@article {2624, title = {Chemically Induced Cell Wall Stapling in Bacteria.}, journal = {Cell Chem Biol}, volume = {28}, year = {2021}, month = {2021 Feb 18}, pages = {213-220.e4}, abstract = {

Transpeptidation reinforces the structure of cell-wall peptidoglycan, an extracellular heteropolymer that protects bacteria from osmotic lysis. The clinical success of transpeptidase-inhibiting \β-lactam antibiotics illustrates the essentiality of these cross-linkages for cell-wall integrity, but the presence of multiple, seemingly redundant transpeptidases in many species makes it challenging to determine cross-link function. Here, we present a technique to link peptide strands by chemical rather than enzymatic reaction. We employ biocompatible click chemistry to induce triazole formation between azido- and alkynyl-d-alanine residues that are metabolically installed in the peptidoglycan of Gram-positive or Gram-negative bacteria. Synthetic triazole cross-links can be visualized using azidocoumarin-d-alanine, an amino acid derivative that undergoes fluorescent enhancement upon reaction with terminal alkynes. Cell-wall stapling protects Escherichia coli from treatment with the broad-spectrum \β-lactams ampicillin and carbenicillin. Chemical control of cell-wall structure in live bacteria can provide functional insights that are orthogonal to those obtained by genetics.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2020.11.006}, author = {Rivera, Sylvia L and Espaillat, Akbar and Aditham, Arjun K and Shieh, Peyton and Muriel-Mundo, Chris and Kim, Justin and Cava, Felipe and Siegrist, M Sloan} } @article {3063, title = {Correlation of Key Physiological Properties of Isolates with Environment of Origin.}, journal = {Appl Environ Microbiol}, volume = {87}, year = {2021}, month = {2021 Jun 11}, pages = {e0073121}, abstract = {

It is known that the physiology of species can differ significantly, but the ecological impact of these differences is unclear. We recovered two strains of from two different ecosystems with a similar enrichment and isolation method. Both strains had the same ability to metabolize organic substrates and participate in direct interspecies electron transfer but also had major physiological differences. Strain DH-1, which was isolated from an anaerobic digester, used H as an electron donor. Genome analysis indicated that it lacks an Rnf complex and conserves energy from acetate metabolism via intracellular H cycling. In contrast, strain DH-2, a subsurface isolate, lacks hydrogenases required for H uptake and cycling and has an Rnf complex for energy conservation when growing on acetate. Further analysis of the genomes of previously described isolates, as well as phylogenetic and metagenomic data on uncultured in anaerobic digesters and diverse soils and sediments, revealed a physiological dichotomy that corresponded with environment of origin. The physiology of type I revolves around H production and consumption. In contrast, type II species eschew H and have genes for an Rnf complex and the multiheme, membrane-bound -type cytochrome MmcA, shown to be essential for extracellular electron transfer. The distribution of species in diverse environments suggests that the type I H-based physiology is well suited for high-energy environments, like anaerobic digesters, whereas type II Rnf/cytochrome-based physiology is an adaptation to the slower, steady-state carbon and electron fluxes common in organic-poor anaerobic soils and sediments. Biogenic methane is a significant greenhouse gas, and the conversion of organic wastes to methane is an important bioenergy process. species play an important role in methane production in many methanogenic soils and sediments as well as anaerobic waste digesters. The studies reported here emphasize that the genus is composed of two physiologically distinct groups. This is important to recognize when interpreting the role of in methanogenic environments, especially regarding H metabolism. Furthermore, the finding that type I species predominate in environments with high rates of carbon and electron flux and that type II species predominate in lower-energy environments suggests that evaluating the relative abundance of type I and type II may provide further insights into rates of carbon and electron flux in methanogenic environments.

}, keywords = {Acetates, Anaerobiosis, Bioreactors, Ecosystem, Electron Transport, Ethanol, Genome, Archaeal, Hydrogen, Methane, Methanosarcina, Phylogeny}, issn = {1098-5336}, doi = {10.1128/AEM.00731-21}, author = {Zhou, Jinjie and Holmes, Dawn E and Tang, Hai-Yan and Lovley, Derek R} } @article {3067, title = { Capable of Direct Interspecies Electron Transfer.}, journal = {Environ Sci Technol}, volume = {54}, year = {2020}, month = {2020 Dec 01}, pages = {15347-15354}, abstract = {

Direct interspecies electron transfer (DIET) from bacteria to methanogens is a revolutionary concept for syntrophic metabolism in methanogenic soils/sediments and anaerobic digestion. Previous studies have indicated that the potential for DIET is limited to methanogens in the , leading to the assumption that an abundance of other types of methanogens, such as species, indicates a lack of DIET. We report here on a strain of , designated strain YSL, that grows via DIET in defined cocultures with . The cocultures formed aggregates, in which cells of strain YSL and were uniformly dispersed throughout. This close association of the two species is the likely explanation for the ability of a strain of that could not express electrically conductive pili to grow in coculture with strain YSL. Granular activated carbon promoted the initial formation of the DIET-based cocultures. The discovery of DIET in , the genus of methanogens that has been the exemplar for interspecies electron transfer H, suggests that the capacity for DIET is much more broadly distributed among methanogens than previously considered. More innovative approaches to microbial isolation and characterization are needed in order to better understand how methanogenic communities function.

}, keywords = {Electron Transport, Electrons, Geobacter, Methane, Methanobacterium}, issn = {1520-5851}, doi = {10.1021/acs.est.0c05525}, author = {Zheng, Shiling and Liu, Fanghua and Wang, Bingchen and Zhang, Yuechao and Lovley, Derek R} } @article {3070, title = {An Chassis for Production of Electrically Conductive Protein Nanowires.}, journal = {ACS Synth Biol}, volume = {9}, year = {2020}, month = {2020 Mar 20}, pages = {647-654}, abstract = {

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

}, keywords = {Electric Conductivity, Escherichia coli, Fimbriae Proteins, Fimbriae, Bacterial, Geobacter, Graphite, Microorganisms, Genetically-Modified, Microscopy, Atomic Force, Nanowires, Operon, Protein Engineering}, issn = {2161-5063}, doi = {10.1021/acssynbio.9b00506}, author = {Ueki, Toshiyuki and Walker, David J F and Woodard, Trevor L and Nevin, Kelly P and Nonnenmann, Stephen S and Lovley, Derek R} } @article {2622, title = {Chemical Biology Tools for Examining the Bacterial Cell Wall.}, journal = {Cell Chem Biol}, volume = {27}, year = {2020}, month = {2020 08 20}, pages = {1052-1062}, abstract = {

Bacteria surround themselves with cell walls to maintain cell rigidity and protect against environmental insults. Here we review chemical and biochemical techniques employed to study bacterial cell wall biogenesis. Recent advances including the ability to isolate critical intermediates, metabolic approaches for probe incorporation, and isotopic labeling techniques have provided critical insight into the biochemistry of cell walls. Fundamental manuscripts that have used these techniques to discover cell wall-interacting proteins, flippases, and cell wall stoichiometry are discussed in detail. The review highlights that these powerful methods and techniques have exciting potential to identify and characterize new targets for antibiotic development.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2020.07.024}, author = {Brown, Ashley R and Gordon, Rebecca A and Hyland, Stephen N and Siegrist, M Sloan and Grimes, Catherine L} } @article {3076, title = {Cryo-EM reveals the structural basis of long-range electron transport in a cytochrome-based bacterial nanowire.}, journal = {Commun Biol}, volume = {2}, year = {2019}, month = {2019}, pages = {219}, abstract = {

Electrically conductive pili from species, termed bacterial nanowires, are intensely studied for their biological significance and potential in the development of new materials. Using cryo-electron microscopy, we have characterized nanowires from conductive pili preparations that are composed solely of head-to-tail stacked monomers of the six-heme C-type cytochrome OmcS. The unique fold of OmcS - closely wrapped around a continuous stack of hemes that can serve as an uninterrupted path for electron transport - generates a scaffold that supports the unbranched chain of hemes along the central axis of the filament. We present here, at 3.4 {\r A} resolution, the structure of this cytochrome-based filament and discuss its possible role in long-range biological electron transport.

}, keywords = {Cryoelectron Microscopy, Cytochromes c, Electron Transport, Fimbriae, Bacterial, Geobacter, Nanowires}, issn = {2399-3642}, doi = {10.1038/s42003-019-0448-9}, author = {Filman, David J and Marino, Stephen F and Ward, Joy E and Yang, Lu and Mester, Zolt{\'a}n and Bullitt, Esther and Lovley, Derek R and Strauss, Mike} } @article {2372, title = {Cell cycle localization dynamics of mitochondrial DNA polymerase IC in African trypanosomes.}, journal = {Mol Biol Cell}, volume = {29}, year = {2018}, month = {10/2018}, pages = {2540-2552}, abstract = {

Trypanosoma brucei has a unique catenated mitochondrial DNA (mtDNA) network called kinetoplast DNA (kDNA). Replication of kDNA occurs once per cell cycle in near synchrony with nuclear S phase and requires the coordination of many proteins. Among these are three essential DNA polymerases (TbPOLIB, IC, and ID). Localization dynamics of these proteins with respect to kDNA replication stages and how they coordinate their functions during replication are not well understood. We previously demonstrated that TbPOLID undergoes dynamic localization changes that are coupled to kDNA replication events. Here, we report the localization of TbPOLIC, a second essential DNA polymerase, and demonstrate the accumulation of TbPOLIC foci at active kDNA replication sites (antipodal sites) during stage II of the kDNA duplication cycle. While TbPOLIC was undetectable by immunofluorescence during other cell cycle stages, steady-state protein levels measured by Western blot remained constant. TbPOLIC foci colocalized with the fraction of TbPOLID that localized to the antipodal sites. However, the partial colocalization of the two essential DNA polymerases suggests a highly dynamic environment at the antipodal sites to coordinate the trafficking of replication proteins during kDNA synthesis. These data indicate that cell cycle-dependent localization is a major regulatory mechanism for essential mtDNA polymerases during kDNA replication.

}, keywords = {Cell Cycle, DNA Replication, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Gene Knockdown Techniques, Gene Silencing, Mitochondria, Protozoan Proteins, S Phase, Trypanosoma brucei brucei}, issn = {1939-4586}, doi = {10.1091/mbc.E18-02-0127}, author = {Concepci{\'o}n-Acevedo, Jeniffer and Miller, Jonathan C and Boucher, Michael J and Klingbeil, Michele M} } @article {3081, title = {Conductive Composite Materials Fabricated from Microbially Produced Protein Nanowires.}, journal = {Small}, volume = {14}, year = {2018}, month = {2018 Nov}, pages = {e1802624}, abstract = {

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

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

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

}, issn = {1664-302X}, doi = {10.3389/fmicb.2018.01512}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Aklujkar, Muktak A and Holmes, Dawn E and Lovley, Derek R} } @article {3121, title = {Centimeter-long electron transport in marine sediments via conductive minerals.}, journal = {ISME J}, volume = {9}, year = {2015}, month = {2015 Feb}, pages = {527-31}, abstract = {

Centimeter-long electron conduction through marine sediments, in which electrons derived from sulfide in anoxic sediments are transported to oxygen in surficial sediments, may have an important influence on sediment geochemistry. Filamentous bacteria have been proposed to mediate the electron transport, but the filament conductivity could not be verified and other mechanisms are possible. Surprisingly, previous investigations have never actually measured the sediment conductivity or its basic physical properties. Here we report direct measurements that demonstrate centimeter-long electron flow through marine sediments, with conductivities sufficient to account for previously estimated electron fluxes. Conductivity was lost for oxidized sediments, which contrasts with the previously described increase in the conductivity of microbial biofilms upon oxidation. Adding pyrite to the sediments significantly enhanced the conductivity. These results suggest that the role of conductive minerals, which are more commonly found in sediments than centimeter-long microbial filaments, need to be considered when modeling marine sediment biogeochemistry.

}, keywords = {Electron Transport, Geologic Sediments, Iron, Minerals, Oxidation-Reduction, Sulfides}, issn = {1751-7370}, doi = {10.1038/ismej.2014.131}, author = {Malvankar, Nikhil S and King, Gary M and Lovley, Derek R} } @article {1728, title = {Complete genome sequence of the hyperthermophilic methanogen Methanocaldococcus bathoardescens JH146(T) isolated from the basalt subseafloor.}, journal = {Mar Genomics}, volume = {24 Pt 3}, year = {2015}, month = {2015 Dec}, pages = {229-30}, abstract = {

Methanocaldococcus bathoardescens JH146(T) is a hyperthermophilic and obligate hydrogenotrophic methanogen isolated from low-temperature (26 {\textdegree}C) hydrothermal vent fluid at Axial Seamount in the northeastern Pacific Ocean. It is most closely related to the N2-fixing methanogen Methanocaldococcus sp. FS406-22; however, they differ in that JH146 cannot fix N2 or reductively assimilate nitrate. In this study, we present the complete genome sequence of strain JH146(T) (1,607,556 bp) with its 1635 protein coding genes, and 41 RNA genes. Our analysis focuses on its methane production via the acetyl-CoA pathway and its deleted gene clusters related to nitrogen assimilation. This study extends our understanding of methanogenesis at high temperatures and the impact of these organisms on the biogeochemistry of subseafloor hydrothermal environments and the deep sea.

}, issn = {1876-7478}, doi = {10.1016/j.margen.2015.06.002}, author = {Kim, You-Tae and Jung, Jong-Hyun and Stewart, Lucy C and Kwon, Soon-Wo and Holden, James F and Park, Cheon-Seok} } @article {3117, title = {Carbon cloth stimulates direct interspecies electron transfer in syntrophic co-cultures.}, journal = {Bioresour Technol}, volume = {173}, year = {2014}, month = {2014 Dec}, pages = {82-86}, abstract = {

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

}, keywords = {Carbon, Cell Communication, Coculture Techniques, Electric Conductivity, Electron Transport, Geobacter, Materials Testing, Membranes, Artificial, Microbial Consortia, Oxidation-Reduction, Symbiosis}, issn = {1873-2976}, doi = {10.1016/j.biortech.2014.09.009}, author = {Chen, Shanshan and Rotaru, Amelia-Elena and Liu, Fanghua and Philips, Jo and Woodard, Trevor L and Nevin, Kelly P and Lovley, Derek R} } @article {1965, title = {CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding}, journal = {PLoS ONE}, volume = {9}, year = {2014}, month = {mar}, pages = {e92581}, doi = {10.1371/journal.pone.0092581}, url = {https://doi.org/10.1371/journal.pone.0092581}, author = {Mandy Muller and Caroline Demeret}, editor = {Lawrence Banks} } @article {1200, title = {Complete genome sequence of hyperthermophilic archaeon Thermococcus sp. ES1.}, journal = {J Biotechnol}, volume = {174C}, year = {2014}, month = {2014 Jan 25}, pages = {14-15}, abstract = {

Thermococcus sp. strain ES1 is an anaerobic, hyperthermophilic archaeon from a hydrothermal vent that catabolizes sugars and peptides and produces H2S from S{\textdegree}, H2, acetate and CO2 as its primary metabolites. We present the complete genome sequence of this strain (1,957,742bp) with a focus on its substrate utilization and metabolite production capabilities. The sequence will contribute to the development of heterotrophic archaea for bioenergy production and biogeochemical modeling in hydrothermal environments.

}, issn = {1873-4863}, doi = {10.1016/j.jbiotec.2014.01.022}, author = {Jung, Jong-Hyun and Kim, You-Tae and Jeon, Eun-Jung and Seo, Dong-Ho and Hensley, Sarah A and Holden, James F and Lee, Ju-Hoon and Park, Cheon-Seok} } @article {3127, title = {Constraint-based modeling of carbon fixation and the energetics of electron transfer in Geobacter metallireducens.}, journal = {PLoS Comput Biol}, volume = {10}, year = {2014}, month = {2014 Apr}, pages = {e1003575}, abstract = {

Geobacter species are of great interest for environmental and biotechnology applications as they can carry out direct electron transfer to insoluble metals or other microorganisms and have the ability to assimilate inorganic carbon. Here, we report on the capability and key enabling metabolic machinery of Geobacter metallireducens GS-15 to carry out CO2 fixation and direct electron transfer to iron. An updated metabolic reconstruction was generated, growth screens on targeted conditions of interest were performed, and constraint-based analysis was utilized to characterize and evaluate critical pathways and reactions in G. metallireducens. The novel capability of G. metallireducens to grow autotrophically with formate and Fe(III) was predicted and subsequently validated in vivo. Additionally, the energetic cost of transferring electrons to an external electron acceptor was determined through analysis of growth experiments carried out using three different electron acceptors (Fe(III), nitrate, and fumarate) by systematically isolating and examining different parts of the electron transport chain. The updated reconstruction will serve as a knowledgebase for understanding and engineering Geobacter and similar species.

}, keywords = {Carbon, Electron Transport, Energy Metabolism, Genome, Bacterial, Geobacter, Models, Biological}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.1003575}, author = {Feist, Adam M and Nagarajan, Harish and Rotaru, Amelia-Elena and Tremblay, Pier-Luc and Zhang, Tian and Nevin, Kelly P and Lovley, Derek R and Zengler, Karsten} } @article {3115, title = {Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii.}, journal = {mBio}, volume = {5}, year = {2014}, month = {2014 Oct 21}, pages = {e01636-14}, abstract = {

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

}, keywords = {Acetyl Coenzyme A, Butyrates, Carbon Dioxide, Clostridium, Metabolic Engineering, Metabolic Flux Analysis, Metabolic Networks and Pathways, Recombinant Proteins}, issn = {2150-7511}, doi = {10.1128/mBio.01636-14}, author = {Ueki, Toshiyuki and Nevin, Kelly P and Woodard, Trevor L and Lovley, Derek R} } @article {3113, title = {Correlation between microbial community and granule conductivity in anaerobic bioreactors for brewery wastewater treatment.}, journal = {Bioresour Technol}, volume = {174}, year = {2014}, month = {2014 Dec}, pages = {306-10}, abstract = {

Prior investigation of an upflow anaerobic sludge blanket (UASB) reactor treating brewery wastes suggested that direct interspecies electron transfer (DIET) significantly contributed to interspecies electron transfer to methanogens. To investigate DIET in granules further, the electrical conductivity and bacterial community composition of granules in fourteen samples from four different UASB reactors treating brewery wastes were investigated. All of the UASB granules were electrically conductive whereas control granules from ANAMMOX (ANaerobic AMMonium OXidation) reactors and microbial granules from an aerobic bioreactor designed for phosphate removal were not. There was a moderate correlation (r=0.67) between the abundance of Geobacter species in the UASB granules and granule conductivity, suggesting that Geobacter contributed to granule conductivity. These results, coupled with previous studies, which have demonstrated that Geobacter species can donate electrons to methanogens that are typically predominant in anaerobic digesters, suggest that DIET may be a widespread phenomenon in UASB reactors treating brewery wastes.

}, keywords = {Alcoholic Beverages, Anaerobiosis, Bacteria, Bioreactors, Electric Conductivity, Ethanol, Sequence Analysis, DNA, Sewage, Waste Disposal, Fluid, Wastewater, Water Purification}, issn = {1873-2976}, doi = {10.1016/j.biortech.2014.10.004}, author = {Shrestha, Pravin Malla and Malvankar, Nikhil S and Werner, Jeffrey J and Franks, Ashley E and Elena-Rotaru, Amelia and Shrestha, Minita and Liu, Fanghua and Nevin, Kelly P and Angenent, Largus T and Lovley, Derek R} } @article {1452, title = {Changes in microbial dynamics during long-term decomposition in tropical forests.}, journal = {Soil Biology and Biochemistry}, volume = {66}, year = {2013}, month = {2013 Jun}, pages = {60-68}, doi = {10.1016/j.soilbio.2013.06.010}, author = {DeAngelis, KM and Chivian, D and Fortney, J and Simmons, BA and Hazen, TC and Silver, WL} } @article {3133, title = {Characterization and modelling of interspecies electron transfer mechanisms and microbial community dynamics of a syntrophic association.}, journal = {Nat Commun}, volume = {4}, year = {2013}, month = {2013}, pages = {2809}, abstract = {

Syntrophic associations are central to microbial communities and thus have a fundamental role in the global carbon cycle. Despite biochemical approaches describing the physiological activity of these communities, there has been a lack of a mechanistic understanding of the relationship between complex nutritional and energetic dependencies and their functioning. Here we apply a multi-omic modelling workflow that combines genomic, transcriptomic and physiological data with genome-scale models to investigate dynamics and electron flow mechanisms in the syntrophic association of Geobacter metallireducens and Geobacter sulfurreducens. Genome-scale modelling of direct interspecies electron transfer reveals insights into the energetics of electron transfer mechanisms. While G. sulfurreducens adapts to rapid syntrophic growth by changes at the genomic and transcriptomic level, G. metallireducens responds only at the transcriptomic level. This multi-omic approach enhances our understanding of adaptive responses and factors that shape the evolution of syntrophic communities.

}, keywords = {Adaptation, Biological, Biological Evolution, Electron Transport, Genome, Bacterial, Geobacter, Microbial Interactions, Models, Biological, Symbiosis, Transcriptome}, issn = {2041-1723}, doi = {10.1038/ncomms3809}, author = {Nagarajan, Harish and Embree, Mallory and Rotaru, Amelia-Elena and Shrestha, Pravin M and Feist, Adam M and Palsson, Bernhard {\O} and Lovley, Derek R and Zengler, Karsten} } @article {3154, title = {Characterization and transcription of arsenic respiration and resistance genes during in situ uranium bioremediation.}, journal = {ISME J}, volume = {7}, year = {2013}, month = {2013 Feb}, pages = {370-83}, abstract = {

The possibility of arsenic release and the potential role of Geobacter in arsenic biogeochemistry during in situ uranium bioremediation was investigated because increased availability of organic matter has been associated with substantial releases of arsenic in other subsurface environments. In a field experiment conducted at the Rifle, CO study site, groundwater arsenic concentrations increased when acetate was added. The number of transcripts from arrA, which codes for the α-subunit of dissimilatory As(V) reductase, and acr3, which codes for the arsenic pump protein Acr3, were determined with quantitative reverse transcription-PCR. Most of the arrA (>60\%) and acr3-1 (>90\%) sequences that were recovered were most similar to Geobacter species, while the majority of acr3-2 (>50\%) sequences were most closely related to Rhodoferax ferrireducens. Analysis of transcript abundance demonstrated that transcription of acr3-1 by the subsurface Geobacter community was correlated with arsenic concentrations in the groundwater. In contrast, Geobacter arrA transcript numbers lagged behind the major arsenic release and remained high even after arsenic concentrations declined. This suggested that factors other than As(V) availability regulated the transcription of arrA in situ, even though the presence of As(V) increased the transcription of arrA in cultures of Geobacter lovleyi, which was capable of As(V) reduction. These results demonstrate that subsurface Geobacter species can tightly regulate their physiological response to changes in groundwater arsenic concentrations. The transcriptomic approach developed here should be useful for the study of a diversity of other environments in which Geobacter species are considered to have an important influence on arsenic biogeochemistry.

}, keywords = {Acetates, Arsenate Reductases, Arsenic, Biodegradation, Environmental, Colorado, Gene Expression Regulation, Bacterial, Genes, Bacterial, Geobacter, Groundwater, Transcriptome, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2012.109}, author = {Giloteaux, Ludovic and Holmes, Dawn E and Williams, Kenneth H and Wrighton, Kelly C and Wilkins, Michael J and Montgomery, Alison P and Smith, Jessica A and Orellana, Roberto and Thompson, Courtney A and Roper, Thomas J and Long, Philip E and Lovley, Derek R} } @article {3132, title = {Characterizing acetogenic metabolism using a genome-scale metabolic reconstruction of Clostridium ljungdahlii.}, journal = {Microb Cell Fact}, volume = {12}, year = {2013}, month = {2013 Nov 25}, pages = {118}, abstract = {

BACKGROUND: The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow autotrophically on H2/CO2, and more importantly on synthesis gas (H2/CO/CO2) make them very attractive candidates as production hosts for biofuels and biocommodities. Acetogenic metabolism is considered one of the earliest modes of bacterial metabolism. A thorough understanding of various factors governing the metabolism, in particular energy conservation mechanisms, is critical for metabolic engineering of acetogens for targeted production of desired chemicals.

RESULTS: Here, we present the genome-scale metabolic network of Clostridium ljungdahlii, the first such model for an acetogen. This genome-scale model (iHN637) consisting of 637 genes, 785 reactions, and 698 metabolites captures all the major central metabolic and biosynthetic pathways, in particular pathways involved in carbon fixation and energy conservation. A combination of metabolic modeling, with physiological and transcriptomic data provided insights into autotrophic metabolism as well as aided the characterization of a nitrate reduction pathway in C. ljungdahlii. Analysis of the iHN637 metabolic model revealed that flavin based electron bifurcation played a key role in energy conservation during autotrophic growth and helped identify genes for some of the critical steps in this mechanism.

CONCLUSIONS: iHN637 represents a predictive model that recapitulates experimental data, and provides valuable insights into the metabolic response of C. ljungdahlii to genetic perturbations under various growth conditions. Thus, the model will be instrumental in guiding metabolic engineering of C. ljungdahlii for the industrial production of biocommodities and biofuels.

}, keywords = {Acetates, Biofuels, Clostridium, Genome, Metabolic Engineering, Metabolic Networks and Pathways}, issn = {1475-2859}, doi = {10.1186/1475-2859-12-118}, author = {Nagarajan, Harish and Sahin, Merve and Nogales, Juan and Latif, Haythem and Lovley, Derek R and Ebrahim, Ali and Zengler, Karsten} } @article {3140, title = {Characterizing the interplay between multiple levels of organization within bacterial sigma factor regulatory networks.}, journal = {Nat Commun}, volume = {4}, year = {2013}, month = {2013}, pages = {1755}, abstract = {

Bacteria contain multiple sigma factors, each targeting diverse, but often overlapping sets of promoters, thereby forming a complex network. The layout and deployment of such a sigma factor network directly impacts global transcriptional regulation and ultimately dictates the phenotype. Here we integrate multi-omic data sets to determine the topology, the operational, and functional states of the sigma factor network in Geobacter sulfurreducens, revealing a unique network topology of interacting sigma factors. Analysis of the operational state of the sigma factor network shows a highly modular structure with σ(N) being the major regulator of energy metabolism. Surprisingly, the functional state of the network during the two most divergent growth conditions is nearly static, with sigma factor binding profiles almost invariant to environmental stimuli. This first comprehensive elucidation of the interplay between different levels of the sigma factor network organization is fundamental to characterize transcriptional regulatory mechanisms in bacteria.

}, keywords = {Energy Metabolism, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Gene Regulatory Networks, Genes, Bacterial, Geobacter, Models, Biological, Regulon, Sigma Factor}, issn = {2041-1723}, doi = {10.1038/ncomms2743}, author = {Qiu, Yu and Nagarajan, Harish and Embree, Mallory and Shieu, Wendy and Abate, Elisa and Ju{\'a}rez, Katy and Cho, Byung-Kwan and Elkins, James G and Nevin, Kelly P and Barrett, Christian L and Lovley, Derek R and Palsson, Bernhard O and Zengler, Karsten} } @article {1966, title = {A Comparative Approach to Characterize the Landscape of Host-Pathogen Protein-Protein Interactions}, journal = {Journal of Visualized Experiments}, year = {2013}, month = {jul}, doi = {10.3791/50404}, url = {https://doi.org/10.3791/50404}, author = {Mandy Muller and Patricia Cassonnet and Michel Favre and Yves Jacob and Caroline Demeret} } @article {1423, title = {Conversion of the Amazon rainforest to agriculture results in biotic homogenization of soil bacterial communities.}, journal = {Proc Natl Acad Sci U S A}, volume = {110}, year = {2013}, month = {2013 Jan 15}, pages = {988-93}, abstract = {

The Amazon rainforest is the Earth\&$\#$39;s largest reservoir of plant and animal diversity, and it has been subjected to especially high rates of land use change, primarily to cattle pasture. This conversion has had a strongly negative effect on biological diversity, reducing the number of plant and animal species and homogenizing communities. We report here that microbial biodiversity also responds strongly to conversion of the Amazon rainforest, but in a manner different from plants and animals. Local taxonomic and phylogenetic diversity of soil bacteria increases after conversion, but communities become more similar across space. This homogenization is driven by the loss of forest soil bacteria with restricted ranges (endemics) and results in a net loss of diversity. This study shows homogenization of microbial communities in response to human activities. Given that soil microbes represent the majority of biodiversity in terrestrial ecosystems and are intimately involved in ecosystem functions, we argue that microbial biodiversity loss should be taken into account when assessing the impact of land use change in tropical forests.

}, keywords = {Agriculture, Animals, Bacteria, Biodiversity, Brazil, Cattle, Ecosystem, Humans, Phylogeny, Rain, Soil Microbiology, Trees, Tropical Climate}, issn = {1091-6490}, doi = {10.1073/pnas.1220608110}, author = {Rodrigues, Jorge L M and Pellizari, Vivian H and Mueller, Rebecca and Baek, Kyunghwa and Jesus, Ederson da C and Paula, Fabiana S and Mirza, Babur and Hamaoui, George S and Tsai, Siu Mui and Feigl, Brigitte and Tiedje, James M and Bohannan, Brendan J M and N{\"u}sslein, Klaus} } @article {935, title = {Critical roles for lipomannan and lipoarabinomannan in cell wall integrity of mycobacteria and pathogenesis of tuberculosis.}, journal = {MBio}, volume = {4}, year = {2013}, month = {2013}, abstract = {

ABSTRACT Lipomannan (LM) and lipoarabinomannan (LAM) are mycobacterial glycolipids containing a long mannose polymer. While they are implicated in immune modulations, the significance of LM and LAM as structural components of the mycobacterial cell wall remains unknown. We have previously reported that a branch-forming mannosyltransferase plays a critical role in controlling the sizes of LM and LAM and that deletion or overexpression of this enzyme results in gross changes in LM/LAM structures. Here, we show that such changes in LM/LAM structures have a significant impact on the cell wall integrity of mycobacteria. In Mycobacterium smegmatis, structural defects in LM and LAM resulted in loss of acid-fast staining, increased sensitivity to \β-lactam antibiotics, and faster killing by THP-1 macrophages. Furthermore, equivalent Mycobacterium\ tuberculosis mutants became more sensitive to \β-lactams, and one mutant showed attenuated virulence in mice. Our results revealed previously unknown structural roles for LM and LAM and further demonstrated that they are important for the pathogenesis of tuberculosis. IMPORTANCE Tuberculosis (TB) is a global burden, affecting millions of people worldwide. Mycobacterium tuberculosis is a causative agent of TB, and understanding the biology of M.\ tuberculosis is essential for tackling this devastating disease. The cell wall of M.\ tuberculosis is highly impermeable and plays a protective role in establishing infection. Among the cell wall components, LM and LAM are major glycolipids found in all Mycobacterium species, show various immunomodulatory activities, and have been thought to play roles in TB pathogenesis. However, the roles of LM and LAM as integral parts of the cell wall structure have not been elucidated. Here we show that LM and LAM play critical roles in the integrity of mycobacterial cell wall and the pathogenesis of TB. These findings will now allow us to seek the possibility that the LM/LAM biosynthetic pathway is a chemotherapeutic target.

}, issn = {2150-7511}, doi = {10.1128/mBio.00472-12}, author = {Fukuda, Takeshi and Matsumura, Takayuki and Ato, Manabu and Hamasaki, Maho and Nishiuchi, Yukiko and Murakami, Yoshiko and Maeda, Yusuke and Yoshimori, Tamotsu and Matsumoto, Sohkichi and Kobayashi, Kazuo and Kinoshita, Taroh and Morita, Yasu S} } @article {1969, title = {Comparative analysis of virus{\textendash}host interactomes with a mammalian high-throughput protein complementation assay based on Gaussia princeps luciferase}, journal = {Methods}, volume = {58}, year = {2012}, month = {dec}, pages = {349{\textendash}359}, doi = {10.1016/j.ymeth.2012.07.029}, url = {https://doi.org/10.1016/j.ymeth.2012.07.029}, author = {Gr{\'e}gory Neveu and Patricia Cassonnet and Pierre-Olivier Vidalain and Caroline Rolloy and Jos{\'e} Mendoza and Louis Jones and Fr{\'e}d{\'e}ric Tangy and Mandy Muller and Caroline Demeret and Lionel Tafforeau and Vincent Lotteau and Chantal Rabourdin-Combe and Gilles Trav{\'e} and Am{\'e}lie Dricot and David E. Hill and Marc Vidal and Michel Favre and Yves Jacob} } @article {3156, title = {Comparative genomic analysis of Geobacter sulfurreducens KN400, a strain with enhanced capacity for extracellular electron transfer and electricity production.}, journal = {BMC Genomics}, volume = {13}, year = {2012}, month = {2012 Sep 12}, pages = {471}, abstract = {

BACKGROUND: A new strain of Geobacter sulfurreducens, strain KN400, produces more electrical current in microbial fuel cells and reduces insoluble Fe(III) oxides much faster than the wildtype strain, PCA. The genome of KN400 was compared to wildtype with the goal of discovering how the network for extracellular electron transfer has changed and how these two strains evolved.

RESULTS: Both genomes were re-annotated, resulting in 14 fewer genes (net) in the PCA genome; 28 fewer (net) in the KN400 genome; and ca. 400 gene start and stop sites moved. 96\% of genes in KN400 had clear orthologs with conserved synteny in PCA. Most of the remaining genes were in regions of genomic mobility and were strain-specific or conserved in other Geobacteraceae, indicating that the changes occurred post-divergence. There were 27,270 single nucleotide polymorphisms (SNP) between the genomes. There was significant enrichment for SNP locations in non-coding or synonymous amino acid sites, indicating significant selective pressure since the divergence. 25\% of orthologs had sequence differences, and this set was enriched in phosphorylation and ATP-dependent enzymes. Substantial sequence differences (at least 12 non-synonymous SNP/kb) were found in 3.6\% of the orthologs, and this set was enriched in cytochromes and integral membrane proteins. Genes known to be involved in electron transport, those used in the metabolic cell model, and those that exhibit changes in expression during growth in microbial fuel cells were examined in detail.

CONCLUSIONS: The improvement in external electron transfer in the KN400 strain does not appear to be due to novel gene acquisition, but rather to changes in the common metabolic network. The increase in electron transfer rate and yield in KN400 may be due to changes in carbon flux towards oxidation pathways and to changes in ATP metabolism, both of which indicate that the overall energy state of the cell may be different. The electrically conductive pili appear to be unchanged, but cytochrome folding, localization, and redox potentials may all be affected, which would alter the electrical connection between the cell and the substrate.

}, keywords = {Bioelectric Energy Sources, Comparative Genomic Hybridization, Electron Transport, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Metabolic Networks and Pathways, Molecular Sequence Annotation, Polymorphism, Single Nucleotide}, issn = {1471-2164}, doi = {10.1186/1471-2164-13-471}, author = {Butler, Jessica E and Young, Nelson D and Aklujkar, Muktak and Lovley, Derek R} } @article {1207, title = {Complete genome sequence of the hyperthermophilic archaeon Pyrococcus sp. strain ST04, isolated from a deep-sea hydrothermal sulfide chimney on the Juan de Fuca Ridge.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Aug}, pages = {4434-5}, abstract = {

Pyrococcus sp. strain ST04 is a hyperthermophilic, anaerobic, and heterotrophic archaeon isolated from a deep-sea hydrothermal sulfide chimney on the Endeavour Segment of the Juan de Fuca Ridge in the northeastern Pacific Ocean. To further understand the distinct characteristics of this archaeon at the genome level (polysaccharide utilization at high temperature and ATP generation by a Na(+) gradient), the genome of strain ST04 was completely sequenced and analyzed. Here, we present the complete genome sequence analysis results of Pyrococcus sp. ST04 and report the major findings from the genome annotation, with a focus on its saccharolytic and metabolite production potential.

}, keywords = {Adenosine Triphosphate, Anaerobiosis, DNA, Archaeal, Genome, Archaeal, Heterotrophic Processes, Hydrothermal Vents, Molecular Sequence Data, Pacific Ocean, Polysaccharides, Pyrococcus, Seawater, Sequence Analysis, DNA, Sodium Chloride, Sulfides}, issn = {1098-5530}, doi = {10.1128/JB.00824-12}, author = {Jung, Jong-Hyun and Lee, Ju-Hoon and Holden, James F and Seo, Dong-Ho and Shin, Hakdong and Kim, Hae-Yeong and Kim, Wooki and Ryu, Sangryeol and Park, Cheon-Seok} } @article {1205, title = {Complete genome sequence of the hyperthermophilic archaeon Thermococcus sp. strain CL1, isolated from a Paralvinella sp. polychaete worm collected from a hydrothermal vent.}, journal = {J Bacteriol}, volume = {194}, year = {2012}, month = {2012 Sep}, pages = {4769-70}, abstract = {

Thermococcus sp. strain CL1 is a hyperthermophilic, anaerobic, and heterotrophic archaeon isolated from a Paralvinella sp. polychaete worm living on an active deep-sea hydrothermal sulfide chimney on the Cleft Segment of the Juan de Fuca Ridge. To further understand the distinct characteristics of this archaeon at the genome level, its genome was completely sequenced and analyzed. Here, we announce the complete genome sequence (1,950,313 bp) of Thermococcus sp. strain CL1, with a focus on H(2)- and energy-producing capabilities and its amino acid biosynthesis and acquisition in an extreme habitat.

}, keywords = {Animals, Base Sequence, Chromosome Mapping, DNA, Archaeal, DNA, Ribosomal, Genome, Bacterial, Hydrothermal Vents, Molecular Sequence Data, Phylogeny, Polychaeta, Sequence Analysis, DNA, Thermococcus}, issn = {1098-5530}, doi = {10.1128/JB.01016-12}, author = {Jung, Jong-Hyun and Holden, James F and Seo, Dong-Ho and Park, Kwan-Hwa and Shin, Hakdong and Ryu, Sangryeol and Lee, Ju-Hoon and Park, Cheon-Seok} } @article {1567, title = {Characterization of a novel facultative Methylocystis species capable of growth on methane, acetate and ethanol.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {174-81}, abstract = {

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

}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00204.x}, author = {Im, Jeongdae and Lee, Sung-Woo and Yoon, Sukhwan and Dispirito, Alan A and Semrau, Jeremy D} } @article {375, title = {Characterization of trapped lignin-degrading microbes in tropical forest soil.}, journal = {PLoS One}, volume = {6}, year = {2011}, month = {2011}, pages = {e19306}, abstract = {Lignin is often the most difficult portion of plant biomass to degrade, with fungi generally thought to dominate during late stage decomposition. Lignin in feedstock plant material represents a barrier to more efficient plant biomass conversion and can also hinder enzymatic access to cellulose, which is critical for biofuels production. Tropical rain forest soils in Puerto Rico are characterized by frequent anoxic conditions and fluctuating redox, suggesting the presence of lignin-degrading organisms and mechanisms that are different from known fungal decomposers and oxygen-dependent enzyme activities. We explored microbial lignin-degraders by burying bio-traps containing lignin-amended and unamended biosep beads in the soil for 1, 4, 13 and 30 weeks. At each time point, phenol oxidase and peroxidase enzyme activity was found to be elevated in the lignin-amended versus the unamended beads, while cellulolytic enzyme activities were significantly depressed in lignin-amended beads. Quantitative PCR of bacterial communities showed more bacterial colonization in the lignin-amended compared to the unamended beads after one and four weeks, suggesting that the lignin supported increased bacterial abundance. The microbial community was analyzed by small subunit 16S ribosomal RNA genes using microarray (PhyloChip) and by high-throughput amplicon pyrosequencing based on universal primers targeting bacterial, archaeal, and eukaryotic communities. Community trends were significantly affected by time and the presence of lignin on the beads. Lignin-amended beads have higher relative abundances of representatives from the phyla Actinobacteria, Firmicutes, Acidobacteria and Proteobacteria compared to unamended beads. This study suggests that in low and fluctuating redox soils, bacteria could play a role in anaerobic lignin decomposition.}, keywords = {Biodiversity, Biomass, Ecosystem, Gases, Lignin, Oligonucleotide Array Sequence Analysis, Phylogeny, Plants, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Soil Microbiology, Trees}, issn = {1932-6203}, doi = {10.1371/journal.pone.0019306}, author = {Deangelis, Kristen M and Allgaier, Martin and Chavarria, Yaucin and Fortney, Julian L and Hugenholtz, Phillip and Simmons, Blake and Sublette, Kerry and Silver, Whendee L and Hazen, Terry C} } @article {851, title = {Colonization of paediatric lower respiratory tract with genital Mycoplasma species.}, journal = {Respirology}, volume = {16}, year = {2011}, month = {2011 Oct}, pages = {1081-7}, abstract = {BACKGROUND AND OBJECTIVE: Recently, much attention has been given to the possible role played by pathogens that colonize neonatal or paediatric airway and their potential involvement in chronic respiratory disease. The goal of the current study was to evaluate the prevalence of Mycoplasma organisms in the BAL fluid of paediatric patients suffering from a variety of chronic respiratory diseases to determine if there was any clear disease association with bacterial presence. METHODS: We examined 319 paediatric BAL samples for the presence of M.genitalium, M.hominis, U.urealyticum, U.parvum and M.pneumoniae DNA with species-specific PCR. RESULTS: Mycoplasma DNA was found in 32.6\% (104/319) of patient samples; 10\% (32/319) for M.pneumoniae, 8.8\% for U.parvum, 2.8\% for U.urealyticum; 4.7\% for M.hominis and 9.1\% for M.genitalium. There were no significant clinical and laboratory differences except serum IgE in asthmatics according to Mycoplasma colonization or not. Elevated levels of IgE were found more often in Mycoplasma DNA-negative patients than patients with bacterial DNA, 85/109 versus 24/109 respectively (P<0.0001). There was no difference in the frequency of Mycoplasmas between the asthmatics and the non-asthmatics; 30.6\% (69/225) versus 37.2\% (35/94) for Mycoplasma 16S DNA, and 8\% versus 14.9\% for M.pneumoniae, respectively. CONCLUSIONS: Our data indicate that in addition to M.pneumoniae, urogenital Mycoplasma species may colonize the airway of patients with chronic respiratory diseases. There was, however, no association between chronic asthma diagnosis and Mycoplasma colonization in this study.}, keywords = {Adolescent, Asthma, Child, Child, Preschool, Cohort Studies, DNA, Bacterial, Female, Humans, Infant, Infant, Newborn, Male, Mycoplasma, Mycoplasma genitalium, Mycoplasma hominis, Mycoplasma Infections, Mycoplasma pneumoniae, Respiratory System, Species Specificity, Ureaplasma, Ureaplasma Infections, Ureaplasma urealyticum, Young Adult}, issn = {1440-1843}, doi = {10.1111/j.1440-1843.2011.02016.x}, author = {Patel, Katir K and Salva, Paul S and Webley, Wilmore C} } @article {371, title = {Complete genome sequence of "Enterobacter lignolyticus" SCF1.}, journal = {Stand Genomic Sci}, volume = {5}, year = {2011}, month = {2011 Oct 15}, pages = {69-85}, abstract = {In an effort to discover anaerobic bacteria capable of lignin degradation, we isolated "Enterobacter lignolyticus" SCF1 on minimal media with alkali lignin as the sole source of carbon. This organism was isolated anaerobically from tropical forest soils collected from the Short Cloud Forest site in the El Yunque National Forest in Puerto Rico, USA, part of the Luquillo Long-Term Ecological Research Station. At this site, the soils experience strong fluctuations in redox potential and are net methane producers. Because of its ability to grow on lignin anaerobically, we sequenced the genome. The genome of "E. lignolyticus" SCF1 is 4.81 Mbp with no detected plasmids, and includes a relatively small arsenal of lignocellulolytic carbohydrate active enzymes. Lignin degradation was observed in culture, and the genome revealed two putative laccases, a putative peroxidase, and a complete 4-hydroxyphenylacetate degradation pathway encoded in a single gene cluster.}, issn = {1944-3277}, doi = {10.4056/sigs.2104875}, author = {Deangelis, Kristen M and D{\textquoteright}haeseleer, Patrik and Chivian, Dylan and Fortney, Julian L and Khudyakov, Jane and Simmons, Blake and Woo, Hannah and Arkin, Adam P and Davenport, Karen Walston and Goodwin, Lynne and Chen, Amy and Ivanova, Natalia and Kyrpides, Nikos C and Mavromatis, Konstantinos and Woyke, Tanja and Hazen, Terry C} } @article {1566, title = {Constitutive expression of pMMO by Methylocystis strain SB2 when grown on multi-carbon substrates: implications for biodegradation of chlorinated ethenes.}, journal = {Environ Microbiol Rep}, volume = {3}, year = {2011}, month = {2011 Apr}, pages = {182-8}, abstract = {

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

}, issn = {1758-2229}, doi = {10.1111/j.1758-2229.2010.00205.x}, author = {Yoon, Sukhwan and Im, Jeongdae and Bandow, Nathan and Dispirito, Alan A and Semrau, Jeremy D} } @article {441, title = {A c-type cytochrome and a transcriptional regulator responsible for enhanced extracellular electron transfer in Geobacter sulfurreducens revealed by adaptive evolution.}, journal = {Environ Microbiol}, volume = {13}, year = {2011}, month = {2011 Jan}, pages = {13-23}, abstract = {The stimulation of subsurface microbial metabolism often associated with engineered bioremediation of groundwater contaminants presents subsurface microorganisms, which are adapted for slow growth and metabolism in the subsurface, with new selective pressures. In order to better understand how Geobacter species might adapt to selective pressure for faster metal reduction in the subsurface, Geobacter sulfurreducens was put under selective pressure for rapid Fe(III) oxide reduction. The genomes of two resultant strains with rates of Fe(III) oxide reduction that were 10-fold higher than those of the parent strain were resequenced. Both strains contain either a single base-pair change or a 1 nucleotide insertion in a GEMM riboswitch upstream of GSU1761, a gene coding for the periplasmic c-type cytochrome designated PgcA. GSU1771, a gene coding for a SARP regulator, was also mutated in both strains. Introduction of either of the GEMM riboswitch mutations upstream of pgcA in the wild-type increased the abundance of pgcA transcripts, consistent with increased expression of pgcA in the adapted strains. One of the mutations doubled the rate of Fe(III) oxide reduction. Interruption of GSU1771 doubled the Fe(III) oxide reduction rate. This was associated with an increased in expression of pilA, the gene encoding the structural protein for the pili thought to function as microbial nanowires. The combination of the GSU1771 interruption with either of the pgcA mutations resulted in a strain that reduced Fe(III) as fast as the comparable adapted strain. These results suggest that the accumulation of a small number of beneficial mutations under selective pressure, similar to that potentially present during bioremediation, can greatly enhance the capacity for Fe(III) oxide reduction in G. sulfurreducens. Furthermore, the results emphasize the importance of the c-type cytochrome PgcA and pili in Fe(III) oxide reduction and demonstrate how adaptive evolution studies can aid in the elucidation of complex mechanisms, such as extracellular electron transfer.}, keywords = {Adaptation, Physiological, Biodegradation, Environmental, Cytochrome c Group, DNA, Bacterial, Electron Transport, Evolution, Molecular, Ferric Compounds, Gene Expression Profiling, Genes, Bacterial, Genome, Bacterial, Geobacter, Mutagenesis, Insertional, Mutation, Oligonucleotide Array Sequence Analysis, Oxidation-Reduction, Riboswitch, Sequence Analysis, DNA}, issn = {1462-2920}, doi = {10.1111/j.1462-2920.2010.02302.x}, author = {Tremblay, Pier-Luc and Summers, Zarath M and Glaven, Richard H and Nevin, Kelly P and Zengler, Karsten and Barrett, Christian L and Qiu, Yu and Palsson, Bernhard O and Lovley, Derek R} } @article {807, title = {Clinical applications of biomarkers in pediatric traumatic brain injury.}, journal = {Childs Nerv Syst}, volume = {26}, year = {2010}, month = {2010 Feb}, pages = {205-13}, abstract = {INTRODUCTION: The diagnosis, treatment, and prediction of outcome in pediatric traumatic brain injury (TBI) present significant challenges to the treating clinician. Clinical and radiological tools for assessing injury severity and predicting outcome, in particular, lack sensitivity and specificity. In patients with mild TBI, often there is uncertainty about which patients should undergo radiological imaging and who is at risk for long term neurological sequelae. In severe TBI, often there is uncertainty about which patients will experience secondary insults and what the outcome for individual patients will be. In several other clinical specialties, biomarkers are used to diagnose disease, direct treatment, and prognosticate. However, an ideal biomarker for brain injury has not been found. METHODS: In this review, we examine the various factors that must be taken into account in the search for a reliable biomarker in brain injury. We review the important studies that have investigated common biomarkers of structural brain injury, in particular S100B, neuron-specific enolase, myelin basic protein, and glial fibrillary acid protein. DISCUSSION: The potential uses and limitations of these biomarkers in the context of TBI are discussed.}, keywords = {Animals, Biological Markers, Brain Injuries, Child, Humans}, issn = {1433-0350}, doi = {10.1007/s00381-009-1009-1}, author = {Sandler, Simon J I and Figaji, Anthony A and Adelson, P David} } @article {428, title = {Constraint-based modeling analysis of the metabolism of two Pelobacter species.}, journal = {BMC Syst Biol}, volume = {4}, year = {2010}, month = {2010}, pages = {174}, abstract = {BACKGROUND: Pelobacter species are commonly found in a number of subsurface environments, and are unique members of the Geobacteraceae family. They are phylogenetically intertwined with both Geobacter and Desulfuromonas species. Pelobacter species likely play important roles in the fermentative degradation of unusual organic matters and syntrophic metabolism in the natural environments, and are of interest for applications in bioremediation and microbial fuel cells. RESULTS: In order to better understand the physiology of Pelobacter species, genome-scale metabolic models for Pelobacter carbinolicus and Pelobacter propionicus were developed. Model development was greatly aided by the availability of models of the closely related Geobacter sulfurreducens and G. metallireducens. The reconstructed P. carbinolicus model contains 741 genes and 708 reactions, whereas the reconstructed P. propionicus model contains 661 genes and 650 reactions. A total of 470 reactions are shared among the two Pelobacter models and the two Geobacter models. The different reactions between the Pelobacter and Geobacter models reflect some unique metabolic capabilities such as fermentative growth for both Pelobacter species. The reconstructed Pelobacter models were validated by simulating published growth conditions including fermentations, hydrogen production in syntrophic co-culture conditions, hydrogen utilization, and Fe(III) reduction. Simulation results matched well with experimental data and indicated the accuracy of the models. CONCLUSIONS: We have developed genome-scale metabolic models of P. carbinolicus and P. propionicus. These models of Pelobacter metabolism can now be incorporated into the growing repertoire of genome scale models of the Geobacteraceae family to aid in describing the growth and activity of these organisms in anoxic environments and in the study of their roles and interactions in the subsurface microbial community.}, keywords = {Anaerobiosis, Citric Acid Cycle, Desulfuromonas, Electron Transport, Energy Metabolism, Gene Expression Regulation, Bacterial, Models, Biological, Reproducibility of Results, Sulfur}, issn = {1752-0509}, doi = {10.1186/1752-0509-4-174}, author = {Sun, Jun and Haveman, Shelley A and Bui, Olivia and Fahland, Tom R and Lovley, Derek R} } @article {677, title = {ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids.}, journal = {Nucleic Acids Res}, volume = {38}, year = {2010}, month = {2010 Jul}, pages = {W529-33}, abstract = {It is informative to detect highly conserved positions in proteins and nucleic acid sequence/structure since they are often indicative of structural and/or functional importance. ConSurf (http://consurf.tau.ac.il) and ConSeq (http://conseq.tau.ac.il) are two well-established web servers for calculating the evolutionary conservation of amino acid positions in proteins using an empirical Bayesian inference, starting from protein structure and sequence, respectively. Here, we present the new version of the ConSurf web server that combines the two independent servers, providing an easier and more intuitive step-by-step interface, while offering the user more flexibility during the process. In addition, the new version of ConSurf calculates the evolutionary rates for nucleic acid sequences. The new version is freely available at: http://consurf.tau.ac.il/.}, keywords = {Amino Acid Sequence, Base Sequence, Conserved Sequence, Evolution, Molecular, Internet, Nucleic Acid Conformation, Protein Conformation, Sequence Analysis, Sequence Homology, Amino Acid, Software}, issn = {1362-4962}, doi = {10.1093/nar/gkq399}, author = {Ashkenazy, Haim and Erez, Elana and Martz, Eric and Pupko, Tal and Ben-Tal, Nir} } @article {689, title = {Controlled expression of branch-forming mannosyltransferase is critical for mycobacterial lipoarabinomannan biosynthesis.}, journal = {J Biol Chem}, volume = {285}, year = {2010}, month = {2010 Apr 30}, pages = {13326-36}, abstract = {Lipomannan (LM) and lipoarabinomannan (LAM) are phosphatidylinositol-anchored glycans present in the mycobacterial cell wall. In Mycobacterium smegmatis, the mannan core of LM/LAM constitutes a linear chain of 20-25 alpha1,6-mannoses elaborated by 8-9 alpha1,2-monomannose side branches. At least two alpha1,6-mannosyltransferases mediate the linear mannose chain elongation, and one branching alpha1,2-mannosyltransferase (encoded by MSMEG_4247) transfers monomannose branches. An MSMEG_4247 deletion mutant accumulates branchless LAM and interestingly fails to accumulate LM, suggesting an unexpected role of mannose branching for LM synthesis or maintenance. To understand the roles of MSMEG_4247-mediated branching more clearly, we analyzed the MSMEG_4247 deletion mutant in detail. Our study showed that the deletion mutant restored the synthesis of wild-type LM and LAM upon the expression of MSMEG_4247 at wild-type levels. In striking contrast, overexpression of MSMEG_4247 resulted in the accumulation of dwarfed LM/LAM, although monomannose branching was restored. The dwarfed LAM carried a mannan chain less than half the length of wild-type LAM and was elaborated by an arabinan that was about 4 times smaller. Induced overexpression of an elongating alpha1,6-mannosyltransferase competed with the overexpressed branching enzyme, alleviating the dwarfing effect of the branching enzyme. In wild-type cells, LM and LAM decreased in quantity in the stationary phase, and the expression levels of branching and elongating mannosyltransferases were reduced in concert, presumably to avoid producing abnormal LM/LAM. These data suggest that the coordinated expressions of branching and elongating mannosyltransferases are critical for mannan backbone elongation.}, keywords = {Bacterial Proteins, Cell Wall, Gene Deletion, Lipopolysaccharides, Mannose, Mannosyltransferases, Mycobacterium smegmatis}, issn = {1083-351X}, doi = {10.1074/jbc.M109.077297}, author = {Sena, Chubert B C and Fukuda, Takeshi and Miyanagi, Kana and Matsumoto, Sohkichi and Kobayashi, Kazuo and Murakami, Yoshiko and Maeda, Yusuke and Kinoshita, Taroh and Morita, Yasu S} } @article {739, title = {Comparison of facially amphiphilic versus segregated monomers in the design of antibacterial copolymers.}, journal = {Chemistry}, volume = {15}, year = {2009}, month = {2009}, pages = {433-9}, abstract = {A direct comparison of two strategies for designing antimicrobial polymers is presented. Previously, we published several reports on the use of facially amphiphilic (FA) monomers which led to polynorbornenes with excellent antimicrobial activities and selectivities. Our polymers obtained by copolymerization of structurally similar segregated monomers, in which cationic and non-polar moieties reside on separate repeat units, led to polymers with less pronounced activities. A wide range of polymer amphiphilicities was surveyed by pairing a cationic oxanorbornene with eleven different non-polar monomers and varying the comonomer feed ratios. Their properties were tested using antimicrobial assays and copolymers possessing intermediate hydrophobicities were the most active. Polymer-induced leakage of dye-filled liposomes and microscopy of polymer-treated bacteria support a membrane-based mode of action. From these results there appears to be profound differences in how a polymer made from FA monomers interacts with the phospholipid bilayer compared with copolymers from segregated monomers. We conclude that a well-defined spatial relationship of the whole polymer is crucial to obtain synthetic mimics of antimicrobial peptides (SMAMPs): charged and non-polar moieties need to be balanced locally, for example, at the monomer level, and not just globally. We advocate the use of FA monomers for better control of biological properties. It is expected that this principle will be usefully applied to other backbones such as the polyacrylates, polystyrenes, and non-natural polyamides.}, keywords = {Anti-Bacterial Agents, Bacteria, Cell Membrane, Drug Design, Hemolysis, Plastics, Polymers, Surface-Active Agents}, issn = {1521-3765}, doi = {10.1002/chem.200801233}, author = {Gabriel, Gregory J and Maegerlein, Janet A and Nelson, Christopher F and Dabkowski, Jeffrey M and Eren, Tarik and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {768, title = {Correlation between tick density and pathogen endemicity, New Hampshire.}, journal = {Emerg Infect Dis}, volume = {15}, year = {2009}, month = {2009 Apr}, pages = {585-7}, abstract = {To assess the endemicity of tick-borne pathogens in New Hampshire, we surveyed adult tick vectors. Pathogens were more prevalent in areas of high tick density, suggesting a correlation between tick establishment and pathogen endemicity. Infection rates in ticks correlated with disease frequency in humans.}, keywords = {Anaplasma phagocytophilum, Animals, Arachnid Vectors, Babesia microti, Babesiosis, Biosurveillance, Borrelia burgdorferi, Communicable Diseases, Emerging, Ehrlichiosis, Humans, Ixodes, Lyme Disease, New Hampshire}, issn = {1080-6059}, doi = {10.3201/eid1504.080940}, author = {Walk, Seth T and Xu, Guang and Stull, Jason W and Rich, Stephen M} } @article {464, title = {Coupling a genome-scale metabolic model with a reactive transport model to describe in situ uranium bioremediation.}, journal = {Microb Biotechnol}, volume = {2}, year = {2009}, month = {2009 Mar}, pages = {274-86}, abstract = {The increasing availability of the genome sequences of microorganisms involved in important bioremediation processes makes it feasible to consider developing genome-scale models that can aid in predicting the likely outcome of potential subsurface bioremediation strategies. Previous studies of the in situ bioremediation of uranium-contaminated groundwater have demonstrated that Geobacter species are often the dominant members of the groundwater community during active bioremediation and the primary organisms catalysing U(VI) reduction. Therefore, a genome-scale, constraint-based model of the metabolism of Geobacter sulfurreducens was coupled with the reactive transport model HYDROGEOCHEM in an attempt to model in situ uranium bioremediation. In order to simplify the modelling, the influence of only three growth factors was considered: acetate, the electron donor added to stimulate U(VI) reduction; Fe(III), the electron acceptor primarily supporting growth of Geobacter; and ammonium, a key nutrient. The constraint-based model predicted that growth yields of Geobacter varied significantly based on the availability of these three growth factors and that there are minimum thresholds of acetate and Fe(III) below which growth and activity are not possible. This contrasts with typical, empirical microbial models that assume fixed growth yields and the possibility for complete metabolism of the substrates. The coupled genome-scale and reactive transport model predicted acetate concentrations and U(VI) reduction rates in a field trial of in situ uranium bioremediation that were comparable to the predictions of a calibrated conventional model, but without the need for empirical calibration, other than specifying the initial biomass of Geobacter. These results suggest that coupling genome-scale metabolic models with reactive transport models may be a good approach to developing models that can be truly predictive, without empirical calibration, for evaluating the probable response of subsurface microorganisms to possible bioremediation approaches prior to implementation.}, keywords = {Acetates, Biodegradation, Environmental, Biological Transport, Genome, Bacterial, Geobacter, Iron, Models, Biological, Uranium}, issn = {1751-7915}, doi = {10.1111/j.1751-7915.2009.00087.x}, author = {Scheibe, Timothy D and Mahadevan, Radhakrishnan and Fang, Yilin and Garg, Srinath and Long, Philip E and Lovley, Derek R} } @article {486, title = {Characterization of extracellular minerals produced during dissimilatory Fe(III) and U(VI) reduction at 100 degrees C by Pyrobaculum islandicum.}, journal = {Geobiology}, volume = {6}, year = {2008}, month = {2008 Mar}, pages = {147-54}, abstract = {In order to gain insight into the significance of biotic metal reduction and mineral formation in hyperthermophilic environments, metal mineralization as a result of the dissimilatory reduction of poorly crystalline Fe(III) oxide, and U(VI) reduction at 100 degrees C by Pyrobaculum islandicum was investigated. When P. islandicum was grown in a medium with poorly crystalline Fe(III) oxide as an electron acceptor and hydrogen as an electron donor, the Fe(III) oxide was reduced to an extracellular, ultrafine-grained magnetite with characteristics similar to that found in some hot environments and that was previously thought to be of abiotic origin. Furthermore, cell suspensions of P. islandicum rapidly reduced the soluble and oxidized form of uranium, U(VI), to extracellular precipitates of the highly insoluble U(IV) mineral, uraninite (UO(2)). The reduction of U(VI) was dependent on the presence of hydrogen as the electron donor. These findings suggest that microbes may play a key role in metal deposition in hyperthermophilic environments and provide a plausible explanation for such phenomena as magnetite accumulation and formation of uranium deposits at ca. 100 degrees C.}, keywords = {Ferric Compounds, Ferumoxytol, Hot Temperature, Hydrogen, Minerals, Oxidation-Reduction, Pyrobaculum, Uranium, X-Ray Diffraction}, issn = {1472-4669}, doi = {10.1111/j.1472-4669.2007.00142.x}, author = {Kashefi, K and Moskowitz, B M and Lovley, D R} } @article {700, title = {Characterizing regulation of metabolism in Geobacter sulfurreducens through genome-wide expression data and sequence analysis.}, journal = {OMICS}, volume = {12}, year = {2008}, month = {2008 Mar}, pages = {33-59}, abstract = {Geobacteraceae are a family of metal reducing bacteria with important applications in bioremediation and electricity generation. G. sulfurreducens is a representative of Geobacteraceae that has been extensively studied with the goal of extending the understanding of this family of organisms for optimizing their practical applications. Here, we have analyzed gene expression data from 10 experiments involving environmental and genetic perturbations and have identified putative transcription factor binding sites (TFBS) involved in regulating key aspects of metabolism. Specifically, we considered data from both a subset of 10 microarray experiments (7 of 10) and all 10 experiments. The expression data from these two sets were independently clustered, and the upstream regions of genes and operons from the clusters in both sets were used to identify TFBS using the AlignACE program. This analysis resulted in the identification of motifs upstream of several genes involved in central metabolism, sulfate assimilation, and energy metabolism, as well as genes potentially encoding acetate permease. Further, similar TFBS were identified from the analysis of both sets, suggesting that these TFBS are significant in the regulation of metabolism in G. sulfurreducens. In addition, we have utilized microarray data to derive condition specific constraints on the capacity of key enzymes in central metabolism. We have incorporated these constraints into the metabolic model of G. sulfurreducens and simulated Fe(II)-limited growth. The resulting prediction was consistent with data, suggesting that regulatory constraints are important for simulating growth phenotypes in nonoptimal environments.}, keywords = {Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Models, Genetic, Oligonucleotide Array Sequence Analysis, Sequence Analysis, DNA, Transcription, Genetic}, issn = {1536-2310}, doi = {10.1089/omi.2007.0043}, author = {Mahadevan, Radhakrishnan and Yan, Bin and Postier, Brad and Nevin, Kelly P and Woodard, Trevor L and O{\textquoteright}Neil, Regina and Coppi, Maddalena V and Meth{\'e}, Barbara A and Krushkal, Julia} } @article {470, title = {Comparative genomics of Geobacter chemotaxis genes reveals diverse signaling function.}, journal = {BMC Genomics}, volume = {9}, year = {2008}, month = {2008}, pages = {471}, abstract = {BACKGROUND: Geobacter species are delta-Proteobacteria and are often the predominant species in a variety of sedimentary environments where Fe(III) reduction is important. Their ability to remediate contaminated environments and produce electricity makes them attractive for further study. Cell motility, biofilm formation, and type IV pili all appear important for the growth of Geobacter in changing environments and for electricity production. Recent studies in other bacteria have demonstrated that signaling pathways homologous to the paradigm established for Escherichia coli chemotaxis can regulate type IV pili-dependent motility, the synthesis of flagella and type IV pili, the production of extracellular matrix material, and biofilm formation. The classification of these pathways by comparative genomics improves the ability to understand how Geobacter thrives in natural environments and better their use in microbial fuel cells. RESULTS: The genomes of G. sulfurreducens, G. metallireducens, and G. uraniireducens contain multiple (approximately 70) homologs of chemotaxis genes arranged in several major clusters (six, seven, and seven, respectively). Unlike the single gene cluster of E. coli, the Geobacter clusters are not all located near the flagellar genes. The probable functions of some Geobacter clusters are assignable by homology to known pathways; others appear to be unique to the Geobacter sp. and contain genes of unknown function. We identified large numbers of methyl-accepting chemotaxis protein (MCP) homologs that have diverse sensing domain architectures and generate a potential for sensing a great variety of environmental signals. We discuss mechanisms for class-specific segregation of the MCPs in the cell membrane, which serve to maintain pathway specificity and diminish crosstalk. Finally, the regulation of gene expression in Geobacter differs from E. coli. The sequences of predicted promoter elements suggest that the alternative sigma factors sigma28 and sigma54 play a role in regulating the Geobacter chemotaxis gene expression. CONCLUSION: The numerous chemoreceptors and chemotaxis-like gene clusters of Geobacter appear to be responsible for a diverse set of signaling functions in addition to chemotaxis, including gene regulation and biofilm formation, through functionally and spatially distinct signaling pathways.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Chemotaxis, Escherichia coli, Gene Expression Regulation, Bacterial, Genome, Bacterial, Genomics, Geobacter, Membrane Proteins, Molecular Sequence Data, Multigene Family, Promoter Regions, Genetic, Sequence Alignment, Sequence Homology, Amino Acid}, issn = {1471-2164}, doi = {10.1186/1471-2164-9-471}, author = {Tran, Hoa T and Krushkal, Julia and Antommattei, Frances M and Lovley, Derek R and Weis, Robert M} } @article {489, title = {Computational and experimental analysis of redundancy in the central metabolism of Geobacter sulfurreducens.}, journal = {PLoS Comput Biol}, volume = {4}, year = {2008}, month = {2008 Feb}, pages = {e36}, abstract = {Previous model-based analysis of the metabolic network of Geobacter sulfurreducens suggested the existence of several redundant pathways. Here, we identified eight sets of redundant pathways that included redundancy for the assimilation of acetate, and for the conversion of pyruvate into acetyl-CoA. These equivalent pathways and two other sub-optimal pathways were studied using 5 single-gene deletion mutants in those pathways for the evaluation of the predictive capacity of the model. The growth phenotypes of these mutants were studied under 12 different conditions of electron donor and acceptor availability. The comparison of the model predictions with the resulting experimental phenotypes indicated that pyruvate ferredoxin oxidoreductase is the only activity able to convert pyruvate into acetyl-CoA. However, the results and the modeling showed that the two acetate activation pathways present are not only active, but needed due to the additional role of the acetyl-CoA transferase in the TCA cycle, probably reflecting the adaptation of these bacteria to acetate utilization. In other cases, the data reconciliation suggested additional capacity constraints that were confirmed with biochemical assays. The results demonstrate the need to experimentally verify the activity of key enzymes when developing in silico models of microbial physiology based on sequence-based reconstruction of metabolic networks.}, keywords = {Bacterial Proteins, Base Sequence, Computer Simulation, Gene Expression Regulation, Bacterial, Geobacter, Models, Biological, Molecular Sequence Data, Multienzyme Complexes, Signal Transduction}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.0040036}, author = {Segura, Daniel and Mahadevan, Radhakrishnan and Ju{\'a}rez, Katy and Lovley, Derek R} } @article {1210, title = {Constraints on anaerobic respiration in the hyperthermophilic Archaea Pyrobaculum islandicum and Pyrobaculum aerophilum.}, journal = {Appl Environ Microbiol}, volume = {74}, year = {2008}, month = {2008 Jan}, pages = {396-402}, abstract = {

Pyrobaculum islandicum uses iron, thiosulfate, and elemental sulfur for anaerobic respiration, while Pyrobaculum aerophilum uses iron and nitrate; however, the constraints on these processes and their physiological mechanisms for iron and sulfur reduction are not well understood. Growth rates on sulfur compounds are highest at pH 5 to 6 and highly reduced (<-420-mV) conditions, while growth rates on nitrate and iron are highest at pH 7 to 9 and more-oxidized (>-210-mV) conditions. Growth on iron expands the known pH range of growth for both organisms. P. islandicum differs from P. aerophilum in that it requires direct contact with insoluble iron oxide for growth, it did not produce any extracellular compounds when grown on insoluble iron, and it lacked 2,6-anthrahydroquinone disulfonate oxidase activity. Furthermore, iron reduction in P. islandicum appears to be completely independent of c-type cytochromes. Like that in P. aerophilum, NADH-dependent ferric reductase activity in P. islandicum increased significantly in iron-grown cultures relative to that in non-iron-grown cultures. Proteomic analyses showed that there were significant increases in the amounts of a putative membrane-bound thiosulfate reductase in P. islandicum cultures grown on thiosulfate relative to those in cultures grown on iron and elemental sulfur. This is the first evidence of this enzyme being used in either a hyperthermophile or an archaeon. Pyrobaculum arsenaticum and Pyrobaculum calidifontis also grew on Fe(III) citrate and insoluble iron oxide, but only P. arsenaticum could grow on insoluble iron without direct contact.

}, keywords = {Anaerobiosis, FMN Reductase, Hydrogen-Ion Concentration, Iron, Nitrates, Oxidation-Reduction, Pyrobaculum, Species Specificity, Sulfur, Sulfurtransferases, Thiosulfates}, issn = {1098-5336}, doi = {10.1128/AEM.02033-07}, author = {Feinberg, Lawrence F and Srikanth, R and Vachet, Richard W and Holden, James F} } @article {1212, title = {Characterization of malate dehydrogenase from the hyperthermophilic archaeon Pyrobaculum islandicum.}, journal = {Extremophiles}, volume = {11}, year = {2007}, month = {2007 Sep}, pages = {741-6}, abstract = {

Native and recombinant malate dehydrogenase (MDH) was characterized from the hyperthermophilic, facultatively autotrophic archaeon Pyrobaculum islandicum. The enzyme is a homotetramer with a subunit mass of 33 kDa. The activity kinetics of the native and recombinant proteins are the same. The apparent K ( m ) values of the recombinant protein for oxaloacetate (OAA) and NADH (at 80 degrees C and pH 8.0) were 15 and 86 microM, respectively, with specific activity as high as 470 U mg(-1). Activity decreased more than 90\% when NADPH was used. The catalytic efficiency of OAA reduction by P. islandicum MDH using NADH was significantly higher than that reported for any other archaeal MDH. Unlike other archaeal MDHs, specific activity of the P. islandicum MDH back-reaction also decreased more than 90\% when malate and NAD(+) were used as substrates and was not detected with NADP(+). A phylogenetic tree of 31 archaeal MDHs shows that they fall into 5 distinct groups separated largely along taxonomic lines suggesting minimal lateral mdh transfer between Archaea.

}, keywords = {Citric Acid Cycle, Hydrogen-Ion Concentration, Kinetics, Malate Dehydrogenase, Malates, Molecular Weight, NAD, NADP, Oxaloacetic Acid, Phylogeny, Protein Conformation, Protein Subunits, Pyrobaculum, Recombinant Proteins, Sequence Analysis, Protein, Substrate Specificity, Temperature}, issn = {1431-0651}, doi = {10.1007/s00792-007-0081-2}, author = {Yennaco, Lynda J and Hu, Yajing and Holden, James F} } @article {746, title = {Comparative bacterial diversity in recent Hawaiian volcanic deposits of different ages.}, journal = {FEMS Microbiol Ecol}, volume = {60}, year = {2007}, month = {2007 Apr}, pages = {60-73}, abstract = {Volcanic activity creates new landforms that can change dramatically over time as a consequence of biotic succession. Nonetheless, volcanic deposits present severe constraints for microbial colonization and activity. We have characterized bacterial diversity on four recent deposits at Kilauea volcano, Hawaii (KVD). Much of the diversity was either closely related to uncultured organisms or distinct from any reported 16S rRNA gene sequences. Diversity indices suggested that diversity was highest in a moderately vegetated 210-year-old ash deposit (1790-KVD), and lowest for a 79-year-old lava flow (1921-KVD). Diversity for a 41-year-old tephra deposit (1959-KVD) and a 300-year-old rainforest (1700-KVD) reached intermediate values. The 1959-KVD and 1790-KVD communities were dominated by Acidobacteria, Alpha- and Gammaproteobacteria, Actinobacteria, Cyanobacteria, and many unclassified phylotypes. The 1921-KVD, an unvegetated low pH deposit, was dominated by unclassified phylotypes. In contrast, 1700-KVD was primarily populated by Alphaproteobacteria with very few unclassified phylotypes. Similar diversity indices and levels of trace gas flux were found for 1959-KVD and 1790-KVD; however, statistical analyses indicated significantly different communities. This study not only showed that microorganisms colonize recent volcanic deposits and are able to establish diverse communities, but also that their composition is governed by variations in local deposit parameters.}, keywords = {Bacteria, DNA, Bacterial, Ecosystem, Genes, rRNA, Genetic Variation, Hawaii, Molecular Sequence Data, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Soil Microbiology, Time Factors, Volcanic Eruptions}, issn = {0168-6496}, doi = {10.1111/j.1574-6941.2006.00253.x}, author = {Gomez-Alvarez, Vicente and King, Gary M and N{\"u}sslein, Klaus} } @article {1214, title = {Characterization of dissimilatory Fe(III) versus NO3- reduction in the hyperthermophilic archaeon Pyrobaculum aerophilum.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Jan}, pages = {525-31}, abstract = {

The hyperthermophilic archaeon Pyrobaculum aerophilum used 20 mM Fe(III) citrate, 100 mM poorly crystalline Fe(III) oxide, and 10 mM KNO3 as terminal electron acceptors. The two forms of iron were reduced at different rates but with equal growth yields. The insoluble iron was reduced when segregated spatially by dialysis tubing, indicating that direct contact with the iron was not necessary for growth. When partitioned, there was no detectable Fe(III) or Fe(II) outside of the tubing after growth, suggesting that an electron shuttle, not a chelator, may be used as an extracellular mediator of iron reduction. The addition of 25 and 50\% (vol vol(-1)) cell-free spent insoluble iron media to fresh media led to growth without a lag phase. Liquid chromatography analysis of spent media showed that cultures grown in iron, especially insoluble iron, produced soluble extracellular compounds that were absent or less abundant in spent nitrate medium. NADH-dependent ferric reductase activity increased approximately 100-fold, while nitrate reductase activity decreased 10-fold in whole-cell extracts from iron-grown cells relative to those from nitrate-grown cells, suggesting that dissimilatory iron reduction was regulated. A novel 2,6-anthrahydroquinone disulfonate oxidase activity was more than 580-fold higher in iron-grown cells than in nitrate-grown cells. The activity was primarily (>95\%) associated with the membrane cellular fraction, but its physiological function is unknown. Nitrate-grown cultures produced two membrane-bound, c-type cytochromes that are predicted to be monoheme and part of nitrite reductase and a bc1 complex using genome analyses. Only one cytochrome was present in cells grown on Fe(III) citrate whose relative abundance was unchanged.

}, keywords = {Cytochromes c, Ferric Compounds, FMN Reductase, Hot Temperature, NAD, Nitrates, Oxidation-Reduction, Pyrobaculum}, issn = {0021-9193}, doi = {10.1128/JB.188.2.525-531.2006}, author = {Feinberg, Lawrence F and Holden, James F} } @article {524, title = {Characterization of metabolism in the Fe(III)-reducing organism Geobacter sulfurreducens by constraint-based modeling.}, journal = {Appl Environ Microbiol}, volume = {72}, year = {2006}, month = {2006 Feb}, pages = {1558-68}, abstract = {Geobacter sulfurreducens is a well-studied representative of the Geobacteraceae, which play a critical role in organic matter oxidation coupled to Fe(III) reduction, bioremediation of groundwater contaminated with organics or metals, and electricity production from waste organic matter. In order to investigate G. sulfurreducens central metabolism and electron transport, a metabolic model which integrated genome-based predictions with available genetic and physiological data was developed via the constraint-based modeling approach. Evaluation of the rates of proton production and consumption in the extracellular and cytoplasmic compartments revealed that energy conservation with extracellular electron acceptors, such as Fe(III), was limited relative to that associated with intracellular acceptors. This limitation was attributed to lack of cytoplasmic proton consumption during reduction of extracellular electron acceptors. Model-based analysis of the metabolic cost of producing an extracellular electron shuttle to promote electron transfer to insoluble Fe(III) oxides demonstrated why Geobacter species, which do not produce shuttles, have an energetic advantage over shuttle-producing Fe(III) reducers in subsurface environments. In silico analysis also revealed that the metabolic network of G. sulfurreducens could synthesize amino acids more efficiently than that of Escherichia coli due to the presence of a pyruvate-ferredoxin oxidoreductase, which catalyzes synthesis of pyruvate from acetate and carbon dioxide in a single step. In silico phenotypic analysis of deletion mutants demonstrated the capability of the model to explore the flexibility of G. sulfurreducens central metabolism and correctly predict mutant phenotypes. These results demonstrate that iterative modeling coupled with experimentation can accelerate the understanding of the physiology of poorly studied but environmentally relevant organisms and may help optimize their practical applications.}, keywords = {Amino Acids, Electron Transport, Escherichia coli, Fumarates, Geobacter, Iron, Models, Biological, Mutation, Oxidation-Reduction, Phenotype, Protons, Quinones, Species Specificity}, issn = {0099-2240}, doi = {10.1128/AEM.72.2.1558-1568.2006}, author = {Mahadevan, R and Bond, D R and Butler, J E and Esteve-Nu{\~n}ez, A and Coppi, M V and Palsson, B O and Schilling, C H and Lovley, D R} } @article {1213, title = {Citric acid cycle in the hyperthermophilic archaeon Pyrobaculum islandicum grown autotrophically, heterotrophically, and mixotrophically with acetate.}, journal = {J Bacteriol}, volume = {188}, year = {2006}, month = {2006 Jun}, pages = {4350-5}, abstract = {

The hyperthermophilic archaeon Pyrobaculum islandicum uses the citric acid cycle in the oxidative and reductive directions for heterotrophic and autotrophic growth, respectively, but the control of carbon flow is poorly understood. P. islandicum was grown at 95 degrees C autotrophically, heterotrophically, and mixotrophically with acetate, H2, and small amounts of yeast extract and with thiosulfate as the terminal electron acceptor. The autotrophic growth rates and maximum concentrations of cells were significantly lower than those in other media. The growth rates on H2 and 0.001\% yeast extract with and without 0.05\% acetate were the same, but the maximum concentration of cells was fourfold higher with acetate. There was no growth with acetate if 0.001\% yeast extract was not present, and addition of H2 to acetate-containing medium greatly increased the growth rates and maximum concentrations of cells. P. islandicum cultures assimilated 14C-labeled acetate in the presence of H2 and yeast extract with an efficiency of 55\%. The activities of 11 of 19 enzymes involved in the central metabolism of P. islandicum were regulated under the three different growth conditions. Pyruvate synthase and acetate:coenzyme A (CoA) ligase (ADP-forming) activities were detected only in heterotrophically grown cultures. Citrate synthase activity decreased in autotrophic and acetate-containing cultures compared to the activity in heterotrophic cultures. Acetylated citrate lyase, acetate:CoA ligase (AMP forming), and phosphoenolpyruvate carboxylase activities increased in autotrophic and acetate-containing cultures. Citrate lyase activity was higher than ATP citrate synthase activity in autotrophic cultures. These data suggest that citrate lyase and AMP-forming acetate:CoA ligase, but not ATP citrate synthase, work opposite citrate synthase to control the direction of carbon flow in the citric acid cycle.

}, keywords = {Acetates, Carbon Dioxide, Citric Acid, Citric Acid Cycle, Culture Media, Deuterium, Ligases, Lyases, Pyrobaculum, Pyruvate Synthase, Substrate Specificity}, issn = {0021-9193}, doi = {10.1128/JB.00138-06}, author = {Hu, Yajing and Holden, James F} } @article {512, title = {Computational prediction of RpoS and RpoD regulatory sites in Geobacter sulfurreducens using sequence and gene expression information.}, journal = {Gene}, volume = {384}, year = {2006}, month = {2006 Dec 15}, pages = {73-95}, abstract = {RpoS, the sigma S subunit of RNA polymerase, is vital during the growth and survival of Geobacter sulfurreducens under conditions typically encountered in its native subsurface environments. We investigated the conservation of sites that may be important for RpoS function in G. sulfurreducens. We also employed sequence information and expression microarray data to predict G. sulfurreducens genome sites that may be related to RpoS regulation. Hierarchical clustering identified three clusters of significantly downregulated genes in the rpoS deletion mutant. The search for conserved overrepresented motifs in co-regulated operons identified likely -35 and -10 promoter elements upstream of a number of functionally important G. sulfurreducens operons that were downregulated in the rpoS deletion mutant. Putative -35/-10 promoter elements were also identified in the G. sulfurreducens genome using sequence similarity searches to matrices of -35/-10 promoter elements found in G. sulfurreducens and in Escherichia coli. Due to a sufficient degree of sequence similarity between -35/-10 promoter elements for RpoS, RpoD, and other sigma factors, both the sequence similarity searches and the search for conserved overrepresented motifs using microarray data may identify promoter elements for both RpoS and other sigma factors.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Base Sequence, Citrates, Computational Biology, Conserved Sequence, DNA-Directed RNA Polymerases, Escherichia coli, Escherichia coli Proteins, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Genes, Bacterial, Genome, Bacterial, Geobacter, Molecular Sequence Data, Oligonucleotide Array Sequence Analysis, Operon, Promoter Regions, Genetic, Sigma Factor, Transcription, Genetic}, issn = {0378-1119}, doi = {10.1016/j.gene.2006.06.025}, author = {Yan, Bin and N{\'u}{\~n}ez, Cinthia and Ueki, Toshiyuki and Esteve-N{\'u}{\~n}ez, Abraham and Puljic, Marko and Adkins, Ronald M and Meth{\'e}, Barbara A and Lovley, Derek R and Krushkal, Julia} } @article {515, title = {c-Type cytochromes in Pelobacter carbinolicus.}, journal = {Appl Environ Microbiol}, volume = {72}, year = {2006}, month = {2006 Nov}, pages = {6980-5}, abstract = {Previous studies failed to detect c-type cytochromes in Pelobacter species despite the fact that other close relatives in the Geobacteraceae, such as Geobacter and Desulfuromonas species, have abundant c-type cytochromes. Analysis of the recently completed genome sequence of Pelobacter carbinolicus revealed 14 open reading frames that could encode c-type cytochromes. Transcripts for all but one of these open reading frames were detected in acetoin-fermenting and/or Fe(III)-reducing cells. Three putative c-type cytochrome genes were expressed specifically during Fe(III) reduction, suggesting that the encoded proteins may participate in electron transfer to Fe(III). One of these proteins was a periplasmic triheme cytochrome with a high level of similarity to PpcA, which has a role in Fe(III) reduction in Geobacter sulfurreducens. Genes for heme biosynthesis and system II cytochrome c biogenesis were identified in the genome and shown to be expressed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels of protein extracted from acetoin-fermenting P. carbinolicus cells contained three heme-staining bands which were confirmed by mass spectrometry to be among the 14 predicted c-type cytochromes. The number of cytochrome genes, the predicted amount of heme c per protein, and the ratio of heme-stained protein to total protein were much smaller in P. carbinolicus than in G. sulfurreducens. Furthermore, many of the c-type cytochromes that genetic studies have indicated are required for optimal Fe(III) reduction in G. sulfurreducens were not present in the P. carbinolicus genome. These results suggest that further evaluation of the functions of c-type cytochromes in the Geobacteraceae is warranted.}, keywords = {Bacterial Proteins, Cytochromes c, Deltaproteobacteria, Heme, Polymerase Chain Reaction, Proteomics, Reverse Transcriptase Polymerase Chain Reaction, RNA, Messenger}, issn = {0099-2240}, doi = {10.1128/AEM.01128-06}, author = {Haveman, Shelley A and Holmes, Dawn E and Ding, Yan-Huai R and Ward, Joy E and Didonato, Raymond J and Lovley, Derek R} } @article {717, title = {The caveolae-mediated sv40 entry pathway bypasses the golgi complex en route to the endoplasmic reticulum.}, journal = {Virol J}, volume = {2}, year = {2005}, month = {2005}, pages = {38}, abstract = {BACKGROUND: Simian virus 40 (SV40) enters cells via an atypical caveolae-mediated endocytic pathway, which delivers the virus to a new intermediary compartment, the caveosome. The virus then is believed to go directly from the caveosome to the endoplasmic reticulum. Cholera toxin likewise enters via caveolae and traffics to caveosomes. But, in contrast to SV40, cholera toxin is transported from caveosomes to the endoplasmic reticulum via the Golgi. For that reason, and because the caveosome and Golgi may have some common markers, we revisited the issue of whether SV40 might access the endoplasmic reticulum via the Golgi. RESULTS: We confirmed our earlier finding that SV40 co localizes with the Golgi marker beta-COP. However, we show that the virus does not co localize with the more discriminating Golgi markers, golgin 97 and BODIPY-ceramide. CONCLUSION: The caveolae-mediated SV40 entry pathway does not intersect the Golgi. SV40 is seen to co localize with beta-COP because that protein is a marker for caveosomes as well as the Golgi. Moreover, these results are consistent with the likelihood that the caveosome is a sorting organelle. In addition, there are at least two distinct but related routes by which a ligand might traffic from the caveosome to the ER; one route involving transport through the Golgi, and another pathway that does not involve the Golgi.}, keywords = {Animals, Caveolae, Cercopithecus aethiops, Endoplasmic Reticulum, Fibroblasts, Golgi Apparatus, Immunohistochemistry, Simian virus 40}, issn = {1743-422X}, doi = {10.1186/1743-422X-2-38}, author = {Norkin, Leonard C and Kuksin, Dmitry} } @article {533, title = {Characterization of citrate synthase from Geobacter sulfurreducens and evidence for a family of citrate synthases similar to those of eukaryotes throughout the Geobacteraceae.}, journal = {Appl Environ Microbiol}, volume = {71}, year = {2005}, month = {2005 Jul}, pages = {3858-65}, abstract = {Members of the family Geobacteraceae are commonly the predominant Fe(III)-reducing microorganisms in sedimentary environments, as well as on the surface of energy-harvesting electrodes, and are able to effectively couple the oxidation of acetate to the reduction of external electron acceptors. Citrate synthase activity of these organisms is of interest due to its key role in acetate metabolism. Prior sequencing of the genome of Geobacter sulfurreducens revealed a putative citrate synthase sequence related to the citrate synthases of eukaryotes. All citrate synthase activity in G. sulfurreducens could be resolved to a single 49-kDa protein via affinity chromatography. The enzyme was successfully expressed at high levels in Escherichia coli with similar properties as the native enzyme, and kinetic parameters were comparable to related citrate synthases (kcat= 8.3 s(-1); Km= 14.1 and 4.3 microM for acetyl coenzyme A and oxaloacetate, respectively). The enzyme was dimeric and was slightly inhibited by ATP (Ki= 1.9 mM for acetyl coenzyme A), which is a known inhibitor for many eukaryotic, dimeric citrate synthases. NADH, an allosteric inhibitor of prokaryotic hexameric citrate synthases, did not affect enzyme activity. Unlike most prokaryotic dimeric citrate synthases, the enzyme did not have any methylcitrate synthase activity. A unique feature of the enzyme, in contrast to citrate synthases from both eukaryotes and prokaryotes, was a lack of stimulation by K+ ions. Similar citrate synthase sequences were detected in a diversity of other Geobacteraceae members. This first characterization of a eukaryotic-like citrate synthase from a prokaryote provides new insight into acetate metabolism in Geobacteraceae members and suggests a molecular target for tracking the presence and activity of these organisms in the environment.}, keywords = {Amino Acid Sequence, Citrate (si)-Synthase, Culture Media, Deltaproteobacteria, DNA, Bacterial, Eukaryotic Cells, Geobacter, Kinetics, Molecular Sequence Data, Phylogeny, Sequence Analysis, DNA}, issn = {0099-2240}, doi = {10.1128/AEM.71.7.3858-3865.2005}, author = {Bond, Daniel R and Mester, T{\"u}nde and Nesb{\o}, Camilla L and Izquierdo-Lopez, Andrea V and Collart, Frank L and Lovley, Derek R} } @article {396, title = {Characterization of TetD as a transcriptional activator of a subset of genes of the Escherichia coli SoxS/MarA/Rob regulon.}, journal = {Mol Microbiol}, volume = {56}, year = {2005}, month = {2005 May}, pages = {1103-17}, abstract = {In Escherichia coli, SoxS, MarA and Rob form a closely related subset of the AraC/XylS family of positive regulators, sharing approximately 42\% amino acid sequence identity over the length of SoxS and the ability to activate transcription of a common set of target genes that provide resistance to redox-cycling compounds and antibiotics. On the basis of its approximately 43\% amino acid sequence identity with SoxS, MarA and Rob, TetD, encoded by transposon Tn10, appears to be a fourth member of the subset. However, although its expression has been shown to be negatively regulated by TetC and not inducible by tetracycline, the physiological function of TetD is unknown. Accordingly, in the work presented here, we initiate a molecular characterization of TetD. We show that expression of TetD activates transcription of a subset of the SoxS/MarA/Rob regulon genes and confers resistance to redox-cycling compounds and antibiotics. We show that mutations in the putative TetD binding site of a TetD-activatable promoter and a mutation in the protein{\textquoteright}s N-terminal DNA recognition helix interfere with transcription activation, thereby indicating that TetD directly activates target gene transcription. Finally, we show that TetD, like SoxS and MarA, is intrinsically unstable; however, unlike SoxS and MarA, TetD is not degraded by Lon or any of the cell{\textquoteright}s known cytoplasmic ATP-dependent proteases. Thus, we conclude that TetD is a bona fide member of the SoxS/MarA/Rob subfamily of positive regulators.}, keywords = {Amino Acid Sequence, Binding Sites, DNA, DNA-Binding Proteins, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Multigene Family, Promoter Regions, Genetic, Regulon, Tetracycline Resistance, Trans-Activators, Transcription Factors, Transcription, Genetic}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2005.04599.x}, author = {Griffith, Kevin L and Becker, Stephen M and Wolf, Richard E} } @article {696, title = {Compartmentalization of lipid biosynthesis in mycobacteria.}, journal = {J Biol Chem}, volume = {280}, year = {2005}, month = {2005 Jun 3}, pages = {21645-52}, abstract = {The plasma membrane of Mycobacterium sp. is the site of synthesis of several distinct classes of lipids that are either retained in the membrane or exported to the overlying cell envelope. Here, we provide evidence that enzymes involved in the biosynthesis of two major lipid classes, the phosphatidylinositol mannosides (PIMs) and aminophospholipids, are compartmentalized within the plasma membrane. Enzymes involved in the synthesis of early PIM intermediates were localized to a membrane subdomain termed PMf, that was clearly resolved from the cell wall by isopyknic density centrifugation and amplified in rapidly dividing Mycobacterium smegmatis. In contrast, the major pool of apolar PIMs and enzymes involved in polar PIM biosynthesis were localized to a denser fraction that contained both plasma membrane and cell wall markers (PM-CW). Based on the resistance of the PIMs to solvent extraction in live but not lysed cells, we propose that polar PIM biosynthesis occurs in the plasma membrane rather than the cell wall component of the PM-CW. Enzymes involved in phosphatidylethanolamine biosynthesis also displayed a highly polarized distribution between the PMf and PM-CW fractions. The PMf was greatly reduced in non-dividing cells, concomitant with a reduction in the synthesis and steady-state levels of PIMs and amino-phospholipids and the redistribution of PMf marker enzymes to non-PM-CW fractions. The formation of the PMf and recruitment of enzymes to this domain may thus play a role in regulating growth-specific changes in the biosynthesis of membrane and cell wall lipids.}, keywords = {Bacterial Proteins, Biochemistry, Cell Membrane, Cell Wall, Hemagglutinins, Lipid Metabolism, Lipids, Mannosides, Mannosyltransferases, Microscopy, Electron, Models, Biological, Mycobacterium smegmatis, Phosphatidylethanolamines, Phosphatidylinositols, Phospholipids, Protein Structure, Tertiary, Subcellular Fractions}, issn = {0021-9258}, doi = {10.1074/jbc.M414181200}, author = {Morita, Yasu S and Velasquez, Ren{\'e} and Taig, Ellen and Waller, Ross F and Patterson, John H and Tull, Dedreia and Williams, Spencer J and Billman-Jacobe, Helen and McConville, Malcolm J} } @article {683, title = {ConSurf 2005: the projection of evolutionary conservation scores of residues on protein structures.}, journal = {Nucleic Acids Res}, volume = {33}, year = {2005}, month = {2005 Jul 1}, pages = {W299-302}, abstract = {Key amino acid positions that are important for maintaining the 3D structure of a protein and/or its function(s), e.g. catalytic activity, binding to ligand, DNA or other proteins, are often under strong evolutionary constraints. Thus, the biological importance of a residue often correlates with its level of evolutionary conservation within the protein family. ConSurf (http://consurf.tau.ac.il/) is a web-based tool that automatically calculates evolutionary conservation scores and maps them on protein structures via a user-friendly interface. Structurally and functionally important regions in the protein typically appear as patches of evolutionarily conserved residues that are spatially close to each other. We present here version 3.0 of ConSurf. This new version includes an empirical Bayesian method for scoring conservation, which is more accurate than the maximum-likelihood method that was used in the earlier release. Various additional steps in the calculation can now be controlled by a number of advanced options, thus further improving the accuracy of the calculation. Moreover, ConSurf version 3.0 also includes a measure of confidence for the inferred amino acid conservation scores.}, keywords = {Amino Acid Substitution, Amino Acids, Bacterial Proteins, Bayes Theorem, Evolution, Molecular, Internet, Models, Molecular, Potassium Channels, Protein Conformation, Software}, issn = {1362-4962}, doi = {10.1093/nar/gki370}, author = {Landau, Meytal and Mayrose, Itay and Rosenberg, Yossi and Glaser, Fabian and Martz, Eric and Pupko, Tal and Ben-Tal, Nir} } @article {539, title = {Crystal ball. In silico biology meets in situ phenomenology.}, journal = {Environ Microbiol}, volume = {7}, year = {2005}, month = {2005 Apr}, pages = {478-9}, keywords = {Bacterial Physiological Phenomena, Computational Biology, Models, Biological}, issn = {1462-2912}, doi = {10.1111/j.1462-2920.2005.803_8.x}, author = {Lovley, Derek R} } @article {716, title = {Caveolin-2 associates with intracellular chlamydial inclusions independently of caveolin-1.}, journal = {BMC Infect Dis}, volume = {4}, year = {2004}, month = {2004 Jul 22}, pages = {23}, abstract = {BACKGROUND: Lipid raft domains form in plasma membranes of eukaryotic cells by the tight packing of glycosphingolipids and cholesterol. Caveolae are invaginated structures that form in lipid raft domains when the protein caveolin-1 is expressed. The Chlamydiaceae are obligate intracellular bacterial pathogens that replicate entirely within inclusions that develop from the phagocytic vacuoles in which they enter. We recently found that host cell caveolin-1 is associated with the intracellular vacuoles and inclusions of some chlamydial strains and species, and that entry of those strains depends on intact lipid raft domains. Caveolin-2 is another member of the caveolin family of proteins that is present in caveolae, but of unknown function. METHODS: We utilized a caveolin-1 negative/caveolin-2 positive FRT cell line and laser confocal immunofluorescence techniques to visualize the colocalization of caveolin-2 with the chlamydial inclusions. RESULTS: We show here that in infected HeLa cells, caveolin-2, as well as caveolin-1, colocalizes with inclusions of C. pneumoniae (Cp), C. caviae (GPIC), and C. trachomatis serovars E, F and K. In addition, caveolin-2 also associates with C. trachomatis serovars A, B and C, although caveolin-1 did not colocalize with these organisms. Moreover, caveolin-2 appears to be specifically, or indirectly, associated with the pathogens at the inclusion membranes. Using caveolin-1 deficient FRT cells, we show that although caveolin-2 normally is not transported out of the Golgi in the absence of caveolin-1, it nevertheless colocalizes with chlamydial inclusions in these cells. However, our results also show that caveolin-2 did not colocalize with UV-irradiated Chlamydia in FRT cells, suggesting that in these caveolin-1 negative cells, pathogen viability and very likely pathogen gene expression are necessary for the acquisition of caveolin-2 from the Golgi. CONCLUSION: Caveolin-2 associates with the chlamydial inclusion independently of caveolin-1. The function of caveolin-2, either in the uninfected cell or in the chlamydial developmental cycle, remains to be elucidated. Nevertheless, this second caveolin protein can now be added to the small number of host proteins that are associated with the inclusions of this obligate intracellular pathogen.}, keywords = {Animals, Blotting, Western, Caveolin 1, Caveolin 2, Caveolins, Cells, Cultured, Chlamydia, Fluorescent Antibody Technique, Golgi Apparatus, Guinea Pigs, HeLa Cells, Humans, Mice, Microscopy, Confocal, Rats, Vacuoles, Virulence}, issn = {1471-2334}, doi = {10.1186/1471-2334-4-23}, author = {Webley, Wilmore C and Norkin, Leonard C and Stuart, Elizabeth S} } @article {857, title = {Cell surface display of the chlamydial glycolipid exoantigen (GLXA) demonstrated by antibody-dependent complement-mediated cytotoxicity.}, journal = {Curr Microbiol}, volume = {49}, year = {2004}, month = {2004 Jul}, pages = {13-21}, abstract = {The chlamydial species are Gram-negative bacterial pathogens critical to human health. Their developmental cycle is associated with the formation and release of the broadly conserved glycolipid exoantigen (GLXA), which has been implicated in the chlamydial elementary body-host cell interaction. This study examines the potential surface display of this glycolipid by chlamydiae-infected cells and the ability of the GLXA they secrete to associate with the plasma membranes of uninfected cells, a prerequisite for exerting influence on them. The sequential incubation of anti-GLXA antibody and complement with Chlamydia trachomatis serovar K or C. pneumoniae AR-39-infected HeLa 229 or macrophage cells resulted in significant cellular cytotoxicity, which preceded the formation of mature elementary bodies. For uninfected cells, co-incubation of GLXA, purified from supernatants of either C. trachomatis or C. pneumoniae-infected HeLa 229 cells, followed by the successive addition of mouse anti-GLXA antibody and complement, yielded similar levels of cellular cytotoxicity. Thus, GLXA indeed is displayed on the surface of infected cells and, therefore, if antibody of appropriate specificity were present, this GLXA could serve to target these infected cells for elimination. Furthermore, released GLXA can associate with uninfected cells and therefore would be positioned to influence their behavior, especially in the context of infection.}, keywords = {Antibody Specificity, Chlamydia, Complement System Proteins, Glycolipids, HeLa Cells, Humans, Polysaccharides, Bacterial}, issn = {0343-8651}, doi = {10.1007/s00284-003-4181-7}, author = {Webley, Wilmore C and Vora, Gary J and Stuart, Elizabeth S} } @article {385, title = {Closing the gaps in kinetoplast DNA network replication.}, journal = {Proc Natl Acad Sci U S A}, volume = {101}, year = {2004}, month = {2004 Mar 30}, pages = {4333-4}, keywords = {Animals, Crithidia fasciculata, DNA Ligases, DNA Replication, DNA, Kinetoplast, DNA, Mitochondrial, Microscopy, Electron, Trypanosoma brucei brucei}, issn = {0027-8424}, doi = {10.1073/pnas.0401400101}, author = {Klingbeil, Michele M and Englund, Paul T} } @article {547, title = {Comparison of 16S rRNA, nifD, recA, gyrB, rpoB and fusA genes within the family Geobacteraceae fam. nov.}, journal = {Int J Syst Evol Microbiol}, volume = {54}, year = {2004}, month = {2004 Sep}, pages = {1591-9}, abstract = {The sequences of five conserved genes, in addition to the 16S rRNA gene, were investigated in 30 members of the Geobacteraceae fam. nov. All members of the Geobacteraceae examined contained nifD, suggesting that they are capable of nitrogen fixation, which may explain their ability to compete effectively in nitrogen-poor subsurface environments undergoing remediation for petroleum or metal contamination. The phylogenies predicted from rpoB, gyrB, fusA, recA and nifD were generally in agreement with the phylogeny predicted from 16S rRNA gene sequences. Furthermore, phylogenetic analysis of concatemers constructed from all five protein-coding genes corresponded closely with the 16S rRNA gene-based phylogeny. This study demonstrated that the Geobacteraceae is a phylogenetically coherent family within the delta-subclass of the Proteobacteria that is composed of three distinct phylogenetic clusters: Geobacter, Desulfuromonas and Desulfuromusa. The sequence data provided here will make it possible to discriminate better between physiologically distinct members of the Geobacteraceae, such as Pelobacter propionicus and Geobacter species, in geobacteraceae-dominated microbial communities and greatly expands the potential to identify geobacteraceae sequences in libraries of environmental genomic DNA.}, keywords = {Bacterial Proteins, Deltaproteobacteria, Desulfuromonas, DNA Gyrase, DNA, Bacterial, DNA, Ribosomal, DNA-Directed RNA Polymerases, Genes, Bacterial, Genes, rRNA, Geobacter, Molecular Sequence Data, Nitrogen Fixation, Peptide Elongation Factor G, Phylogeny, Rec A Recombinases, RNA, Bacterial, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1466-5026}, doi = {10.1099/ijs.0.02958-0}, author = {Holmes, Dawn E and Nevin, Kelly P and Lovley, Derek R} } @article {550, title = {Computational prediction of conserved operons and phylogenetic footprinting of transcription regulatory elements in the metal-reducing bacterial family Geobacteraceae.}, journal = {J Theor Biol}, volume = {230}, year = {2004}, month = {2004 Sep 7}, pages = {133-44}, abstract = {Members of the family Geobacteraceae are an important group of microorganisms from the delta subdivision of Proteobacteria that couple the oxidation of organic compounds to metal reduction. In order to uncover transcription regulatory interactions in these organisms, we used computational methods to identify conserved operons and putative cis-regulatory transcription elements. We identified 26 putative operons with gene order and function conserved among two species of Geobacteraceae, Geobacter sulfurreducens and Geobacter metallireducens. Most of these operons were also conserved in Desulfovibrio vulgaris, an additional metal reducing organism from family Desulfovibrionaceae of the delta subdivision of Proteobacteria. The predicted conserved operons were investigated for the presence of transcription factor binding sites by two different methods, (i) comparison of non-coding regions in conserved operons, and (ii) neural network promoter prediction. Predicted motifs were screened to identify most likely transcription factor binding sites and ribosome-binding sites. We provide information on motifs in Geobacteraceae similar to known transcription factor binding sites in Escherichia coli, conserved motifs in other bacterial species, putative palindromic sites, and predicted ribosome-binding sites. These predictions will aid in further elucidation of regulatory networks of gene interactions in Geobacteraceae.}, keywords = {Animals, Computational Biology, Conserved Sequence, DNA Footprinting, Genes, Regulator, Genome, Bacterial, Geobacter, Models, Genetic, Operon, Phylogeny}, issn = {0022-5193}, doi = {10.1016/j.jtbi.2004.04.022}, author = {Yan, Bin and Meth{\'e}, Barbara A and Lovley, Derek R and Krushkal, Julia} } @article {558, title = {Cleaning up with genomics: applying molecular biology to bioremediation.}, journal = {Nat Rev Microbiol}, volume = {1}, year = {2003}, month = {2003 Oct}, pages = {35-44}, abstract = {Bioremediation has the potential to restore contaminated environments inexpensively yet effectively, but a lack of information about the factors controlling the growth and metabolism of microorganisms in polluted environments often limits its implementation. However, rapid advances in the understanding of bioremediation are on the horizon. Researchers now have the ability to culture microorganisms that are important in bioremediation and can evaluate their physiology using a combination of genome-enabled experimental and modelling techniques. In addition, new environmental genomic techniques offer the possibility for similar studies on as-yet-uncultured organisms. Combining models that can predict the activity of microorganisms that are involved in bioremediation with existing geochemical and hydrological models should transform bioremediation from a largely empirical practice into a science.}, keywords = {Bacteria, Bacterial Physiological Phenomena, Biodegradation, Environmental, Ecology, Environmental Microbiology, Fungi, Genetic Vectors, Genome, Bacterial, Genome, Fungal, Genomics, Models, Biological, Molecular Biology}, issn = {1740-1526}, doi = {10.1038/nrmicro731}, author = {Lovley, Derek R} } @article {684, title = {ConSurf: identification of functional regions in proteins by surface-mapping of phylogenetic information.}, journal = {Bioinformatics}, volume = {19}, year = {2003}, month = {2003 Jan}, pages = {163-4}, abstract = {We recently developed algorithmic tools for the identification of functionally important regions in proteins of known three dimensional structure by estimating the degree of conservation of the amino-acid sites among their close sequence homologues. Projecting the conservation grades onto the molecular surface of these proteins reveals patches of highly conserved (or occasionally highly variable) residues that are often of important biological function. We present a new web server, ConSurf, which automates these algorithmic tools. ConSurf may be used for high-throughput characterization of functional regions in proteins. AVAILABILITY: The ConSurf web server is available at:http://consurf.tau.ac.il. SUPPLEMENTARY INFORMATION: A set of examples is available at http://consurf.tau.ac.il under {\textquoteright}GALLERY{\textquoteright}.}, keywords = {Amino Acids, bcl-X Protein, Conserved Sequence, Databases, Protein, Evolution, Molecular, Internet, Phylogeny, Protein Conformation, Proteins, Proto-Oncogene Proteins c-bcl-2, Sequence Alignment, Sequence Analysis, Protein, User-Computer Interface}, issn = {1367-4803}, author = {Glaser, Fabian and Pupko, Tal and Paz, Inbal and Bell, Rachel E and Bechor-Shental, Dalit and Martz, Eric and Ben-Tal, Nir} } @article {776, title = {Cryptosporidium parvum in children with diarrhea in Mulago Hospital, Kampala, Uganda.}, journal = {Am J Trop Med Hyg}, volume = {68}, year = {2003}, month = {2003 Jun}, pages = {710-5}, abstract = {A cross-sectional case-control study (ratio = 3:1) was conducted over a 15-month period to determine the prevalence and consequences of cryptosporidiosis in hospitalized diarrheic children (0-5 years old) at Mulago Hospital in Kampala, Uganda. Cryptosporidium parvum was detected and genotyped among 2,446 children of whom 1,779 (72.7\%) had diarrhea, and 667 (27.3\%) were age- and sex-matched controls. Of the 1,779 children with diarrhea, 532 (29.9\%) had persistent (> 14 days) diarrhea and 1,247 (70.1\%) had acute diarrhea. Overall, 444 (25.0\%) of the 1,779 children with diarrhea had C. parvum, compared with only 57 (8.5\%) of the 667 children without diarrhea (chi2 = 80.2, P < or = 0.0001). Within this group of infected children, 72.8\% were infected with genotype 1, 18.4\% with genotype 2, and 4.1\% with a mixture of both genotypes, and 4.1\% isolates were either unclassified or C. meleagridis. The prevalence was highest during the rainy months of April to June. Of the 532 children with persistent diarrhea, 166 (31.2\%) had C. parvum compared with 278 (22.3\%) of the 1,247 children with acute diarrhea (chi2 = 15.8, P < or = 0.0001). There was a significant association between C. parvum and malnutrition including stunting, being underweight, and wasting. Unfavorable outcome (death or failure to resolve within 14 days) occurred in 139 (72.8\%) of the 191 children with C. parvum, and in only 65.1\% of the 545 without (odds ratio = 1.117, 95\% confidence interval = 1.005-1.243, P = 0.05), Of the 191 children with C. parvum, 24 (12.6\%) died, compared with 34 (6.2\%) of the 545 without C. parvum (P = 0.005). Mortality rates were higher among children with severe dehydration and persistent diarrhea, and in stunted or underweight children infected with C. parvum. Among Ugandan children, cryptosporidiosis, which remains untreatable, is frequently associated with diarrhea and other serious and unfavorable consequences.}, keywords = {Animals, Case-Control Studies, Child, Preschool, Cross-Sectional Studies, Cryptosporidiosis, Cryptosporidium parvum, Diarrhea, Diarrhea, Infantile, DNA, Protozoan, Female, Hospitalization, Humans, Infant, Infant, Newborn, Male, Nutrition Disorders, Polymerase Chain Reaction, Polymorphism, Restriction Fragment Length, Prevalence, Uganda}, issn = {0002-9637}, author = {Tumwine, James K and Kekitiinwa, Addy and Nabukeera, Nicolette and Akiyoshi, Donna E and Rich, Stephen M and Widmer, Giovanni and Feng, Xiaochuan and Tzipori, Saul} } @article {714, title = {Caveolar endocytosis of simian virus 40 is followed by brefeldin A-sensitive transport to the endoplasmic reticulum, where the virus disassembles.}, journal = {J Virol}, volume = {76}, year = {2002}, month = {2002 May}, pages = {5156-66}, abstract = {Simian virus 40 (SV40) enters cells by atypical endocytosis mediated by caveolae that transports the virus to the endoplasmic reticulum (ER) instead of to the endosomal-lysosomal compartment, which is the usual destination for viruses and other cargo that enter by endocytosis. We show here that SV4O is transported to the ER via an intermediate compartment that contains beta-COP, which is best known as a component of the COPI coatamer complexes that are required for the retrograde retrieval pathway from the Golgi to the ER. Additionally, transport of SV40 to the ER, as well as infection, is sensitive to brefeldin A. This drug acts by specifically inhibiting the ARF1 GTPase, which is known to regulate assembly of COPI coat complexes on Golgi cisternae. Moreover, some beta-COP colocalizes with intracellular caveolin-1, which was previously shown to be present on a new organelle (termed the caveosome) that is an intermediate in the transport of SV40 to the ER (L. Pelkmans, J. Kartenbeck, and A. Helenius, Nat. Cell Biol. 3:473-483, 2001). We also show that the internal SV40 capsid proteins VP2 and VP3 become accessible to immunostaining starting at about 5 h. Most of that immunostaining overlays the ER, with some appearing outside of the ER. In contrast, immunostaining with anti-SV40 antisera remains confined to the ER.}, keywords = {Animals, Antiviral Agents, Biological Transport, Brefeldin A, Capsid, Caveolae, Coatomer Protein, Endocytosis, Endoplasmic Reticulum, Simian virus 40, Virus Replication}, issn = {0022-538X}, author = {Norkin, Leonard C and Anderson, Howard A and Wolfrom, Scott A and Oppenheim, Ariella} } @article {399, title = {A comprehensive alanine scanning mutagenesis of the Escherichia coli transcriptional activator SoxS: identifying amino acids important for DNA binding and transcription activation.}, journal = {J Mol Biol}, volume = {322}, year = {2002}, month = {2002 Sep 13}, pages = {237-57}, abstract = {SoxS is the direct transcriptional activator of the superoxide regulon. SoxS recognizes a highly degenerate "soxbox" DNA sequence, and activates transcription from class I and class II promoters. SoxS is the smallest member of the AraC/XylS family of transcription regulators whose hallmark is dual helix-turn-helix (HTH) DNA-binding motifs. Evidence suggests that the N-terminal HTH motif of SoxS interacts with a highly conserved region of the soxbox termed recognition element 1 (RE1), while the C-terminal HTH motif interacts with the less conserved recognition element 2 (RE2). In the work described here, we prepared a complete library of 101 SoxS mutants containing single alanine substitutions of SoxS, and we characterized the mutant proteins in vivo and in vitro. With SoxS being closely related to MarA, we analyzed the effects of the SoxS mutations in the context of the MarA-mar crystal structure and with respect to the NMR study of MarA-DNA complexes in solution. From the properties of the alanine substitutions, we conclude the following. (1) Surface-exposed residues of helix 3 and helix 6, the recognition helices of the dual HTH motifs, are important to DNA binding and transcription activation; however, substitutions of residues predicted from the MarA-mar crystal structure to make contact with the sugar-phosphate backbone are more detrimental to DNA binding than mutations predicted to make base-specific contacts. (2) Substitution of several residues within the recognition helix predicted to make base-specific contacts with RE2 have relatively little effect on DNA-binding, suggesting the possibility of alternative protein-DNA interactions than those inferred from the MarA-mar crystal structure. (3) DNA binding and transcription activation were reduced by substitution of conserved amino acid residues comprising the hydrophobic core, presumably because they disrupt the structural integrity of SoxS. (4) Mutant K30A appears to be a positive control mutant defective in a protein-protein interaction with RNA polymerase that is required for transcription activation at all SoxS-dependent promoters because it binds and bends DNA normally but fails to activate transcription from both classes of promoters. Alanine substitutions of surface-exposed residues H3, K5, D9, S31, and V45 confer a similar phenotype. Since these residues are near K30 on the surface of the protein, the surface formed by the six residues may be used to make protein-protein interactions with RNA polymerase that are required for transcription activation at both class I and class II SoxS-dependent promoters. (5) Mutants F74A, D75A, M78A, D79A and Q85A appear to define a surface required for protein-protein interaction with RNA polymerase specifically at class II promoters because these positive control mutants bind and bend DNA normally but are defective in activation of class II promoters but not class I promoters. These SoxS mutants that bind and bend DNA normally but are defective in transcription activation represent the first positive control mutants with putative defects in protein-protein interactions with RNA polymerase among the SoxS/MarA/Rob subset of the AraC/XylS family of transcription regulators.}, keywords = {Alanine, Amino Acid Substitution, Base Sequence, Binding Sites, DNA, DNA-Binding Proteins, Electrophoretic Mobility Shift Assay, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genes, Lethal, Hydrophobic and Hydrophilic Interactions, Lac Operon, Models, Molecular, Mutagenesis, Nucleic Acid Conformation, Phenotype, Phosphates, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, Structure-Activity Relationship, Trans-Activators, Transcriptional Activation}, issn = {0022-2836}, author = {Griffith, Kevin L and Wolf, Richard E} } @article {713, title = {Caveolae in the uptake and targeting of infectious agents and secreted toxins.}, journal = {Adv Drug Deliv Rev}, volume = {49}, year = {2001}, month = {2001 Jul 28}, pages = {301-15}, abstract = {A variety of microbial pathogens, including viruses, intracellular bacteria, and prions, as well as certain secreted bacterial toxins, can now be added to the list of ligands that enter cells via caveolae or caveolae-like membrane domains. In general, the caveolae-mediated entry pathway results in transport of these microbes and toxins to intracellular destinations that are different from that of cargo entering by other means. As a result, the caveolae-mediated entry pathway can profoundly affect the host cell-pathogen interaction long after entry has occurred. Furthermore, some microbes such as SV40 that enter via cavolae will be valuable as probes to analyze certain poorly understood intracellular trafficking pathways, such as retrograde transport to the ER. Also, viruses that enter via caveolae may have unique potential as gene and drug delivery vectors. In addition, some extracellular microbial pathogens, such as Pneumocystis carinii, may also interact with host cells via caveolae. Finally, caveolae may play a role in host immune defense mechanisms.}, keywords = {Animals, Bacterial Infections, Caveolae, Humans, Toxins, Biological, Virus Diseases}, issn = {0169-409X}, author = {Norkin, L C} } @article {600, title = {Comment on "Abiotic controls on H2 production from basalt-water reactions and implications for aquifer biogeochemistry".}, journal = {Environ Sci Technol}, volume = {35}, year = {2001}, month = {2001 Apr 1}, pages = {1556-9}, keywords = {Hydrogen, Minerals, Organic Chemicals, Oxidation-Reduction, Silicates, Water, Water Microbiology}, issn = {0013-936X}, author = {Anderson, R T and Chapelle, F H and Lovley, D R} } @article {866, title = {Comparative study of the relationship between monomer structure and reactivity for two polyhydroxyalkanoate synthases.}, journal = {Appl Microbiol Biotechnol}, volume = {56}, year = {2001}, month = {2001 Jul}, pages = {131-6}, abstract = {Using organically synthesized hydroxyalkanoate coenzyme A thioesters, the activities of two short-chain polyhydroxalkanoate (PHA) synthases were investigated--Ralstonia eutropha PHA synthase (a type I PHA synthase) and Ectothiorhodospira shaposhnikovii PHA synthase (a type III synthase). The results indicate that the two synthases have similar activities towards most of the monomers tested. 3-Hydroxybutyryl CoA was found to be the most efficient substrate for both synthases. Changes in the side-chain length of the monomers affect monomer reactivity, with shortening of the side-chain length having the more severe effect. Hydrophobicity in the side chain appears to play an important role in the catalytic reaction. The configuration and the position of the hydroxyl group also affect the reactivity of a monomer. Monomers with the [S] configuration can not be recognized by either synthase. Moving the hydroxyl group from the beta carbon to the alpha carbon has a much more severe effect on the reactivity of the monomer than moving the hydroxyl group to the gamma carbon. The results demonstrate that the in vitro system can be used to prepare entirely novel polymers that may not be obtainable from living cells because of metabolic restrictions.}, keywords = {Acyl Coenzyme A, Acyltransferases, Structure-Activity Relationship, Substrate Specificity}, issn = {0175-7598}, author = {Zhang, S and Kamachi, M and Takagi, Y and Lenz, R W and Goodwin, S} } @article {613, title = {Characterization of a membrane-bound NADH-dependent Fe(3+) reductase from the dissimilatory Fe(3+)-reducing bacterium Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {185}, year = {2000}, month = {2000 Apr 15}, pages = {205-11}, abstract = {Geobacter sulfurreducens produces a single, membrane-associated Fe(3+) reductase activity when grown on fumarate or Fe(3+). The activity was initially isolated by solubilization of membranes with the non-ionic detergent dodecyl-beta-D-maltoside, and partially purified by a combination of ion exchange chromatography and preparative non-denaturing gel electrophoresis. Molecular mass of the reductase, as determined by gel filtration chromatography, was approximately 300 kDa. Cofactor analysis of the purified reductase demonstrates that it contains a hemoprotein and flavin adenine dinucleotide. Kinetic and inhibitor studies show that the reductase is specific for NADH as electron donor, and confirm that the reductase enzymatically reduces Fe(3+). The cytochrome associated with the complex undergoes a reoxidation upon addition of Fe(3+) compounds, indicating an ability to pass reducing equivalents to Fe(3+). This is the first description of a purified NADH-dependent Fe(3+) reductase from a microorganism capable of coupling Fe(3+) reduction to growth.}, keywords = {Cell Membrane, Deltaproteobacteria, Ferric Compounds, FMN Reductase, NADH, NADPH Oxidoreductases, Oxidation-Reduction}, issn = {0378-1097}, author = {Magnuson, T S and Hodges-Myerson, A L and Lovley, D R} } @article {868, title = {Characterization, seasonal occurrence, and diel fluctuation of poly(hydroxyalkanoate) in photosynthetic microbial mats.}, journal = {Appl Environ Microbiol}, volume = {66}, year = {2000}, month = {2000 Oct}, pages = {4279-91}, abstract = {In situ poly(hydroxyalkanoate) (PHA) levels and repeating-unit compositions were examined in stratified photosynthetic microbial mats from Great Sippewissett Salt Marsh, Mass., and Ebro Delta, Spain. Unlike what has been observed in pure cultures of phototrophic bacteria, the prevalence of hydroxyvalerate (HV) repeating units relative to hydroxybutyrate (HB) repeating units was striking. In the cyanobacteria-dominated green material of Sippewissett mats, the mole percent ratio of repeating units was generally 1HB:1HV. In the purple sulfur bacteria-dominated pink material the relationship was typically 1HB:2HV. In Sippewissett mats, PHA contributed about 0.5 to 1\% of the organic carbon in the green layer and up to 6\% in the pink layer. In Ebro Delta mats, PHA of approximately 1HB:2HV-repeating-unit distribution contributed about 2\% of the organic carbon of the composite photosynthetic layers (the green and pink layers were not separated). Great Sippewissett Salt Marsh mats were utilized for more extensive investigation of seasonal, diel, and exogenous carbon effects. When the total PHA content was normalized to organic carbon, there was little seasonal variation in PHA levels. However, routine daily variation was evident at all sites and seasons. In every case, PHA levels increased during the night and decreased during the day. This phenomenon was conspicuous in the pink layer, where PHA levels doubled overnight. The daytime declines could be inhibited by artificial shading. Addition of exogenous acetate, lactate, and propionate induced two- to fivefold increases in the total PHA levels when applied in the daylight but had no effect when applied at night. The distinct diel pattern of in situ PHA accumulation at night appears to be related, in some phototrophs, to routine dark energy metabolism and is not influenced by the availability of organic nutrients.}, keywords = {Alkanes, Biofilms, Hydroxybutyrates, Massachusetts, Photosynthesis, Seasons, Spain, Valerates, Water, Water Microbiology}, issn = {0099-2240}, author = {Rothermich, M M and Guerrero, R and Lenz, R W and Goodwin, S} } @article {45, title = {Combinatorial regulation of phospholipid biosynthetic gene expression by the UME6, SIN3 and RPD3 genes.}, journal = {Nucleic Acids Res}, volume = {28}, year = {2000}, month = {2000 Aug 15}, pages = {3160-7}, abstract = {The Ume6p-Sin3p-Rpd3p complex negatively regulates expression of genes containing a Ume6p binding site. However, these regulatory proteins also function independently to regulate gene expression both negatively and positively. The model system for this combinatorial regulation is the yeast phospholipid biosynthetic pathway. Sin3p negatively regulates the INO1, CHO1, CHO2 and OPI3 genes while Ume6p negatively regulates the INO1 gene and positively regulates the other genes. We have suggested that the positive regulation results from indirect effects on expression of the INO2 transcriptional activator gene. Here, we demonstrate that the effect of Ume6p on INO2 gene expression is also indirect. We also show that Rpd3p is a negative regulator of phospholipid biosynthetic gene expression. The ability of Ume6p, Sin3p and Rpd3p to differentially regulate expression of the phospholipid biosynthetic genes affects phospholipid composition. A sin3 mutant strain lacks detectable levels of phosphatidylethanolamine and elevated levels of phosphatidylcholine (PC) and a rpd3 mutant strain has reduced levels of PC. These alterations in membrane composition suggest that there may exist additional differences in regulation of phospholipid biosynthetic gene expression and that membrane compositions may be coordinated with other biological processes regulated by Ume6p, Sin3p and Rpd3p.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, DNA-Binding Proteins, Fungal Proteins, Gene Expression Regulation, Fungal, Helix-Loop-Helix Motifs, Histone Deacetylases, Kinetics, Phospholipids, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors, Transcription, Genetic}, issn = {1362-4962}, author = {Elkhaimi, M and Kaadige, M R and Kamath, D and Jackson, J C and Biliran, H and Lopes, J M} } @article {516, title = {Carbohydrate oxidation coupled to Fe(III) reduction, a novel form of anaerobic metabolism.}, journal = {Anaerobe}, volume = {4}, year = {1998}, month = {1998 Dec}, pages = {277-82}, abstract = {An isolate, designated GC-29, that could incompletely oxidize glucose to acetate and carbon dioxide with Fe(III) serving as the electron acceptor was recovered from freshwater sediments of the Potomac River, Maryland. This metabolism yielded energy to support cell growth. Strain GC-29 is a facultatively anaerobic, gram-negative motile rod which, in addition to glucose, also used sucrose, lactate, pyruvate, yeast extract, casamino acids or H2 as alternative electron donors for Fe(III) reduction. Stain GC-29 could reduce NO3(-), Mn(IV), U(VI), fumarate, malate, S2O3(2-), and colloidal S0 as well as the humics analog, 2,6-anthraquinone disulfonate. Analysis of the almost complete 16S rRNA sequence indicated that strain GC-29 belongs in the Shewanella genus in the epsilon subdivision of the Proteobacteria. The name Shewanella saccharophilia is proposed. Shewanella saccharophilia differs from previously described fermentative microorganisms that metabolize glucose with the reduction of Fe(III) because it transfers significantly more electron equivalents to Fe(III); acetate and carbon dioxide are the only products of glucose metabolism; energy is conserved from Fe(III) reduction; and glucose is not metabolized in the absence of Fe(III). The metabolism of organisms like S. saccharophilia may account for the fact that glucose is metabolized primarily to acetate and carbon dioxide in a variety of sediments in which Fe(III) reduction is the terminal electron accepting process.}, issn = {1075-9964}, doi = {10.1006/anae.1998.0172}, author = {Coates, J D and Councell, T and Ellis, D J and Lovley, D R} } @article {754, title = {Characterization of the dominant and rare members of a young Hawaiian soil bacterial community with small-subunit ribosomal DNA amplified from DNA fractionated on the basis of its guanine and cytosine composition.}, journal = {Appl Environ Microbiol}, volume = {64}, year = {1998}, month = {1998 Apr}, pages = {1283-9}, abstract = {The small-subunit ribosomal DNA (rDNA) diversity was found to be very high in a Hawaiian soil community that might be expected to have lower diversity than the communities in continental soils because the Hawaiian soil is geographically isolated and only 200 years old, is subjected to a constant climate, and harbors low plant diversity. Since an underlying community structure could not be revealed by analyzing the total eubacterial rDNA, we first fractionated the DNA on the basis of guanine-plus-cytosine (G + C) content by using bisbenzimidazole and equilibrium centrifugation and then analyzed the bacterial rDNA amplified from a fraction with a high biomass (63\% G + C fraction) and a fraction with a low biomass (35\% G + C fraction). The rDNA clone libraries were screened by amplified rDNA restriction analysis to determine phylotype distribution. The dominant biomass reflected by the 63\% G + C fraction contained several dominant phylotypes, while the community members that were less successful (35\% G + C fraction) did not show dominance but there was a very high diversity of phylotypes. Nucleotide sequence analysis revealed taxa belonging to the groups expected for the G + C contents used. The dominant phylotypes in the 63\% G + C fraction were members of the Pseudomonas, Rhizobium-Agrobacterium, and Rhodospirillum assemblages, while all of the clones sequenced from the 35\% G + C fraction were affiliated with several Clostridium assemblages. The two-step rDNA analysis used here uncovered more diversity than can be detected by direct rDNA analysis of total community DNA. The G + C separation step is also a way to detect some of the less dominant organisms in a community.}, keywords = {Base Composition, Clostridium, DNA, Bacterial, DNA, Ribosomal, Ecosystem, Genetic Variation, Hawaii, Phylogeny, Pseudomonas, Rhizobium, Rhodospirillum, Soil Microbiology}, issn = {0099-2240}, author = {N{\"u}sslein, K and Tiedje, J M} } @article {392, title = {Characterization of cDNA clones for the 2-methyl branched-chain enoyl-CoA reductase. An enzyme involved in branched-chain fatty acid synthesis in anaerobic mitochondria of the parasitic nematode Ascaris suum.}, journal = {J Biol Chem}, volume = {268}, year = {1993}, month = {1993 Oct 25}, pages = {22391-6}, abstract = {The 2-methyl branched-chain enoyl-CoA reductase plays a pivotal role in the reversal of beta-oxidation operating in anaerobic mitochondria of the parasitic nematode Ascaris suum. An affinity-purified polyclonal anti-serum against the reductase was used to screen a cDNA library constructed in lambda gt11 with poly(A)+ RNA from adult A. suum muscle. A 1.2-kilobase partial cDNA clone was isolated, subcloned, and sequenced in both directions. Additional sequence at the 5{\textquoteright} end of the mRNA was determined by the RACE (rapid amplification of cDNA ends) procedure. Nucleotide sequence analysis of the cDNAs revealed the 22-nucleotide trans-spliced leader sequence characteristic of many nematode mRNAs, an open reading frame of 1236 nucleotides and a 3{\textquoteright}-untranslated sequence of 109 nucleotides including a short poly(A) tail 14 nucleotides from a polyadenylation signal (AATAAA). The open reading frame encoded a 396-amino acid sequence (M(r) 43,046) including a 16-amino acid leader peptide. Two-dimensional gel electrophoresis of the purified reductase yielded multiple spots with two distinct but overlapping amino-terminal amino acid sequences. Both sequences overlapped with the sequence predicted from the mRNA, and one of the sequences was identical to the predicted sequence. Comparison of the ascarid sequence with that of mammalian acyl-CoA dehydrogenases revealed a high degree of sequence identity, suggesting that these enzymes may have evolved from a common ancestral gene even though the ascarid enzyme functions as a reductase, not as a dehydrogenase. Immunoblotting of A. suum larval stages and adult tissues with antisera that cross-reacted with each of the spots separated on two-dimensional gels suggested that the reductase was only found in adult muscle. Northern blotting using the partial cDNA revealed a hybridization band of about 1.5 kilobases and also suggested that the enzyme was tissue-specific and developmentally regulated in agreement with the results of the immunoblotting.}, keywords = {Amino Acid Sequence, Anaerobiosis, Animals, Ascaris suum, Base Sequence, Cloning, Molecular, Consensus Sequence, DNA, DNA Primers, DNA, Complementary, Fatty Acid Desaturases, Gene Library, Humans, Mitochondria, Molecular Sequence Data, Oligonucleotides, Antisense, Oxidoreductases, Oxidoreductases Acting on CH-CH Group Donors, Poly A, RNA, RNA, Messenger, Sequence Homology, Amino Acid}, issn = {0021-9258}, author = {Duran, E and Komuniecki, R W and Komuniecki, P R and Wheelock, M J and Klingbeil, M M and Ma, Y C and Johnson, K R} } @article {801, title = {Competence of Peromyscus maniculatus (Rodentia: Cricetidae) as a reservoir host for Borrelia burgdorferi (Spirochaetares: Spirochaetaceae) in the wild.}, journal = {J Med Entomol}, volume = {30}, year = {1993}, month = {1993 May}, pages = {614-8}, abstract = {Although capable of maintaining and transmitting Borrelia burgdorferi Johnson, Schmidt, Hyde, Steigerwalt \& Brenner, the causative spirochete of Lyme disease, in the laboratory, the specific ability of deer mice, Peromyscus maniculatus Le Conte, to support this zoonosis has not been established. Demonstration that P. maniculatus is a competent reservoir host in the wild would indicate that the spread of Lyme disease is not limited to the range of the primary reservoir host, P. leucopus Rafinesque. Isle au Haut, an offshore Maine island upon which the vector tick Ixodes dammini Spielman, Clifford, Piesman \& Corwin has become established, supports an isolated population of mice that are exclusively P. maniculatus. We examined the reservoir competence of this species by comparing infection rates of B. burgdorferi among juvenile ticks removed from livetrapped mice on this island with those removed from P. leucopus obtained at a mainland site endemic for Lyme disease. Equivalent rates of infection among engorged larval ticks, survival of infection through the larval-nymphal molt, and the isolation of B. burgdorferi from mice at both sites attest to the reservoir competence of P. maniculatus.}, keywords = {Animals, Borrelia burgdorferi Group, Host-Parasite Interactions, Lyme Disease, Peromyscus, Ticks}, issn = {0022-2585}, author = {Rand, P W and Lacombe, E H and Smith, R P and Rich, S M and Kilpatrick, C W and Dragoni, C A and Caporale, D} } @article {648, title = {Composition of Non-Microbially Reducible Fe(III) in Aquatic Sediments.}, journal = {Appl Environ Microbiol}, volume = {59}, year = {1993}, month = {1993 Aug}, pages = {2727-9}, abstract = {The production of small quantities of Fe(II) during the initial phase of microbial Fe(III) reduction greatly increased the amount of Fe(III) that could be extracted from freshwater sediments with oxalate. This finding and other evidence suggest that the oxalate-extractable Fe(III) that is unavailable for microbial reduction in anoxic sediments is not in the form of mixed Fe(III)-Fe(II) forms, as was previously suggested, but rather is in the form of highly crystalline Fe(III) oxides.}, issn = {0099-2240}, author = {Phillips, E J and Lovley, D R and Roden, E E} } @article {703, title = {Class I major histocompatibility proteins are an essential component of the simian virus 40 receptor.}, journal = {J Virol}, volume = {66}, year = {1992}, month = {1992 Apr}, pages = {2037-45}, abstract = {The class I molecules encoded by the major histocompatibility complex (MHC) present endogenously synthesized antigenic peptide fragments to cytotoxic T lymphocytes. We show here that these proteins are an essential component of the cell surface receptor for simian virus 40 (SV40). First, SV40 binding to cells can be blocked by two monoclonal antibodies against class I human lymphocyte antigen (HLA) proteins but not by monoclonal antibodies specific for other cell surface proteins. Second, SV40 does not bind to cells of two different human lymphoblastoid cell lines which do not express surface class I MHC proteins because of genetic defects in the beta 2-microglobulin gene in one line and in the HLA complex in the other. Transfection of these cell lines with cloned genes for beta 2-microglobulin and HLA-B8, respectively, restored expression of their surface class I MHC proteins and resulted in concomitant SV40 binding. Finally, SV40 binds to purified HLA proteins in vitro and selectively binds to class I MHC proteins in a cell surface extract.}, keywords = {Animals, Binding, Competitive, Blotting, Western, Cell Line, Flow Cytometry, Histocompatibility Antigens Class I, Humans, Precipitin Tests, Receptors, Virus, Simian virus 40, Transfection}, issn = {0022-538X}, author = {Breau, W C and Atwood, W J and Norkin, L C} } @article {663, title = {Competitive mechanisms for inhibition of sulfate reduction and methane production in the zone of ferric iron reduction in sediments.}, journal = {Appl Environ Microbiol}, volume = {53}, year = {1987}, month = {1987 Nov}, pages = {2636-41}, abstract = {Mechanisms for inhibition of sulfate reduction and methane production in the zone of Fe(III) reduction in sediments were investigated. Addition of amorphic iron(III) oxyhydroxide to sediments in which sulfate reduction was the predominant terminal electron-accepting process inhibited sulfate reduction 86 to 100\%. The decrease in electron flow to sulfate reduction was accompanied by a corresponding increase in electron flow to Fe(III) reduction. In a similar manner, Fe(III) additions also inhibited methane production in sulfate-depleted sediments. The inhibition of sulfate reduction and methane production was the result of substrate limitation, because the sediments retained the potential for sulfate reduction and methane production in the presence of excess hydrogen and acetate. Sediments in which Fe(III) reduction was the predominant terminal electron-accepting process had much lower concentrations of hydrogen and acetate than sediments in which sulfate reduction or methane production was the predominant terminal process. The low concentrations of hydrogen and acetate in the Fe(III)-reducing sediments were the result of metabolism by Fe(III)-reducing organisms of hydrogen and acetate at concentrations lower than sulfate reducers or methanogens could metabolize them. The results indicate that when Fe(III) is in a form that Fe(III)-reducing organisms can readily reduce, Fe(III)-reducing organisms can inhibit sulfate reduction and methane production by outcompeting sulfate reducers and methanogens for electron donors.}, issn = {0099-2240}, author = {Lovley, D R and Phillips, E J} } @article {1707, title = {Citrate synthase.}, journal = {Curr Top Cell Regul}, volume = {10}, year = {1976}, month = {1976}, pages = {161-204}, keywords = {Adenosine Diphosphate, Adenosine Monophosphate, Adenosine Triphosphate, Amino Acids, Animals, Bacteria, Binding Sites, Cations, Divalent, Cations, Monovalent, Citrate (si)-Synthase, Dithionitrobenzoic Acid, Ketoglutaric Acids, Kinetics, Molecular Weight, NAD, NADP, Oxo-Acid-Lyases, Protein Binding, Species Specificity}, issn = {0070-2137}, author = {Weitzman, P D and Danson, M J} } @article {1711, title = {Comparison between procaine and isocarboxazid metabolism in vitro by a liver microsomal amidase-esterase.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Aug 15}, pages = {1517-21}, keywords = {Amidohydrolases, Animals, Esterases, Hydrogen-Ion Concentration, In Vitro Techniques, Isocarboxazid, Kinetics, Male, Metals, Microsomes, Liver, Phospholipids, Procaine, Proteins, Rats, Subcellular Fractions, Temperature}, issn = {0006-2952}, author = {Moroi, K and Sato, T} } @article {1725, title = {A comparison of the substrate specificities of endo-beta-N-acetylglucosaminidases from Streptomyces griseus and Diplococcus Pneumoniae.}, journal = {Biochem Biophys Res Commun}, volume = {67}, year = {1975}, month = {1975 Nov 3}, pages = {455-62}, keywords = {Acetylglucosaminidase, Fucose, Hexosaminidases, Kinetics, Oligosaccharides, Species Specificity, Streptococcus pneumoniae, Streptomyces griseus, Structure-Activity Relationship}, issn = {0006-291X}, author = {Tarentino, A L and Maley, F} }