with the cells. Electron transfer from the inner membrane to the outer membrane in Geobacter and Shewanella species appears to involve an electron transport chain of inner-membrane, periplasmic, and outer-membrane c-type cytochromes, but the cytochromes involved in these processes in the two organisms are different. In addition, Geobacter species specifically express flagella and pili during growth on Fe(III) and Mn(IV) oxides and are chemotactic to Fe(II) and Mn(II), which may lead Geobacter species to the oxides under anoxic conditions. The physiological characteristics of Geobacter species appear to explain why they have consistently been found to be the predominant Fe(III)- and Mn(IV)-reducing microorganisms in a variety of sedimentary environments. In comparison with other respiratory processes, the study of Fe(III) and Mn(IV) reduction is in its infancy, but genome-enabled approaches are rapidly advancing our understanding of this environmentally significant physiology.}, keywords = {Archaea, Biodegradation, Environmental, Ferric Compounds, Geobacter, Manganese, Manganese Compounds, Oxidation-Reduction, Oxides, Shewanella, Soil Microbiology, Soil Pollutants}, issn = {0065-2911}, doi = {10.1016/S0065-2911(04)49005-5}, author = {Lovley, Derek R and Holmes, Dawn E and Nevin, Kelly P} } @article {824, title = {A dnaT mutant with phenotypes similar to those of a priA2::kan mutant in Escherichia coli K-12.}, journal = {Genetics}, volume = {167}, year = {2004}, month = {2004 Jun}, pages = {569-78}, abstract = {The ability to repair damaged replication forks and restart them is important for cell survival. DnaT is essential for replication restart in vitro and yet no definite genetic analysis has been done in Escherichia coli K-12. To begin, dnaT822, an in-frame six-codon (87-92) deletion was constructed. DnaT822 mutants show colony size, cell morphology, inability to properly partition nucleoids, UV sensitivity, and basal SOS expression similar to priA2::kan mutants. DnaT822 priA2::kan double mutants had phenotypes similar to those of the single mutants. DnaT822 and dnaT822 priA2::kan mutant phenotypes were fully suppressed by dnaC809. Previously, a dominant temperature-sensitive lethal mutation, dnaT1, had been isolated in E. coli 15T(-). DnaT1 was found to have a base-pair change relative to the E. coli 15T(-) and E. coli K-12 dnaT genes that led to a single amino acid change: R152C. A plasmid-encoded E. coli K-12 mutant dnaT gene with the R152C amino acid substitution did not display a dominant temperature-sensitive lethal phenotype in a dnaT(+) strain of E. coli K-12. Instead, this mutant dnaT gene was found to complement the E. coli K-12 dnaT822 mutant phenotypes. The significance of these results is discussed in terms of models for replication restart.}, keywords = {Base Sequence, Codon, DNA Primers, DNA Repair, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli K12, Escherichia coli Proteins, Molecular Sequence Data, Mutation, Plasmids, Polymerase Chain Reaction, Sequence Deletion}, issn = {0016-6731}, doi = {10.1534/genetics.103.025296}, author = {McCool, Jesse D and Ford, Christopher C and Sandler, Steven J} } @article {560, title = {Diversity of Geobacteraceae species inhabiting metal-polluted freshwater lake sediments ascertained by 16S rDNA analyses.}, journal = {Microb Ecol}, volume = {46}, year = {2003}, month = {2003 Aug}, pages = {257-69}, abstract = {The abundance, distribution, and phylogenetic diversity of members of the Fe(III)-reducing family Geobacteraceae were studied along a gradient of metal contaminants in Lake Coeur d{\textquoteright}Alene, Idaho. Partial 16S rRNA gene fragments were amplified by PCR using primers directed toward conserved regions of the gene within the family Geobacteraceae. Analysis of amplicons separated by denaturing gradient gel electrophoresis (DGGE) suggested within-site variation was as great as between-site variation. Amplicons were cloned and grouped by RFLP type and DGGE migration distance and representatives were sequenced. Grouping clones with 3\% or less sequence dissimilarity, 15 distinct phylotypes were identified compared to 16 distinct DGGE bands. Only 1 phylotype was recovered from all sites. This clone, B14, is most closely related to Geobacter metallireducens and constituted a greater portion of the pristine community than of the contaminated communities. A second phylotype, Q2, predominated in the contaminated communities and was notably absent from the pristine libraries. Clone Q2 presents a high degree of sequence similarity to two Geobacter spp. previously isolated from this region of Lake Coeur d{\textquoteright}Alene. Six phylotypes were unique to the contaminated sediments, whereas two were found only in the pristine sediments. Indices of diversity (Shannon and Simpson) were consistently higher when calculated with DGGE data than when clone library data were used. Most-probable-number PCR and real-time PCR suggested that the Geobacteraceae phylotypes were spread relatively evenly across all three sites along the gradient. Our data indicate that the Geobacteraceae are diverse and abundant in Lake Coeur d{\textquoteright}Alene sediments, regardless of metals content. These results provide insight into the ability of dissimilatory Fe(III)-reducing bacteria to colonize habitats with elevated metal concentrations, and they have important implications for the management and remediation of metal-contaminated sites.}, keywords = {Cloning, Molecular, Geobacter, Geologic Sediments, Phylogeny, Polymerase Chain Reaction, Population Dynamics, RNA, Ribosomal, 16S, Water Pollutants, Water Supply}, issn = {0095-3628}, author = {Cummings, D E and Snoeyenbos-West, O L and Newby, D T and Niggemyer, A M and Lovley, D R and Achenbach, L A and Rosenzweig, R F} } @article {574, title = {Desulfitobacterium metallireducens sp. nov., an anaerobic bacterium that couples growth to the reduction of metals and humic acids as well as chlorinated compounds.}, journal = {Int J Syst Evol Microbiol}, volume = {52}, year = {2002}, month = {2002 Nov}, pages = {1929-35}, abstract = {Strain 853-15A(T) was enriched and isolated from uranium-contaminated aquifer sediment by its ability to grow under anaerobic conditions via the oxidation of lactate coupled to the reduction of anthraquinone-2,6-disulfonate (AQDS) to anthrahydroquinone-2,6-disulfonate (AHQDS). Lactate was oxidized incompletely to acetate and carbon dioxide according to the reaction CH3CHOHCOO(-)+ 2AQDS+H2O --> CH3COO(-)+ 2AHQDS+CO2. Additional electron donors utilized included formate, ethanol, butanol, butyrate, malate and pyruvate. Lactate also supported growth with Fe(III) citrate, Mn(IV) oxide, humic substances, elemental sulfur, 3-chloro-4-hydroxyphenylacetate, trichloroethylene or tetrachloroethylene serving as the electron acceptor. Growth was not observed with sulfate, sulfite, nitrate or fumarate as the terminal electron acceptor. The temperature optimum for growth was 30 degrees C, but growth was also observed at 20 and 37 degrees C. The pH optimum was approximately 7.0. The 16S rDNA sequence of strain 853-15A(T) suggested that it was most closely related to Desulfitobacterium dehalogenans and closely related to Desulfitobacterium chlororespirans and Desulfitobacterium frappieri. The phylogenetic and physiological properties exhibited by strain 853-15A(T) (= ATCC BAA-636(T)) place it within the genus Desulfitobacterium as the type strain of a novel species, Desulfitobacterium metallireducens sp. nov.}, keywords = {Anaerobiosis, DNA, Bacterial, DNA, Ribosomal, Ecosystem, Electron Transport, Geologic Sediments, Humic Substances, Metals, Microscopy, Electron, Molecular Sequence Data, Oxidation-Reduction, Peptococcaceae, Phenotype, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Species Specificity}, issn = {1466-5026}, author = {Finneran, Kevin T and Forbush, Heather M and VanPraagh, Catherine V Gaw and Lovley, Derek R} } @article {1218, title = {Determination of polydextrose in foods by ion chromatography: collaborative study.}, journal = {J AOAC Int}, volume = {84}, year = {2001}, month = {2001 Mar-Apr}, pages = {472-8}, abstract = {

Eight collaborating laboratories assayed 7 blind duplicate pairs of foods for polydextrose content. The 7 test sample pairs ranged from low (2\%) to high (95\%) levels. The following foods were prepared with polydextrose mixed into the other ingredients and then baked, cooked, or otherwise prepared: milk chocolate candy, iced tea, sugar cookie, grape jelly, soft jellied candy, and powdered drink mix. Collaborators received a polydextrose standard to develop a calibration curve. The method determined polydextrose by ion chromatography, after removal of interfering food components (high molecular weight solubles). Repeatability standard deviations (RSDr) ranged from 3.93 to 9.04\%; reproducibility standard deviations (RSDR) ranged from 4.48 to 14.06\%. The average recovery was 94\%.

}, keywords = {Algorithms, Beverages, Cacao, Candy, Chromatography, Ion Exchange, Food Analysis, Glucans, Indicators and Reagents, Reference Standards, Tea, Ultracentrifugation}, issn = {1060-3271}, author = {Craig, S A and Holden, J F and Khaled, M Y} } @article {595, title = {Development of a genetic system for Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {67}, year = {2001}, month = {2001 Jul}, pages = {3180-7}, abstract = {Members of the genus Geobacter are the dominant metal-reducing microorganisms in a variety of anaerobic subsurface environments and have been shown to be involved in the bioremediation of both organic and metal contaminants. To facilitate the study of the physiology of these organisms, a genetic system was developed for Geobacter sulfurreducens. The antibiotic sensitivity of this organism was characterized, and optimal conditions for plating it at high efficiency were established. A protocol for the introduction of foreign DNA into G. sulfurreducens by electroporation was also developed. Two classes of broad-host-range vectors, IncQ and pBBR1, were found to be capable of replication in G. sulfurreducens. In particular, the IncQ plasmid pCD342 was found to be a suitable expression vector for this organism. When the information and novel methods described above were utilized, the nifD gene of G. sulfurreducens was disrupted by the single-step gene replacement method. Insertional mutagenesis of this key gene in the nitrogen fixation pathway impaired the ability of G. sulfurreducens to grow in medium lacking a source of fixed nitrogen. Expression of the nifD gene in trans complemented this phenotype. This paper constitutes the first report of genetic manipulation of a member of the Geobacter genus.}, keywords = {Anti-Bacterial Agents, Bacterial Proteins, Blotting, Southern, Culture Media, Deltaproteobacteria, Electroporation, Fimbriae Proteins, Genetic Complementation Test, Genetic Vectors, Microbial Sensitivity Tests, Mutagenesis, Insertional, Nitrogen Fixation, Plasmids, Transformation, Bacterial}, issn = {0099-2240}, doi = {10.1128/AEM.67.7.3180-3187.2001}, author = {Coppi, M V and Leang, C and Sandler, S J and Lovley, D R} } @article {606, title = {Differences in Fe(III) reduction in the hyperthermophilic archaeon, Pyrobaculum islandicum, versus mesophilic Fe(III)-reducing bacteria.}, journal = {FEMS Microbiol Lett}, volume = {195}, year = {2001}, month = {2001 Feb 20}, pages = {253-8}, abstract = {The discovery that all hyperthermophiles that have been evaluated have the capacity to reduce Fe(III) has raised the question of whether mechanisms for dissimilatory Fe(III) reduction have been conserved throughout microbial evolution. Many studies have suggested that c-type cytochromes are integral components in electron transport to Fe(III) in mesophilic dissimilatory Fe(III)-reducing microorganisms. However, Pyrobaculum islandicum, the hyperthermophile in which Fe(III) reduction has been most intensively studied, did not contain c-type cytochromes. NADPH was a better electron donor for the Fe(III) reductase activity in P. islandicum than NADH. This is the opposite of what has been observed with mesophiles. Thus, if previous models for dissimilatory Fe(III) reduction by mesophilic bacteria are correct, then it is unlikely that a single strategy for electron transport to Fe(III) is present in all dissimilatory Fe(III)-reducing microorganisms.}, keywords = {Cytochrome c Group, Electron Transport, Ferric Compounds, FMN Reductase, NADH, NADPH Oxidoreductases, NADP, Oxidation-Reduction, Temperature, Thermoproteaceae}, issn = {0378-1097}, author = {Childers, S E and Lovley, D R} } @article {1217, title = {Diversity among three novel groups of hyperthermophilic deep-sea Thermococcus species from three sites in the northeastern Pacific Ocean.}, journal = {FEMS Microbiol Ecol}, volume = {36}, year = {2001}, month = {2001 Jun}, pages = {51-60}, abstract = {

Eight new strains of deep-sea hyperthermophilic sulfur reducers were isolated from hydrothermal vent fields at 9 degrees 50{\textquoteright}N East Pacific Rise (EPR) and at the Cleft and CoAxial segments along the Juan de Fuca Ridge (JdFR). 16S rRNA gene sequence analysis showed that each strain belongs to the genus Thermococcus. Restriction fragment length polymorphism patterns of the 16S/23S rRNA intergenic spacer region revealed that these isolates fell into three groups: those from the EPR, those from fluid and rock sources on the JdFR, and those isolated from Paralvinella spp. polychaete vent worms from the JdFR. The optimum-temperature specific growth rates and the temperature ranges for growth were significantly higher and broader for those strains isolated from worms relative to those isolated from low-temperature diffuse hydrothermal fluids. Furthermore, the worm-derived isolates generally produced a larger array of proteases and amylases based on zymogram analyses. The zymogram patterns also changed with growth temperature suggesting that these organisms alter their lytic protein suites in response to changes in temperature. This study suggests that there is significant phenotypic diversity in Thermococcus that is not apparent from their highly conserved 16S rRNA nucleotide sequences.

}, issn = {1574-6941}, author = {Holden, J F. and Takai, K and Summit, M and Bolton, S and Zyskowski, J and Baross, J A.} } @article {1198, title = {Determination of polydextrose as dietary fiber in foods.}, journal = {J AOAC Int}, volume = {83}, year = {2000}, month = {2000 Jul-Aug}, pages = {1006-12}, abstract = {

Polydextrose (Litesse) provides physiological effects consistent with dietary fiber. However, AOAC methods for measuring total dietary fiber (TDF) in foods include an ethanol precipitation step in which polydextrose and similar carbohydrates are discarded and therefore not quantitated. This study describes a method developed to quantitate polydextrose in foods. The new method includes water extraction, centrifugal ultrafiltration, multienzyme hydrolysis, and anion exchange chromatography with electrochemical detection. Six foods were prepared with 4 levels of polydextrose to test the ruggedness of the method. Internal validation demonstrated the ruggedness of the method with recoveries ranging from 83 to 104\% with an average of 95\% (n = 24) and relative standard deviation of recoveries ranging from 0.7 to 13\% with an average of 3.3\% (n = 24). The value is added to that obtained for dietary fiber content of foods using the AOAC methods, to determine the TDF content of the food.

}, keywords = {Animals, Anions, Bacterial Proteins, Beverages, Cacao, Candy, Chromatography, Ion Exchange, Dietary Fiber, Ethanol, Food Analysis, Glucan 1,4-alpha-Glucosidase, Glucans, Glycoside Hydrolases, Humans, Hydrolysis, Isoamylase, Tea, Ultrafiltration}, issn = {1060-3271}, author = {Craig, S A and Holden, J F and Khaled, M Y and Craig, S A and Holden, J F and Khaled, M Y} } @article {607, title = {Direct and Fe(II)-mediated reduction of technetium by Fe(III)-reducing bacteria.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Sep}, pages = {3743-9}, abstract = {The dissimilatory Fe(III)-reducing bacterium Geobacter sulfurreducens reduced and precipitated Tc(VII) by two mechanisms. Washed cell suspensions coupled the oxidation of hydrogen to enzymatic reduction of Tc(VII) to Tc(IV), leading to the precipitation of TcO(2) at the periphery of the cell. An indirect, Fe(II)-mediated mechanism was also identified. Acetate, although not utilized efficiently as an electron donor for direct cell-mediated reduction of technetium, supported the reduction of Fe(III), and the Fe(II) formed was able to transfer electrons abiotically to Tc(VII). Tc(VII) reduction was comparatively inefficient via this indirect mechanism when soluble Fe(III) citrate was supplied to the cultures but was enhanced in the presence of solid Fe(III) oxide. The rate of Tc(VII) reduction was optimal, however, when Fe(III) oxide reduction was stimulated by the addition of the humic analog and electron shuttle anthaquinone-2,6-disulfonate, leading to the rapid formation of the Fe(II)-bearing mineral magnetite. Under these conditions, Tc(VII) was reduced and precipitated abiotically on the nanocrystals of biogenic magnetite as TcO(2) and was removed from solution to concentrations below the limit of detection by scintillation counting. Cultures of Fe(III)-reducing bacteria enriched from radionuclide-contaminated sediment using Fe(III) oxide as an electron acceptor in the presence of 25 microM Tc(VII) contained a single Geobacter sp. detected by 16S ribosomal DNA analysis and were also able to reduce and precipitate the radionuclide via biogenic magnetite. Fe(III) reduction was stimulated in aquifer material, resulting in the formation of Fe(II)-containing minerals that were able to reduce and precipitate Tc(VII). These results suggest that Fe(III)-reducing bacteria may play an important role in immobilizing technetium in sediments via direct and indirect mechanisms.}, keywords = {Culture Media, Deltaproteobacteria, Ferric Compounds, Ferrous Compounds, Fresh Water, Oxidation-Reduction, Technetium}, issn = {0099-2240}, author = {Lloyd, J R and Sole, V A and Van Praagh, C V and Lovley, D R} } @article {832, title = {Diversity of radA genes from cultured and uncultured archaea: comparative analysis of putative RadA proteins and their use as a phylogenetic marker.}, journal = {J Bacteriol}, volume = {181}, year = {1999}, month = {1999 Feb}, pages = {907-15}, abstract = {Archaea-specific radA primers were used with PCR to amplify fragments of radA genes from 11 cultivated archaeal species and one marine sponge tissue sample that contained essentially an archaeal monoculture. The amino acid sequences encoded by the PCR fragments, three RadA protein sequences previously published (21), and two new complete RadA sequences were aligned with representative bacterial RecA proteins and eucaryal Rad51 and Dmc1 proteins. The alignment supported the existence of four insertions and one deletion in the archaeal and eucaryal sequences relative to the bacterial sequences. The sizes of three of the insertions were found to have taxonomic and phylogenetic significance. Comparative analysis of the RadA sequences, omitting amino acids in the insertions and deletions, shows a cladal distribution of species which mimics to a large extent that obtained by a similar analysis of archaeal 16S rRNA sequences. The PCR technique also was used to amplify fragments of 15 radA genes from uncultured natural sources. Phylogenetic analysis of the amino acid sequences encoded by these fragments reveals several clades with affinity, sometimes only distant, to the putative RadA proteins of several species of Crenarcheota. The two most deeply branching archaeal radA genes found had some amino acid deletion and insertion patterns characteristic of bacterial recA genes. Possible explanations are discussed. Finally, signature codons are presented to distinguish among RecA protein family members.}, keywords = {Amino Acid Sequence, Archaea, Archaeal Proteins, Bacteria, Bacterial Proteins, Cloning, Molecular, DNA Primers, DNA Repair, DNA-Binding Proteins, Evolution, Molecular, Humans, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, Recombinant Proteins, RNA, Ribosomal, 16S, Saccharomyces cerevisiae, Sequence Alignment}, issn = {0021-9193}, author = {Sandler, S J and Hugenholtz, P and Schleper, C and DeLong, E F and Pace, N R and Clark, A J} } @article {830, title = {dnaC mutations suppress defects in DNA replication- and recombination-associated functions in priB and priC double mutants in Escherichia coli K-12.}, journal = {Mol Microbiol}, volume = {34}, year = {1999}, month = {1999 Oct}, pages = {91-101}, abstract = {PriA, PriB and PriC were originally discovered as proteins essential for the PhiX174 in vitro DNA replication system. Recent studies have shown that PriA mutants are poorly viable, have high basal levels of SOS expression (SOSH), are recombination deficient (Rec-), sensitive to UV irradiation (UVS) and sensitive to rich media. These data suggest that priA{\textquoteright}s role may be more complex than previously thought and may involve both DNA replication and homologous recombination. Based on the PhiX174 system, mutations in priB and priC should cause phenotypes like those seen in priA2:kan mutants. To test this, mutations in priB and priC were constructed. We found that, contrary to the PhiX174 model, del(priB)302 and priC303:kan mutants have almost wild-type phenotypes. Most unexpectedly, we then found that the priBC double mutant had very poor viability and/or a slow growth rate (even less than a priA2:kan mutant). This suggests that priB and priC have a redundant and important role in Escherichia coli. The priA2:kan suppressor, dnaC809, partially suppressed the poor viability/slow growth phenotype of the priBC double mutant. The resulting triple mutant (priBC dnaC809 ) had small colony size, recombination deficiency and levels of SOS expression similar to a priA2:kan mutant. The priBC dnaC809 mutant, however, was moderately UVR and had good viability, unlike a priA2:kan mutant. Additional mutations in the triple mutant were selected to suppress the slow growth phenotype. One suppressor restored all phenotypes tested to nearly wild-type levels. This mutation was identified as dnaC820 (K178N) [mapping just downstream of dnaC809 (E176G)]. Experiments suggest that dnaC820 makes dnaC809 suppression of priA and or priBC mutants priB and or priC independent. A model is proposed for the roles of these proteins in terms of restarting collapsed replication forks from recombinational intermediates.}, keywords = {Bacterial Proteins, Bacteriophage mu, Cell Division, DNA Replication, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Mutation, Phenotype, Recombination, Genetic, Replication Protein A, SOS Response (Genetics), Substrate Specificity, Suppression, Genetic}, issn = {0950-382X}, author = {Sandler, S J and Marians, K J and Zavitz, K H and Coutu, J and Parent, M A and Clark, A J} } @article {627, title = {Dissimilatory arsenate and sulfate reduction in Desulfotomaculum auripigmentum sp. nov.}, journal = {Arch Microbiol}, volume = {168}, year = {1997}, month = {1997 Nov}, pages = {380-8}, abstract = {A newly discovered arsenate-reducing bacterium, strain OREX-4, differed significantly from strains MIT-13 and SES-3, the previously described arsenate-reducing isolates, which grew on nitrate but not on sulfate. In contrast, strain OREX-4 did not respire nitrate but grew on lactate, with either arsenate or sulfate serving as the electron acceptor, and even preferred arsenate. Both arsenate and sulfate reduction were inhibited by molybdate. Strain OREX-4, a gram-positive bacterium with a hexagonal S-layer on its cell wall, metabolized compounds commonly used by sulfate reducers. Scorodite (FeAsO42. H2O) an arsenate-containing mineral, provided micromolar concentrations of arsenate that supported cell growth. Physiologically and phylogenetically, strain OREX-4 was far-removed from strains MIT-13 and SES-3: strain OREX-4 grew on different electron donors and electron acceptors, and fell within the gram-positive group of the Bacteria, whereas MIT-13 and SES-3 fell together in the epsilon-subdivision of the Proteobacteria. Together, these results suggest that organisms spread among diverse bacterial phyla can use arsenate as a terminal electron acceptor, and that dissimilatory arsenate reduction might occur in the sulfidogenic zone at arsenate concentrations of environmental interest. 16S rRNA sequence analysis indicated that strain OREX-4 is a new species of the genus Desulfotomaculum, and accordingly, the name Desulfotomaculum auripigmentum is proposed.}, keywords = {Arsenates, Bacteria, Anaerobic, Biotransformation, Geologic Sediments, Gram-Positive Endospore-Forming Rods, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Substrate Specificity, Sulfates, Sulfides, Sulfur-Reducing Bacteria, Water Microbiology}, issn = {0302-8933}, author = {Newman, D K and Kennedy, E K and Coates, J D and Ahmann, D and Ellis, D J and Lovley, D R and Morel, F M} } @article {836, title = {Differential suppression of priA2::kan phenotypes in Escherichia coli K-12 by mutations in priA, lexA, and dnaC.}, journal = {Genetics}, volume = {143}, year = {1996}, month = {1996 May}, pages = {5-13}, abstract = {First identified as an essential component of the phi X174 in vitro DNA replication system, PriA has ATPase, helicase, translocase, and primosome-assembly activities. priA1::kan strains of Escherichia coli are sensitive to UV irradiation, deficient in homologous recombination following transduction, and filamentous. priA2::kan strains have eightfold higher levels of uninduced SOS expression than wild type. We show that (1) priA1::kan strains have eightfold higher levels of uninduced SOS expression, (2) priA2::kan strains are UVS and Rec-, (3) lexA3 suppresses the high basal levels of SOS expression of a priA2::kan strain, and (4) plasmid-encoded priA300 (K230R), a mutant allele retaining only the primosome-assembly activity of priA+, restores both UVR and Rec+ phenotypes to a priA2::kan strain. Finally, we have isolated 17 independent UVR Rec+ revertants of priA2::kan strains that carry extragenic suppressors. All 17 map in the C-terminal half of the dnaC gene. DnaC loads the DnaB helicase onto DNA as a prelude for primosome assembly and DNA replication. We conclude that priA{\textquoteright}s primosome-assembly activity is essential for DNA repair and recombination and that the dnaC suppressor mutations allow these processes to occur in the absence of priA.}, keywords = {Bacterial Proteins, Bacteriophage phi X 174, beta-Galactosidase, Chromosomes, Bacterial, DNA Helicases, DNA Replication, DNA-Binding Proteins, Dose-Response Relationship, Radiation, Escherichia coli, Escherichia coli Proteins, Genes, Bacterial, Genetic Markers, Mutagenesis, Phenotype, Recombination, Genetic, Replication Protein A, Repressor Proteins, Serine Endopeptidases, Suppression, Genetic, Transduction, Genetic, Ultraviolet Rays}, issn = {0016-6731}, author = {Sandler, S J and Samra, H S and Clark, A J} } @article {637, title = {Desulfuromonas palmitatis sp. nov., a marine dissimilatory Fe(III) reducer that can oxidize long-chain fatty acids.}, journal = {Arch Microbiol}, volume = {164}, year = {1995}, month = {1995 Dec}, pages = {406-13}, abstract = {Studies on the microorganisms living in hydrocarbon-contaminated sediments in San Diego Bay, California led to the isolation of a novel Fe(III)-reducing microorganism. This organism, designated strain SDBY1, was an obligately anaerobic, non-motile, non-flagellated, gram-negative rod. Strain SDBY1 conserves energy to support growth from the oxidation of acetate, lactate, succinate, fumarate, laurate, palmitate, or stearate. H2 was also oxidized with the reduction of Fe(III), but growth with H2 as the sole electron donor was not observed. In addition to various forms of soluble and insoluble Fe(III), strain SDBY1 also coupled growth to the reduction of fumarate, Mn(IV), or S0. Air-oxidized minus dithionite-reduced difference spectra exhibited peaks at 552.8, 523.6, and 422.8 nm, indicative of c-type cytochrome(s). Strain SDBY1 shares physiological characteristics with organisms in the genera Geobacter, Pelobacter, and Desulfuromonas. Detailed analysis of the 16S rRNA sequence indicated that strain SDBY1 should be placed in the genus Desulfuromonas. The new species name Desulfuromonas palmitatis is proposed. D. palmitatis is only the second marine organism found (after D. acetoxidans) to oxidize multicarbon organic compounds completely to carbon dioxide with Fe(III) as an electron acceptor and provides the first pure culture model for the oxidation of long-chain fatty acids coupled to Fe(III) reduction.}, keywords = {Electron Transport, Fatty Acids, Gram-Negative Anaerobic Bacteria, Iron, Microscopy, Electron, Scanning, Oxidation-Reduction, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Water Microbiology}, issn = {0302-8933}, author = {Coates, J D and Lonergan, D J and Philips, E J and Jenter, H and Lovley, D R} } @article {798, title = {Discriminating between Ixodes ticks by means of mitochondrial DNA sequences.}, journal = {Mol Phylogenet Evol}, volume = {4}, year = {1995}, month = {1995 Dec}, pages = {361-5}, abstract = {Ticks of the genus Ixodes have recently assumed prominence because they frequently serve as vectors of important zoonoses, including Lyme disease and babesiosis. The morphological characteristics that have been used in their identification often are ambiguous and are useful solely at a particular stage of development. Here we report the DNA sequence of the mitochondrially encoded 16S rRNA gene of nine different Ixodes ticks and an outgroup from another genus, Dermacentor. The sequences readily discriminate between these ticks. Samples of I. dammini from the northeastern and upper midwestern United States differ from southeastern I. scapularis at about 2\% of the nucleotides. This difference is about half that separating other members of the I. ricinus group of species, but exceeds typical levels of intraspecific variation. Two major clades exist within the I. ricinus complex. One includes I. cookei, I. hexagonus, and I. angustus. The other includes I. persulcatus, I. pacificus, I. muris, I. ricinus, I. scapularis, and I. dammini. We conclude that mtDNA sequences are useful for unravelling the systematics of these important vectors of human disease.}, keywords = {Animals, Arachnid Vectors, Base Sequence, Dermacentor, DNA Primers, DNA, Mitochondrial, Genes, Humans, Ixodes, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Species Specificity}, issn = {1055-7903}, doi = {10.1006/mpev.1995.1033}, author = {Caporale, D A and Rich, S M and Spielman, A and Telford, S R and Kocher, T D} } @article {799, title = {Distribution of the Ixodes ricinus-like ticks of eastern North America.}, journal = {Proc Natl Acad Sci U S A}, volume = {92}, year = {1995}, month = {1995 Jul 3}, pages = {6284-8}, abstract = {We analyzed the geographic distribution of the Ixodes ricinus-like ticks in eastern North America by comparing the mitochondrial 16S rDNA sequences of specimens sampled directly from the field during the 1990s. Two distinct lineages are evident. The southern clade includes ticks from the southeastern and middle-eastern regions of the United States. The range of the northern clade, which appears to have been restricted to the northeastern region until the mid-1900s, now extends throughout the northeastern and middle-eastern regions. These phyletic units correspond to northern and southern taxa that have previously been assigned specific status as Ixodes dammini and Ixodes scapularis, respectively. The expanding range of I. dammini appears to drive the present outbreaks of zoonotic disease in eastern North America that include Lyme disease and human babesiosis.}, keywords = {Animals, Base Sequence, Demography, DNA Primers, DNA, Mitochondrial, DNA, Ribosomal, Geography, Mitochondria, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, Population, RNA, Ribosomal, 16S, Ticks, United States}, issn = {0027-8424}, author = {Rich, S M and Caporale, D A and Telford, S R and Kocher, T D and Hartl, D L and Spielman, A} } @article {650, title = {Dissimilatory Fe(III) Reduction by the Marine Microorganism Desulfuromonas acetoxidans.}, journal = {Appl Environ Microbiol}, volume = {59}, year = {1993}, month = {1993 Mar}, pages = {734-42}, abstract = {The ability of the marine microorganism Desulfuromonas acetoxidans to reduce Fe(III) was investigated because of its close phylogenetic relationship with the freshwater dissimilatory Fe(III) reducer Geobacter metallireducens. Washed cell suspensions of the type strain of D. acetoxidans reduced soluble Fe(III)-citrate and Fe(III) complexed with nitriloacetic acid. The c-type cytochrome(s) of D. acetoxidans was oxidized by Fe(III)-citrate and Mn(IV)-oxalate, as well as by two electron acceptors known to support growth, colloidal sulfur and malate. D. acetoxidans grew in defined anoxic, bicarbonate-buffered medium with acetate as the sole electron donor and poorly crystalline Fe(III) or Mn(IV) as the sole electron acceptor. Magnetite (Fe(3)O(4)) and siderite (FeCO(3)) were the major end products of Fe(III) reduction, whereas rhodochrosite (MnCO(3)) was the end product of Mn(IV) reduction. Ethanol, propanol, pyruvate, and butanol also served as electron donors for Fe(III) reduction. In contrast to D. acetoxidans, G. metallireducens could only grow in freshwater medium and it did not conserve energy to support growth from colloidal S reduction. D. acetoxidans is the first marine microorganism shown to conserve energy to support growth by coupling the complete oxidation of organic compounds to the reduction of Fe(III) or Mn(IV). Thus, D. acetoxidans provides a model enzymatic mechanism for Fe(III) or Mn(IV) oxidation of organic compounds in marine and estuarine sediments. These findings demonstrate that 16S rRNA phylogenetic analyses can suggest previously unrecognized metabolic capabilities of microorganisms.}, issn = {0099-2240}, author = {Roden, E E and Lovley, D R} } @article {652, title = {Dissimilatory metal reduction.}, journal = {Annu Rev Microbiol}, volume = {47}, year = {1993}, month = {1993}, pages = {263-90}, abstract = {Microorganisms can enzymatically reduce a variety of metals in metabolic processes that are not related to metal assimilation. Some microorganisms can conserve energy to support growth by coupling the oxidation of simple organic acids and alcohols, H2, or aromatic compounds to the reduction of Fe(III) or Mn(IV). This dissimilatory Fe(III) and Mn(IV) reduction influences the organic as well as the inorganic geochemistry of anaerobic aquatic sediments and ground water. Microorganisms that use U(VI) as a terminal electron acceptor play an important role in uranium geochemistry and may be a useful tool for removing uranium from contaminated environments. Se(VI) serves as a terminal electron acceptor to support anaerobic growth of some microorganisms. Reduction of Se(VI) to Se(O) is an important mechanism for the precipitation of selenium from contaminated waters. Enzymatic reduction of Cr(VI) to the less mobile and less toxic Cr(III), and reduction of soluble Hg(II) to volatile Hg(O) may affect the fate of these compounds in the environment and might be used as a remediation strategy. Microorganisms can also enzymatically reduce other metals such as technetium, vanadium, molybdenum, gold, silver, and copper, but reduction of these metals has not been studied extensively.}, keywords = {Bacteria, Metals, Oxidation-Reduction}, issn = {0066-4227}, doi = {10.1146/annurev.mi.47.100193.001403}, author = {Lovley, D R} } @article {656, title = {Dissimilatory Fe(III) and Mn(IV) reduction.}, journal = {Microbiol Rev}, volume = {55}, year = {1991}, month = {1991 Jun}, pages = {259-87}, abstract = {The oxidation of organic matter coupled to the reduction of Fe(III) or Mn(IV) is one of the most important biogeochemical reactions in aquatic sediments, soils, and groundwater. This process, which may have been the first globally significant mechanism for the oxidation of organic matter to carbon dioxide, plays an important role in the oxidation of natural and contaminant organic compounds in a variety of environments and contributes to other phenomena of widespread significance such as the release of metals and nutrients into water supplies, the magnetization of sediments, and the corrosion of metal. Until recently, much of the Fe(III) and Mn(IV) reduction in sedimentary environments was considered to be the result of nonenzymatic processes. However, microorganisms which can effectively couple the oxidation of organic compounds to the reduction of Fe(III) or Mn(IV) have recently been discovered. With Fe(III) or Mn(IV) as the sole electron acceptor, these organisms can completely oxidize fatty acids, hydrogen, or a variety of monoaromatic compounds. This metabolism provides energy to support growth. Sugars and amino acids can be completely oxidized by the cooperative activity of fermentative microorganisms and hydrogen- and fatty-acid-oxidizing Fe(III) and Mn(IV) reducers. This provides a microbial mechanism for the oxidation of the complex assemblage of sedimentary organic matter in Fe(III)- or Mn(IV)-reducing environments. The available evidence indicates that this enzymatic reduction of Fe(III) or Mn(IV) accounts for most of the oxidation of organic matter coupled to reduction of Fe(III) and Mn(IV) in sedimentary environments. Little is known about the diversity and ecology of the microorganisms responsible for Fe(III) and Mn(IV) reduction, and only preliminary studies have been conducted on the physiology and biochemistry of this process.}, keywords = {Bacteria, Electron Transport, Ferric Compounds, Fungi, Geological Phenomena, Geology, Manganese, Oxidation-Reduction, Soil Microbiology, Water Microbiology}, issn = {0146-0749}, author = {Lovley, D R} } @article {1731, title = {Disposition of (15,16-3H)naltrexone in the central nervous system of the rat.}, journal = {Drug Metab Dispos}, volume = {4}, year = {1976}, month = {1976 May-Jun}, pages = {276-80}, abstract = {

After injection of (15,16-3H)naltrexone (10 mg/kg s.c.) in male Wistar rats, peak concentrations of drug occurred in brain and plasma within 0.5 hr. Levels of naltrexone were sustained in brain between 2 and 24 hr and were barely detectable at 48 hr. Significant amounts of metabolities were present in brain and plasma at longer time periods. The t1/2 of naltrexone in brain and plasma were approximately 8.0 and 11.4 hr. respectively. The brain/plasma ratios of naltrexone at earlier times (0.5-1 hr) were higher than those at later times. The binding of naltrexone in vitro with rat plasma proteins in concentrations of 1-10 mug/ml ranged between 41 and 59\% 6beta-Naltrexol was present in very small amounts in brain but not in plasma. In addition to 7,8-dihydro-14-hydroxynormophinone and 7,8-dihydro-14-hydroxynormophine, tentative evidence was obtained for three other metabolites of naltrexone in brain. These metabolites were also present in plasma in addition to free and conjugated naltrexone and its N-dealkylated metabolites.

}, keywords = {Animals, Blood-Brain Barrier, Brain, Chromatography, Thin Layer, Male, Naloxone, Naltrexone, Rats}, issn = {0090-9556}, author = {Misra, A L and Bloch, R and Vardy, J and Mul{\'e}, S J and Verebely, K} } @article {1710, title = {Delineation of the intimate details of the backbone conformation of pyridine nucleotide coenzymes in aqueous solution.}, journal = {Biochem Biophys Res Commun}, volume = {66}, year = {1975}, month = {1975 Oct 27}, pages = {1173-9}, keywords = {Fourier Analysis, Magnetic Resonance Spectroscopy, Models, Molecular, Molecular Conformation, NAD, NADP, Structure-Activity Relationship, Temperature}, issn = {0006-291X}, author = {Bose, K S and Sarma, R H} } @article {1706, title = {Digitoxin metabolism by rat liver microsomes.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Sep 1}, pages = {1639-41}, keywords = {Animals, Chromatography, Thin Layer, Digitoxigenin, Digitoxin, Hydroxylation, In Vitro Techniques, Male, Microsomes, Liver, NADP, Rats, Time Factors}, issn = {0006-2952}, author = {Schmoldt, A and Benthe, H F and Haberland, G} }