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1دورية أكاديمية
المؤلفون: Sang Up Lee, Jin Hwan Park, Seh Hee Jang, Nielsen, Lars K., Jaehyun Kim, Kwang S.Jung
المساهمون: Douglas S. Clark
مصطلحات موضوعية: butanol • metabolic engineering • Clostridium • systems biotechnology, C1, 8699 Other Manufacturing, 100303 Fermentation, 100306 Industrial Molecular Engineering of Nucleic Acids and Proteins
الوصف: Butanol is an aliphatic saturated alcohol having the molecular formula of C4H9OH. Butanol can be used as an intermediate in chemical synthesis and as a solvent for a wide variety of chemical and textile industry applications. Moreover, butanol has been considered as a potential fuel or fuel additive. Biological production of butanol (with acetone and ethanol) was one of the largest industrial fermentation processes early in the 20th century. However, fermentative production of butanol had lost its competitiveness by 1960s due to increasing substrate costs and the advent of more efficient petrochemical processes. Recently, increasing demand for the use of renewable resources as feedstock for the production of chemicals combined with advances in biotechnology through omics, systems biology, metabolic engineering and innovative process developments is generating a renewed interest in fermentative butanol production. This article reviews biotechnological production of butanol by clostridia and some relevant fermentation and downstream processes. The strategies for strain improvement by metabolic engineering and further requirements to make fermentative butanol production a successful industrial process are also discussed. Biotechnol. Bioeng. 2008;101: 209-228. © 2008 Wiley Periodicals, Inc.
العلاقة: orcid:0000-0001-8191-3511; 09-03-01066-a
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2دورية أكاديمية
المؤلفون: Lake-ee Quek, Nielsen, Lars K.
المساهمون: J. Arthur, S-K Ng
مصطلحات موضوعية: systems biology, metabolism, computational model, mouse, C1, 970106 Expanding Knowledge in the Biological Sciences, 060114 Systems Biology
الوصف: Genome-scale metabolic modeling is a systems-based approach that attempts to capture the metabolic complexity of the whole cell, for the purpose of gaining insight into metabolic function and regulation. This is achieved by organizing the metabolic components and their corresponding interactions into a single context. The reconstruction process is a challenging and laborious task, especially during the stage of manual curation. For the mouse genome-scale metabolic model, however, we were able to rapidly reconstruct a compartmentalized model from well-curated metabolic databases online. The prototype model was comprehensive. Apart from minor compound naming and compartmentalization issues, only nine additional reactions without gene associations were added during model curation before the model was able to simulate growth in silico. Further curation led to a metabolic model that consists of 1399 genes mapped to 1757 reactions, with a total of 2037 reactions compartmentalized into the cytoplasm and mitochondria, capable of reproducing metabolic functions inferred from literatures. The reconstruction is made more tractable by developing a formal system to update the model against online databases. Effectively, we can focus our curation efforts into establishing better model annotations and gene–protein–reaction associations within the core metabolism, while relying on genome and proteome databases to build new annotations for peripheral pathways, which may bear less relevance to our modeling interest
العلاقة: orcid:0000-0001-8191-3511
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3دورية أكاديمية
المؤلفون: Lars M. Blank, Philip Hugenholtz, Nielsen, Lars K.
المساهمون: Martin Kreitman
مصطلحات موضوعية: Biochemistry & Molecular Biology, Evolutionary Biology, Genetics & Heredity, C1, 970106 Expanding Knowledge in the Biological Sciences, 060409 Molecular Evolution
الوصف: Hyaluronic acid (HA) is a ubiquitous linear polysaccharide in vertebrates and also is the capsule material of some pathogenic bacteria including group A and C streptococci. In bacteria, the HA synthase occurs in an operon (has) coding for enzymes involved in the production of HA precursors. We report two new members of the has operon family from Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) and Streptococcus equi subsp. equi (S. equi). The has operon of S. zooepidemicus contains, in order, hasA, hasB, hasC, glum, and pgi, whereas these genes are separated on two operons in S. equi (hasA, hasB, hasC and hasC, glmU, pgi). The transcription start site and a σ70 promoter were experimentally identified 50 bp upstream of hasA in S. zooepidemicus. We performed a phylogenetic analysis of each of the has operon genes to determine the evolutionary origin(s) of the streptococcal has operon. In contrast to other capsular and exopolysaccharide operons, has operons have undergone no detectable interspecies lateral gene transfers in their construction, instead relying on intragenome gene duplication for their assembly. Specifically, hasC and glmU appear to have been duplicated into the S. zooepidemicus has operon from remotely located but near-identical paralogues most likely to improve HA productivity by gene dosage in this streptococcus. The intragene rearrangements appear to be ongoing events and the two has operons of the S. equi subspecies represent two alternatives of the same gene arrangement. A scenario for the evolution of streptococcal has operons is proposed.
العلاقة: orcid:0000-0001-5386-7925; orcid:0000-0001-8191-3511
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4دورية أكاديمية
المؤلفون: Gnanasambandam, Annathurai, Anderson, David J., Purnell, Matthew P., Nielsen, Lars K., Brumbley, Stevens M.
مصطلحات موضوعية: mitochondrial presequence, protein targeting, confocal laser scanning microscopy, C1, 829999 Plant Production and Plant Primary Products not elsewhere classified, 060199 Biochemistry and Cell Biology not elsewhere classified, 0607 Plant Biology
الوصف: Approximately 10-15% of plant nuclear genes appear to encode mitochondrial proteins that are directed to mitochondria by specific targeting signals. Reports on the heterologous function of these targeting signals are generally limited to one or a few species, with an emphasis on model plants such as tobacco and Arabidopsis. Given their sequence diversity and their insufficient testing in commercially important crops (including monocotyledonous crops), the extent to which these signals can be relied on for biotechnological purposes across species remains to be established. This study provides the experimental verification of a mitochondrial signal that is functional across diverse crop species, including five monocots (sugarcane, wheat, corn, sorghum and onion) and seven dicots ( cucumber, cauliflower, tomato, capsicum, pumpkin, coriander and sunflower). In all 12 crops, transient assays following microprojectile bombardment showed that the N-terminal mitochondrial presequence from F-1-ATPase beta-subunit (ATPase-beta) of Nicotiana plumbaginifolia Viv. targeted green fluorescent fusion protein to the mitochondria. The transient assay results in sugarcane were confirmed in stably transformed root cells. The ATPase-beta signal should be a useful metabolic engineering tool for directing recombinant proteins to the mitochondrial matrix in diverse plant species of commercial interest.
العلاقة: orcid:0000-0003-1660-0118; orcid:0000-0001-8191-3511
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5دورية أكاديمية
المؤلفون: Michael Hines, Nielsen, Lars K., Cooper-White, Justin J.
المساهمون: J. Melling
مصطلحات موضوعية: Biotechnology & Applied Microbiology, Chemistry, Multidisciplinary, Engineering, Environmental, Chemical, C1, 860899 Human Pharmaceutical Products not elsewhere classified, 100404 Regenerative Medicine (incl. Stem Cells and Tissue Engineering)
الوصف: The pluripotent nature and self-renewal capability of hematopoietic stem cells (HSCs) has resulted in these cells being seen as the ideal cell source for the on-demand production of blood and blood products. This potential to save many thousands of lives is as yet unmet, owing to our inability to reliably expand this cell source ex vivo to clinically relevant numbers. This is, to a large extent, due to the absence of a single cell surface antigen to identify them and an optimum set of in vitro conditions sufficient to expand them while maintaining stemness. This review briefly summarizes the current research efforts aimed at defining the appropriate in vivo conditions to be mimicked in ex vivo culture. In particular, it focuses on the molecules known to participate in the functionality of the bone marrow stem cell niche. These niche molecules include adhesion molecules, extracellular matrix molecules, and soluble and bound growth factors. Finally, this review proposes an approach to find the optimum conditions for this expansion utilizing smart surfaces and a criterion for developing these surfaces. Copyright © 2008 Society of Chemical Industry
العلاقة: orcid:0000-0001-8191-3511; orcid:0000-0002-1920-8229
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6دورية أكاديمية
المؤلفون: Koebmann, Brian, Blank, Lars Mathias, Solem, Christian, Petranovic, Dina, Nielsen, Lars K., Jensen, Peter Ruhdal
المساهمون: Parviz Shamlou
مصطلحات موضوعية: cytochrome, energy metabolism, haemin, Lactococcus lactis, redox cofactor metabolism, respiration, C1, 970110 Expanding Knowledge in Technology, 1003 Industrial Biotechnology
الوصف: Lactococcus lactis is known to be capable of respiration under aerobic conditions in the presence of haemin. In the present study the effect of respiration on ATP production during growth on different sugars was examined. With glucose as the sole carbon source, respiratory conditions in L. lactis MG1363 resulted in only a minor increase, 21%, in biomass yield. Since ATP production through substrate-level phosphorylation was essentially identical with and without respiration, the increased biomass yield was a result of energy-saving under respiratory conditions estimated to be 0.4 mol of ATP/mol of glucose. With maltose as the energy source, the increase in biomass yield amounted to 51% compared with an aerobic culture that lacked haemin. This higher ATP yield was obtained by redirecting pyruvate metabolism from lactate to acetate production, and from savings through respiration. However, even after subtracting these contributions, approx. 0.3 mol of ATP/mol of glucose remained unaccounted for. A similar response to respiratory conditions (0.2 mol of ATP/mol of glucose) was observed in a mutant that had a decreased glucose uptake rate during growth on glucose caused by disruption of the PTSmannose (glucose/mannose-specific phosphotransferase system). Amino acid catabolism could be excluded as the source of the additional ATP. Since mutants without a functional H+-ATPase produced less ATP under sugar starvation and respiratory conditions, the additional ATP yield appears to come partly from energy saved on proton pumping through the H+-ATPase due to respiration and partly from a reversed function of the H+-ATPase towards oxidative phosphorylation. These results may contribute to the design and implementation of carbon-efficient high-cell-density cultures of this industrially important species of bacterium.
العلاقة: orcid:0000-0001-8191-3511
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7دورية أكاديمية
المؤلفون: Song, Hyohak, Kim, Tae Yong, Choi, Bo-Kyeong, Choi, Seong Jun, Nielsen, Lars K., Chang, Ho Nam, Lee, Sang Yup
المساهمون: Alexander Steinbuechel
مصطلحات موضوعية: Chemically defined medium, Mannheimia succiniciproducens, Metabolic network, Flux balance analysis, Succinic acid, C1, 970106 Expanding Knowledge in the Biological Sciences, 060114 Systems Biology, 100303 Fermentation
الوصف: This study presents a novel methodology for the development of a chemically defined medium (CDM) using genome-scale metabolic network and flux balance analysis. The genome-based in silico analysis identified two amino acids and four vitamins as non-substitutable essential compounds to be supplemented to a minimal medium for the sustainable growth of Mannheimia succiniciproducens, while no substitutable essential compounds were identified. The in silico predictions were verified by cultivating the cells on a CDM containing the six non-substitutable essential compounds, and it was further demonstrated by observing no cell growth on the CDM lacking any one of the non-substitutable essentials. An optimal CDM for the enhancement of cell growth and succinic acid production, as a target product, was formulated with a single-addition technique. The fermentation on the optimal CDM increased the succinic acid productivity by 36%, the final succinic acid concentration by 17%, and the succinic acid yield on glucose by 15% compared to the cultivation using a complex medium. The optimal CDM also lowered the sum of the amounts of by-products (acetic, formic, and lactic acids) by 30%. The strategy reported in this paper should be generally applicable to the development of CDMs for other organisms, whose genome sequences are available.
العلاقة: orcid:0000-0001-8191-3511; orcid:0000-0003-0599-3091
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8دورية أكاديمية
مصطلحات موضوعية: Biotechnology & Applied Microbiology, Plant Sciences, biofactory, polyhydroxybutyrate, real-time polymerase chain reaction, Saccharum, transgenic sugarcane, Poly-beta-hydroxybutyrate, Alcaligenes-eutrophus H16, Transplastomic Tobacco, Transgenic Tobacco, Expression, Plants, Gene, Ketothiolase, Arabidopsis, Biosynthesis, 270899 Biotechnology not elsewhere classified, C1, 780105 Biological sciences
الوصف: We report here the results from a glasshouse trial of several transgenic sugarcane (Saccharum spp. hybrids) lines accumulating the bacterial polyester polyhydroxybutyrate (PHB) in plastids. The aims of the trial were to characterize the spatio-temporal pattern of PHB accumulation at a whole-plant level, to identify factors limiting PHB production and to determine whether agronomic performance was affected adversely by PHB accumulation. Statistical analysis showed that a vertical PHB concentration gradient existed throughout the plant, the polymer concentration being lowest in the youngest leaves and increasing with leaf age. In addition, there was a horizontal gradient along the length of a leaf, with the PHB concentration increasing from the youngest part of the leaf (the base) to the oldest (the tip). The rank order of the lines did not change over time. Moreover, there was a uniform spatio-temporal pattern of relative PHB accumulation among the lines, despite the fact that they showed marked differences in absolute PHB concentration. Molecular analysis revealed that the expression of the transgenes encoding the PHB biosynthesis enzymes was apparently coordinated, and that there were good correlations between PHB concentration and the abundance of the PHB biosynthesis enzymes. The maximum recorded PHB concentration, 1.77% of leaf dry weight, did not confer an agronomic penalty. The plant height, total aerial biomass and culm-internode sugar content were not affected relative to controls. Although moderate PHB concentrations were achieved in leaves, the maximum total-plant PHB yield was only 0.79% (11.9 g PHB in 1.51 kg dry weight). We combine the insights from our statistical and molecular analyses to discuss possible strategies for increasing the yield of PHB in sugarcane.
العلاقة: orcid:0000-0001-8191-3511
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9دورية أكاديمية
المؤلفون: Peng, Judy C., Hyde, Claire, Pai, Saparna, O'Sullivan, Brendan J., Nielsen, Lars K., Thomas, Ranjeny
المساهمون: S. A. Rosenberg
مصطلحات موضوعية: Oncology, Immunology, Medicine, Research & Experimental, human dendritic cells, hyperthermia, IL-12, CD40, Heat-shock-protein, Dendritic Cell Maturation, Whole-body Hyperthermia, Fever-range Hyperthermia, Serum-free Conditions, Nf-kappa-b, In-vivo, T-cells, Thermal-stress, Cd40 Ligation, 320202 Cellular Immunology, C1, 730102 Immune system and allergy, 1107 Immunology, 110704 Cellular Immunology
الوصف: Fever is an evolutionarily conserved mechanism to improve survival during infection. Previous studies have shown that feverlike temperatures directly enhance the function of murine bone marrow-derived dendritic cells (DCs). In the present study, we examined the response of human monocyte-derived DC to 39.5 degrees C hyperthermia. When primed with toll-like receptor agonists or bacterial extract but not proinflammatory cytokines, hyperthermia specifically enhanced secretion of interleukin (IL)-12p70 by DC, without altering the secretion of IL-10, tumor necrosis factor a or IL-1 beta. These DC induced significantly higher levels of T-cell proliferation and interferon gamma production in assays of antigen presentation and MLR. Endogenous heat-sock protein 70 colocalized with CD40 in DC exposed to hyperthermic conditions. Recombinant CD40Fc fusion protein blocked the increase in IL-12p70 secretion by DC primed with bacterial extract and hyperthermia. Thus, DC primed with toll-like receptor-agonists respond to hyperthermia with increased IL-12p70 secretion, mediated by heat-shock protein binding and activation of CD40. The data have important applications for clinical immunotherapy and the mechanism of fever
العلاقة: orcid:0000-0001-9773-2571; orcid:0000-0001-8191-3511; orcid:0000-0002-0518-8386
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10دورية أكاديمية
المؤلفون: Peng, Judy C., Thomas, Ranjeny, Nielsen, Lars K.
المساهمون: Steven A. Rosenberg
مصطلحات موضوعية: Dendritic Cells, Immunotherapy, Serum-free Medium, Anti-interleukin-10, Oncology, Immunology, Medicine, Research & Experimental, Nf-kappa-b, Interferon-gamma, Il-12 Production, Cd40 Ligand, T-cells, Reciprocal Regulation, Metastatic Melanoma, Interleukin (il)-4, Immune-responses, In-vitro, 270303 Virology, 320202 Cellular Immunology, C1, 730102 Immune system and allergy, 770804 Control of pests and exotic species, 110322 Rheumatology and Arthritis
الوصف: Monocyte-derived dendritic cells (MoDCs) in clinical use for cancer immunotherapy are ideally generated in serum-free medium (SFM) with inclusion of a suitable maturation factor toward the end of the incubation period. Three good manfacturing practice (GMP) grade SFMs (AIM-V, X-VIVO 15, and X-VIVO 20) were compared with RPMI-1640, supplemented with 10% fetal bovine serum or 10% human serum. DCs generated for 7 days in SFM were less mature and secreted less interleukin (IL) 12p70 and IL-10 than DCs generated in 10% serum. DC yield was comparable in SFMs, and a greater proportion of cells was viable after maturation. Toll-like receptor (TLR) ligands were compared for their ability to induce cytokine secretion under serum-free conditions in the presence of interferon (IFN) gamma. With the exception of Poly I:C, TLR ligands stimulated high levels of IL-10 secretion. High levels of IL-12p70 were induced by two TLR4-mediated stimuli, lipopolysaccharide and Ribomunyl, a clinical-grade bacterial extract. When T-cell responses were compared in allogeneic mixed leukocyte reaction, DCs stimulated with Ribomunyl induced higher levels of IFN gamma than DCs stimulated with the cytokine cocktail: tumor necrosis factor-alpha, IL-1 beta, IL-6, and prostaglandin E-2. In the presence of IL-10 neutralizing antibodies, DC IL-12p70 production and T-cell IFN gamma were increased in vitro. Similarly, DCs stimulated with Ribomunyl, IFN gamma, and anti-IL-10 induced high levels of tetanus toxoid-specific T-cell proliferation and IFN gamma secretion. Thus, MoDCs generated ill SFM efficiently stimulate T-cell IFN gamma production after maturation in the presence of a clinical-grade TLR4 agonist and IL-10 neutralization.
العلاقة: orcid:0000-0002-0518-8386; orcid:0000-0001-8191-3511
النص الكامل