@article {3065, title = {Intrinsically Conductive Microbial Nanowires for {\textquoteright}Green{\textquoteright} Electronics with Novel Functions.}, journal = {Trends Biotechnol}, volume = {39}, year = {2021}, month = {2021 Sep}, pages = {940-952}, abstract = {

Intrinsically conductive protein nanowires, microbially produced from inexpensive, renewable feedstocks, are a sustainable alternative to traditional nanowire electronic materials, which require high energy inputs and hazardous conditions/chemicals for fabrication and can be highly toxic. Pilin-based nanowires can be tailored for specific functions via the design of synthetic pilin genes to tune wire conductivity or introduce novel functionalities. Other microbially produced nanowire options for electronics may include cytochrome wires, curli fibers, and the conductive fibers of cable bacteria. Proof-of-concept protein nanowire electronics that have been successfully demonstrated include biomedical sensors, neuromorphic devices, and a device that generates electricity from ambient humidity. Further development of applications will require interdisciplinary teams of engineers, biophysicists, and synthetic biologists.

}, keywords = {Electric Conductivity, Electronics, Nanowires, Proteins}, issn = {1879-3096}, doi = {10.1016/j.tibtech.2020.12.005}, author = {Lovley, Derek R and Yao, Jun} } @article {3077, title = {Iron Corrosion via Direct Metal-Microbe Electron Transfer.}, journal = {mBio}, volume = {10}, year = {2019}, month = {2019 May 14}, abstract = {

The concept that anaerobic microorganisms can directly accept electrons from Fe(0) has been controversial because direct metal-microbe electron transfer has previously only been indirectly inferred. Fe(0) oxidation was studied with strain ACL, an autotrophic strain that was previously shown to grow with electrons derived from a graphite cathode as the sole electron donor. Strain ACL grew with Fe(0) as the sole electron donor and fumarate as the electron acceptor. However, it appeared that at least a portion of the electron transfer was via H produced nonenzymatically from the oxidation of Fe(0) to Fe(II). H, which accumulated in abiotic controls, was consumed during the growth of strain ACL, the cells were predominately planktonic, and genes for the uptake hydrogenase were highly expressed. Strain ACL was constructed to prevent growth on H or formate by deleting the genes for the uptake of hydrogenase and formate dehydrogenases from strain ACL. Strain ACL also grew with Fe(0) as the sole electron donor, but H accumulated in the culture, and cells heavily colonized Fe(0) surfaces with no visible planktonic growth. Transcriptomics suggested that the outer surface -type cytochromes OmcS and OmcZ were important during growth of strain ACL on Fe(0). Strain ACL did not grow on Fe(0) if the gene for either of these cytochromes was deleted. The specific attachment of strain ACL to Fe(0), coupled with requirements for known extracellular electrical contacts, suggest that direct metal-microbe electron transfer is the most likely option for Fe(0) serving as an electron donor. The anaerobic corrosion of iron structures is expensive to repair and can be a safety and environmental concern. It has been known for over 100 years that the presence of anaerobic respiratory microorganisms can accelerate iron corrosion. Multiple studies have suggested that there are sulfate reducers, methanogens, and acetogens that can directly accept electrons from Fe(0) to support sulfate or carbon dioxide reduction. However, all of the strains studied can also use H as an electron donor for growth, which is known to be abiotically produced from Fe(0). Furthermore, no proteins definitely shown to function as extracellular electrical contacts with Fe(0) were identified. The studies described here demonstrate that direct electron transfer from Fe(0) can support anaerobic respiration. They also map out a simple genetic approach to the study of iron corrosion mechanisms in other microorganisms. A better understanding of how microorganisms promote iron corrosion is expected to lead to the development of strategies that can help reduce adverse impacts from this process.

}, keywords = {Anaerobiosis, Corrosion, Cytochromes, Electron Transport, Formate Dehydrogenases, Geobacter, Iron, Oxidation-Reduction, Oxidoreductases, Transcriptome}, issn = {2150-7511}, doi = {10.1128/mBio.00303-19}, author = {Tang, Hai-Yan and Holmes, Dawn E and Ueki, Toshiyuki and Palacios, Paola A and Lovley, Derek R} } @article {1559, title = {Identification of 4-Hydroxycumyl Alcohol As the Major MnO2-Mediated Bisphenol A Transformation Product and Evaluation of Its Environmental Fate.}, journal = {Environ Sci Technol}, volume = {49}, year = {2015}, month = {2015 May 19}, pages = {6214-21}, abstract = {

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

}, issn = {1520-5851}, doi = {10.1021/acs.est.5b00372}, author = {Im, Jeongdae and Prevatte, Carson W and Campagna, Shawn R and L{\"o}ffler, Frank E} } @article {1437, title = {Illumination of growth, division and secretion by metabolic labeling of the bacterial cell surface.}, journal = {FEMS Microbiol Rev}, volume = {39}, year = {2015}, month = {2015 Mar}, pages = {184-202}, abstract = {

The cell surface is the essential interface between a bacterium and its surroundings. Composed primarily of molecules that are not directly genetically encoded, this highly dynamic structure accommodates the basic cellular processes of growth and division as well as the transport of molecules between the cytoplasm and the extracellular milieu. In this review, we describe aspects of bacterial growth, division and secretion that have recently been uncovered by metabolic labeling of the cell envelope. Metabolite derivatives can be used to label a variety of macromolecules, from proteins to non-genetically-encoded glycans and lipids. The embedded metabolite enables precise tracking in time and space, and the versatility of newer chemoselective detection methods offers the ability to execute multiple experiments concurrently. In addition to reviewing the discoveries enabled by metabolic labeling of the bacterial cell envelope, we also discuss the potential of these techniques for translational applications. Finally, we offer some guidelines for implementing this emerging technology.

}, issn = {1574-6976}, doi = {10.1093/femsre/fuu012}, author = {Siegrist, M Sloan and Swarts, Benjamin M and Fox, Douglas M and Lim, Shion An and Bertozzi, Carolyn R} } @article {1201, title = {Identification and Characterization of an Archaeal Kojibiose Catabolic Pathway in the Hyperthermophilic Pyrococcus sp. Strain ST04.}, journal = {J Bacteriol}, volume = {196}, year = {2014}, month = {2014 Mar}, pages = {1122-31}, abstract = {

A unique gene cluster responsible for kojibiose utilization was identified in the genome of Pyrococcus sp. strain ST04. The proteins it encodes hydrolyze kojibiose, a disaccharide product of glucose caramelization, and form glucose-6-phosphate (G6P) in two steps. Heterologous expression of the kojibiose-related enzymes in Escherichia coli revealed that two genes, Py04_1502 and Py04_1503, encode kojibiose phosphorylase (designated PsKP, for Pyrococcus sp. strain ST04 kojibiose phosphorylase) and β-phosphoglucomutase (PsPGM), respectively. Enzymatic assays show that PsKP hydrolyzes kojibiose to glucose and β-glucose-1-phosphate (β-G1P). The Km values for kojibiose and phosphate were determined to be 2.53 {\textpm} 0.21 mM and 1.34 {\textpm} 0.04 mM, respectively. PsPGM then converts β-G1P into G6P in the presence of 6 mM MgCl2. Conversion activity from β-G1P to G6P was 46.81 {\textpm} 3.66 U/mg, and reverse conversion activity from G6P to β-G1P was 3.51 {\textpm} 0.13 U/mg. The proteins are highly thermostable, with optimal temperatures of 90{\textdegree}C for PsKP and 95{\textdegree}C for PsPGM. These results indicate that Pyrococcus sp. strain ST04 converts kojibiose into G6P, a substrate of the glycolytic pathway. This is the first report of a disaccharide utilization pathway via phosphorolysis in hyperthermophilic archaea.

}, issn = {1098-5530}, doi = {10.1128/JB.01222-13}, author = {Jung, Jong-Hyun and Seo, Dong-Ho and Holden, James F and Park, Cheon-Seok} } @article {3122, title = {Identification of genes specifically required for the anaerobic metabolism of benzene in Geobacter metallireducens.}, journal = {Front Microbiol}, volume = {5}, year = {2014}, month = {2014}, pages = {245}, abstract = {

Although the biochemical pathways for the anaerobic degradation of many of the hydrocarbon constituents in petroleum reservoirs have been elucidated, the mechanisms for anaerobic activation of benzene, a very stable molecule, are not known. Previous studies have demonstrated that Geobacter metallireducens can anaerobically oxidize benzene to carbon dioxide with Fe(III) as the sole electron acceptor and that phenol is an intermediate in benzene oxidation. In an attempt to identify enzymes that might be involved in the conversion of benzene to phenol, whole-genome gene transcript abundance was compared in cells metabolizing benzene and cells metabolizing phenol. Eleven genes had significantly higher transcript abundance in benzene-metabolizing cells. Five of these genes had annotations suggesting that they did not encode proteins that could be involved in benzene metabolism and were not further studied. Strains were constructed in which one of the remaining six genes was deleted. The strain in which the monocistronic gene Gmet 0232 was deleted metabolized phenol, but not benzene. Transcript abundance of the adjacent monocistronic gene, Gmet 0231, predicted to encode a zinc-containing oxidoreductase, was elevated in cells metabolizing benzene, although not at a statistically significant level. However, deleting Gmet 0231 also yielded a strain that could metabolize phenol, but not benzene. Although homologs of Gmet 0231 and Gmet 0232 are found in microorganisms not known to anaerobically metabolize benzene, the adjacent localization of these genes is unique to G. metallireducens. The discovery of genes that are specifically required for the metabolism of benzene, but not phenol in G. metallireducens is an important step in potentially identifying the mechanisms for anaerobic benzene activation.

}, issn = {1664-302X}, doi = {10.3389/fmicb.2014.00245}, author = {Zhang, Tian and Tremblay, Pier-Luc and Chaurasia, Akhilesh K and Smith, Jessica A and Bain, Timothy S and Lovley, Derek R} } @article {1442, title = {Imaging bacterial peptidoglycan with near-infrared fluorogenic azide probes.}, journal = {Proc Natl Acad Sci U S A}, volume = {111}, year = {2014}, month = {2014 Apr 15}, pages = {5456-61}, abstract = {

Fluorescent probes designed for activation by bioorthogonal chemistry have enabled the visualization of biomolecules in living systems. Such activatable probes with near-infrared (NIR) emission would be ideal for in vivo imaging but have proven difficult to engineer. We present the development of NIR fluorogenic azide probes based on the Si-rhodamine scaffold that undergo a fluorescence enhancement of up to 48-fold upon reaction with terminal or strained alkynes. We used the probes for mammalian cell surface imaging and, in conjunction with a new class of cyclooctyne D-amino acids, for visualization of bacterial peptidoglycan without the need to wash away unreacted probe.

}, keywords = {Azides, Fluorescent Dyes, Molecular Imaging, Molecular Probe Techniques, Molecular Structure, Peptidoglycan}, issn = {1091-6490}, doi = {10.1073/pnas.1322727111}, author = {Shieh, Peyton and Siegrist, M Sloan and Cullen, Andrew J and Bertozzi, Carolyn R} } @article {1422, title = {Identification and characterization of a NaCl-responsive genetic locus involved in survival during desiccation in Sinorhizobium meliloti.}, journal = {Appl Environ Microbiol}, volume = {79}, year = {2013}, month = {2013 Sep}, pages = {5693-700}, abstract = {

The Rhizobiaceae are a bacterial family of enormous agricultural importance due to the ability of its members to fix atmospheric nitrogen in an intimate relationship with plants. Their survival as naturally occurring soil bacteria in agricultural soils as well as popular seed inocula is affected directly by drought and salinity. Survival after desiccation in the presence of NaCl is enabled by underlying genetic mechanisms in the model organism Sinorhizobium meliloti 1021. Since salt stress parallels a loss in water activity, the identification of NaCl-responsive loci may identify loci involved in survival during desiccation. This approach enabled identification of the loci asnO and ngg by their reduced ability to grow on increased NaCl concentrations, likely due to their inability to produce the osmoprotectant N-acetylglutaminylglutamine (NAGGN). In addition, the mutant harboring ngg::Tn5luxAB was affected in its ability to survive desiccation and responded to osmotic stress. The desiccation sensitivity may have been due to secondary functions of Ngg (N-acetylglutaminylglutamine synthetase)-like cell wall metabolism as suggested by the presence of a d-alanine-d-alanine ligase (dAla-dAla) domain and by sensitivity of the mutant to \β-lactam antibiotics. asnO::Tn5luxAB is expressed during the stationary phase under normal growth conditions. Amino acid sequence similarity to enzymes producing \β-lactam inhibitors and increased resistance to \β-lactam antibiotics may indicate that asnO is involved in the production of a \β-lactam inhibitor.

}, keywords = {Desiccation, DNA Transposable Elements, Gene Expression Regulation, Bacterial, Gene Knockout Techniques, Genes, Bacterial, Genetic Loci, Microbial Viability, Mutagenesis, Insertional, Osmotic Pressure, Sinorhizobium meliloti, Sodium Chloride, Stress, Physiological}, issn = {1098-5336}, doi = {10.1128/AEM.01037-13}, author = {Vriezen, Jan A C and de Bruijn, Frans J and N{\"u}sslein, Klaus} } @article {3139, title = {Improved cathode for high efficient microbial-catalyzed reduction in microbial electrosynthesis cells.}, journal = {Phys Chem Chem Phys}, volume = {15}, year = {2013}, month = {2013 Sep 14}, pages = {14290-4}, abstract = {

Microbial electrosynthesis cells (MECs) are devices wherein microorganisms can electrochemically interact with electrodes, directly donating or accepting electrons from electrode surfaces. Here, we developed a novel cathode by using nickel nanowires anchored to graphite for the improvement of microbial-catalyzed reduction in MEC cathode chamber. This porous nickel-nanowire-network-coated graphite electrode increased the interfacial area and interfacial interactions between the cathode surface and the microbial biofilm. A 2.3 fold increase in bio-reduction rate over the untreated graphite was observed. Around 282 mM day(-1) m(-2) of acetate resulting from the bio-reduction of carbon dioxide by Sporomusa was produced with 82 {\textpm} 14\% of the electrons consumed being recovered in acetate.

}, keywords = {Biocatalysis, Bioelectric Energy Sources, Biofilms, Carbon Dioxide, Electrochemical Techniques, Electrodes, Graphite, Nanowires, Nickel, Oxidation-Reduction, Veillonellaceae}, issn = {1463-9084}, doi = {10.1039/c3cp52697f}, author = {Nie, Huarong and Zhang, Tian and Cui, Mengmeng and Lu, Haiyun and Lovley, Derek R and Russell, Thomas P} } @article {1563, title = {Interference of ferric ions with ferrous iron quantification using the ferrozine assay.}, journal = {J Microbiol Methods}, volume = {95}, year = {2013}, month = {2013 Dec}, pages = {366-7}, abstract = {

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

}, keywords = {Chemistry Techniques, Analytical, Colorimetry, Darkness, Diagnostic Errors, Ferric Compounds, Ferrous Compounds, Ferrozine, Ions, Iron, Time Factors}, issn = {1872-8359}, doi = {10.1016/j.mimet.2013.10.005}, author = {Im, Jeongdae and Lee, Jaejin and L{\"o}ffler, Frank E} } @article {412, title = {Identification of multicomponent histidine-aspartate phosphorelay system controlling flagellar and motility gene expression in Geobacter species.}, journal = {J Biol Chem}, volume = {287}, year = {2012}, month = {2012 Mar 30}, pages = {10958-66}, abstract = {Geobacter species play an important role in the natural biogeochemical cycles of aquatic sediments and subsurface environments as well as in subsurface bioremediation by oxidizing organic compounds with the reduction of insoluble Fe(III) oxides. Flagellum-based motility is considered to be critical for Geobacter species to locate fresh sources of Fe(III) oxides. Functional and comparative genomic approaches, coupled with genetic and biochemical methods, identified key regulators for flagellar gene expression in Geobacter species. A master transcriptional regulator, designated FgrM, is a member of the enhancer-binding protein family. The fgrM gene in the most studied strain of Geobacter species, Geobacter sulfurreducens strain DL-1, is truncated by a transposase gene, preventing flagellar biosynthesis. Integrating a functional FgrM homolog restored flagellar biosynthesis and motility in G. sulfurreducens DL-1 and enhanced the ability to reduce insoluble Fe(III) oxide. Interrupting the fgrM gene in G. sulfurreducens strain KN400, which is motile, removed the capacity for flagellar production and inhibited Fe(III) oxide reduction. FgrM, which is also a response regulator of the two-component His-Asp phosphorelay system, was phosphorylated by histidine kinase GHK4, which was essential for flagellar production and motility. GHK4, which is a hybrid kinase with a receiver domain at the N terminus, was phosphorylated by another histidine kinase, GHK3. Therefore, the multicomponent His-Asp phosphorelay system appears to control flagellar gene expression in Geobacter species.}, issn = {1083-351X}, doi = {10.1074/jbc.M112.345041}, author = {Ueki, Toshiyuki and Leang, Ching and Inoue, Kengo and Lovley, Derek R} } @article {848, title = {In vitro assessment of halobacterial gas vesicles as a Chlamydia vaccine display and delivery system.}, journal = {Vaccine}, volume = {30}, year = {2012}, month = {2012 Sep 7}, pages = {5942-8}, abstract = {Chlamydia trachomatis is the leading cause of bacterial sexually transmitted disease worldwide and while antibiotic treatment is effective in eliminating the pathogen, up to 70\% of all infections are asymptomatic. Despite sustained efforts over the past 2 decades, an effective chlamydial vaccine remains elusive, due in large part to the lack of an effective delivery system. We explored the use of gas vesicles derived from Halobacterium salinarium as a potential display and delivery vehicle for chlamydial antigens of vaccine interest. Various size gene fragments coding for the major outer membrane protein (MOMP), outer membrane complex B (OmcB) and polymorphic outer membrane protein D (PompD) were integrated into and expressed as part of the gas vesicle protein C (gvpC) on the surface of these stable structures. The presence of the recombinant proteins was confirmed by Western blots probed using anti-gvpC and anti-Chlamydia antibodies as well as sera from Chlamydia-positive patients. Tissue culture evaluation revealed stability and a time-dependent degradation of recombinant gas vesicles (r-Gv) in human and animal cell lines. In vitro assessment using human foreskin fibroblasts (HFF) confirmed Toll-like receptor (TLR) 4 and 5 engagement by wild type and r-Gv, leading to MyD88 activation, TNF-α, IL-6 and IL-12 production. The data suggest that r-GV could be an effective, naturally adjuvanting, time-release antigen delivery system for immunologically relevant Chlamydia vaccine antigens which are readily recognized by human immune sera.}, issn = {1873-2518}, doi = {10.1016/j.vaccine.2012.07.038}, author = {Childs, Tawanna S and Webley, Wilmore C} } @article {404, title = {Interspecies electron transfer via hydrogen and formate rather than direct electrical connections in cocultures of Pelobacter carbinolicus and Geobacter sulfurreducens.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Nov}, pages = {7645-51}, abstract = {

Direct interspecies electron transfer (DIET) is an alternative to interspecies H(2)/formate transfer as a mechanism for microbial species to cooperatively exchange electrons during syntrophic metabolism. To understand what specific properties contribute to DIET, studies were conducted with Pelobacter carbinolicus, a close relative of Geobacter metallireducens, which is capable of DIET. P. carbinolicus grew in coculture with Geobacter sulfurreducens with ethanol as the electron donor and fumarate as the electron acceptor, conditions under which G. sulfurreducens formed direct electrical connections with G. metallireducens. In contrast to the cell aggregation associated with DIET, P. carbinolicus and G. sulfurreducens did not aggregate. Attempts to initiate cocultures with a genetically modified strain of G. sulfurreducens incapable of both H(2) and formate utilization were unsuccessful, whereas cocultures readily grew with mutant strains capable of formate but not H(2) uptake or vice versa. The hydrogenase mutant of G. sulfurreducens compensated, in cocultures, with significantly increased formate dehydrogenase gene expression. In contrast, the transcript abundance of a hydrogenase gene was comparable in cocultures with that for the formate dehydrogenase mutant of G. sulfurreducens or the wild type, suggesting that H(2) was the primary electron carrier in the wild-type cocultures. Cocultures were also initiated with strains of G. sulfurreducens that could not produce pili or OmcS, two essential components for DIET. The finding that P. carbinolicus exchanged electrons with G. sulfurreducens via interspecies transfer of H(2)/formate rather than DIET demonstrates that not all microorganisms that can grow syntrophically are capable of DIET and that closely related microorganisms may use significantly different strategies for interspecies electron exchange.

}, keywords = {Coculture Techniques, Deltaproteobacteria, Electricity, Electron Transport, Electrons, Formates, Geobacter, Hydrogen, Microbial Interactions}, issn = {1098-5336}, doi = {10.1128/AEM.01946-12}, author = {Rotaru, Amelia-Elena and Shrestha, Pravin M and Liu, Fanghua and Ueki, Toshiyuki and Nevin, Kelly and Summers, Zarath M and Lovley, Derek R} } @article {357, title = {Identification of a nucleopolyhedrovirus in winter moth populations from Massachusetts.}, journal = {J Invertebr Pathol}, volume = {108}, year = {2011}, month = {2011 Nov}, pages = {217-9}, abstract = {Winter moth, Operophtera brumata, originally from Europe, has invaded eastern Massachusetts causing widespread defoliation and damage to many deciduous tree species and a variety of crop plants in the infested area. We identified O. brumata nucleopolyhedrovirus (OpbuNPV) in winter moth larvae collected from field sites in Massachusetts by using PCR to amplify a 482 bp region of the baculovirus polyhedrin gene. Viral sequences were also detected in winter moth pupae that failed to emerge, suggesting that these insects may have died as a result of viral infection. This represents the first report of OpbuNPV in winter moth populations in the US.}, keywords = {Animals, Base Sequence, Environmental Monitoring, Larva, Massachusetts, Molecular Sequence Data, Moths, Nucleopolyhedrovirus, Viral Proteins, Virus Diseases}, issn = {1096-0805}, doi = {10.1016/j.jip.2011.08.005}, author = {Burand, John P and Kim, Woojin and Welch, Anna and Elkinton, Joseph S} } @article {686, title = {Identification of a second catalytically active trans-sialidase in Trypanosoma brucei.}, journal = {Biochem Biophys Res Commun}, volume = {415}, year = {2011}, month = {2011 Nov 18}, pages = {421-5}, abstract = {The procyclic stage of Trypanosoma brucei is covered by glycosylphosphatidylinositol (GPI)-anchored surface proteins called procyclins. The procyclin GPI anchor contains a side chain of N-acetyllactosamine repeats terminated by sialic acids. Sialic acid modification is mediated by trans-sialidases expressed on the parasite{\textquoteright}s cell surface. Previous studies suggested the presence of more than one active trans-sialidases, but only one has so far been reported. Here we cloned and examined enzyme activities of four additional trans-sialidase homologs, and show that one of them, Tb927.8.7350, encodes another active trans-sialidase, designated as TbSA C2. In an in vitro assay, TbSA C2 utilized α2-3 sialyllactose as a donor, and produced an α2-3-sialylated product, suggesting that it is an α2-3 trans-sialidase. We suggest that TbSA C2 plays a role in the sialic acid modification of the trypanosome cell surface.}, keywords = {Amino Sugars, Catalysis, Cell Membrane, Cloning, Molecular, Glycoproteins, Glycosylphosphatidylinositols, Mutation, N-Acetylneuraminic Acid, Neuraminidase, Trypanosoma brucei brucei}, issn = {1090-2104}, doi = {10.1016/j.bbrc.2011.10.085}, author = {Nakatani, Fumiki and Morita, Yasu S and Ashida, Hisashi and Nagamune, Kisaburo and Maeda, Yusuke and Kinoshita, Taroh} } @article {430, title = {In situ to in silico and back: elucidating the physiology and ecology of Geobacter spp. using genome-scale modelling.}, journal = {Nat Rev Microbiol}, volume = {9}, year = {2011}, month = {2011 Jan}, pages = {39-50}, abstract = {There is a wide diversity of unexplored metabolism encoded in the genomes of microorganisms that have an important environmental role. Genome-scale metabolic modelling enables the individual reactions that are encoded in annotated genomes to be organized into a coherent whole, which can then be used to predict metabolic fluxes that will optimize cell function under a range of conditions. In this Review, we summarize a series of studies in which genome-scale metabolic modelling of Geobacter spp. has resulted in an in-depth understanding of their central metabolism and ecology. A similar iterative modelling and experimental approach could accelerate elucidation of the physiology and ecology of other microorganisms inhabiting a diversity of environments, and could guide optimization of the practical applications of these species.}, keywords = {Computer Simulation, Environment, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Models, Biological}, issn = {1740-1534}, doi = {10.1038/nrmicro2456}, author = {Mahadevan, Radhakrishnan and Palsson, Bernhard {\O} and Lovley, Derek R} } @article {687, title = {Inositol lipid metabolism in mycobacteria: biosynthesis and regulatory mechanisms.}, journal = {Biochim Biophys Acta}, volume = {1810}, year = {2011}, month = {2011 Jun}, pages = {630-41}, abstract = {BACKGROUND: The genus Mycobacterium includes a number of medically important pathogens. The cell walls of these bacteria have many unique features, including the abundance of various inositol lipids, such as phosphatidylinositol mannosides (PIMs), lipomannan (LM), and lipoarabinomannan (LAM). The biosynthesis of these lipids is believed to be prime drug targets, and has been clarified in detail over the past several years. SCOPE OF REVIEW: Here we summarize our current understanding of the inositol lipid metabolism in mycobacteria. We will highlight unsolved issues and future directions especially in the context of metabolic regulation. MAJOR CONCLUSIONS: Inositol is a building block of phosphatidylinositol (PI), which is further elaborated to become PIMs, LM and LAM. d-myo-inositol 3-phosphate is an intermediate of the de novo inositol synthesis, but it is also the starting substrate for mycothiol synthesis. Controlling the level of d-myo-inositol 3-phosphate appears to be important for maintaining the steady state levels of mycothiol and inositol lipids. Several additional control mechanisms must exist to control the complex biosynthetic pathways of PI, PIMs, LM and LAM. These may include regulatory proteins such as a lipoprotein LpqW, and spatial separation of enzymes, such as the amphipathic PimA mannosyltransferase and later enzymes in the PIMs/LM biosynthetic pathway. Finally, we discuss mechanisms that underlie control of LM/LAM glycan polymer elongation. GENERAL SIGNIFICANCE: Mycobacteria have evolved a complex network of inositol metabolism. Clarifying its metabolism will not only provide better understanding of bacterial pathogenesis, but also understanding of the evolution and general functions of inositol lipids in nature.}, keywords = {Inositol, Lipid Metabolism, Lipids, Models, Biological, Mycobacterium, Phosphatidylinositols}, issn = {0006-3002}, doi = {10.1016/j.bbagen.2011.03.017}, author = {Morita, Yasu S and Fukuda, Takeshi and Sena, Chubert B C and Yamaryo-Botte, Yoshiki and McConville, Malcolm J and Kinoshita, Taroh} } @article {852, title = {Infectious Chlamydia pneumoniae is associated with elevated interleukin-8 and airway neutrophilia in children with refractory asthma.}, journal = {Pediatr Infect Dis J}, volume = {29}, year = {2010}, month = {2010 Dec}, pages = {1093-8}, abstract = {BACKGROUND: Neutrophilic asthma is thought to be less responsive than eosinophilic asthma to anti-inflammatory therapies including corticosteroids. Chlamydia pneumoniae has been implicated in asthma, possibly by induction of interleukin (IL-8). We hypothesized that IL-8 is increased in the bronchoalveolar lavage (BAL) fluid from children with asthma and C. pneumoniae. METHODS: BAL fluid was analyzed for C. pneumoniae and IL-8 using polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay from 2 asthma patient populations in the Bronx, NY and Massachusetts with an average age of 8 and 8.7 years old, respectively. For comparison, samples were also analyzed for C. trachomatis and Mycoplasma 16s DNA. RESULTS: Of 18 Bronx samples analyzed, 6 (33\%) were PCR-positive for C. pneumoniae, 10 (56\%) for C. trachomatis, and 8 (44\%) for Mycoplasma 16s DNA. IL-8 from C. pneumoniae-positive samples was 3.3-fold higher compared with negative samples (P = 0.003). There was no difference between patients tested for C. trachomatis or Mycoplasma. Of 84 Massachusetts samples analyzed, 42 (50\%) were PCR-positive for C. pneumoniae, 42 (50\%) for C. trachomatis, and 13 (16\%) for Mycoplasma. IL-8 concentration from C. pneumoniae-positive samples was 10.49-fold higher compared with negative samples (P = 0.0001). As in the Bronx cohort, there were no differences between patients tested for C. trachomatis or Mycoplasma. Lastly, BAL neutrophilia predicted the presence of C. pneumoniae but not Mycoplasma or C. trachomatis. CONCLUSIONS: Children with asthma who were PCR-positive for C. pneumoniae demonstrated elevated concentrations of IL-8 and neutrophils in BAL fluid compared with similar patients who were positive for C. trachomatis or Mycoplasma organisms, but PCR-negative for C. pneumoniae. Undiagnosed C. pneumoniae infection in children may therefore contribute to poorly controlled asthma via induction of IL-8.}, keywords = {Asthma, Bronchoalveolar Lavage Fluid, Child, Chlamydia trachomatis, Chlamydophila Infections, Chlamydophila pneumoniae, DNA, Bacterial, DNA, Ribosomal, Enzyme-Linked Immunosorbent Assay, Female, Humans, Interleukin-8, Male, Massachusetts, Mycoplasma, Neutrophils, New York City, Polymerase Chain Reaction, Respiratory System, RNA, Ribosomal, 16S}, issn = {1532-0987}, author = {Patel, Katir K and Vicencio, Alfin G and Du, Zhongfang and Tsirilakis, Kalliope and Salva, Paul S and Webley, Wilmore C} } @article {439, title = {Interference with histidyl-tRNA synthetase by a CRISPR spacer sequence as a factor in the evolution of Pelobacter carbinolicus.}, journal = {BMC Evol Biol}, volume = {10}, year = {2010}, month = {2010}, pages = {230}, abstract = {BACKGROUND: Pelobacter carbinolicus, a bacterium of the family Geobacteraceae, cannot reduce Fe(III) directly or produce electricity like its relatives. How P. carbinolicus evolved is an intriguing problem. The genome of P. carbinolicus contains clustered regularly interspaced short palindromic repeats (CRISPR) separated by unique spacer sequences, which recent studies have shown to produce RNA molecules that interfere with genes containing identical sequences. RESULTS: CRISPR spacer $\#$1, which matches a sequence within hisS, the histidyl-tRNA synthetase gene of P. carbinolicus, was shown to be expressed. Phylogenetic analysis and genetics demonstrated that a gene paralogous to hisS in the genomes of Geobacteraceae is unlikely to compensate for interference with hisS. Spacer $\#$1 inhibited growth of a transgenic strain of Geobacter sulfurreducens in which the native hisS was replaced with that of P. carbinolicus. The prediction that interference with hisS would result in an attenuated histidyl-tRNA pool insufficient for translation of proteins with multiple closely spaced histidines, predisposing them to mutation and elimination during evolution, was investigated by comparative genomics of P. carbinolicus and related species. Several ancestral genes with high histidine demand have been lost or modified in the P. carbinolicus lineage, providing an explanation for its physiological differences from other Geobacteraceae. CONCLUSIONS: The disappearance of multiheme c-type cytochromes and other genes typical of a metal-respiring ancestor from the P. carbinolicus lineage may be the consequence of spacer $\#$1 interfering with hisS, a condition that can be reproduced in a heterologous host. This is the first successful co-introduction of an active CRISPR spacer and its target in the same cell, the first application of a chimeric CRISPR construct consisting of a spacer from one species in the context of repeats of another species, and the first report of a potential impact of CRISPR on genome-scale evolution by interference with an essential gene.}, keywords = {Base Sequence, Comparative Genomic Hybridization, Computational Biology, Deltaproteobacteria, DNA, Bacterial, DNA, Intergenic, Evolution, Molecular, Genes, Bacterial, Genome, Bacterial, Geobacillus, Histidine-tRNA Ligase, Inverted Repeat Sequences, Molecular Sequence Data, Phylogeny, Sequence Alignment, Sequence Analysis, DNA}, issn = {1471-2148}, doi = {10.1186/1471-2148-10-230}, author = {Aklujkar, Muktak and Lovley, Derek R} } @article {459, title = {Influence of heterogeneous ammonium availability on bacterial community structure and the expression of nitrogen fixation and ammonium transporter genes during in situ bioremediation of uranium-contaminated groundwater.}, journal = {Environ Sci Technol}, volume = {43}, year = {2009}, month = {2009 Jun 15}, pages = {4386-92}, abstract = {The influence of ammonium availability on bacterial community structure and the physiological status of Geobacter species during in situ bioremediation of uranium-contaminated groundwater was evaluated. Ammonium concentrations varied by 2 orders of magnitude (< 4 to 400 microM) across th study site. Analysis of 16S rRNA sequences suggested that ammonium may have been one factor influencing the community composition prior to acetate amendment with Rhodoferax species predominating over Geobacter species with higher ammonium and Dechloromonas species dominating at the site with lowest ammonium. However, once acetate was added and dissimilatory metal reduction was stimulated, Geobacter species became the predominant organisms at all locations. Rates of U(VI) reduction appeared to be more related to acetate concentrations rather than ammonium levels. In situ mRNA transcript abundance of the nitrogen fixation gene, nifD, and the ammonium transporter gene, amtB, in Geobacter species indicated that ammonium was the primary source of nitrogen during uranium reduction. The abundance of amtB was inversely correlated to ammonium levels, whereas nifD transcript levels were similar across all sites examined. These results suggest that nifD and amtB expression are closely regulated in response to ammonium availability to ensure an adequate supply of nitrogen while conserving cell resources. Thus, quantifying nifD and amtB transcript expression appears to be a useful approach for monitoring the nitrogen-related physiological status of subsurface Geobacter species. This study also emphasizes the need for more detailed analysis of geochemical and physiological interactions at the field scale in order to adequately model subsurface microbial processes during bioremediation.}, keywords = {Carrier Proteins, DNA, Bacterial, Environmental Remediation, Gene Expression Regulation, Bacterial, Gene Library, Geobacter, Nitrogen Fixation, Quaternary Ammonium Compounds, Time Factors, Uranium, Water, Water Pollutants, Radioactive}, issn = {0013-936X}, author = {Mouser, Paula J and N{\textquoteright}guessan, Lucie A and Elifantz, Hila and Holmes, Dawn E and Williams, Kenneth H and Wilkins, Michael J and Long, Philip E and Lovley, Derek R} } @article {394, title = {Inducible protein degradation in Bacillus subtilis using heterologous peptide tags and adaptor proteins to target substrates to the protease ClpXP.}, journal = {Mol Microbiol}, volume = {70}, year = {2008}, month = {2008 Nov}, pages = {1012-25}, abstract = {The ability to manipulate protein levels is useful for dissecting regulatory pathways, elucidating gene function and constructing synthetic biological circuits. We engineered an inducible protein degradation system for use in Bacillus subtilis based on Escherichia coli and Caulobacter crescentusssrA tags and SspB adaptors that deliver proteins to ClpXP for proteolysis. In this system, modified ssrA degradation tags are fused onto the 3{\textquoteright} end of the genes of interest. Unlike wild-type ssrA, these modified tags require the adaptor protein SspB to target tagged proteins for proteolysis. In the absence of SspB, the tagged proteins accumulate to near physiological levels. By inducing SspB expression from a regulated promoter, the tagged substrates are rapidly delivered to the B. subtilis ClpXP protease for degradation. We used this system to degrade the reporter GFP and several native B. subtilis proteins, including, the transcription factor ComA, two sporulation kinases (KinA, KinB) and the sporulation and chromosome partitioning protein Spo0J. We also used modified E. coli and C. crescentus ssrA tags to independently control the degradation of two different proteins in the same cell. These tools will be useful for studying biological processes in B. subtilis and can potentially be modified for use in other bacteria.}, keywords = {Alleles, Bacillus subtilis, Bacterial Proteins, Carrier Proteins, Caulobacter crescentus, DNA-Binding Proteins, Endopeptidase Clp, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Genetic Vectors, Green Fluorescent Proteins, Plasmids, Promoter Regions, Genetic, Protein Engineering, RNA, Bacterial, Substrate Specificity}, issn = {1365-2958}, doi = {10.1111/j.1365-2958.2008.06467.x}, author = {Griffith, Kevin L and Grossman, Alan D} } @article {480, title = {Insights into genes involved in electricity generation in Geobacter sulfurreducens via whole genome microarray analysis of the OmcF-deficient mutant.}, journal = {Bioelectrochemistry}, volume = {73}, year = {2008}, month = {2008 Jun}, pages = {70-5}, abstract = {Geobacter sulfurreducens effectively produces electricity in microbial fuel cells by oxidizing acetate with an electrode serving as the sole electron acceptor. Deletion of the gene encoding OmcF, a monoheme outer membrane c-type cytochrome, substantially decreased current production. Previous studies demonstrated that inhibition of Fe(III) reduction in the OmcF-deficient mutant could be attributed to poor transcription of the gene for OmcB, an outer membrane c-type cytochrome that is required for Fe(III) reduction. However, a mutant in which omcB was deleted produced electricity as well as wild type. Microarray analysis of the OmcF-deficient mutant versus the wild type revealed that many of the genes with the greatest decreases in transcript levels were genes whose expression was previously reported to be upregulated in cells grown with an electrode as the sole electron acceptor. These included genes with putative functions related to metal efflux and/or type I secretion and two hypothetical proteins. The outer membrane cytochromes, OmcS and OmcE, which previous studies have demonstrated are required for optimal current generation, were not detected on the outer surface of the OmcF-deficient mutant even though the omcS and omcE genes were still transcribed, suggesting that the putative secretion system could be involved in the export of outer membrane proteins necessary for electron transfer to the fuel cell anode. These results suggest that the requirement for OmcF for optimal current production is not because OmcF is directly involved in extracellular electron transfer but because OmcF is required for the appropriate transcription of other genes either directly or indirectly involved in electricity production.}, keywords = {Bacterial Outer Membrane Proteins, Cytochromes c, Down-Regulation, Electricity, Gene Expression Regulation, Bacterial, Genome, Bacterial, Geobacter, Mutation, Oligonucleotide Array Sequence Analysis, Transcription, Genetic}, issn = {1567-5394}, doi = {10.1016/j.bioelechem.2008.04.023}, author = {Kim, Byoung-Chan and Postier, Bradley L and Didonato, Raymond J and Chaudhuri, Swades K and Nevin, Kelly P and Lovley, Derek R} } @article {741, title = {Investigating the effect of increasing charge density on the hemolytic activity of synthetic antimicrobial polymers.}, journal = {Biomacromolecules}, volume = {9}, year = {2008}, month = {2008 Oct}, pages = {2805-10}, abstract = {The current study is aimed at investigating the effect of fine-tuning the cationic character of synthetic mimics of antimicrobial peptides (SMAMPs) on the hemolytic and antibacterial activities. A series of novel norbornene monomers that carry one, two, or three Boc-protected amine functionalities was prepared. Ring-opening metathesis polymerization (ROMP) of the monomers, followed by deprotection of the amine groups resulted in cationic antimicrobial polynorbornenes that carry one, two, and three charges per monomer repeat unit. Increasing the number of amine groups on the most hydrophobic polymer reduced its hemolytic activity significantly. To understand the membrane activity of these polymers, we conducted dye leakage experiments on lipid vesicles that mimic bacteria and red blood cell membranes, and these results showed a strong correlation with the hemolysis data.}, keywords = {Amines, Anti-Infective Agents, Antimicrobial Cationic Peptides, Chemistry, Pharmaceutical, Drug Design, Erythrocytes, Escherichia coli, Hemolysis, Humans, Lipids, Magnetic Resonance Spectroscopy, Models, Chemical, Peptides, Polymers, Staphylococcus aureus}, issn = {1526-4602}, doi = {10.1021/bm800569x}, author = {Al-Badri, Zoha M and Som, Abhigyan and Lyon, Sarah and Nelson, Christopher F and N{\"u}sslein, Klaus and Tew, Gregory N} } @article {478, title = {Investigation of direct vs. indirect involvement of the c-type cytochrome MacA in Fe(III) reduction by Geobacter sulfurreducens.}, journal = {FEMS Microbiol Lett}, volume = {286}, year = {2008}, month = {2008 Sep}, pages = {39-44}, abstract = {The electron transfer pathway to Fe(III) reduction in Geobacter sulfurreducens has been hypothesized to consist of a series of c-type cytochromes. Previous genetic studies suggested that the inner membrane-associated, c-type cytochrome, MacA, was a component of the electron transfer chain leading to Fe(III) reduction in the dissimilatory Fe(III)-reducer, G. sulfurreducens. However, investigation of the expression of OmcB, an outer-membrane c-type cytochrome demonstrated previously to be critical for optimal Fe(III) reduction, revealed that both omcB transcript and protein levels were dramatically reduced in the MacA-deficient mutant. Expression of the omcB gene in trans enabled the MacA-deficient mutant to reduce Fe(III) at a rate that was proportional to the level of omcB expression. These results suggest that MacA is not directly involved in electron transfer to Fe(III) and further confirm the importance of OmcB in Fe(III) reduction by G. sulfurreducens.}, keywords = {ATP-Binding Cassette Transporters, Bacterial Proteins, Cytochromes c, Ferric Compounds, Gene Expression Regulation, Bacterial, Geobacter, Oxidation-Reduction}, issn = {0378-1097}, doi = {10.1111/j.1574-6968.2008.01252.x}, author = {Kim, Byoung-Chan and Lovley, Derek R} } @article {505, title = {Importance of c-Type cytochromes for U(VI) reduction by Geobacter sulfurreducens.}, journal = {BMC Microbiol}, volume = {7}, year = {2007}, month = {2007}, pages = {16}, abstract = {BACKGROUND: In order to study the mechanism of U(VI) reduction, the effect of deleting c-type cytochrome genes on the capacity of Geobacter sulfurreducens to reduce U(VI) with acetate serving as the electron donor was investigated. RESULTS: The ability of several c-type cytochrome deficient mutants to reduce U(VI) was lower than that of the wild type strain. Elimination of two confirmed outer membrane cytochromes and two putative outer membrane cytochromes significantly decreased (ca. 50-60\%) the ability of G. sulfurreducens to reduce U(VI). Involvement in U(VI) reduction did not appear to be a general property of outer membrane cytochromes, as elimination of two other confirmed outer membrane cytochromes, OmcB and OmcC, had very little impact on U(VI) reduction. Among the periplasmic cytochromes, only MacA, proposed to transfer electrons from the inner membrane to the periplasm, appeared to play a significant role in U(VI) reduction. A subpopulation of both wild type and U(VI) reduction-impaired cells, 24-30\%, accumulated amorphous uranium in the periplasm. Comparison of uranium-accumulating cells demonstrated a similar amount of periplasmic uranium accumulation in U(VI) reduction-impaired and wild type G. sulfurreducens. Assessment of the ability of the various suspensions to reduce Fe(III) revealed no correlation between the impact of cytochrome deletion on U(VI) reduction and reduction of Fe(III) hydroxide and chelated Fe(III). CONCLUSION: This study indicates that c-type cytochromes are involved in U(VI) reduction by Geobacter sulfurreducens. The data provide new evidence for extracellular uranium reduction by G. sulfurreducens but do not rule out the possibility of periplasmic uranium reduction. Occurrence of U(VI) reduction at the cell surface is supported by the significant impact of elimination of outer membrane cytochromes on U(VI) reduction and the lack of correlation between periplasmic uranium accumulation and the capacity for uranium reduction. Periplasmic uranium accumulation may reflect the ability of uranium to penetrate the outer membrane rather than the occurrence of enzymatic U(VI) reduction. Elimination of cytochromes rarely had a similar impact on both Fe(III) and U(VI) reduction, suggesting that there are differences in the routes of electron transfer to U(VI) and Fe(III). Further studies are required to clarify the pathways leading to U(VI) reduction in G. sulfurreducens.}, keywords = {Biodegradation, Environmental, Cytochrome c Group, Ferric Compounds, Geobacter, Microscopy, Electron, Transmission, Mutation, Oxidation-Reduction, Periplasm, Uranium}, issn = {1471-2180}, doi = {10.1186/1471-2180-7-16}, author = {Shelobolina, Evgenya S and Coppi, Maddalena V and Korenevsky, Anton A and DiDonato, Laurie N and Sullivan, Sara A and Konishi, Hiromi and Xu, Huifang and Leang, Ching and Butler, Jessica E and Kim, Byoung-Chan and Lovley, Derek R} } @article {815, title = {Intracranial pressure and cerebral oxygenation changes after decompressive craniectomy in a child with traumatic brain swelling.}, journal = {Childs Nerv Syst}, volume = {23}, year = {2007}, month = {2007 Nov}, pages = {1331-5}, abstract = {CASE REPORT: The authors present the case of a 5-year-old child with severe traumatic brain injury in whom decompressive hemicraniectomy was performed for progressive increased intracranial pressure (ICP) unresponsive to medical treatment. Data from ICP and cerebral tissue oxygenation monitoring in the contralateral hemisphere were recorded, which demonstrated the immediate and delayed mechanical and physiological changes occurring after bony and dural decompression. DISCUSSION: The role of the procedure and that of the monitoring approach are discussed.}, keywords = {Brain Edema, Brain Injuries, Cerebrum, Child, Preschool, Craniotomy, Decompression, Surgical, Functional Laterality, Humans, Hypoxia, Brain, Intracranial Hypertension, Male, Oxygen, Treatment Outcome}, issn = {0256-7040}, doi = {10.1007/s00381-007-0388-4}, author = {Figaji, A A and Fieggen, A G and Sandler, S J I and Argent, A C and Le Roux, P D and Peter, J C} } @article {499, title = {Involvement of Geobacter sulfurreducens SfrAB in acetate metabolism rather than intracellular, respiration-linked Fe(III) citrate reduction.}, journal = {Microbiology}, volume = {153}, year = {2007}, month = {2007 Oct}, pages = {3572-85}, abstract = {A soluble ferric reductase, SfrAB, which catalysed the NADPH-dependent reduction of chelated Fe(III), was previously purified from the dissimilatory Fe(III)-reducing micro-organism Geobacter sulfurreducens, suggesting that reduction of chelated forms of Fe(III) might be cytoplasmic. However, metabolically active spheroplast suspensions could not catalyse acetate-dependent Fe(III) citrate reduction, indicating that periplasmic and/or outer-membrane components were required for Fe(III) citrate reduction. Furthermore, phenotypic analysis of an SfrAB knockout mutant suggested that SfrAB was involved in acetate metabolism rather than respiration-linked Fe(III) reduction. The mutant could not grow via the reduction of either Fe(III) citrate or fumarate when acetate was the electron donor but could grow with either acceptor if either hydrogen or formate served as the electron donor. Following prolonged incubation in acetate : fumarate medium in the absence of hydrogen and formate, an {\textquoteright}acetate-adapted{\textquoteright} SfrAB-null strain was isolated that was capable of growth on acetate : fumarate medium but not acetate : Fe(III) citrate medium. Comparison of gene expression in this strain with that of the wild-type revealed upregulation of a potential NADPH-dependent ferredoxin oxidoreductase as well as genes involved in energy generation and amino acid uptake, suggesting that NADPH homeostasis and the tricarboxylic acid (TCA) cycle were perturbed in the {\textquoteright}acetate-adapted{\textquoteright} SfrAB-null strain. Membrane and soluble fractions prepared from the {\textquoteright}acetate-adapted{\textquoteright} strain were depleted of NADPH-dependent Fe(III), viologen and quinone reductase activities. These results indicate that cytoplasmic, respiration-linked reduction of Fe(III) by SfrAB in vivo is unlikely and suggest that deleting SfrAB may interfere with growth via acetate oxidation by interfering with NADP regeneration.}, keywords = {Acetates, Amino Acid Transport Systems, Bacterial Proteins, Cell Membrane, Citric Acid Cycle, Cytoplasm, Energy Metabolism, Ferric Compounds, Formic Acids, Fumarates, Gene Deletion, Gene Expression Profiling, Geobacter, Hydrogen, NADH, NADPH Oxidoreductases, Oligonucleotide Array Sequence Analysis}, issn = {1350-0872}, doi = {10.1099/mic.0.2007/006478-0}, author = {Coppi, Maddalena V and O{\textquoteright}Neil, Regina A and Leang, Ching and Kaufmann, Franz and Meth{\'e}, Barbara A and Nevin, Kelly P and Woodard, Trevor L and Liu, Anna and Lovley, Derek R} } @article {694, title = {Identification of a novel protein with a role in lipoarabinomannan biosynthesis in mycobacteria.}, journal = {J Biol Chem}, volume = {281}, year = {2006}, month = {2006 Apr 7}, pages = {9011-7}, abstract = {All species of Mycobacteria synthesize distinctive cell walls that are rich in phosphatidylinositol mannosides (PIMs), lipomannan (LM), and lipoarabinomannan (LAM). PIM glycolipids, having 2-4 mannose residues, can either be channeled into polar PIM species (with 6 Man residues) or hypermannosylated to form LM and LAM. In this study, we have identified a Mycobacterium smegmatis gene, termed lpqW, that is required for the conversion of PIMs to LAM and is highly conserved in all mycobacteria. A transposon mutant, Myco481, containing an insertion near the 3{\textquoteright} end of lpqW exhibited altered colony morphology on complex agar medium. This mutant was unstable and was consistently overgrown by a second mutant, represented by Myco481.1, that had normal growth and colony characteristics. Biochemical analysis and metabolic labeling studies showed that Myco481 synthesized the complete spectrum of apolar and polar PIMs but was unable to make LAM. LAM biosynthesis was restored to near wild type levels in Myco481.1. However, this mutant was unable to synthesize the major polar PIM (AcPIM6) and accumulated a smaller intermediate, AcPIM4. Targeted disruption of the lpqW gene and complementation of the initial Myco481 mutant with the wild type gene confirmed that the phenotype of this mutant was due to loss of LpqW. These studies suggest that LpqW has a role in regulating the flux of early PIM intermediates into polar PIM or LAM biosynthesis. They also suggest that AcPIM4 is the likely branch point intermediate in polar PIM and LAM biosynthesis.}, keywords = {Cell Membrane, Gene Expression Regulation, Bacterial, Genes, Bacterial, Lipopolysaccharides, Mutation, Mycobacterium smegmatis, Phenotype, Phosphatidylinositols, Virulence}, issn = {0021-9258}, doi = {10.1074/jbc.M511709200}, author = {Kovacevic, Svetozar and Anderson, Dianne and Morita, Yasu S and Patterson, John and Haites, Ruth and McMillan, Benjamin N I and Coppel, Ross and McConville, Malcolm J and Billman-Jacobe, Helen} } @article {361, title = {Infection with the insect virus Hz-2v alters mating behavior and pheromone production in female Helicoverpa zea moths.}, journal = {J Insect Sci}, volume = {5}, year = {2005}, month = {2005}, pages = {6}, abstract = {The effect of Hz-2V virus infection on the reproductive physiology and behavior of infected Helicoverpa zea female moths was examined. In the absence of males, infected females exhibited calling behavior and called as often but for shorter periods on average than control females. As expected, control females mated with males for extend periods when they were present and did not call after mating, while virus-infected females made many frequent contacts with males and continued to call even after these contacts. Virus-infected females were found to produce five to seven times more pheromone than control females and attracted twice as many males as did control females in flight tunnel experiments. The ability of Hz-2V to alter the physiology and behavior of infected females observed here may serve to facilitate the transmission of virus in insect populations.}, keywords = {Animals, Female, Infectious Disease Transmission, Vertical, Insect Viruses, Male, Moths, Pheromones, Sexual Behavior, Animal, Time Factors, Vocalization, Animal}, issn = {1536-2442}, author = {Burand, John P and Tan, Weijia and Kim, Woojin and Nojima, Satoshi and Roelofs, Wendell} } @article {555, title = {Identification of an uptake hydrogenase required for hydrogen-dependent reduction of Fe(III) and other electron acceptors by Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {186}, year = {2004}, month = {2004 May}, pages = {3022-8}, abstract = {Geobacter sulfurreducens, a representative of the family Geobacteraceae that predominates in Fe(III)-reducing subsurface environments, can grow by coupling the oxidation of hydrogen to the reduction of a variety of electron acceptors, including Fe(III), fumarate, and quinones. An examination of the G. sulfurreducens genome revealed two operons, hya and hyb, which appeared to encode periplasmically oriented respiratory uptake hydrogenases. In order to assess the roles of these two enzymes in hydrogen-dependent growth, Hya- and Hyb-deficient mutants were generated by gene replacement. Hyb was found to be required for hydrogen-dependent reduction of Fe(III), anthraquinone-2,6-disulfonate, and fumarate by resting cell suspensions and to be essential for growth with hydrogen and these three electron acceptors. Hya, in contrast, was not. These findings suggest that Hyb is an essential respiratory hydrogenase in G. sulfurreducens.}, keywords = {Anthraquinones, Ferric Compounds, Fumarates, Geobacter, Hydrogen, Operon, Oxidation-Reduction, Oxidoreductases, Phenotype}, issn = {0021-9193}, author = {Coppi, Maddalena V and O{\textquoteright}Neil, Regina A and Lovley, Derek R} } @article {41, title = {Identification of novel dominant INO2c mutants with an Opi- phenotype.}, journal = {Mol Microbiol}, volume = {52}, year = {2004}, month = {2004 Jun}, pages = {1271-80}, abstract = {The INO2 gene of Saccharomyces cerevisiae is required for derepression of the phospholipid biosynthetic genes in response to inositol depletion. Conversely, the OPI1 gene is required for repression in response to inositol supplementation. Results of an in vitro assay have led to a model in which Opi1p interacts with Ino2p. However, there is no in vivo evidence to support this model. Additionally, most of the previously isolated ino2 mutants offer little insight into this model. Here, we report the isolation of a new class of dominant mutations in the INO2 gene, which yield constitutive expression of a target gene (i.e. an Opi(-) mutant phenotype). Two mutations reside in a region of the Ino2p required for interaction with Opi1p in vitro. Three other mutations are at the amino-terminus in a transcriptional activation domain.}, keywords = {Basic Helix-Loop-Helix Transcription Factors, Gene Expression Regulation, Fungal, Inositol, Mutation, Phenotype, Repressor Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Transcription Factors}, issn = {0950-382X}, doi = {10.1111/j.1365-2958.2004.04069.x}, author = {Gardenour, Kyle R and Levy, Jamie and Lopes, John M} } @article {543, title = {In situ expression of nifD in Geobacteraceae in subsurface sediments.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 Dec}, pages = {7251-9}, abstract = {In order to determine whether the metabolic state of Geobacteraceae involved in bioremediation of subsurface sediments might be inferred from levels of mRNA for key genes, in situ expression of nifD, a highly conserved gene involved in nitrogen fixation, was investigated. When Geobacter sulfurreducens was grown without a source of fixed nitrogen in chemostats with acetate provided as the limiting electron donor and Fe(III) as the electron acceptor, levels of nifD transcripts were 4 to 5 orders of magnitude higher than in chemostat cultures provided with ammonium. In contrast, the number of transcripts of recA and the 16S rRNA gene were slightly lower in the absence of ammonium. The addition of acetate to organic- and nitrogen-poor subsurface sediments stimulated the growth of Geobacteraceae and Fe(III) reduction, as well as the expression of nifD in Geobacteraceae. Levels of nifD transcripts in Geobacteraceae decreased more than 100-fold within 2 days after the addition of 100 microM ammonium, while levels of recA and total bacterial 16S rRNA in Geobacteraceae remained relatively constant. Ammonium amendments had no effect on rates of Fe(III) reduction in acetate-amended sediments or toluene degradation in petroleum-contaminated sediments, suggesting that other factors, such as the rate that Geobacteraceae could access Fe(III) oxides, limited Fe(III) reduction. These results demonstrate that it is possible to monitor one aspect of the in situ metabolic state of Geobacteraceae species in subsurface sediments via analysis of mRNA levels, which is the first step toward a more global analysis of in situ gene expression related to nutrient status and stress response during bioremediation by Geobacteraceae.}, keywords = {Acetates, Biodegradation, Environmental, Culture Media, DNA, Ribosomal, Fresh Water, Gene Expression Regulation, Bacterial, Geobacter, Geologic Sediments, Nitrogenase, Petroleum, Phylogeny, Polymerase Chain Reaction, Quaternary Ammonium Compounds, Rec A Recombinases, RNA, Ribosomal, 16S, Water Pollutants, Chemical}, issn = {0099-2240}, doi = {10.1128/AEM.70.12.7251-7259.2004}, author = {Holmes, Dawn E and Nevin, Kelly P and Lovley, Derek R} } @article {362, title = {In vivo dose-response of insects to Hz-2V infection.}, journal = {Virol J}, volume = {1}, year = {2004}, month = {2004}, pages = {15}, abstract = {BACKGROUND: Hz-2V infection of female Helicoverpa zea moths is manifested as insects that are either sterile "agonadal" individuals with malformed reproductive tissues or fertile asymptomatic carriers which are capable of transmitting virus on to their progeny. Virus infected progeny arising from eggs laid by asymptomatic carrier females may themselves be either sterile agonadals or asymptomatic carriers. RESULTS: By injecting virus into female moths, a correlation was established between virus doses administered to the females and the levels of resulting asymptomatic and sterile progeny. CONCLUSIONS: The results of these experiments indicate that high virus doses produced a higher level of agonadal progeny and lower doses produced higher levels of asymptomatic carriers.}, keywords = {Animals, Female, Insect Viruses, Male, Moths, Ovum, Virus Diseases}, issn = {1743-422X}, doi = {10.1186/1743-422X-1-15}, author = {Burand, John P and Rallis, Christopher P} } @article {554, title = {Isolation, characterization, and U(VI)-reducing potential of a facultatively anaerobic, acid-resistant Bacterium from Low-pH, nitrate- and U(VI)-contaminated subsurface sediment and description of Salmonella subterranea sp. nov.}, journal = {Appl Environ Microbiol}, volume = {70}, year = {2004}, month = {2004 May}, pages = {2959-65}, abstract = {A facultatively anaerobic, acid-resistant bacterium, designated strain FRCl, was isolated from a low-pH, nitrate- and U(VI)-contaminated subsurface sediment at site FW-024 at the Natural and Accelerated Bioremediation Research Field Research Center in Oak Ridge, Tenn. Strain FRCl was enriched at pH 4.5 in minimal medium with nitrate as the electron acceptor, hydrogen as the electron donor, and acetate as the carbon source. Clones with 16S ribosomal DNA (rDNA) sequences identical to the sequence of strain FRCl were also detected in a U(VI)-reducing enrichment culture derived from the same sediment. Cells of strain FRCl were gram-negative motile regular rods 2.0 to 3.4 micro m long and 0.7 to 0.9 microm in diameter. Strain FRCl was positive for indole production, by the methyl red test, and for ornithine decarboxylase; it was negative by the Voges-Proskauer test (for acetylmethylcarbinol production), for urea hydrolysis, for arginine dihydrolase, for lysine decarboxylase, for phenylalanine deaminase, for H(2)S production, and for gelatin hydrolysis. Strain FRCl was capable of using O(2), NO(3)(-), S(2)O(3)(2-), fumarate, and malate as terminal electron acceptors and of reducing U(VI) in the cell suspension. Analysis of the 16S rDNA sequence of the isolate indicated that this strain was 96.4\% similar to Salmonella bongori and 96.3\% similar to Enterobacter cloacae. Physiological and phylogenetic analyses suggested that strain FRCl belongs to the genus Salmonella and represents a new species, Salmonella subterranea sp. nov.}, keywords = {Anaerobiosis, Culture Media, DNA, Ribosomal, Fresh Water, Geologic Sediments, Hydrogen-Ion Concentration, Molecular Sequence Data, Nitrates, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, Salmonella, Sequence Analysis, DNA, Uranium, Water Pollution, Chemical}, issn = {0099-2240}, author = {Shelobolina, Evgenya S and Sullivan, Sara A and O{\textquoteright}Neill, Kathleen R and Nevin, Kelly P and Lovley, Derek R} } @article {1215, title = {Identification of 2D-gel proteins: a comparison of MALDI/TOF peptide mass mapping to mu LC-ESI tandem mass spectrometry.}, journal = {J Am Soc Mass Spectrom}, volume = {14}, year = {2003}, month = {2003 Sep}, pages = {957-70}, abstract = {

A comparative analysis of protein identification for a total of 162 protein spots separated by two-dimensional gel electrophoresis from two fully sequenced archaea, Methanococcus jannaschii and Pyrococcus furiosus, using MALDI-TOF peptide mass mapping (PMM) and mu LC-MS/MS is presented. 100\% of the gel spots analyzed were successfully matched to the predicted proteins in the two corresponding open reading frame databases by mu LC-MS/MS while 97\% of them were identified by MALDI-TOF PMM. The high success rate from the PMM resulted from sample desalting/concentrating with ZipTip(C18) and optimization of several PMM search parameters including a 25 ppm average mass tolerance and the application of two different protein molecular weight search windows. By using this strategy, low-molecular weight (<23 kDa) proteins could be identified unambiguously with less than 5 peptide matches. Nine percent of spots were identified as containing multiple proteins. By using mu LC-MS/MS, 50\% of the spots analyzed were identified as containing multiple proteins. mu LC-MS/MS demonstrated better protein sequence coverage than MALDI-TOF PMM over the entire mass range of proteins identified. MALDI-TOF and PMM produced unique peptide molecular weight matches that were not identified by mu LC-MS/MS. By incorporating amino acid sequence modifications into database searches, combined sequence coverage obtained from these two complimentary ionization methods exceeded 50\% for approximately 70\% of the 162 spots analyzed. This improved sequence coverage in combination with enzymatic digestions of different specificity is proposed as a method for analysis of post-translational modification from 2D-gel separated proteins.

}, keywords = {Amino Acid Sequence, Chromatography, Liquid, Databases, Protein, Electrophoresis, Gel, Two-Dimensional, Methanococcus, Molecular Sequence Data, Molecular Weight, Peptide Mapping, Proteins, Pyrococcus furiosus, Sensitivity and Specificity, Software, Spectrometry, Mass, Electrospray Ionization, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Trypsin}, issn = {1044-0305}, author = {Lim, Hanjo and Eng, Jimmy and Yates, John R and Tollaksen, Sandra L and Giometti, Carol S and Holden, James F and Adams, Michael W W and Reich, Claudia I and Olsen, Gary J and Hays, Lara G} } @article {779, title = {Infectivity of a Cryptosporidium parvum isolate of cervine origin for healthy adults and interferon-gamma knockout mice.}, journal = {J Infect Dis}, volume = {185}, year = {2002}, month = {2002 May 1}, pages = {1320-5}, abstract = {The infectivity of a Cryptosporidium parvum isolate of cervine origin (type 2, Moredun) propagated in calves was investigated simultaneously in healthy adult human volunteers and in interferon-gamma knockout (GKO) mice. After exposure to 100-3000 oocysts, 16 volunteers recorded, for a duration of 6 weeks, the number and form of stools that they passed and any symptoms that they experienced. Oocyst excretion was assessed by enzyme-linked immunosorbent assay and direct immunofluorescence assay. Eleven subjects (69\%) became ill, and 8 subjects (50\%) shed oocysts in stool. The median duration of illness was 169 h, and the median number of unformed stools passed was 24. The duration and intensity of symptoms were more severe than were those associated with previously studied isolates. The median infectious dose was estimated to be 300 oocysts for humans and 1 oocyst for the GKO mouse model. The Moredun isolate was more pathogenic than the reference GCH-1 isolate. The GKO mouse model of cryptosporidiosis is useful for discerning isolate-specific differences in pathogenicity.}, keywords = {Adult, Animals, Body Weight, Cryptosporidiosis, Cryptosporidium parvum, Deer, Diarrhea, Disease Susceptibility, Female, Humans, Interferon-gamma, Mice, Mice, Knockout}, issn = {0022-1899}, doi = {10.1086/340132}, author = {Okhuysen, Pablo C and Rich, Stephen M and Chappell, Cynthia L and Grimes, Kevin A and Widmer, Giovanni and Feng, Xiaochuan and Tzipori, Saul} } @article {1211, title = {Identification of membrane proteins in the hyperthermophilic archaeon pyrococcus furiosus using proteomics and prediction programs.}, journal = {Comp Funct Genomics}, volume = {2}, year = {2001}, month = {2001}, pages = {275-88}, abstract = {

Cell-free extracts from the hyperthermophilic archaeon Pyrococcus furiosus were separated into membrane and cytoplasmic fractions and each was analyzed by 2D-gel electrophoresis. A total of 66 proteins were identified, 32 in the membrane fraction and 34 in the cytoplasmic fraction. Six prediction programs were used to predict the subcellular locations of these proteins. Three were based on signal-peptides (SignalP, TargetP, and SOSUISignal) and three on transmembrane-spanning alpha-helices (TSEG, SOSUI, and PRED-TMR2). A consensus of the six programs predicted that 23 of the 32 proteins (72\%) from the membrane fraction should be in the membrane and that all of the proteins from the cytoplasmic fraction should be in the cytoplasm. Two membrane-associated proteins predicted to be cytoplasmic by the programs are also predicted to consist primarily of transmembrane-spanning beta-sheets using porin protein models, suggesting that they are, in fact, membrane components. An ATPase subunit homolog found in the membrane fraction, although predicted to be cytoplasmic, is most likely complexed with other ATPase subunits in the membrane fraction. An additional three proteins predicted to be cytoplasmic but found in the membrane fraction, may be cytoplasmic contaminants. These include a chaperone homolog that may have attached to denatured membrane proteins during cell fractionation. Omitting these three proteins would boost the membrane-protein predictability of the models to near 80\%. A consensus prediction using all six programs for all 2242 ORFs in the P. furiosus genome estimates that 24\% of the ORF products are found in the membrane. However, this is likely to be a minimum value due to the programs{\textquoteright} inability to recognize certain membrane-related proteins, such as subunits associated with membrane complexes and porin-type proteins.

}, issn = {1531-6912}, doi = {10.1002/cfg.110}, author = {Holden, J F and Poole Ii, F L and Tollaksen, S L and Giometti, C S and Lim, H and Yates Iii, J R and Adams, M W} } @article {592, title = {Isolation and characterization of a soluble NADPH-dependent Fe(III) reductase from Geobacter sulfurreducens.}, journal = {J Bacteriol}, volume = {183}, year = {2001}, month = {2001 Aug}, pages = {4468-76}, abstract = {NADPH is an intermediate in the oxidation of organic compounds coupled to Fe(III) reduction in Geobacter species, but Fe(III) reduction with NADPH as the electron donor has not been studied in these organisms. Crude extracts of Geobacter sulfurreducens catalyzed the NADPH-dependent reduction of Fe(III)-nitrilotriacetic acid (NTA). The responsible enzyme, which was recovered in the soluble protein fraction, was purified to apparent homogeneity in a four-step procedure. Its specific activity for Fe(III) reduction was 65 micromol. min(-1). mg(-1). The soluble Fe(III) reductase was specific for NADPH and did not utilize NADH as an electron donor. Although the enzyme reduced several forms of Fe(III), Fe(III)-NTA was the preferred electron acceptor. The protein possessed methyl viologen:NADP(+) oxidoreductase activity and catalyzed the reduction of NADP(+) with reduced methyl viologen as electron donor at a rate of 385 U/mg. The enzyme consisted of two subunits with molecular masses of 87 and 78 kDa and had a native molecular mass of 320 kDa, as determined by gel filtration. The purified enzyme contained 28.9 mol of Fe, 17.4 mol of acid-labile sulfur, and 0.7 mol of flavin adenine dinucleotide per mol of protein. The genes encoding the two subunits were identified in the complete sequence of the G. sulfurreducens genome from the N-terminal amino acid sequences derived from the subunits of the purified protein. The sequences of the two subunits had about 30\% amino acid identity to the respective subunits of the formate dehydrogenase from Moorella thermoacetica, but the soluble Fe(III) reductase did not possess formate dehydrogenase activity. This soluble Fe(III) reductase differs significantly from previously characterized dissimilatory and assimilatory Fe(III) reductases in its molecular composition and cofactor content.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Deltaproteobacteria, Ferric Compounds, FMN Reductase, Molecular Sequence Data, NADH, NADPH Oxidoreductases, NADP, Nitrilotriacetic Acid, Sequence Analysis, Protein, Sequence Homology, Amino Acid, Solubility}, issn = {0021-9193}, doi = {10.1128/JB.183.15.4468-4476.2001}, author = {Kaufmann, F and Lovley, D R} } @article {590, title = {Isolation, characterization and gene sequence analysis of a membrane-associated 89 kDa Fe(III) reducing cytochrome c from Geobacter sulfurreducens.}, journal = {Biochem J}, volume = {359}, year = {2001}, month = {2001 Oct 1}, pages = {147-52}, abstract = {Geobacter sulfurreducens is capable of anaerobic respiration with Fe(III) as a terminal electron acceptor via a membrane-bound Fe(III) reductase activity associated with a large molecular mass cytochrome c. This cytochrome was purified by detergent extraction of the membrane fraction, Q-Sepharose ion-exchange chromatography, preparative electrophoresis, and MonoQ ion-exchange chromatography. Spectrophotometric analysis of the purified cytochrome reveals a c-type haem, with no evidence of haem a, haem b or sirohaem. The cytochrome has an M(r) of 89000 as determined by denaturing PAGE, and has an isoelectric point of 5.2 as determined by analytical isoelectric focusing. Dithionite-reduced cytochrome can donate electrons to Fe(III)-nitrilotriacetic acid and synthetic ferrihydrite, thus demonstrating that the cytochrome has redox and thermodynamic properties required for reduction of Fe(III). Analysis using cyclic voltammetry confirmed that the reduced cytochrome can catalytically transfer electrons to ferrihydrite, further demonstrating its ability to be an electron transport mediator in anaerobic Fe(III) respiration. Sequence analysis of a cloned chromosomal DNA fragment revealed a 2307 bp open reading frame (ferA) encoding a 768 amino acid protein corresponding to the 89 kDa cytochrome. The deduced amino acid sequence (FerA) translated from the open reading frame contained 12 putative haem-binding motifs, as well as a hydrophobic N-terminal membrane anchor sequence, a lipid-attachment site and an ATP/GTP-binding site. FerA displayed 20\% or less identity with amino acid sequences of other known cytochromes, although it does share some features with characterized polyhaem cytochromes c.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Chromatography, Ion Exchange, Cloning, Molecular, Cytochrome c Group, Deltaproteobacteria, DNA Primers, Electron Transport, Electrophoresis, Polyacrylamide Gel, Ferric Compounds, Gene Expression Regulation, Bacterial, Molecular Sequence Data, Molecular Weight, Nitrilotriacetic Acid, Oxidation-Reduction, Polymerase Chain Reaction, Sequence Homology, Amino Acid}, issn = {0264-6021}, author = {Magnuson, T S and Isoyama, N and Hodges-Myerson, A L and Davidson, G and Maroney, M J and Geesey, G G and Lovley, D R} } @article {828, title = {The importance of repairing stalled replication forks.}, journal = {Nature}, volume = {404}, year = {2000}, month = {2000 Mar 2}, pages = {37-41}, abstract = {The bacterial SOS response to unusual levels of DNA damage has been recognized and studied for several decades. Pathways for re-establishing inactivated replication forks under normal growth conditions have received far less attention. In bacteria growing aerobically in the absence of SOS-inducing conditions, many replication forks encounter DNA damage, leading to inactivation. The pathways for fork reactivation involve the homologous recombination systems, are nonmutagenic, and integrate almost every aspect of DNA metabolism. On a frequency-of-use basis, these pathways represent the main function of bacterial DNA recombination systems, as well as the main function of a number of other enzymatic systems that are associated with replication and site-specific recombination.}, keywords = {Bacteria, Bacterial Proteins, Chromosomes, Bacterial, DNA Replication, DNA, Bacterial, Escherichia coli, Recombination, Genetic, Replication Origin, SOS Response (Genetics)}, issn = {0028-0836}, doi = {10.1038/35003501}, author = {Cox, M M and Goodman, M F and Kreuzer, K N and Sherratt, D J and Sandler, S J and Marians, K J} } @article {864, title = {In vitro polymerization and copolymerization of 3-hydroxypropionyl-CoA with the PHB synthase from Ralstonia eutropha.}, journal = {Biomacromolecules}, volume = {1}, year = {2000}, month = {2000 Fall}, pages = {433-9}, abstract = {The poly(3-hydroxybutyrate) (PHB) synthase of Ralstonia eutropha, which was produced by a recombinant strain of Escherichia coli and purified in one step with a methyl-HIC column to a purity of more than 90\%, was used to polymerize 3-hydroxypropionyl-CoA (3HPCoA) and to copolymerize 3HPCoA with 3-hydroxybutyryl-CoA (3HBCoA). A Km of 189 microM and a kcat of 10 s-1 were determined for the activity of the enzyme in the polymerization reaction of 3HPCoA based on the assumption that the dimer form of PHB synthase was the active form. Free coenzyme A was found to be a very effective competitive inhibitor for the polymerization of 3HPCoA with a Ki of 85 microM. The maximum degree of conversion of 3HPCoA to polymer was less than 40\%. In the simultaneous copolymerization reactions of these two monomers, both the turnover number for the copolymerization reaction and the maximum degree of conversion of 3HPCoA and 3HBCoA to copolymers increased with an increase in the amount of 3HBCoA in the monomer mixture. However, the maximum conversion of 3HPCoA to copolymer was always less than 35\%, regardless of the ratio of 3HPCoA to 3HBCoA. Block copolymers were obtained by the sequential copolymerization of the two monomers and these copolymers had a much narrower molecular weight distribution than those obtained by the simultaneous copolymerization for the same molar ratio of 3HPCoA to 3HBCoA.}, keywords = {Acyltransferases, Coenzyme A, Escherichia coli, Kinetics, Magnetic Resonance Spectroscopy, Molecular Weight}, issn = {1525-7797}, author = {Song, J J and Zhang, S and Lenz, R W and Goodwin, S} } @article {403, title = {Interdependence of the position and orientation of SoxS binding sites in the transcriptional activation of the class I subset of Escherichia coli superoxide-inducible promoters.}, journal = {Mol Microbiol}, volume = {34}, year = {1999}, month = {1999 Nov}, pages = {414-30}, abstract = {SoxS is the direct transcriptional activator of the member genes of the Escherichia coli superoxide regulon. At class I SoxS-dependent promoters, e.g. zwf and fpr, whose SoxS binding sites ({\textquoteright}soxbox{\textquoteright}) lie upstream of the -35 region of the promoter, activation requires the C-terminal domain of the RNA polymerase alpha-subunit, while at class II SoxS-dependent promoters, e.g. fumC and micF, whose binding sites overlap the -35 region, activation is independent of the alpha-CTD. To determine whether SoxS activation of its class I promoters shows the same helical phase-dependent spacing requirement as class I promoters activated by catabolite gene activator protein, we increased the 7 bp distance between the 20 bp zwf soxbox and the zwf -35 promoter hexamer by 5 bp and 11 bp, and we decreased the 15 bp distance between the 20 bp fpr soxbox and the fpr -35 promoter hexamer by the same amounts. In both cases, displacement of the binding site by a half or full turn of the DNA helix prevented transcriptional activation. With constructs containing the binding site of one gene fused to the promoter of the other, we demonstrated that the positional requirements are a function of the specific binding site, not the promoter. Supposing that opposite orientation of the SoxS binding site at the two promoters might account for the positional requirements, we placed the zwf and fpr soxboxes in the reverse orientation at the various positions upstream of the promoters and determined the effect of orientation on transcription activation. We found that reversing the orientation of the zwf binding site converts its positional requirement to that of the fpr binding site in its normal orientation, and vice versa. Analysis by molecular information theory of DNA sequences known to bind SoxS in vitro is consistent with the opposite orientation of the zwf and fpr soxboxes.}, keywords = {Bacterial Proteins, Base Sequence, Binding Sites, Escherichia coli, Escherichia coli Proteins, Molecular Sequence Data, Plasmids, Promoter Regions, Genetic, Sequence Analysis, DNA, Superoxides, Trans-Activators, Transcription Factors, Transcription, Genetic, Transcriptional Activation}, issn = {0950-382X}, author = {Wood, T I and Griffith, K L and Fawcett, W P and Jair, K W and Schneider, T D and Wolf, R E} } @article {391, title = {Identification of a novel dihydrolipoyl dehydrogenase-binding protein in the pyruvate dehydrogenase complex of the anaerobic parasitic nematode, Ascaris suum.}, journal = {J Biol Chem}, volume = {271}, year = {1996}, month = {1996 Mar 8}, pages = {5451-7}, abstract = {A novel dihydrolipoyl dehydrogenase-binding protein (E3BP) which lacks an amino-terminal lipoyl domain, p45, has been identified in the pyruvate dehydrogenase complex (PDC) of the adult parasitic nematode, Ascaris suum. Sequence at the amino terminus of p45 exhibited significant similarity with internal E3-binding domains of dihydrolipoyl transacetylase (E2) and E3BP. Dissociation and resolution of a pyruvate dehydrogenase-depleted adult A. suum PDC in guanidine hydrochloride resulted in two E3-depleted E2 core preparations which were either enriched or substantially depleted of p45. Following reconstitution, the p45-enriched E2 core exhibited enhanced E3 binding, whereas, the p45-depleted E2 core exhibited dramatically reduced E3 binding. Reconstitution of either the bovine kidney or A. suum PDCs with the A. suum E3 suggested that the ascarid E3 was more sensitive to NADH inhibition when bound to the bovine kidney core. The expression of p45 was developmentally regulated and p45 was most abundant in anaerobic muscle. In contrast, E3s isolated from anaerobic muscle or aerobic second-stage larvae were identical. These results suggest that during the transition to anaerobic metabolism, E3 remains unchanged, but it appears that a novel E3BP, p45, is expressed which may help to maintain the activity of the PDC in the face of the elevated intramitochondrial NADH/NAD+ ratios associated with anaerobiosis.}, keywords = {Amino Acid Sequence, Anaerobiosis, Animals, Ascaris suum, Binding Sites, Carrier Proteins, Dihydrolipoamide Dehydrogenase, Electrophoresis, Polyacrylamide Gel, Flavin-Adenine Dinucleotide, Helminth Proteins, Kinetics, Larva, Molecular Sequence Data, NAD, Oxidation-Reduction, Pyruvate Dehydrogenase Complex, Sequence Homology, Amino Acid}, issn = {0021-9258}, author = {Klingbeil, M M and Walker, D J and Arnette, R and Sidawy, E and Hayton, K and Komuniecki, P R and Komuniecki, R} } @article {633, title = {Isolation of Geobacter species from diverse sedimentary environments.}, journal = {Appl Environ Microbiol}, volume = {62}, year = {1996}, month = {1996 May}, pages = {1531-6}, abstract = {In an attempt to better understand the microorganisms responsible for Fe(III) reduction in sedimentary environments, Fe(III)-reducing microorganisms were enriched for and isolated from freshwater aquatic sediments, a pristine deep aquifer, and a petroleum-contaminated shallow aquifer. Enrichments were initiated with acetate or toluene as the electron donor and Fe(III) as the electron acceptor. Isolations were made with acetate or benzoate. Five new strains which could obtain energy for growth by dissimilatory Fe(III) reduction were isolated. All five isolates are gram-negative strict anaerobes which grow with acetate as the electron donor and Fe(III) as the electron acceptor. Analysis of the 16S rRNA sequence of the isolated organisms demonstrated that they all belonged to the genus Geobacter in the delta subdivision of the Proteobacteria. Unlike the type strain, Geobacter metallireducens, three of the five isolates could use H2 as an electron donor for Fe(III) reduction. The deep subsurface isolate is the first Fe(III) reducer shown to completely oxidize lactate to carbon dioxide, while one of the freshwater sediment isolates is only the second Fe(III) reducer known that can oxidize toluene. The isolation of these organisms demonstrates that Geobacter species are widely distributed in a diversity of sedimentary environments in which Fe(III) reduction is an important process.}, keywords = {Base Sequence, Ferric Compounds, Gram-Negative Anaerobic Bacteria, Molecular Sequence Data, Phylogeny, RNA, Bacterial, RNA, Ribosomal, 16S, Water Microbiology}, issn = {0099-2240}, author = {Coates, J D and Phillips, E J and Lonergan, D J and Jenter, H and Lovley, D R} } @article {871, title = {Influence of mass transfer limitations on determination of the half saturation constant for hydrogen uptake in a mixed-culture CH(4)-producing enrichment.}, journal = {Biotechnol Bioeng}, volume = {40}, year = {1992}, month = {1992 Oct 5}, pages = {768-76}, abstract = {There is strong evidence in the literature supporting the existence of significant mass transfer limitations on the kinetics of exogenous H(2) consumption by methanogens. The half saturation constant for H (2) uptake by a mixed-culture, CH(4) producing enrichment was measured using an experimental protocol that avoided internal mass transfer limitations. The value obtained was two orders of magnitude smaller than any other previously reported. A mathematical model for acetogenic syntrophic associations was developed to check the capacity of H(2) as electron transporter between syntrophic partners. It was found that H(2) diffusion could account for the rate of transport of electrons between the syntrophic microorganisms and that formate is not a necessary intermediate. The possibility that formate may be an intermediate in this system was not ruled out. A Monod-type kinetic equation was modified to include the observed H(2) threshold effect. This modified equation was used to predict the CH(4)-production rate in a batch-fed digester. The results show that the external and internal H(2) pools are kinetically coupled.}, issn = {0006-3592}, doi = {10.1002/bit.260400704}, author = {Giraldo-Gomez, E and Goodwin, S and Switzenbaum, M S} } @article {669, title = {Identification of methyl coenzyme M as an intermediate in methanogenesis from acetate in Methanosarcina spp.}, journal = {J Bacteriol}, volume = {160}, year = {1984}, month = {1984 Nov}, pages = {521-5}, abstract = {The transfer of the methyl group of acetate to coenzyme M (2-mercaptoethanesulfonic acid; HS-CoM) during the metabolism of acetate to methane was investigated in cultures of Methanosarcina strain TM-1. The organism metabolized CD3COO- to 83\% CD3H and 17\% CD2H2 and produced no CDH3 or CH4. The isotopic composition of coenzyme M in cells grown on CD3COO- was analyzed with a novel gas chromatography-mass spectrometry technique. The cells contained CD3-D-CoM and CD2H-S-CoM) in a proportion similar to that of CD3H to CD2H2. These results, in conjunction with a report (J.K. Nelson and J.G. Ferry, J. Bacteriol. 160:526-532, 1984) that extracts of acetate-grown strain TM-1 contain high levels of CH3-S-CoM methylreductase, indicate that CH3-S-CoM is an intermediate in the metabolism of acetate to methane in this organism.}, keywords = {Acetates, Deuterium, Euryarchaeota, Gas Chromatography-Mass Spectrometry, Mercaptoethanol, Mesna, Methane, Methylation}, issn = {0021-9193}, author = {Lovley, D R and White, R H and Ferry, J G} } @article {675, title = {Intermediary metabolism of organic matter in the sediments of a eutrophic lake.}, journal = {Appl Environ Microbiol}, volume = {43}, year = {1982}, month = {1982 Mar}, pages = {552-60}, abstract = {The rates, products, and controls of the metabolism of fermentation intermediates in the sediments of a eutrophic lake were examined. C-fatty acids were directly injected into sediment subcores for turnover rate measurements. The highest rates of acetate turnover were in surface sediments (0- to 2-cm depth). Methane was the dominant product of acetate metabolism at all depths. Simultaneous measurements of acetate, propionate, and lactate turnover in surface sediments gave turnover rates of 159, 20, and 3 muM/h, respectively. [2-C]propionate and [U-C]lactate were metabolized to [C]acetate, CO(2), and CH(4). [C]formate was completely converted to CO(2) in less than 1 min. Inhibition of methanogenesis with chloroform resulted in an immediate accumulation of volatile fatty acids and hydrogen. Hydrogen inhibited the metabolism of C(3)-C(5) volatile fatty acids. The rates of fatty acid production were estimated from the rates of fatty acid accumulation in the presence of chloroform or hydrogen. The mean molar rates of production were acetate, 82\%; propionate, 13\%; butyrates, 2\%; and valerates, 3\%. A working model for carbon and electron flow is presented which illustrates that fermentation and methanogenesis are the predominate steps in carbon flow and that there is a close interaction between fermentative bacteria, acetogenic hydrogen-producing bacteria, and methanogens.}, issn = {0099-2240}, author = {Lovley, D R and Klug, M J} } @article {1722, title = {Influence of a new virostatic compound on the induction of enzymes in rat liver.}, journal = {Arzneimittelforschung}, volume = {25}, year = {1975}, month = {1975 Sep}, pages = {1426-9}, abstract = {

The virostatic compound N,N-diethyl-4-[2-(2-oxo-3-tetradecyl-1-imidazolidinyl)-ethyl]-1-piperazinecarboxamide-hydrochloride (5531) was analyzed as to its effect on the induction of tryptophan-pyrrolase and tyrosineaminotransferase in rat liver. 1. The basic activity of the enzymes was not influenced by the substance either in normal or in adrenalectomized animals. 2. The induction of the enzymes by cortisone increased in the presence of the compound whereas the substrate induction remained unchanged. 3. The induction of tyrosine-aminotransferase by dexamethasonephosphate in tissue culture is inhibited if the dose of compound 5531 is higher than 5 mug/ml.

}, keywords = {Adrenalectomy, Animals, Antiviral Agents, Cortisone, Culture Techniques, Dose-Response Relationship, Drug, Enzyme Induction, Female, Imidazoles, Liver, Piperazines, Rats, Time Factors, Tryptophan Oxygenase, Tyrosine, Tyrosine Transaminase}, issn = {0004-4172}, author = {Kr{\"o}ger, H and Donner, I and Skiello, G} } @article {1720, title = {[Influence of beta-blocking agents on experimental atherosclerosis of cocks (author{\textquoteright}s transl)].}, journal = {Cas Lek Cesk}, volume = {114}, year = {1975}, month = {1975 Oct 24}, pages = {1320-5}, keywords = {Adrenergic beta-Antagonists, Animals, Arteriosclerosis, Chickens, Cholesterol, Dietary, Disease Models, Animal, Male}, issn = {0008-7335}, author = {Reinis, Z and Lojda, Z and Heyrovsk{\'y}, A and Hor{\'a}kov{\'a}, D and Reisenauer, R} } @article {1727, title = {Inhibition of aldehyde reductase isoenzymes in human and rat brain.}, journal = {Biochem Pharmacol}, volume = {24}, year = {1975}, month = {1975 Oct 15}, pages = {1865-9}, keywords = {Aldehyde Oxidoreductases, Animals, Anticonvulsants, Barbiturates, Brain, Glutethimide, Humans, Hydantoins, In Vitro Techniques, Isoenzymes, Kinetics, NAD, NADP, Rats, Succinimides}, issn = {0006-2952}, author = {Ris, M M and Deitrich, R A and Von Wartburg, J P} } @article {1714, title = {[Intermediate plateaux in kinetics of the reaction catalyzed by biodegradative L-threonine dehydratase from Escherichia coli].}, journal = {Biokhimiia}, volume = {40}, year = {1975}, month = {1975 May-Jun}, pages = {645-51}, abstract = {

It has been shown that for the reaction catalyzed by "biodegradative" L-threonine dehydratase from E. coli strains K-12 and 980 in 0.5 M phosphate-carbonate buffer, pH 8.4 and pH 9.5, the plots of initial reaction rate (v) versus the initial substrate concentration ([S]0 are characterized by several inflection points, i. e. an intermediate plateau. The plot of v versus the allosteric activator (AMP) concentration have very complicated shapes: there are several inflection points, and also the maximum at L-threonine concentration equal to 3-10(2) and 5-10(-2) M. High AMP concentrations inhibit the enzyme at high substrate concentrations. The reduced glutathion dose not influence the enzyme and does not alter the activating effect of AMP. On the basis of the data obtained it is proposed that the substrate and AMP shift the equilibrium between multiple oligomeric enzyme forms differing in catalytic activity and kinetic manifestations of allosteric interactions between the active and allosteric AMP-binding sites towards polymerization. Thus, the functioning the enzyme under study is discussed in the frames of the model of dissociating regulatory enzymes with multiple intermediate oligomeric forms.

}, keywords = {Adenosine Monophosphate, Allosteric Regulation, Allosteric Site, Binding Sites, Escherichia coli, Hydro-Lyases, Hydrogen-Ion Concentration, Kinetics, Protein Binding, Threonine Dehydratase}, issn = {0320-9725}, author = {Sinelnikova, E M and Dvoretskova, T V and Kagan, Z S} }