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Sample records for metabolic engineering approach

  1. Production of amino acids - Genetic and metabolic engineering approaches.

    Lee, Jin-Ho; Wendisch, Volker F

    2017-12-01

    The biotechnological production of amino acids occurs at the million-ton scale and annually about 6milliontons of l-glutamate and l-lysine are produced by Escherichia coli and Corynebacterium glutamicum strains. l-glutamate and l-lysine production from starch hydrolysates and molasses is very efficient and access to alternative carbon sources and new products has been enabled by metabolic engineering. This review focusses on genetic and metabolic engineering of amino acid producing strains. In particular, rational approaches involving modulation of transcriptional regulators, regulons, and attenuators will be discussed. To address current limitations of metabolic engineering, this article gives insights on recent systems metabolic engineering approaches based on functional tools and method such as genome reduction, amino acid sensors based on transcriptional regulators and riboswitches, CRISPR interference, small regulatory RNAs, DNA scaffolding, and optogenetic control, and discusses future prospects. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. The Need for Integrated Approaches in Metabolic Engineering

    Lechner, Anna; Brunk, Elizabeth; Keasling, Jay D.

    2016-08-15

    This review highlights state-of-the-art procedures for heterologous small-molecule biosynthesis, the associated bottlenecks, and new strategies that have the potential to accelerate future accomplishments in metabolic engineering. We emphasize that a combination of different approaches over multiple time and size scales must b e considered for successful pathway engineering in a heterologous host. We have classified these optimization procedures based on the "system" that is being manipulated: transcriptome, translatome, proteome, or reactome. By bridging multiple disciplines, including molecular biology, biochemistry, biophysics, and computational sciences, we can create an integral framework for the discovery and implementation of novel biosynthetic production routes.

  3. Modular co-culture engineering, a new approach for metabolic engineering.

    Zhang, Haoran; Wang, Xiaonan

    2016-09-01

    With the development of metabolic engineering, employment of a selected microbial host for accommodation of a designed biosynthetic pathway to produce a target compound has achieved tremendous success in the past several decades. Yet, increasing requirements for sophisticated microbial biosynthesis call for establishment and application of more advanced metabolic engineering methodologies. Recently, important progress has been made towards employing more than one engineered microbial strains to constitute synthetic co-cultures and modularizing the biosynthetic labor between the co-culture members in order to improve bioproduction performance. This emerging approach, referred to as modular co-culture engineering in this review, presents a valuable opportunity for expanding the scope of the broad field of metabolic engineering. We highlight representative research accomplishments using this approach, especially those utilizing metabolic engineering tools for microbial co-culture manipulation. Key benefits and major challenges associated with modular co-culture engineering are also presented and discussed. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  4. Metabolic engineering approaches for production of biochemicals in food and medicinal plants.

    Wilson, Sarah A; Roberts, Susan C

    2014-04-01

    Historically, plants are a vital source of nutrients and pharmaceuticals. Recent advances in metabolic engineering have made it possible to not only increase the concentration of desired compounds, but also introduce novel biosynthetic pathways to a variety of species, allowing for enhanced nutritional or commercial value. To improve metabolic engineering capabilities, new transformation techniques have been developed to allow for gene specific silencing strategies or stacking of multiple genes within the same region of the chromosome. The 'omics' era has provided a new resource for elucidation of uncharacterized biosynthetic pathways, enabling novel metabolic engineering approaches. These resources are now allowing for advanced metabolic engineering of plant production systems, as well as the synthesis of increasingly complex products in engineered microbial hosts. The status of current metabolic engineering efforts is highlighted for the in vitro production of paclitaxel and the in vivo production of β-carotene in Golden Rice and other food crops. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Metabolic Engineering X Conference

    Flach, Evan [American Institute of Chemical Engineers

    2015-05-07

    The International Metabolic Engineering Society (IMES) and the Society for Biological Engineering (SBE), both technological communities of the American Institute of Chemical Engineers (AIChE), hosted the Metabolic Engineering X Conference (ME-X) on June 15-19, 2014 at the Westin Bayshore in Vancouver, British Columbia. It attracted 395 metabolic engineers from academia, industry and government from around the globe.

  6. Engineering Cellular Metabolism

    Nielsen, Jens; Keasling, Jay

    2016-01-01

    Metabolic engineering is the science of rewiring the metabolism of cells to enhance production of native metabolites or to endow cells with the ability to produce new products. The potential applications of such efforts are wide ranging, including the generation of fuels, chemicals, foods, feeds...... of metabolic engineering and will discuss how new technologies can enable metabolic engineering to be scaled up to the industrial level, either by cutting off the lines of control for endogenous metabolism or by infiltrating the system with disruptive, heterologous pathways that overcome cellular regulation....

  7. Metabolic Engineering VII Conference

    Kevin Korpics

    2012-12-04

    The aims of this Metabolic Engineering conference are to provide a forum for academic and industrial researchers in the field; to bring together the different scientific disciplines that contribute to the design, analysis and optimization of metabolic pathways; and to explore the role of Metabolic Engineering in the areas of health and sustainability. Presentations, both written and oral, panel discussions, and workshops will focus on both applications and techniques used for pathway engineering. Various applications including bioenergy, industrial chemicals and materials, drug targets, health, agriculture, and nutrition will be discussed. Workshops focused on technology development for mathematical and experimental techniques important for metabolic engineering applications will be held for more in depth discussion. This 2008 meeting will celebrate our conference tradition of high quality and relevance to both industrial and academic participants, with topics ranging from the frontiers of fundamental science to the practical aspects of metabolic engineering.

  8. Genome-wide analytical approaches for reverse metabolic engineering of industrially relevant phenotypes in yeast

    Oud, Bart; Maris, Antonius J A; Daran, Jean-Marc; Pronk, Jack T

    2012-01-01

    Successful reverse engineering of mutants that have been obtained by nontargeted strain improvement has long presented a major challenge in yeast biotechnology. This paper reviews the use of genome-wide approaches for analysis of Saccharomyces cerevisiae strains originating from evolutionary engineering or random mutagenesis. On the basis of an evaluation of the strengths and weaknesses of different methods, we conclude that for the initial identification of relevant genetic changes, whole genome sequencing is superior to other analytical techniques, such as transcriptome, metabolome, proteome, or array-based genome analysis. Key advantages of this technique over gene expression analysis include the independency of genome sequences on experimental context and the possibility to directly and precisely reproduce the identified changes in naive strains. The predictive value of genome-wide analysis of strains with industrially relevant characteristics can be further improved by classical genetics or simultaneous analysis of strains derived from parallel, independent strain improvement lineages. PMID:22152095

  9. Genome scale engineering techniques for metabolic engineering.

    Liu, Rongming; Bassalo, Marcelo C; Zeitoun, Ramsey I; Gill, Ryan T

    2015-11-01

    Metabolic engineering has expanded from a focus on designs requiring a small number of genetic modifications to increasingly complex designs driven by advances in genome-scale engineering technologies. Metabolic engineering has been generally defined by the use of iterative cycles of rational genome modifications, strain analysis and characterization, and a synthesis step that fuels additional hypothesis generation. This cycle mirrors the Design-Build-Test-Learn cycle followed throughout various engineering fields that has recently become a defining aspect of synthetic biology. This review will attempt to summarize recent genome-scale design, build, test, and learn technologies and relate their use to a range of metabolic engineering applications. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  10. Genome-wide analytical approaches for reverse metabolic engineering of industrially relevant phenotypes in yeast.

    Oud, Bart; van Maris, Antonius J A; Daran, Jean-Marc; Pronk, Jack T

    2012-03-01

    Successful reverse engineering of mutants that have been obtained by nontargeted strain improvement has long presented a major challenge in yeast biotechnology. This paper reviews the use of genome-wide approaches for analysis of Saccharomyces cerevisiae strains originating from evolutionary engineering or random mutagenesis. On the basis of an evaluation of the strengths and weaknesses of different methods, we conclude that for the initial identification of relevant genetic changes, whole genome sequencing is superior to other analytical techniques, such as transcriptome, metabolome, proteome, or array-based genome analysis. Key advantages of this technique over gene expression analysis include the independency of genome sequences on experimental context and the possibility to directly and precisely reproduce the identified changes in naive strains. The predictive value of genome-wide analysis of strains with industrially relevant characteristics can be further improved by classical genetics or simultaneous analysis of strains derived from parallel, independent strain improvement lineages. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  11. Biotechnology and synthetic biology approaches for metabolic engineering of bioenergy crops.

    Shih, Patrick M; Liang, Yan; Loqué, Dominique

    2016-07-01

    The Green Revolution has fuelled an exponential growth in human population since the mid-20th century. Due to population growth, food and energy demands will soon surpass supply capabilities. To overcome these impending problems, significant improvements in genetic engineering will be needed to complement breeding efforts in order to accelerate the improvement of agronomical traits. The new field of plant synthetic biology has emerged in recent years and is expected to support rapid, precise, and robust engineering of plants. In this review, we present recent advances made in the field of plant synthetic biology, specifically in genome editing, transgene expression regulation, and bioenergy crop engineering, with a focus on traits related to lignocellulose, oil, and soluble sugars. Ultimately, progress and innovation in these fields may facilitate the development of beneficial traits in crop plants to meet society's bioenergy needs. © 2016 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

  12. Systematic Applications of Metabolomics in Metabolic Engineering

    Robert A. Dromms

    2012-12-01

    Full Text Available The goals of metabolic engineering are well-served by the biological information provided by metabolomics: information on how the cell is currently using its biochemical resources is perhaps one of the best ways to inform strategies to engineer a cell to produce a target compound. Using the analysis of extracellular or intracellular levels of the target compound (or a few closely related molecules to drive metabolic engineering is quite common. However, there is surprisingly little systematic use of metabolomics datasets, which simultaneously measure hundreds of metabolites rather than just a few, for that same purpose. Here, we review the most common systematic approaches to integrating metabolite data with metabolic engineering, with emphasis on existing efforts to use whole-metabolome datasets. We then review some of the most common approaches for computational modeling of cell-wide metabolism, including constraint-based models, and discuss current computational approaches that explicitly use metabolomics data. We conclude with discussion of the broader potential of computational approaches that systematically use metabolomics data to drive metabolic engineering.

  13. A modular metabolic engineering approach for the production of 1,2-propanediol from glycerol by Saccharomyces cerevisiae.

    Islam, Zia-Ul; Klein, Mathias; Aßkamp, Maximilian R; Ødum, Anders S R; Nevoigt, Elke

    2017-11-01

    Compared to sugars, a major advantage of using glycerol as a feedstock for industrial bioprocesses is the fact that this molecule is more reduced than sugars. A compound whose biotechnological production might greatly profit from the substrate's higher reducing power is 1,2-propanediol (1,2-PDO). Here we present a novel metabolic engineering approach to produce 1,2-PDO from glycerol in S. cerevisiae. Apart from implementing the heterologous methylglyoxal (MG) pathway for 1,2-PDO formation from dihydroxyacetone phosphate (DHAP) and expressing a heterologous glycerol facilitator, the employed genetic modifications included the replacement of the native FAD-dependent glycerol catabolic pathway by the 'DHA pathway' for delivery of cytosolic NADH and the reduction of triosephosphate isomerase (TPI) activity for increased precursor (DHAP) supply. The choice of the medium had a crucial impact on both the strength of the metabolic switch towards fermentation in general (as indicated by the production of ethanol and 1,2-PDO) and on the ratio at which these two fermentation products were formed. For example, virtually no 1,2-PDO but only ethanol was formed in synthetic glycerol medium with urea as the nitrogen source. When nutrient-limited complex YG medium was used, significant amounts of 1,2-PDO were formed and it became obvious that the concerted supply of NADH and DHAP are essential for boosting 1,2-PDO production. Additionally, optimizing the flux into the MG pathway improved 1,2-PDO formation at the expense of ethanol. Cultivation of the best-performing strain in YG medium and a controlled bioreactor set-up resulted in a maximum titer of > 4gL -1 1,2-PDO which, to the best of our knowledge, has been the highest titer of 1,2-PDO obtained in yeast so far. Surprisingly, significant 1,2-PDO production was also obtained in synthetic glycerol medium after changing the nitrogen source towards ammonium sulfate and adding a buffer. Copyright © 2017 International Metabolic

  14. Engineering of metabolic control

    Liao, James C.

    2004-03-16

    The invention features a method of producing heterologous molecules in cells under the regulatory control of a metabolite and metabolic flux. The method can enhance the synthesis of heterologous polypeptides and metabolites.

  15. Synthetic biology and metabolic engineering.

    Stephanopoulos, Gregory

    2012-11-16

    Metabolic engineering emerged 20 years ago as the discipline occupied with the directed modification of metabolic pathways for the microbial synthesis of various products. As such, it deals with the engineering (design, construction, and optimization) of native as well as non-natural routes of product synthesis, aided in this task by the availability of synthetic DNA, the core enabling technology of synthetic biology. The two fields, however, only partially overlap in their interest in pathway engineering. While fabrication of biobricks, synthetic cells, genetic circuits, and nonlinear cell dynamics, along with pathway engineering, have occupied researchers in the field of synthetic biology, the sum total of these areas does not constitute a coherent definition of synthetic biology with a distinct intellectual foundation and well-defined areas of application. This paper reviews the origins of the two fields and advances two distinct paradigms for each of them: that of unit operations for metabolic engineering and electronic circuits for synthetic biology. In this context, metabolic engineering is about engineering cell factories for the biological manufacturing of chemical and pharmaceutical products, whereas the main focus of synthetic biology is fundamental biological research facilitated by the use of synthetic DNA and genetic circuits.

  16. Cytochrome P450-mediated metabolic engineering

    Renault, Hugues; Bassard, Jean-Étienne André; Hamberger, Björn Robert

    2014-01-01

    for the engineered bioproduction of such compounds. Two ground-breaking developments of commercial products driven by the engineering of P450s are the antimalarial drug precursor artemisinic acid and blue roses or carnations. Tedious optimizations were required to generate marketable products. Hurdles encountered...... in P450 engineering and their potential solutions are summarized here. Together with recent technical developments and novel approaches to metabolic engineering, the lessons from this pioneering work should considerably boost exploitation of the amazing P450 toolkit emerging from accelerated sequencing...

  17. Systems metabolic engineering in an industrial setting.

    Sagt, Cees M J

    2013-03-01

    Systems metabolic engineering is based on systems biology, synthetic biology, and evolutionary engineering and is now also applied in industry. Industrial use of systems metabolic engineering focuses on strain and process optimization. Since ambitious yields, titers, productivities, and low costs are key in an industrial setting, the use of effective and robust methods in systems metabolic engineering is becoming very important. Major improvements in the field of proteomics and metabolomics have been crucial in the development of genome-wide approaches in strain and process development. This is accompanied by a rapid increase in DNA sequencing and synthesis capacity. These developments enable the use of systems metabolic engineering in an industrial setting. Industrial systems metabolic engineering can be defined as the combined use of genome-wide genomics, transcriptomics, proteomics, and metabolomics to modify strains or processes. This approach has become very common since the technology for generating large data sets of all levels of the cellular processes has developed quite fast into robust, reliable, and affordable methods. The main challenge and scope of this mini review is how to translate these large data sets in relevant biological leads which can be tested for strain or process improvements. Experimental setup, heterogeneity of the culture, and sample pretreatment are important issues which are easily underrated. In addition, the process of structuring, filtering, and visualization of data is important, but also, the availability of a genetic toolbox and equipment for medium/high-throughput fermentation is a key success factor. For an efficient bioprocess, all the different components in this process have to work together. Therefore, mutual tuning of these components is an important strategy.

  18. Metabolic engineering in methanotrophic bacteria

    Kalyuzhnaya, MG; Puri, AW; Lidstrom, ME

    2015-05-01

    Methane, as natural gas or biogas, is the least expensive source of carbon for (bio)chemical synthesis. Scalable biological upgrading of this simple alkane to chemicals and fuels can bring new sustainable solutions to a number of industries with large environmental footprints, such as natural gas/petroleum production, landfills, wastewater treatment, and livestock. Microbial biocatalysis with methane as a feedstock has been pursued off and on for almost a half century, with little enduring success. Today, biological engineering and systems biology provide new opportunities for metabolic system modulation and give new optimism to the concept of a methane-based bio-industry. Here we present an overview of the most recent advances pertaining to metabolic engineering of microbial methane utilization. Some ideas concerning metabolic improvements for production of acetyl-CoA and pyruvate, two main precursors for bioconversion, are presented. We also discuss main gaps in the current knowledge of aerobic methane utilization, which must be solved in order to release the full potential of methane-based biosystems. (C) 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  19. Protein design in systems metabolic engineering for industrial strain development.

    Chen, Zhen; Zeng, An-Ping

    2013-05-01

    Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Engineering central metabolism – a grand challenge for plant biologists

    Sweetlove, Lee J.; Nielsen, Jens; Fernie, Alisdair R.

    2017-01-01

    The goal of increasing crop productivity and nutrient-use efficiency is being addressed by a number of ambitious research projects seeking to re-engineer photosynthetic biochemistry. Many of these projects will require the engineering of substantial changes in fluxes of central metabolism. However......, as has been amply demonstrated in simpler systems such as microbes, central metabolism is extremely difficult to rationally engineer. This is because of multiple layers of regulation that operate to maintain metabolic steady state and because of the highly connected nature of central metabolism....... In this review we discuss new approaches for metabolic engineering that have the potential to address these problems and dramatically improve the success with which we can rationally engineer central metabolism in plants. In particular, we advocate the adoption of an iterative ‘design-build-test-learn’ cycle...

  1. Engineering students' sustainability approaches

    Haase, S.

    2014-05-01

    Sustainability issues are increasingly important in engineering work all over the world. This article explores systematic differences in self-assessed competencies, interests, importance, engagement and practices of newly enrolled engineering students in Denmark in relation to environmental and non-environmental sustainability issues. The empirical base of the article is a nation-wide, web-based survey sent to all newly enrolled engineering students in Denmark commencing their education in the fall term 2010. The response rate was 46%. The survey focused on a variety of different aspects of what can be conceived as sustainability. By means of cluster analysis, three engineering student approaches to sustainability are identified and described. The article provides knowledge on the different prerequisites of engineering students in relation to the role of sustainability in engineering. This information is important input to educators trying to target new engineering students and contribute to the provision of engineers equipped to meet sustainability challenges.

  2. Synthetic Biology and Metabolic Engineering Approaches and Its Impact on Non-Conventional Yeast and Biofuel Production

    Madhavan, Aravind [Biotechnology Division, National Institute for Interdisciplinary Science and Technology, Council of Scientific and Industrial Research, Trivandrum (India); Rajiv Gandhi Centre for Biotechnology, Trivandrum (India); Jose, Anju Alphonsa; Binod, Parameswaran; Sindhu, Raveendran, E-mail: sindhurgcb@gmail.com; Sukumaran, Rajeev K. [Biotechnology Division, National Institute for Interdisciplinary Science and Technology, Council of Scientific and Industrial Research, Trivandrum (India); Pandey, Ashok [Biotechnology Division, National Institute for Interdisciplinary Science and Technology, Council of Scientific and Industrial Research, Trivandrum (India); Center for Innovative and Applied Bioprocessing, Mohali, Punjab (India); Castro, Galliano Eulogio [Dpt. Ingeniería Química, Ambiental y de los Materiales Edificio, Universidad de Jaén, Jaén (Spain)

    2017-04-25

    The increasing fossil fuel scarcity has led to an urgent need to develop alternative fuels. Currently microorganisms have been extensively used for the production of first-generation biofuels from lignocellulosic biomass. Yeast is the efficient producer of bioethanol among all existing biofuels option. Tools of synthetic biology have revolutionized the field of microbial cell factories especially in the case of ethanol and fatty acid production. Most of the synthetic biology tools have been developed for the industrial workhorse Saccharomyces cerevisiae. The non-conventional yeast systems have several beneficial traits like ethanol tolerance, thermotolerance, inhibitor tolerance, genetic diversity, etc., and synthetic biology have the power to expand these traits. Currently, synthetic biology is slowly widening to the non-conventional yeasts like Hansenula polymorpha, Kluyveromyces lactis, Pichia pastoris, and Yarrowia lipolytica. Herein, we review the basic synthetic biology tools that can apply to non-conventional yeasts. Furthermore, we discuss the recent advances employed to develop efficient biofuel-producing non-conventional yeast strains by metabolic engineering and synthetic biology with recent examples. Looking forward, future synthetic engineering tools’ development and application should focus on unexplored non-conventional yeast species.

  3. Synthetic Biology and Metabolic Engineering Approaches and Its Impact on Non-Conventional Yeast and Biofuel Production

    Raveendran Sindhu

    2017-04-01

    Full Text Available The increasing fossil fuel scarcity has led to an urgent need to develop alternative fuels. Currently microorganisms have been extensively used for the production of first-generation biofuels from lignocellulosic biomass. Yeast is the efficient producer of bioethanol among all existing biofuels option. Tools of synthetic biology have revolutionized the field of microbial cell factories especially in the case of ethanol and fatty acid production. Most of the synthetic biology tools have been developed for the industrial workhorse Saccharomyces cerevisiae. The non-conventional yeast systems have several beneficial traits like ethanol tolerance, thermotolerance, inhibitor tolerance, genetic diversity, etc., and synthetic biology have the power to expand these traits. Currently, synthetic biology is slowly widening to the non-conventional yeasts like Hansenula polymorpha, Kluyveromyces lactis, Pichia pastoris, and Yarrowia lipolytica. Herein, we review the basic synthetic biology tools that can apply to non-conventional yeasts. Furthermore, we discuss the recent advances employed to develop efficient biofuel-producing non-conventional yeast strains by metabolic engineering and synthetic biology with recent examples. Looking forward, future synthetic engineering tools’ development and application should focus on unexplored non-conventional yeast species.

  4. Synthetic Biology and Metabolic Engineering Approaches and Its Impact on Non-Conventional Yeast and Biofuel Production

    Madhavan, Aravind; Jose, Anju Alphonsa; Binod, Parameswaran; Sindhu, Raveendran; Sukumaran, Rajeev K.; Pandey, Ashok; Castro, Galliano Eulogio

    2017-01-01

    The increasing fossil fuel scarcity has led to an urgent need to develop alternative fuels. Currently microorganisms have been extensively used for the production of first-generation biofuels from lignocellulosic biomass. Yeast is the efficient producer of bioethanol among all existing biofuels option. Tools of synthetic biology have revolutionized the field of microbial cell factories especially in the case of ethanol and fatty acid production. Most of the synthetic biology tools have been developed for the industrial workhorse Saccharomyces cerevisiae. The non-conventional yeast systems have several beneficial traits like ethanol tolerance, thermotolerance, inhibitor tolerance, genetic diversity, etc., and synthetic biology have the power to expand these traits. Currently, synthetic biology is slowly widening to the non-conventional yeasts like Hansenula polymorpha, Kluyveromyces lactis, Pichia pastoris, and Yarrowia lipolytica. Herein, we review the basic synthetic biology tools that can apply to non-conventional yeasts. Furthermore, we discuss the recent advances employed to develop efficient biofuel-producing non-conventional yeast strains by metabolic engineering and synthetic biology with recent examples. Looking forward, future synthetic engineering tools’ development and application should focus on unexplored non-conventional yeast species.

  5. Modeling of Zymomonas mobilis central metabolism for novel metabolic engineering strategies.

    Kalnenieks, Uldis; Pentjuss, Agris; Rutkis, Reinis; Stalidzans, Egils; Fell, David A

    2014-01-01

    Mathematical modeling of metabolism is essential for rational metabolic engineering. The present work focuses on several types of modeling approach to quantitative understanding of central metabolic network and energetics in the bioethanol-producing bacterium Zymomonas mobilis. Combined use of Flux Balance, Elementary Flux Mode, and thermodynamic analysis of its central metabolism, together with dynamic modeling of the core catabolic pathways, can help to design novel substrate and product pathways by systematically analyzing the solution space for metabolic engineering, and yields insights into the function of metabolic network, hardly achievable without applying modeling tools.

  6. Plant metabolic modeling: achieving new insight into metabolism and metabolic engineering.

    Baghalian, Kambiz; Hajirezaei, Mohammad-Reza; Schreiber, Falk

    2014-10-01

    Models are used to represent aspects of the real world for specific purposes, and mathematical models have opened up new approaches in studying the behavior and complexity of biological systems. However, modeling is often time-consuming and requires significant computational resources for data development, data analysis, and simulation. Computational modeling has been successfully applied as an aid for metabolic engineering in microorganisms. But such model-based approaches have only recently been extended to plant metabolic engineering, mainly due to greater pathway complexity in plants and their highly compartmentalized cellular structure. Recent progress in plant systems biology and bioinformatics has begun to disentangle this complexity and facilitate the creation of efficient plant metabolic models. This review highlights several aspects of plant metabolic modeling in the context of understanding, predicting and modifying complex plant metabolism. We discuss opportunities for engineering photosynthetic carbon metabolism, sucrose synthesis, and the tricarboxylic acid cycle in leaves and oil synthesis in seeds and the application of metabolic modeling to the study of plant acclimation to the environment. The aim of the review is to offer a current perspective for plant biologists without requiring specialized knowledge of bioinformatics or systems biology. © 2014 American Society of Plant Biologists. All rights reserved.

  7. Thermodynamics an engineering approach

    Cengel, Yunus A

    2014-01-01

    Thermodynamics, An Engineering Approach, eighth edition, covers the basic principles of thermodynamics while presenting a wealth of real-world engineering examples so students get a feel for how thermodynamics is applied in engineering practice. This text helps students develop an intuitive understanding by emphasizing the physics and physical arguments. Cengel and Boles explore the various facets of thermodynamics through careful explanations of concepts and use of numerous practical examples and figures, having students develop necessary skills to bridge the gap between knowledge and the confidence to properly apply their knowledge. McGraw-Hill is proud to offer Connect with the eighth edition of Cengel/Boles, Thermodynamics, An Engineering Approach. This innovative and powerful new system helps your students learn more efficiently and gives you the ability to assign homework problems simply and easily. Problems are graded automatically, and the results are recorded immediately. Track individual stude...

  8. Metabolic engineering of biosynthetic pathway for production of renewable biofuels.

    Singh, Vijai; Mani, Indra; Chaudhary, Dharmendra Kumar; Dhar, Pawan Kumar

    2014-02-01

    Metabolic engineering is an important area of research that involves editing genetic networks to overproduce a certain substance by the cells. Using a combination of genetic, metabolic, and modeling methods, useful substances have been synthesized in the past at industrial scale and in a cost-effective manner. Currently, metabolic engineering is being used to produce sufficient, economical, and eco-friendly biofuels. In the recent past, a number of efforts have been made towards engineering biosynthetic pathways for large scale and efficient production of biofuels from biomass. Given the adoption of metabolic engineering approaches by the biofuel industry, this paper reviews various approaches towards the production and enhancement of renewable biofuels such as ethanol, butanol, isopropanol, hydrogen, and biodiesel. We have also identified specific areas where more work needs to be done in the future.

  9. Towards systems metabolic engineering in Pichia pastoris.

    Schwarzhans, Jan-Philipp; Luttermann, Tobias; Geier, Martina; Kalinowski, Jörn; Friehs, Karl

    2017-11-01

    The methylotrophic yeast Pichia pastoris is firmly established as a host for the production of recombinant proteins, frequently outperforming other heterologous hosts. Already, a sizeable amount of systems biology knowledge has been acquired for this non-conventional yeast. By applying various omics-technologies, productivity features have been thoroughly analyzed and optimized via genetic engineering. However, challenging clonal variability, limited vector repertoire and insufficient genome annotation have hampered further developments. Yet, in the last few years a reinvigorated effort to establish P. pastoris as a host for both protein and metabolite production is visible. A variety of compounds from terpenoids to polyketides have been synthesized, often exceeding the productivity of other microbial systems. The clonal variability was systematically investigated and strategies formulated to circumvent untargeted events, thereby streamlining the screening procedure. Promoters with novel regulatory properties were discovered or engineered from existing ones. The genetic tractability was increased via the transfer of popular manipulation and assembly techniques, as well as the creation of new ones. A second generation of sequencing projects culminated in the creation of the second best functionally annotated yeast genome. In combination with landmark physiological insights and increased output of omics-data, a good basis for the creation of refined genome-scale metabolic models was created. The first application of model-based metabolic engineering in P. pastoris showcased the potential of this approach. Recent efforts to establish yeast peroxisomes for compartmentalized metabolite synthesis appear to fit ideally with the well-studied high capacity peroxisomal machinery of P. pastoris. Here, these recent developments are collected and reviewed with the aim of supporting the establishment of systems metabolic engineering in P. pastoris. Copyright © 2017. Published

  10. Genome-scale modeling for metabolic engineering.

    Simeonidis, Evangelos; Price, Nathan D

    2015-03-01

    We focus on the application of constraint-based methodologies and, more specifically, flux balance analysis in the field of metabolic engineering, and enumerate recent developments and successes of the field. We also review computational frameworks that have been developed with the express purpose of automatically selecting optimal gene deletions for achieving improved production of a chemical of interest. The application of flux balance analysis methods in rational metabolic engineering requires a metabolic network reconstruction and a corresponding in silico metabolic model for the microorganism in question. For this reason, we additionally present a brief overview of automated reconstruction techniques. Finally, we emphasize the importance of integrating metabolic networks with regulatory information-an area which we expect will become increasingly important for metabolic engineering-and present recent developments in the field of metabolic and regulatory integration.

  11. Engineering the spatial organization of metabolic pathways

    Albertsen, Line; Maury, Jerome; Bach, Lars Stougaard

    One of the goals of metabolic engineering is to optimize the production of valuable metabolites in cell factories. In this context, modulating the gene expression and activity of enzymes are tools that have been extensively used. Another approach that is gaining interest is the engineering...... of the spatial organization of biosynthetic pathways. Several natural systems for ensuring optimal spatial arrangement of biosynthetic enzymes exist. Sequentially acting enzymes can for example be positioned in close proximity by attachment to cellular structures, up-concentration in membrane enclosed organelles...... or assembly into large complexes. The vision is that by positioning sequentially acting enzymes in close proximity, the cell can accelerate reaction rates and thereby prevent loss of intermediates through diffusion, degradation or competing pathways. The production of valuable metabolites in cell factories...

  12. Perspectives in metabolic engineering: understanding cellular regulation towards the control of metabolic routes.

    Zadran, Sohila; Levine, Raphael D

    2013-01-01

    Metabolic engineering seeks to redirect metabolic pathways through the modification of specific biochemical reactions or the introduction of new ones with the use of recombinant technology. Many of the chemicals synthesized via introduction of product-specific enzymes or the reconstruction of entire metabolic pathways into engineered hosts that can sustain production and can synthesize high yields of the desired product as yields of natural product-derived compounds are frequently low, and chemical processes can be both energy and material expensive; current endeavors have focused on using biologically derived processes as alternatives to chemical synthesis. Such economically favorable manufacturing processes pursue goals related to sustainable development and "green chemistry". Metabolic engineering is a multidisciplinary approach, involving chemical engineering, molecular biology, biochemistry, and analytical chemistry. Recent advances in molecular biology, genome-scale models, theoretical understanding, and kinetic modeling has increased interest in using metabolic engineering to redirect metabolic fluxes for industrial and therapeutic purposes. The use of metabolic engineering has increased the productivity of industrially pertinent small molecules, alcohol-based biofuels, and biodiesel. Here, we highlight developments in the practical and theoretical strategies and technologies available for the metabolic engineering of simple systems and address current limitations.

  13. Metabolic Engineering: Techniques for analysis of targets for genetic manipulations

    Nielsen, Jens Bredal

    1998-01-01

    Metabolic engineering has been defined as the purposeful modification of intermediary metabolism using recombinant DNA techniques. With this definition metabolic engineering includes: (1) inserting new pathways in microorganisms with the aim of producing novel metabolites, e.g., production...... of polyketides by Streptomyces; (2) production of heterologous peptides, e.g., production of human insulin, erythropoitin, and tPA; and (3) improvement of both new and existing processes, e.g., production of antibiotics and industrial enzymes. Metabolic engineering is a multidisciplinary approach, which involves...... input from chemical engineers, molecular biologists, biochemists, physiologists, and analytical chemists. Obviously, molecular biology is central in the production of novel products, as well as in the improvement of existing processes. However, in the latter case, input from other disciplines is pivotal...

  14. Metabolic Engineering for Substrate Co-utilization

    Gawand, Pratish

    Production of biofuels and bio-based chemicals is being increasingly pursued by chemical industry to reduce its dependence on petroleum. Lignocellulosic biomass (LCB) is an abundant source of sugars that can be used for producing biofuels and bio-based chemicals using fermentation. Hydrolysis of LCB results in a mixture of sugars mainly composed of glucose and xylose. Fermentation of such a sugar mixture presents multiple technical challenges at industrial scale. Most industrial microorganisms utilize sugars in a sequential manner due to the regulatory phenomenon of carbon catabolite repression (CCR). Due to sequential utilization of sugars, the LCB-based fermentation processes suffer low productivities and complicated operation. Performance of fermentation processes can be improved by metabolic engineering of microorganisms to obtain superior characteristics such as high product yield. With increased computational power and availability of complete genomes of microorganisms, use of model-based metabolic engineering is now a common practice. The problem of sequential sugar utilization, however, is a regulatory problem, and metabolic models have never been used to solve such regulatory problems. The focus of this thesis is to use model-guided metabolic engineering to construct industrial strains capable of co-utilizing sugars. First, we develop a novel bilevel optimization algorithm SimUp, that uses metabolic models to identify reaction deletion strategies to force co-utilization of two sugars. We then use SimUp to identify reaction deletion strategies to force glucose-xylose co-utilization in Escherichia coli. To validate SimUp predictions, we construct three mutants with multiple gene knockouts and test them for glucose-xylose utilization characteristics. Two mutants, designated as LMSE2 and LMSE5, are shown to co-utilize glucose and xylose in agreement with SimUp predictions. To understand the molecular mechanism involved in glucose-xylose co-utilization of the

  15. Homogenization approach in engineering

    Babuska, I.

    1975-10-01

    Homogenization is an approach which studies the macrobehavior of a medium by its microproperties. Problems with a microstructure play an essential role in such fields as mechanics, chemistry, physics, and reactor engineering. Attention is concentrated on a simple specific model problem to illustrate results and problems typical of the homogenization approach. Only the diffusion problem is treated here, but some statements are made about the elasticity of composite materials. The differential equation is solved for linear cases with and without boundaries and for the nonlinear case. 3 figures, 1 table

  16. Microbial Development and Metabolic Engineering | Bioenergy | NREL

    Diversity Our genetically engineered microbes utilize a variety of feedstock including cellulose, xylan , syngas, simple sugars, organic acids, and carbon dioxide (CO2). We have modified the metabolic pathways

  17. Engineering of sugar metabolism in Lactococcus lactis

    Pool, Weia Arianne

    2008-01-01

    Short English Summary Lactococcus lactis is a lactic acid bacterium used in the dairy industry. This thesis decribes the genetic engineering performed on the sugar metabolism of L. lactis. Besides our fundamental interest for sugar metabolism and its regulation in L. lactis, this project had the

  18. A modular metabolic engineering approach for the production of 1,2-propanediol from glycerol by Saccharomyces cerevisiae

    Islam, Zia-Ul; Klein, Mathias; Aßkamp, Maximilian R.

    2017-01-01

    Compared to sugars, a major advantage of using glycerol as a feedstock for industrial bioprocesses is the fact that this molecule is more reduced than sugars. A compound whose biotechnological production might greatly profit from the substrate's higher reducing power is 1,2-propanediol (1,2-PDO...... as the nitrogen source. When nutrient-limited complex YG medium was used, significant amounts of 1,2-PDO were formed and it became obvious that the concerted supply of NADH and DHAP are essential for boosting 1,2-PDO production. Additionally, optimizing the flux into the MG pathway improved 1,2-PDO formation...... on both the strength of the metabolic switch towards fermentation in general (as indicated by the production of ethanol and 1,2-PDO) and on the ratio at which these two fermentation products were formed. For example, virtually no 1,2-PDO but only ethanol was formed in synthetic glycerol medium with urea...

  19. Metabolic engineering tools in model cyanobacteria.

    Carroll, Austin L; Case, Anna E; Zhang, Angela; Atsumi, Shota

    2018-03-26

    Developing sustainable routes for producing chemicals and fuels is one of the most important challenges in metabolic engineering. Photoautotrophic hosts are particularly attractive because of their potential to utilize light as an energy source and CO 2 as a carbon substrate through photosynthesis. Cyanobacteria are unicellular organisms capable of photosynthesis and CO 2 fixation. While engineering in heterotrophs, such as Escherichia coli, has result in a plethora of tools for strain development and hosts capable of producing valuable chemicals efficiently, these techniques are not always directly transferable to cyanobacteria. However, recent efforts have led to an increase in the scope and scale of chemicals that cyanobacteria can produce. Adaptations of important metabolic engineering tools have also been optimized to function in photoautotrophic hosts, which include Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9, 13 C Metabolic Flux Analysis (MFA), and Genome-Scale Modeling (GSM). This review explores innovations in cyanobacterial metabolic engineering, and highlights how photoautotrophic metabolism has shaped their development. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  20. Computer-aided design for metabolic engineering.

    Fernández-Castané, Alfred; Fehér, Tamás; Carbonell, Pablo; Pauthenier, Cyrille; Faulon, Jean-Loup

    2014-12-20

    The development and application of biotechnology-based strategies has had a great socio-economical impact and is likely to play a crucial role in the foundation of more sustainable and efficient industrial processes. Within biotechnology, metabolic engineering aims at the directed improvement of cellular properties, often with the goal of synthesizing a target chemical compound. The use of computer-aided design (CAD) tools, along with the continuously emerging advanced genetic engineering techniques have allowed metabolic engineering to broaden and streamline the process of heterologous compound-production. In this work, we review the CAD tools available for metabolic engineering with an emphasis, on retrosynthesis methodologies. Recent advances in genetic engineering strategies for pathway implementation and optimization are also reviewed as well as a range of bionalytical tools to validate in silico predictions. A case study applying retrosynthesis is presented as an experimental verification of the output from Retropath, the first complete automated computational pipeline applicable to metabolic engineering. Applying this CAD pipeline, together with genetic reassembly and optimization of culture conditions led to improved production of the plant flavonoid pinocembrin. Coupling CAD tools with advanced genetic engineering strategies and bioprocess optimization is crucial for enhanced product yields and will be of great value for the development of non-natural products through sustainable biotechnological processes. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Applications of computational modeling in metabolic engineering of yeast.

    Kerkhoven, Eduard J; Lahtvee, Petri-Jaan; Nielsen, Jens

    2015-02-01

    Generally, a microorganism's phenotype can be described by its pattern of metabolic fluxes. Although fluxes cannot be measured directly, inference of fluxes is well established. In biotechnology the aim is often to increase the capacity of specific fluxes. For this, metabolic engineering methods have been developed and applied extensively. Many of these rely on balancing of intracellular metabolites, redox, and energy fluxes, using genome-scale models (GEMs) that in combination with appropriate objective functions and constraints can be used to predict potential gene targets for obtaining a preferred flux distribution. These methods point to strategies for altering gene expression; however, fluxes are often controlled by post-transcriptional events. Moreover, GEMs are usually not taking into account metabolic regulation, thermodynamics and enzyme kinetics. To facilitate metabolic engineering, tools from synthetic biology have emerged, enabling integration and assembly of naturally nonexistent, but well-characterized components into a living organism. To describe these systems kinetic models are often used and to integrate these systems with the standard metabolic engineering approach, it is necessary to expand the modeling of metabolism to consider kinetics of individual processes. This review will give an overview about models available for metabolic engineering of yeast and discusses their applications. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  2. Balancing cellular redox metabolism in microbial electrosynthesis and electro fermentation - A chance for metabolic engineering.

    Kracke, Frauke; Lai, Bin; Yu, Shiqin; Krömer, Jens O

    2018-01-01

    More and more microbes are discovered that are capable of extracellular electron transfer, a process in which they use external electrodes as electron donors or acceptors for metabolic reactions. This feature can be used to overcome cellular redox limitations and thus optimizing microbial production. The technologies, termed microbial electrosynthesis and electro-fermentation, have the potential to open novel bio-electro production platforms from sustainable energy and carbon sources. However, the performance of reported systems is currently limited by low electron transport rates between microbes and electrodes and our limited ability for targeted engineering of these systems due to remaining knowledge gaps about the underlying fundamental processes. Metabolic engineering offers many opportunities to optimize these processes, for instance by genetic engineering of pathways for electron transfer on the one hand and target product synthesis on the other hand. With this review, we summarize the status quo of knowledge and engineering attempts around chemical production in bio-electrochemical systems from a microbe perspective. Challenges associated with the introduction or enhancement of extracellular electron transfer capabilities into production hosts versus the engineering of target compound synthesis pathways in natural exoelectrogens are discussed. Recent advances of the research community in both directions are examined critically. Further, systems biology approaches, for instance using metabolic modelling, are examined for their potential to provide insight into fundamental processes and to identify targets for metabolic engineering. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  3. Advances in Metabolic Engineering of Cyanobacteria for Photosynthetic Biochemical Production

    Lai, Martin C.; Lan, Ethan I.

    2015-01-01

    Engineering cyanobacteria into photosynthetic microbial cell factories for the production of biochemicals and biofuels is a promising approach toward sustainability. Cyanobacteria naturally grow on light and carbon dioxide, bypassing the need of fermentable plant biomass and arable land. By tapping into the central metabolism and rerouting carbon flux towards desirable compound production, cyanobacteria are engineered to directly convert CO2 into various chemicals. This review discusses the d...

  4. Use of genome-scale microbial models for metabolic engineering

    Patil, Kiran Raosaheb; Åkesson, M.; Nielsen, Jens

    2004-01-01

    Metabolic engineering serves as an integrated approach to design new cell factories by providing rational design procedures and valuable mathematical and experimental tools. Mathematical models have an important role for phenotypic analysis, but can also be used for the design of optimal metaboli...... network structures. The major challenge for metabolic engineering in the post-genomic era is to broaden its design methodologies to incorporate genome-scale biological data. Genome-scale stoichiometric models of microorganisms represent a first step in this direction....

  5. Computational methods in metabolic engineering for strain design.

    Long, Matthew R; Ong, Wai Kit; Reed, Jennifer L

    2015-08-01

    Metabolic engineering uses genetic approaches to control microbial metabolism to produce desired compounds. Computational tools can identify new biological routes to chemicals and the changes needed in host metabolism to improve chemical production. Recent computational efforts have focused on exploring what compounds can be made biologically using native, heterologous, and/or enzymes with broad specificity. Additionally, computational methods have been developed to suggest different types of genetic modifications (e.g. gene deletion/addition or up/down regulation), as well as suggest strategies meeting different criteria (e.g. high yield, high productivity, or substrate co-utilization). Strategies to improve the runtime performances have also been developed, which allow for more complex metabolic engineering strategies to be identified. Future incorporation of kinetic considerations will further improve strain design algorithms. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Metabolic engineering of Bacillus subtilis fueled by systems biology: Recent advances and future directions.

    Liu, Yanfeng; Li, Jianghua; Du, Guocheng; Chen, Jian; Liu, Long

    By combining advanced omics technology and computational modeling, systems biologists have identified and inferred thousands of regulatory events and system-wide interactions of the bacterium Bacillus subtilis, which is commonly used both in the laboratory and in industry. This dissection of the multiple layers of regulatory networks and their interactions has provided invaluable information for unraveling regulatory mechanisms and guiding metabolic engineering. In this review, we discuss recent advances in the systems biology and metabolic engineering of B. subtilis and highlight current gaps in our understanding of global metabolism and global pathway engineering in this organism. We also propose future perspectives in the systems biology of B. subtilis and suggest ways that this approach can be used to guide metabolic engineering. Specifically, although hundreds of regulatory events have been identified or inferred via systems biology approaches, systematic investigation of the functionality of these events in vivo has lagged, thereby preventing the elucidation of regulatory mechanisms and further rational pathway engineering. In metabolic engineering, ignoring the engineering of multilayer regulation hinders metabolic flux redistribution. Post-translational engineering, allosteric engineering, and dynamic pathway analyses and control will also contribute to the modulation and control of the metabolism of engineered B. subtilis, ultimately producing the desired cellular traits. We hope this review will aid metabolic engineers in making full use of available systems biology datasets and approaches for the design and perfection of microbial cell factories through global metabolism optimization. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Genetic and metabolic engineering in diatoms.

    Huang, Weichao; Daboussi, Fayza

    2017-09-05

    Diatoms have attracted considerable attention due to their success in diverse environmental conditions, which probably is a consequence of their complex origins. Studies of their metabolism will provide insight into their adaptation capacity and are a prerequisite for metabolic engineering. Several years of investigation have led to the development of the genome engineering tools required for such studies, and a profusion of appropriate tools is now available for exploring and exploiting the metabolism of these organisms. Diatoms are highly prized in industrial biotechnology, due to both their richness in natural lipids and carotenoids and their ability to produce recombinant proteins, of considerable value in diverse markets. This review provides an overview of recent advances in genetic engineering methods for diatoms, from the development of gene expression cassettes and gene delivery methods, to cutting-edge genome-editing technologies. It also highlights the contributions of these rapid developments to both basic and applied research: they have improved our understanding of key physiological processes; and they have made it possible to modify the natural metabolism to favour the production of specific compounds or to produce new compounds for green chemistry and pharmaceutical applications.This article is part of the themed issue 'The peculiar carbon metabolism in diatoms'. © 2017 The Author(s).

  8. Engineering Students' Sustainability Approaches

    Haase, S.

    2014-01-01

    Sustainability issues are increasingly important in engineering work all over the world. This article explores systematic differences in self-assessed competencies, interests, importance, engagement and practices of newly enrolled engineering students in Denmark in relation to environmental and non-environmental sustainability issues. The…

  9. SYSTEMS BIOLOGY AND METABOLIC ENGINEERING OF ARTHROSPIRA CELL FACTORIES

    Amornpan Klanchui

    2012-10-01

    Full Text Available Arthrospira are attractive candidates to serve as cell factories for production of many valuable compounds useful for food, feed, fuel and pharmaceutical industries. In connection with the development of sustainable bioprocessing, it is a challenge to design and develop efficient Arthrospira cell factories which can certify effective conversion from the raw materials (i.e. CO2 and sun light into desired products. With the current availability of the genome sequences and metabolic models of Arthrospira, the development of Arthrospira factories can now be accelerated by means of systems biology and the metabolic engineering approach. Here, we review recent research involving the use of Arthrospira cell factories for industrial applications, as well as the exploitation of systems biology and the metabolic engineering approach for studying Arthrospira. The current status of genomics and proteomics through the development of the genome-scale metabolic model of Arthrospira, as well as the use of mathematical modeling to simulate the phenotypes resulting from the different metabolic engineering strategies are discussed. At the end, the perspective and future direction on Arthrospira cell factories for industrial biotechnology are presented.

  10. Quantum Chemical Approach to Estimating the Thermodynamics of Metabolic Reactions

    Adrian Jinich; Dmitrij Rappoport; Ian Dunn; Benjamin Sanchez-Lengeling; Roberto Olivares-Amaya; Elad Noor; Arren Bar Even; Alán Aspuru-Guzik

    2014-01-01

    Thermodynamics plays an increasingly important role in modeling and engineering metabolism. We present the first nonempirical computational method for estimating standard Gibbs reaction energies of metabolic reactions based on quantum chemistry, which can help fill in the gaps in the existing thermodynamic data. When applied to a test set of reactions from core metabolism, the quantum chemical approach is comparable in accuracy to group contribution methods for isomerization and group transfe...

  11. Metabolic engineering of cyanobacteria for the synthesis of commodity products

    Angermayr, S.A.; Gorchs Rovira, A.; Hellingwerf, K.J.

    2015-01-01

    Through metabolic engineering cyanobacteria can be employed in biotechnology. Combining the capacity for oxygenic photosynthesis and carbon fixation with an engineered metabolic pathway allows carbon-based product formation from CO2, light, and water directly. Such cyanobacterial 'cell factories'

  12. Metabolic Engineering of Probiotic Saccharomyces boulardii.

    Liu, Jing-Jing; Kong, In Iok; Zhang, Guo-Chang; Jayakody, Lahiru N; Kim, Heejin; Xia, Peng-Fei; Kwak, Suryang; Sung, Bong Hyun; Sohn, Jung-Hoon; Walukiewicz, Hanna E; Rao, Christopher V; Jin, Yong-Su

    2016-04-01

    Saccharomyces boulardiiis a probiotic yeast that has been used for promoting gut health as well as preventing diarrheal diseases. This yeast not only exhibits beneficial phenotypes for gut health but also can stay longer in the gut than Saccharomyces cerevisiae Therefore, S. boulardiiis an attractive host for metabolic engineering to produce biomolecules of interest in the gut. However, the lack of auxotrophic strains with defined genetic backgrounds has hampered the use of this strain for metabolic engineering. Here, we report the development of well-defined auxotrophic mutants (leu2,ura3,his3, and trp1) through clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9-based genome editing. The resulting auxotrophic mutants can be used as a host for introducing various genetic perturbations, such as overexpression or deletion of a target gene, using existing genetic tools forS. cerevisiae We demonstrated the overexpression of a heterologous gene (lacZ), the correct localization of a target protein (red fluorescent protein) into mitochondria by using a protein localization signal, and the introduction of a heterologous metabolic pathway (xylose-assimilating pathway) in the genome ofS. boulardii We further demonstrated that human lysozyme, which is beneficial for human gut health, could be secreted by S. boulardii Our results suggest that more sophisticated genetic perturbations to improveS. boulardii can be performed without using a drug resistance marker, which is a prerequisite for in vivo applications using engineeredS. boulardii. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Key applications of plant metabolic engineering.

    Warren Lau

    2014-06-01

    Full Text Available Great strides have been made in plant metabolic engineering over the last two decades, with notable success stories including Golden rice. Here, we discuss the field's progress in addressing four long-standing challenges: creating plants that satisfy their own nitrogen requirement, so reducing or eliminating the need for nitrogen fertilizer; enhancing the nutrient content of crop plants; engineering biofuel feed stocks that harbor easy-to-access fermentable saccharides by incorporating self-destructing lignin; and increasing photosynthetic efficiency. We also look to the future at emerging areas of research in this field.

  14. Metabolic Engineering of Probiotic Saccharomyces boulardii

    Liu, Jing-Jing; Kong, In Iok; Zhang, Guo-Chang; Jayakody, Lahiru N.; Kim, Heejin; Xia, Peng-Fei; Kwak, Suryang; Sung, Bong Hyun; Sohn, Jung-Hoon; Walukiewicz, Hanna E.; Rao, Christopher V.; Jin, Yong-Su

    2016-01-01

    Saccharomyces boulardii is a probiotic yeast that has been used for promoting gut health as well as preventing diarrheal diseases. This yeast not only exhibits beneficial phenotypes for gut health but also can stay longer in the gut than Saccharomyces cerevisiae. Therefore, S. boulardii is an attractive host for metabolic engineering to produce biomolecules of interest in the gut. However, the lack of auxotrophic strains with defined genetic backgrounds has hampered the use of this strain for...

  15. Essences in Metabolic Engineering of Lignan Biosynthesis

    Honoo Satake

    2015-05-01

    Full Text Available Lignans are structurally and functionally diverse phytochemicals biosynthesized in diverse plant species and have received wide attentions as leading compounds of novel drugs for tumor treatment and healthy diets to reduce of the risks of lifestyle-related non-communicable diseases. However, the lineage-specific distribution and the low-amount of production in natural plants, some of which are endangered species, hinder the efficient and stable production of beneficial lignans. Accordingly, the development of new procedures for lignan production is of keen interest. Recent marked advances in the molecular and functional characterization of lignan biosynthetic enzymes and endogenous and exogenous factors for lignan biosynthesis have suggested new methods for the metabolic engineering of lignan biosynthesis cascades leading to the efficient, sustainable, and stable lignan production in plants, including plant cell/organ cultures. Optimization of light conditions, utilization of a wide range of elicitor treatments, and construction of transiently gene-transfected or transgenic lignan-biosynthesizing plants are mainly being attempted. This review will present the basic and latest knowledge regarding metabolic engineering of lignans based on their biosynthetic pathways and biological activities, and the perspectives in lignan production via metabolic engineering.

  16. Transcriptome data modeling for targeted plant metabolic engineering.

    Yonekura-Sakakibara, Keiko; Fukushima, Atsushi; Saito, Kazuki

    2013-04-01

    The massive data generated by omics technologies require the power of bioinformatics, especially network analysis, for data mining and doing data-driven biology. Gene coexpression analysis, a network approach based on comprehensive gene expression data using microarrays, is becoming a standard tool for predicting gene function and elucidating the relationship between metabolic pathways. Differential and comparative gene coexpression analyses suggest a change in coexpression relationships and regulators controlling common and/or specific biological processes. In conjunction with the newly emerging genome editing technology, network analysis integrated with other omics data should pave the way for robust and practical plant metabolic engineering. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. Genetic Optimization Algorithm for Metabolic Engineering Revisited

    Tobias B. Alter

    2018-05-01

    Full Text Available To date, several independent methods and algorithms exist for exploiting constraint-based stoichiometric models to find metabolic engineering strategies that optimize microbial production performance. Optimization procedures based on metaheuristics facilitate a straightforward adaption and expansion of engineering objectives, as well as fitness functions, while being particularly suited for solving problems of high complexity. With the increasing interest in multi-scale models and a need for solving advanced engineering problems, we strive to advance genetic algorithms, which stand out due to their intuitive optimization principles and the proven usefulness in this field of research. A drawback of genetic algorithms is that premature convergence to sub-optimal solutions easily occurs if the optimization parameters are not adapted to the specific problem. Here, we conducted comprehensive parameter sensitivity analyses to study their impact on finding optimal strain designs. We further demonstrate the capability of genetic algorithms to simultaneously handle (i multiple, non-linear engineering objectives; (ii the identification of gene target-sets according to logical gene-protein-reaction associations; (iii minimization of the number of network perturbations; and (iv the insertion of non-native reactions, while employing genome-scale metabolic models. This framework adds a level of sophistication in terms of strain design robustness, which is exemplarily tested on succinate overproduction in Escherichia coli.

  18. Nonlinear Approaches in Engineering Applications

    Jazar, Reza

    2012-01-01

    Nonlinear Approaches in Engineering Applications focuses on nonlinear phenomena that are common in the engineering field. The nonlinear approaches described in this book provide a sound theoretical base and practical tools to design and analyze engineering systems with high efficiency and accuracy and with less energy and downtime. Presented here are nonlinear approaches in areas such as dynamic systems, optimal control and approaches in nonlinear dynamics and acoustics. Coverage encompasses a wide range of applications and fields including mathematical modeling and nonlinear behavior as applied to microresonators, nanotechnologies, nonlinear behavior in soil erosion,nonlinear population dynamics, and optimization in reducing vibration and noise as well as vibration in triple-walled carbon nanotubes. This book also: Provides a complete introduction to nonlinear behavior of systems and the advantages of nonlinearity as a tool for solving engineering problems Includes applications and examples drawn from the el...

  19. Modularization of genetic elements promotes synthetic metabolic engineering.

    Qi, Hao; Li, Bing-Zhi; Zhang, Wen-Qian; Liu, Duo; Yuan, Ying-Jin

    2015-11-15

    In the context of emerging synthetic biology, metabolic engineering is moving to the next stage powered by new technologies. Systematical modularization of genetic elements makes it more convenient to engineer biological systems for chemical production or other desired purposes. In the past few years, progresses were made in engineering metabolic pathway using synthetic biology tools. Here, we spotlighted the topic of implementation of modularized genetic elements in metabolic engineering. First, we overviewed the principle developed for modularizing genetic elements and then discussed how the genetic modules advanced metabolic engineering studies. Next, we picked up some milestones of engineered metabolic pathway achieved in the past few years. Last, we discussed the rapid raised synthetic biology field of "building a genome" and the potential in metabolic engineering. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Engineering microbial fatty acid metabolism for biofuels and biochemicals

    Marella, Eko Roy; Holkenbrink, Carina; Siewers, Verena

    2017-01-01

    microbial catalysis. This review summarizes the recent advances in the engineering of microbial metabolism for production of fatty acid-derived products. We highlight the efforts in engineering the central carbon metabolism, redox metabolism, controlling the chain length of the products, and obtaining...

  1. Microbial production of antioxidant food ingredients via metabolic engineering.

    Lin, Yuheng; Jain, Rachit; Yan, Yajun

    2014-04-01

    Antioxidants are biological molecules with the ability to protect vital metabolites from harmful oxidation. Due to this fascinating role, their beneficial effects on human health are of paramount importance. Traditional approaches using solvent-based extraction from food/non-food sources and chemical synthesis are often expensive, exhaustive, and detrimental to the environment. With the advent of metabolic engineering tools, the successful reconstitution of heterologous pathways in Escherichia coli and other microorganisms provides a more exciting and amenable alternative to meet the increasing demand of natural antioxidants. In this review, we elucidate the recent progress in metabolic engineering efforts for the microbial production of antioxidant food ingredients - polyphenols, carotenoids, and antioxidant vitamins. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Engineering of metabolic pathways by artificial enzyme channels

    Marlene ePröschel

    2015-10-01

    Full Text Available Application of industrial enzymes for production of valuable chemical compounds has greatly benefited from recent developments in Systems and Synthetic Biology. Both, in vivo and in vitro systems have been established, allowing conversion of simple into complex compounds. Metabolic engineering in living cells needs to be balanced which is achieved by controlling gene expression levels, translation, scaffolding, compartmentation and flux control. In vitro applications are often hampered by limited protein stability/half-life and insufficient rates of substrate conversion. To improve stability and catalytic activity, proteins are post-translationally modified and arranged in artificial metabolic channels. Within the review article we will first discuss the supramolecular organization of enzymes in living systems and secondly summarize current and future approaches to design artificial metabolic channels by additive manufacturing for the efficient production of desired products.

  3. Precision metabolic engineering: The design of responsive, selective, and controllable metabolic systems.

    McNerney, Monica P; Watstein, Daniel M; Styczynski, Mark P

    2015-09-01

    Metabolic engineering is generally focused on static optimization of cells to maximize production of a desired product, though recently dynamic metabolic engineering has explored how metabolic programs can be varied over time to improve titer. However, these are not the only types of applications where metabolic engineering could make a significant impact. Here, we discuss a new conceptual framework, termed "precision metabolic engineering," involving the design and engineering of systems that make different products in response to different signals. Rather than focusing on maximizing titer, these types of applications typically have three hallmarks: sensing signals that determine the desired metabolic target, completely directing metabolic flux in response to those signals, and producing sharp responses at specific signal thresholds. In this review, we will first discuss and provide examples of precision metabolic engineering. We will then discuss each of these hallmarks and identify which existing metabolic engineering methods can be applied to accomplish those tasks, as well as some of their shortcomings. Ultimately, precise control of metabolic systems has the potential to enable a host of new metabolic engineering and synthetic biology applications for any problem where flexibility of response to an external signal could be useful. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  4. Applications of computational modeling in metabolic engineering of yeast

    Kerkhoven, Eduard J.; Lahtvee, Petri-Jaan; Nielsen, Jens

    2015-01-01

    a preferred flux distribution. These methods point to strategies for altering gene expression; however, fluxes are often controlled by post-transcriptional events. Moreover, GEMs are usually not taking into account metabolic regulation, thermodynamics and enzyme kinetics. To facilitate metabolic engineering......, it is necessary to expand the modeling of metabolism to consider kinetics of individual processes. This review will give an overview about models available for metabolic engineering of yeast and discusses their applications....

  5. Yeast metabolic engineering--targeting sterol metabolism and terpenoid formation.

    Wriessnegger, Tamara; Pichler, Harald

    2013-07-01

    Terpenoids comprise various structures conferring versatile functions to eukaryotes, for example in the form of prenyl-anchors they attach proteins to membranes. The physiology of eukaryotic membranes is fine-tuned by another terpenoid class, namely sterols. Evidence is accumulating that numerous membrane proteins require specific sterol structural features for function. Moreover, sterols are intermediates in the synthesis of steroids serving as hormones in higher eukaryotes. Like steroids many compounds of the terpenoid family do not contribute to membrane architecture, but serve as signalling, protective or attractant/repellent molecules. Particularly plants have developed a plenitude of terpenoid biosynthetic routes branching off early in the sterol biosynthesis pathway and, thereby, forming one of the largest groups of naturally occurring organic compounds. Many of these aromatic and volatile molecules are interesting for industrial application ranging from foods to pharmaceuticals. Combining the fortunate situation that sterol biosynthesis is highly conserved in eukaryotes with the amenability of yeasts to genetic and metabolic engineering, basically all naturally occurring terpenoids might be produced involving yeasts. Such engineered yeasts are useful for the study of biological functions and molecular interactions of terpenoids as well as for the large-scale production of high-value compounds, which are unavailable in sufficient amounts from natural sources due to their low abundance. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Modulation of sulfur metabolism enables efficient glucosinolate engineering

    Geu-Flores Fernando

    2011-01-01

    Full Text Available Abstract Background Metabolic engineering in heterologous organisms is an attractive approach to achieve efficient production of valuable natural products. Glucosinolates represent a good example of such compounds as they are thought to be the cancer-preventive agents in cruciferous plants. We have recently demonstrated that it is feasible to engineer benzylglucosinolate (BGLS in the non-cruciferous plant Nicotiana benthamiana by transient expression of five genes from Arabidopsis thaliana. In the same study, we showed that co-expression of a sixth Arabidopsis gene, γ-glutamyl peptidase 1 (GGP1, resolved a metabolic bottleneck, thereby increasing BGLS accumulation. However, the accumulation did not reach the expected levels, leaving room for further optimization. Results To optimize heterologous glucosinolate production, we have in this study performed a comparative metabolite analysis of BGLS-producing N. benthamiana leaves in the presence or absence of GGP1. The analysis revealed that the increased BGLS levels in the presence of GGP1 were accompanied by a high accumulation of the last intermediate, desulfoBGLS, and a derivative thereof. This evidenced a bottleneck in the last step of the pathway, the transfer of sulfate from 3'-phosphoadenosine-5'-phosphosulfate (PAPS to desulfoBGLS by the sulfotransferase AtSOT16. While substitution of AtSOT16 with alternative sulfotransferases did not alleviate the bottleneck, experiments with the three genes involved in the formation and recycling of PAPS showed that co-expression of adenosine 5'-phosphosulfate kinase 2 (APK2 alone reduced the accumulation of desulfoBGLS and its derivative by more than 98% and increased BGLS accumulation 16-fold. Conclusion Adjusting sulfur metabolism by directing sulfur from primary to secondary metabolism leads to a remarkable improvement in BGLS accumulation and thereby represents an important step towards a clean and efficient production of glucosinolates in

  7. The future of metabolic engineering and synthetic biology: towards a systematic practice.

    Yadav, Vikramaditya G; De Mey, Marjan; Lim, Chin Giaw; Ajikumar, Parayil Kumaran; Stephanopoulos, Gregory

    2012-05-01

    Industrial biotechnology promises to revolutionize conventional chemical manufacturing in the years ahead, largely owing to the excellent progress in our ability to re-engineer cellular metabolism. However, most successes of metabolic engineering have been confined to over-producing natively synthesized metabolites in E. coli and S. cerevisiae. A major reason for this development has been the descent of metabolic engineering, particularly secondary metabolic engineering, to a collection of demonstrations rather than a systematic practice with generalizable tools. Synthetic biology, a more recent development, faces similar criticisms. Herein, we attempt to lay down a framework around which bioreaction engineering can systematize itself just like chemical reaction engineering. Central to this undertaking is a new approach to engineering secondary metabolism known as 'multivariate modular metabolic engineering' (MMME), whose novelty lies in its assessment and elimination of regulatory and pathway bottlenecks by re-defining the metabolic network as a collection of distinct modules. After introducing the core principles of MMME, we shall then present a number of recent developments in secondary metabolic engineering that could potentially serve as its facilitators. It is hoped that the ever-declining costs of de novo gene synthesis; the improved use of bioinformatic tools to mine, sort and analyze biological data; and the increasing sensitivity and sophistication of investigational tools will make the maturation of microbial metabolic engineering an autocatalytic process. Encouraged by these advances, research groups across the world would take up the challenge of secondary metabolite production in simple hosts with renewed vigor, thereby adding to the range of products synthesized using metabolic engineering. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Metabolic engineering of volatile isoprenoids in plants and microbes.

    Vickers, Claudia E; Bongers, Mareike; Liu, Qing; Delatte, Thierry; Bouwmeester, Harro

    2014-08-01

    The chemical properties and diversity of volatile isoprenoids lends them to a broad variety of biological roles. It also lends them to a host of biotechnological applications, both by taking advantage of their natural functions and by using them as industrial chemicals/chemical feedstocks. Natural functions include roles as insect attractants and repellents, abiotic stress protectants in pathogen defense, etc. Industrial applications include use as pharmaceuticals, flavours, fragrances, fuels, fuel additives, etc. Here we will examine the ways in which researchers have so far found to exploit volatile isoprenoids using biotechnology. Production and/or modification of volatiles using metabolic engineering in both plants and microorganisms are reviewed, including engineering through both mevalonate and methylerythritol diphosphate pathways. Recent advances are illustrated using several case studies (herbivores and bodyguards, isoprene, and monoterpene production in microbes). Systems and synthetic biology tools with particular utility for metabolic engineering are also reviewed. Finally, we discuss the practical realities of various applications in modern biotechnology, explore possible future applications, and examine the challenges of moving these technologies forward so that they can deliver tangible benefits. While this review focuses on volatile isoprenoids, many of the engineering approaches described here are also applicable to non-isoprenoid volatiles and to non-volatile isoprenoids. © 2014 John Wiley & Sons Ltd.

  9. Patchoulol Production with Metabolically Engineered Corynebacterium glutamicum

    Nadja A. Henke

    2018-04-01

    Full Text Available Patchoulol is a sesquiterpene alcohol and an important natural product for the perfume industry. Corynebacterium glutamicum is the prominent host for the fermentative production of amino acids with an average annual production volume of ~6 million tons. Due to its robustness and well established large-scale fermentation, C. glutamicum has been engineered for the production of a number of value-added compounds including terpenoids. Both C40 and C50 carotenoids, including the industrially relevant astaxanthin, and short-chain terpenes such as the sesquiterpene valencene can be produced with this organism. In this study, systematic metabolic engineering enabled construction of a patchoulol producing C. glutamicum strain by applying the following strategies: (i construction of a farnesyl pyrophosphate-producing platform strain by combining genomic deletions with heterologous expression of ispA from Escherichia coli; (ii prevention of carotenoid-like byproduct formation; (iii overproduction of limiting enzymes from the 2-c-methyl-d-erythritol 4-phosphate (MEP-pathway to increase precursor supply; and (iv heterologous expression of the plant patchoulol synthase gene PcPS from Pogostemon cablin. Additionally, a proof of principle liter-scale fermentation with a two-phase organic overlay-culture medium system for terpenoid capture was performed. To the best of our knowledge, the patchoulol titers demonstrated here are the highest reported to date with up to 60 mg L−1 and volumetric productivities of up to 18 mg L−1 d−1.

  10. The Future of Metabolic Engineering and Synthetic Biology: Towards a Systematic Practice

    Yadav, Vikramaditya G.; De Mey, Marjan; Lim, Chin Giaw; Ajikumar, Parayil Kumaran; Stephanopoulos, Gregory

    2012-01-01

    Industrial biotechnology promises to revolutionize conventional chemical manufacturing in the years ahead, largely owing to the excellent progress in our ability to re-engineer cellular metabolism. However, most successes of metabolic engineering have been confined to over-producing natively synthesized metabolites in E. coli and S. cerevisiae. A major reason for this development has been the descent of metabolic engineering, particularly secondary metabolic engineering, to a collection of demonstrations rather than a systematic practice with generalizable tools. Synthetic biology, a more recent development, faces similar criticisms. Herein, we attempt to lay down a framework around which bioreaction engineering can systematize itself just like chemical reaction engineering. Central to this undertaking is a new approach to engineering secondary metabolism known as ‘multivariate modular metabolic engineering’ (MMME), whose novelty lies in its assessment and elimination of regulatory and pathway bottlenecks by re-defining the metabolic network as a collection of distinct modules. After introducing the core principles of MMME, we shall then present a number of recent developments in secondary metabolic engineering that could potentially serve as its facilitators. It is hoped that the ever-declining costs of de novo gene synthesis; the improved use of bioinformatic tools to mine, sort and analyze biological data; and the increasing sensitivity and sophistication of investigational tools will make the maturation of microbial metabolic engineering an autocatalytic process. Encouraged by these advances, research groups across the world would take up the challenge of secondary metabolite production in simple hosts with renewed vigor, thereby adding to the range of products synthesized using metabolic engineering. PMID:22629571

  11. Interdisciplinary Approach in Engineering Education

    Anda Zeidmane

    2011-04-01

    Full Text Available The analysis of the scientific literature available on the types of general competences and their classification caused the authors to conclude that it is necessary to implement interdisciplinary approach in engineering education to develop competences necessary for engineers to make them competitive in the labour market. The attention should be paid to a professional foreign language, computer literacy and educational psychology recommendations. To improve professional foreign language skills, CLIL (Content and Language Integrated Learning method should be integrated in the study process of engineering education. In order to develop information literacy competence, it is important to create a single e-study environment. The academic staff, developing study subjects for engineering programmes, should focus on the study content and study methods. As regards the content, the compromise should be sought between fundamental acquisition of the knowledge of the subject matter, the know-how of the application of this knowledge as well as the use of brand new software in the calculations. The paper presents the examples of the application of the interdisciplinary approach in the universities, where the authors of the paper are affiliated: the LUA (Latvia University of Agriculture and the RTU (Riga Technical University, respectively.

  12. Engineering yeast metabolism for production of fuels and chemicals

    Nielsen, Jens

    2016-01-01

    faster development of metabolically engineered strains that can be used for production of fuels and chemicals. The yeast Saccharomyces cerevisiae is widely used for production of fuels, chemicals, pharmaceuticals and materials. Through metabolic engineering of this yeast a number of novel industrial...... as for metabolic design. In this lecture it will be demonstrated how the Design-Build-Test cycle of metabolic engineering has allowed for development of yeast cell factories for production of a range of different fuels and chemicals. Some examples of different technologies will be presented together with examples......Metabolic engineering relies on the Design-Build-Test cycle. This cycle includes technologies like mathematical modeling of metabolism, genome editing and advanced tools for phenotypic characterization. In recent years there have been advances in several of these technologies, which has enabled...

  13. Systems biology and metabolic engineering of lactic acid bacteria for improved fermented foods

    Flahaut, N.A.L.; Vos, de W.M.

    2014-01-01

    Lactic acid bacteria have long been used in industrial dairy and other food fermentations that make use of their metabolic activities leading to products with specific organoleptic properties. Metabolic engineering is a rational approach to steer fermentations toward the production of desired

  14. Quantum chemical approach to estimating the thermodynamics of metabolic reactions.

    Jinich, Adrian; Rappoport, Dmitrij; Dunn, Ian; Sanchez-Lengeling, Benjamin; Olivares-Amaya, Roberto; Noor, Elad; Even, Arren Bar; Aspuru-Guzik, Alán

    2014-11-12

    Thermodynamics plays an increasingly important role in modeling and engineering metabolism. We present the first nonempirical computational method for estimating standard Gibbs reaction energies of metabolic reactions based on quantum chemistry, which can help fill in the gaps in the existing thermodynamic data. When applied to a test set of reactions from core metabolism, the quantum chemical approach is comparable in accuracy to group contribution methods for isomerization and group transfer reactions and for reactions not including multiply charged anions. The errors in standard Gibbs reaction energy estimates are correlated with the charges of the participating molecules. The quantum chemical approach is amenable to systematic improvements and holds potential for providing thermodynamic data for all of metabolism.

  15. Next-generation genome-scale models for metabolic engineering

    King, Zachary A.; Lloyd, Colton J.; Feist, Adam M.

    2015-01-01

    Constraint-based reconstruction and analysis (COBRA) methods have become widely used tools for metabolic engineering in both academic and industrial laboratories. By employing a genome-scale in silico representation of the metabolic network of a host organism, COBRA methods can be used to predict...... examples of applying COBRA methods to strain optimization are presented and discussed. Then, an outlook is provided on the next generation of COBRA models and the new types of predictions they will enable for systems metabolic engineering....

  16. Metabolic engineering with plants for a sustainable biobased economy.

    Yoon, Jong Moon; Zhao, Le; Shanks, Jacqueline V

    2013-01-01

    Plants are bona fide sustainable organisms because they accumulate carbon and synthesize beneficial metabolites from photosynthesis. To meet the challenges to food security and health threatened by increasing population growth and depletion of nonrenewable natural resources, recent metabolic engineering efforts have shifted from single pathways to holistic approaches with multiple genes owing to integration of omics technologies. Successful engineering of plants results in the high yield of biomass components for primary food sources and biofuel feedstocks, pharmaceuticals, and platform chemicals through synthetic biology and systems biology strategies. Further discovery of undefined biosynthesis pathways in plants, integrative analysis of discrete omics data, and diversified process developments for production of platform chemicals are essential to overcome the hurdles for sustainable production of value-added biomolecules from plants.

  17. Deriving metabolic engineering strategies from genome-scale modeling with flux ratio constraints.

    Yen, Jiun Y; Nazem-Bokaee, Hadi; Freedman, Benjamin G; Athamneh, Ahmad I M; Senger, Ryan S

    2013-05-01

    Optimized production of bio-based fuels and chemicals from microbial cell factories is a central goal of systems metabolic engineering. To achieve this goal, a new computational method of using flux balance analysis with flux ratios (FBrAtio) was further developed in this research and applied to five case studies to evaluate and design metabolic engineering strategies. The approach was implemented using publicly available genome-scale metabolic flux models. Synthetic pathways were added to these models along with flux ratio constraints by FBrAtio to achieve increased (i) cellulose production from Arabidopsis thaliana; (ii) isobutanol production from Saccharomyces cerevisiae; (iii) acetone production from Synechocystis sp. PCC6803; (iv) H2 production from Escherichia coli MG1655; and (v) isopropanol, butanol, and ethanol (IBE) production from engineered Clostridium acetobutylicum. The FBrAtio approach was applied to each case to simulate a metabolic engineering strategy already implemented experimentally, and flux ratios were continually adjusted to find (i) the end-limit of increased production using the existing strategy, (ii) new potential strategies to increase production, and (iii) the impact of these metabolic engineering strategies on product yield and culture growth. The FBrAtio approach has the potential to design "fine-tuned" metabolic engineering strategies in silico that can be implemented directly with available genomic tools. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Metabolic engineering with systems biology tools to optimize production of prokaryotic secondary metabolites

    Kim, Hyun Uk; Charusanti, Pep; Lee, Sang Yup

    2016-01-01

    Metabolic engineering using systems biology tools is increasingly applied to overproduce secondary metabolites for their potential industrial production. In this Highlight, recent relevant metabolic engineering studies are analyzed with emphasis on host selection and engineering approaches...... for the optimal production of various prokaryotic secondary metabolites: native versus heterologous hosts (e.g., Escherichia coli) and rational versus random approaches. This comparative analysis is followed by discussions on systems biology tools deployed in optimizing the production of secondary metabolites....... The potential contributions of additional systems biology tools are also discussed in the context of current challenges encountered during optimization of secondary metabolite production....

  19. Metabolic Engineering for Probiotics and their Genome-Wide Expression Profiling.

    Yadav, Ruby; Singh, Puneet K; Shukla, Pratyoosh

    2018-01-01

    Probiotic supplements in food industry have attracted a lot of attention and shown a remarkable growth in this field. Metabolic engineering (ME) approaches enable understanding their mechanism of action and increases possibility of designing probiotic strains with desired functions. Probiotic microorganisms generally referred as industrially important lactic acid bacteria (LAB) which are involved in fermenting dairy products, food, beverages and produces lactic acid as final product. A number of illustrations of metabolic engineering approaches in industrial probiotic bacteria have been described in this review including transcriptomic studies of Lactobacillus reuteri and improvement in exopolysaccharide (EPS) biosynthesis yield in Lactobacillus casei LC2W. This review summaries various metabolic engineering approaches for exploring metabolic pathways. These approaches enable evaluation of cellular metabolic state and effective editing of microbial genome or introduction of novel enzymes to redirect the carbon fluxes. In addition, various system biology tools such as in silico design commonly used for improving strain performance is also discussed. Finally, we discuss the integration of metabolic engineering and genome profiling which offers a new way to explore metabolic interactions, fluxomics and probiogenomics using probiotic bacteria like Bifidobacterium spp and Lactobacillus spp. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  20. Recent advances in systems metabolic engineering tools and strategies.

    Chae, Tong Un; Choi, So Young; Kim, Je Woong; Ko, Yoo-Sung; Lee, Sang Yup

    2017-10-01

    Metabolic engineering has been playing increasingly important roles in developing microbial cell factories for the production of various chemicals and materials to achieve sustainable chemical industry. Nowadays, many tools and strategies are available for performing systems metabolic engineering that allows systems-level metabolic engineering in more sophisticated and diverse ways by adopting rapidly advancing methodologies and tools of systems biology, synthetic biology and evolutionary engineering. As an outcome, development of more efficient microbial cell factories has become possible. Here, we review recent advances in systems metabolic engineering tools and strategies together with accompanying application examples. In addition, we describe how these tools and strategies work together in simultaneous and synergistic ways to develop novel microbial cell factories. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. [Improving industrial microbial stress resistance by metabolic engineering: a review].

    Fu, Ruiyan; Li, Yin

    2010-09-01

    Metabolic engineering is a technologic platform for industrial strain improvement and aims not only at modifying microbial metabolic fluxes, but also improving the physiological performance of industrial microbes. Microbes will meet multiple stresses in industrial processes. Consequently, elicited gene responses might result in a decrease in overall cell fitness and the efficiency of biotransformation. Thus, it is crucial to develop robust and productive microbial strains that can be integrated into industrial-scale bioprocesses. In this review, we focus on the progress of these novel methods and strategies for engineering stress-tolerance phenotypes referring to rational metabolic engineering and inverse metabolic engineering in recent years. In addition, we also address problems existing in this area and future research needs of microbial physiological functionality engineering.

  2. 2007 Plant Metabolic Engineering Gordon Conference and Graduate Research Seminar

    Erich Grotewold

    2008-09-15

    Plant Metabolic Engineering is an emerging field that integrates a diverse range of disciplines including plant genetics, genomics, biochemistry, chemistry and cell biology. The Gordon-Kenan Graduate Research Seminar (GRS) in Plant Metabolic Engineering was initiated to provide a unique opportunity for future researcher leaders to present their work in this field. It also creates an environment allowing for peer-review and critical assessment of work without the intimidation usually associated with the presence of senior investigators. The GRS immediately precedes the Plant Metabolic Engineering Gordon Research Conference and will be for and by graduate students and post-docs, with the assistance of the organizers listed.

  3. From pathways to genomes and beyond. The metabolic engineering toolbox and its place in biofuels production

    Liu, Leqian; Reed, Ben; Alper, Hal [Texas Univ., Austin, TX (United States). Dept. of Chemical Engineering

    2011-07-01

    Concerns about the availability of petroleum-derived fuels and chemicals have led to the exploration of metabolically engineered organisms as novel hosts for biofuels and chemicals production. However, the complexity inherent in metabolic and regulatory networks makes this undertaking a complex task. To address these limitations, metabolic engineering has adapted a wide-variety of tools for altering phenotypes. In this review, we will highlight traditional and recent metabolic engineering tools for optimizing cells including pathway-based, global, and genomic-enabled approaches. Specifically, we describe these tools as well as provide demonstrations of their effectiveness in optimizing biofuels production. However, each of these tools provides stepping stones towards the grand goal of biofuels production. Thus, developing methods for large-scale cellular optimization and integrative approaches are invaluable for further cell optimization. This review highlights the challenges that still must be met to accomplish this goal. (orig.)

  4. The Genome-Based Metabolic Systems Engineering to Boost Levan Production in a Halophilic Bacterial Model.

    Aydin, Busra; Ozer, Tugba; Oner, Ebru Toksoy; Arga, Kazim Yalcin

    2018-03-01

    Metabolic systems engineering is being used to redirect microbial metabolism for the overproduction of chemicals of interest with the aim of transforming microbial hosts into cellular factories. In this study, a genome-based metabolic systems engineering approach was designed and performed to improve biopolymer biosynthesis capability of a moderately halophilic bacterium Halomonas smyrnensis AAD6 T producing levan, which is a fructose homopolymer with many potential uses in various industries and medicine. For this purpose, the genome-scale metabolic model for AAD6 T was used to characterize the metabolic resource allocation, specifically to design metabolic engineering strategies for engineered bacteria with enhanced levan production capability. Simulations were performed in silico to determine optimal gene knockout strategies to develop new strains with enhanced levan production capability. The majority of the gene knockout strategies emphasized the vital role of the fructose uptake mechanism, and pointed out the fructose-specific phosphotransferase system (PTS fru ) as the most promising target for further metabolic engineering studies. Therefore, the PTS fru of AAD6 T was restructured with insertional mutagenesis and triparental mating techniques to construct a novel, engineered H. smyrnensis strain, BMA14. Fermentation experiments were carried out to demonstrate the high efficiency of the mutant strain BMA14 in terms of final levan concentration, sucrose consumption rate, and sucrose conversion efficiency, when compared to the AAD6 T . The genome-based metabolic systems engineering approach presented in this study might be considered an efficient framework to redirect microbial metabolism for the overproduction of chemicals of interest, and the novel strain BMA14 might be considered a potential microbial cell factory for further studies aimed to design levan production processes with lower production costs.

  5. Resveratrol biosynthesis: plant metabolic engineering for nutritional improvement of food.

    Giovinazzo, Giovanna; Ingrosso, Ilaria; Paradiso, Annalisa; De Gara, Laura; Santino, Angelo

    2012-09-01

    The plant polyphenol trans-resveratrol (3, 5, 4'-trihydroxystilbene) mainly found in grape, peanut and other few plants, displays a wide range of biological effects. Numerous in vitro studies have described various biological effects of resveratrol. In order to provide more information regarding absorption, metabolism, and bioavailability of resveratrol, various research approaches have been performed, including in vitro, ex vivo, and in vivo models. In recent years, the induction of resveratrol synthesis in plants which normally do not accumulate such polyphenol, has been successfully achieved by molecular engineering. In this context, the ectopic production of resveratrol has been reported to have positive effects both on plant resistance to biotic stress and the enhancement of the nutritional value of several widely consumed fruits and vegetables. The metabolic engineering of plants offers the opportunity to change the content of specific phytonutrients in plant - derived foods. This review focuses on the latest findings regarding on resveratrol bioproduction and its effects on the prevention of the major pathological conditions in man.

  6. Engineering strategy of yeast metabolism for higher alcohol production

    Shimizu Hiroshi

    2011-09-01

    Full Text Available Abstract Background While Saccharomyces cerevisiae is a promising host for cost-effective biorefinary processes due to its tolerance to various stresses during fermentation, the metabolically engineered S. cerevisiae strains exhibited rather limited production of higher alcohols than that of Escherichia coli. Since the structure of the central metabolism of S. cerevisiae is distinct from that of E. coli, there might be a problem in the structure of the central metabolism of S. cerevisiae. In this study, the potential production of higher alcohols by S. cerevisiae is compared to that of E. coli by employing metabolic simulation techniques. Based on the simulation results, novel metabolic engineering strategies for improving higher alcohol production by S. cerevisiae were investigated by in silico modifications of the metabolic models of S. cerevisiae. Results The metabolic simulations confirmed that the high production of butanols and propanols by the metabolically engineered E. coli strains is derived from the flexible behavior of their central metabolism. Reducing this flexibility by gene deletion is an effective strategy to restrict the metabolic states for producing target alcohols. In contrast, the lower yield using S. cerevisiae originates from the structurally limited flexibility of its central metabolism in which gene deletions severely reduced cell growth. Conclusions The metabolic simulation demonstrated that the poor productivity of S. cerevisiae was improved by the introduction of E. coli genes to compensate the structural difference. This suggested that gene supplementation is a promising strategy for the metabolic engineering of S. cerevisiae to produce higher alcohols which should be the next challenge for the synthetic bioengineering of S. cerevisiae for the efficient production of higher alcohols.

  7. Production of L-valine from metabolically engineered Corynebacterium glutamicum.

    Wang, Xiaoyuan; Zhang, Hailing; Quinn, Peter J

    2018-05-01

    L-Valine is one of the three branched-chain amino acids (valine, leucine, and isoleucine) essential for animal health and important in metabolism; therefore, it is widely added in the products of food, medicine, and feed. L-Valine is predominantly produced through microbial fermentation, and the production efficiency largely depends on the quality of microorganisms. In recent years, continuing efforts have been made in revealing the mechanisms and regulation of L-valine biosynthesis in Corynebacterium glutamicum, the most utilitarian bacterium for amino acid production. Metabolic engineering based on the metabolic biosynthesis and regulation of L-valine provides an effective alternative to the traditional breeding for strain development. Industrially competitive L-valine-producing C. glutamicum strains have been constructed by genetically defined metabolic engineering. This article reviews the global metabolic and regulatory networks responsible for L-valine biosynthesis, the molecular mechanisms of regulation, and the strategies employed in C. glutamicum strain engineering.

  8. Protein engineering for metabolic engineering: current and next-generation tools

    Marcheschi, Ryan J.; Gronenberg, Luisa S.; Liao, James C.

    2014-01-01

    Protein engineering in the context of metabolic engineering is increasingly important to the field of industrial biotechnology. As the demand for biologically-produced food, fuels, chemicals, food additives, and pharmaceuticals continues to grow, the ability to design and modify proteins to accomplish new functions will be required to meet the high productivity demands for the metabolism of engineered organisms. This article reviews advances of selecting, modeling, and engineering proteins to improve or alter their activity. Some of the methods have only recently been developed for general use and are just beginning to find greater application in the metabolic engineering community. We also discuss methods of generating random and targeted diversity in proteins to generate mutant libraries for analysis. Recent uses of these techniques to alter cofactor use, produce non-natural amino acids, alcohols, and carboxylic acids, and alter organism phenotypes are presented and discussed as examples of the successful engineering of proteins for metabolic engineering purposes. PMID:23589443

  9. SBOLme: a Repository of SBOL Parts for Metabolic Engineering

    Kuwahara, Hiroyuki; Cui, Xuefeng; Umarov, Ramzan; Grunberg, Raik; Myers, Chris J.; Gao, Xin

    2017-01-01

    The Synthetic Biology Open Language (SBOL) is a community-driven open language to promote standardization in synthetic biology. To support the use of SBOL in metabolic engineering, we developed SBOLme, the first open-access repository of SBOL 2

  10. Advancing metabolic engineering through systems biology of industrial microorganisms.

    Dai, Zongjie; Nielsen, Jens

    2015-12-01

    Development of sustainable processes to produce bio-based compounds is necessary due to the severe environmental problems caused by the use of fossil resources. Metabolic engineering can facilitate the development of highly efficient cell factories to produce these compounds from renewable resources. The objective of systems biology is to gain a comprehensive and quantitative understanding of living cells and can hereby enhance our ability to characterize and predict cellular behavior. Systems biology of industrial microorganisms is therefore valuable for metabolic engineering. Here we review the application of systems biology tools for the identification of metabolic engineering targets which may lead to reduced development time for efficient cell factories. Finally, we present some perspectives of systems biology for advancing metabolic engineering further. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Advancing metabolic engineering through systems biology of industrial microorganisms

    Dai, Zongjie; Nielsen, Jens

    2015-01-01

    resources. The objective of systems biology is to gain a comprehensive and quantitative understanding of living cells and can hereby enhance our ability to characterize and predict cellular behavior. Systems biology of industrial microorganisms is therefore valuable for metabolic engineering. Here we review......Development of sustainable processes to produce bio-based compounds is necessary due to the severe environmental problems caused by the use of fossil resources. Metabolic engineering can facilitate the development of highly efficient cell factories to produce these compounds from renewable...... the application of systems biology tools for the identification of metabolic engineering targets which may lead to reduced development time for efficient cell factories. Finally, we present some perspectives of systems biology for advancing metabolic engineering further....

  12. SBOLme: a Repository of SBOL Parts for Metabolic Engineering

    Kuwahara, Hiroyuki

    2017-01-12

    The Synthetic Biology Open Language (SBOL) is a community-driven open language to promote standardization in synthetic biology. To support the use of SBOL in metabolic engineering, we developed SBOLme, the first open-access repository of SBOL 2-compliant biochemical parts for a wide range of metabolic engineering applications. The URL of our repository is http://www.cbrc.kaust.edu.sa/sbolme.

  13. Evolutionary programming as a platform for in silico metabolic engineering

    Patil, Kiran Raosaheb; Rocha, Isabel; Förster, Jochen

    2005-01-01

    , and it is therefore interesting to develop new faster algorithms. Results In this study we report an evolutionary programming based method to rapidly identify gene deletion strategies for optimization of a desired phenotypic objective function. We illustrate the proposed method for two important design parameters...... of close to optimal solutions. The identified metabolic engineering strategies suggest that non-intuitive genetic modifications span several different pathways and may be necessary for solving challenging metabolic engineering problems....

  14. Impact of systems biology on metabolic engineering of Saccharomyces cerevisiae

    Nielsen, Jens; Jewett, Michael Christopher

    2008-01-01

    in the industrial application of this yeast. Developments in genomics and high-throughput systems biology tools are enhancing one's ability to rapidly characterize cellular behaviour, which is valuable in the field of metabolic engineering where strain characterization is often the bottleneck in strain development...... programmes. Here, the impact of systems biology on metabolic engineering is reviewed and perspectives on the role of systems biology in the design of cell factories are given....

  15. Mini-review: In vitro Metabolic Engineering for Biomanufacturing of High-value Products

    Weihua Guo

    Full Text Available With the breakthroughs in biomolecular engineering and synthetic biology, many valuable biologically active compound and commodity chemicals have been successfully manufactured using cell-based approaches in the past decade. However, because of the high complexity of cell metabolism, the identification and optimization of rate-limiting metabolic pathways for improving the product yield is often difficult, which represents a significant and unavoidable barrier of traditional in vivo metabolic engineering. Recently, some in vitro engineering approaches were proposed as alternative strategies to solve this problem. In brief, by reconstituting a biosynthetic pathway in a cell-free environment with the supplement of cofactors and substrates, the performance of each biosynthetic pathway could be evaluated and optimized systematically. Several value-added products, including chemicals, nutraceuticals, and drug precursors, have been biosynthesized as proof-of-concept demonstrations of in vitro metabolic engineering. This mini-review summarizes the recent progresses on the emerging topic of in vitro metabolic engineering and comments on the potential application of cell-free technology to speed up the “design-build-test” cycles of biomanufacturing. Keywords: Cell-free, Biosynthesis, Metabolic pathways, Design-build-test cycle

  16. Protein engineering approaches to chemical biotechnology.

    Chen, Zhen; Zeng, An-Ping

    2016-12-01

    Protein engineering for the improvement of properties of biocatalysts and for the generation of novel metabolic pathways plays more and more important roles in chemical biotechnology aiming at the production of chemicals from biomass. Although widely used in single-enzyme catalysis process, protein engineering is only being increasingly explored in recent years to achieve more complex in vitro and in vivo biocatalytic processes. This review focuses on major contributions of protein engineering to chemical biotechnology in the field of multi-enzymatic cascade catalysis and metabolic engineering. Especially, we discuss and highlight recent strategies for combining pathway design and protein engineering for the production of novel products. Copyright © 2016. Published by Elsevier Ltd.

  17. Metabolite damage and repair in metabolic engineering design.

    Sun, Jiayi; Jeffryes, James G; Henry, Christopher S; Bruner, Steven D; Hanson, Andrew D

    2017-11-01

    The necessarily sharp focus of metabolic engineering and metabolic synthetic biology on pathways and their fluxes has tended to divert attention from the damaging enzymatic and chemical side-reactions that pathway metabolites can undergo. Although historically overlooked and underappreciated, such metabolite damage reactions are now known to occur throughout metabolism and to generate (formerly enigmatic) peaks detected in metabolomics datasets. It is also now known that metabolite damage is often countered by dedicated repair enzymes that undo or prevent it. Metabolite damage and repair are highly relevant to engineered pathway design: metabolite damage reactions can reduce flux rates and product yields, and repair enzymes can provide robust, host-independent solutions. Herein, after introducing the core principles of metabolite damage and repair, we use case histories to document how damage and repair processes affect efficient operation of engineered pathways - particularly those that are heterologous, non-natural, or cell-free. We then review how metabolite damage reactions can be predicted, how repair reactions can be prospected, and how metabolite damage and repair can be built into genome-scale metabolic models. Lastly, we propose a versatile 'plug and play' set of well-characterized metabolite repair enzymes to solve metabolite damage problems known or likely to occur in metabolic engineering and synthetic biology projects. Copyright © 2017 International Metabolic Engineering Society. All rights reserved.

  18. Genome-scale modeling using flux ratio constraints to enable metabolic engineering of clostridial metabolism in silico.

    McAnulty, Michael J; Yen, Jiun Y; Freedman, Benjamin G; Senger, Ryan S

    2012-05-14

    Genome-scale metabolic networks and flux models are an effective platform for linking an organism genotype to its phenotype. However, few modeling approaches offer predictive capabilities to evaluate potential metabolic engineering strategies in silico. A new method called "flux balance analysis with flux ratios (FBrAtio)" was developed in this research and applied to a new genome-scale model of Clostridium acetobutylicum ATCC 824 (iCAC490) that contains 707 metabolites and 794 reactions. FBrAtio was used to model wild-type metabolism and metabolically engineered strains of C. acetobutylicum where only flux ratio constraints and thermodynamic reversibility of reactions were required. The FBrAtio approach allowed solutions to be found through standard linear programming. Five flux ratio constraints were required to achieve a qualitative picture of wild-type metabolism for C. acetobutylicum for the production of: (i) acetate, (ii) lactate, (iii) butyrate, (iv) acetone, (v) butanol, (vi) ethanol, (vii) CO2 and (viii) H2. Results of this simulation study coincide with published experimental results and show the knockdown of the acetoacetyl-CoA transferase increases butanol to acetone selectivity, while the simultaneous over-expression of the aldehyde/alcohol dehydrogenase greatly increases ethanol production. FBrAtio is a promising new method for constraining genome-scale models using internal flux ratios. The method was effective for modeling wild-type and engineered strains of C. acetobutylicum.

  19. Metabolic engineering of Yarrowia lipolytica for industrial applications.

    Zhu, Quinn; Jackson, Ethel N

    2015-12-01

    Yarrowia lipolytica is a safe and robust yeast that has a history of industrial applications. Its physiological, metabolic and genomic characteristics have made it a superior host for metabolic engineering. The results of optimizing internal pathways and introducing new pathways have demonstrated that Y. lipolytica can be a platform cell factory for cost-effective production of chemicals and fuels derived from fatty acids, lipids and acetyl-CoA. Two products have been commercialized from metabolically engineered Y. lipolytica strains producing high amounts of omega-3 eicosapentaenoic acid, and more products are on the way to be produced at industrial scale. Here we review recent progress in metabolic engineering of Y. lipolytica for production of biodiesel fuel, functional fatty acids and carotenoids. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  20. Nonlinear approaches in engineering applications 2

    Jazar, Reza N

    2013-01-01

    Provides updated principles and applications of the nonlinear approaches in solving engineering and physics problems Demonstrates how nonlinear approaches may open avenues to better, safer, cheaper systems with less energy consumption Has a strong emphasis on the application, physical meaning, and methodologies of nonlinear approaches in different engineering and science problems

  1. Rethinking engineering education the CDIO approach

    Crawley, Edward F; Östlund, Sören; Brodeur, Doris R; Edström, Kristina

    2014-01-01

    This book describes an approach to engineering education that integrates a comprehensive set of personal, interpersonal, and professional engineering skills with engineering disciplinary knowledge in order to prepare innovative and entrepreneurial engineers.  The education of engineers is set in the context of engineering practice, that is, Conceiving, Designing, Implementing, and Operating (CDIO) through the entire lifecycle of engineering processes, products, and systems. The book is both a description of the development and implementation of the CDIO model and a guide to engineering programs worldwide that seek to improve the education of young engineers.   Provides an overview of the CDIO approach, then chapters organized according to the CDIO Standards; Includes in each chapter objectives, discussion questions, case studies and clear diagrams to support key concepts and processes; Avoids the jargon of education specialists and clearly explains education terms in the context of their initial presentatio...

  2. Systems metabolic engineering design: fatty acid production as an emerging case study.

    Tee, Ting Wei; Chowdhury, Anupam; Maranas, Costas D; Shanks, Jacqueline V

    2014-05-01

    Increasing demand for petroleum has stimulated industry to develop sustainable production of chemicals and biofuels using microbial cell factories. Fatty acids of chain lengths from C6 to C16 are propitious intermediates for the catalytic synthesis of industrial chemicals and diesel-like biofuels. The abundance of genetic information available for Escherichia coli and specifically, fatty acid metabolism in E. coli, supports this bacterium as a promising host for engineering a biocatalyst for the microbial production of fatty acids. Recent successes rooted in different features of systems metabolic engineering in the strain design of high-yielding medium chain fatty acid producing E. coli strains provide an emerging case study of design methods for effective strain design. Classical metabolic engineering and synthetic biology approaches enabled different and distinct design paths towards a high-yielding strain. Here we highlight a rational strain design process in systems biology, an integrated computational and experimental approach for carboxylic acid production, as an alternative method. Additional challenges inherent in achieving an optimal strain for commercialization of medium chain-length fatty acids will likely require a collection of strategies from systems metabolic engineering. Not only will the continued advancement in systems metabolic engineering result in these highly productive strains more quickly, this knowledge will extend more rapidly the carboxylic acid platform to the microbial production of carboxylic acids with alternate chain-lengths and functionalities. © 2014 Wiley Periodicals, Inc.

  3. The necessity of a theory of biology for tissue engineering: metabolism-repair systems.

    Ganguli, Suman; Hunt, C Anthony

    2004-01-01

    Since there is no widely accepted global theory of biology, tissue engineering and bioengineering lack a theoretical understanding of the systems being engineered. By default, tissue engineering operates with a "reductionist" theoretical approach, inherited from traditional engineering of non-living materials. Long term, that approach is inadequate, since it ignores essential aspects of biology. Metabolism-repair systems are a theoretical framework which explicitly represents two "functional" aspects of living organisms: self-repair and self-replication. Since repair and replication are central to tissue engineering, we advance metabolism-repair systems as a potential theoretical framework for tissue engineering. We present an overview of the framework, and indicate directions to pursue for extending it to the context of tissue engineering. We focus on biological networks, both metabolic and cellular, as one such direction. The construction of these networks, in turn, depends on biological protocols. Together these concepts may help point the way to a global theory of biology appropriate for tissue engineering.

  4. Advances and prospects in metabolic engineering of Zymomonas mobilis.

    Wang, Xia; He, Qiaoning; Yang, Yongfu; Wang, Jingwen; Haning, Katie; Hu, Yun; Wu, Bo; He, Mingxiong; Zhang, Yaoping; Bao, Jie; Contreras, Lydia M; Yang, Shihui

    2018-04-05

    Biorefinery of biomass-based biofuels and biochemicals by microorganisms is a competitive alternative of traditional petroleum refineries. Zymomonas mobilis is a natural ethanologen with many desirable characteristics, which makes it an ideal industrial microbial biocatalyst for commercial production of desirable bioproducts through metabolic engineering. In this review, we summarize the metabolic engineering progress achieved in Z. mobilis to expand its substrate and product ranges as well as to enhance its robustness against stressful conditions such as inhibitory compounds within the lignocellulosic hydrolysates and slurries. We also discuss a few metabolic engineering strategies that can be applied in Z. mobilis to further develop it as a robust workhorse for economic lignocellulosic bioproducts. In addition, we briefly review the progress of metabolic engineering in Z. mobilis related to the classical synthetic biology cycle of "Design-Build-Test-Learn", as well as the progress and potential to develop Z. mobilis as a model chassis for biorefinery practices in the synthetic biology era. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  5. Software engineering a practitioner's approach

    Pressman, Roger S

    1997-01-01

    This indispensable guide to software engineering exploration enables practitioners to navigate the ins and outs of this rapidly changing field. Pressman's fully revised and updated Fourth Edition provides in-depth coverage of every important management and technical topic in software engineering. Moreover, readers will find the inclusion of the hottest developments in the field such as: formal methods and cleanroom software engineering, business process reengineering, and software reengineering.

  6. Yeast metabolic engineering for hemicellulosic ethanol production

    Jennifer Van Vleet; Thomas W. Jeffries

    2009-01-01

    Efficient fermentation of hemicellulosic sugars is critical for the bioconversion of lignocellulosics to ethanol. Efficient sugar uptake through the heterologous expression of yeast and fungal xylose/glucose transporters can improve fermentation if other metabolic steps are not rate limiting. Rectification of cofactor imbalances through heterologous expression of...

  7. Quantifying complexity in metabolic engineering using the LASER database

    James D. Winkler

    2016-12-01

    Full Text Available We previously introduced the LASER database (Learning Assisted Strain EngineeRing, https://bitbucket.org/jdwinkler/laser_release (Winkler et al. 2015 to serve as a platform for understanding past and present metabolic engineering practices. Over the past year, LASER has been expanded by 50% to include over 600 engineered strains from 450 papers, including their growth conditions, genetic modifications, and other information in an easily searchable format. Here, we present the results of our efforts to use LASER as a means for defining the complexity of a metabolic engineering “design”. We evaluate two complexity metrics based on the concepts of construction difficulty and novelty. No correlation is observed between expected product yield and complexity, allowing minimization of complexity without a performance trade-off. We envision the use of such complexity metrics to filter and prioritize designs prior to implementation of metabolic engineering efforts, thereby potentially reducing the time, labor, and expenses of large-scale projects. Possible future developments based on an expanding LASER database are then discussed. Keywords: Metabolic engineering, Synthetic biology, Standardization, Design tools

  8. Alternative Approach to Power Engineering

    Pedersen, Knud Ole Helgesen; Havemann, Henrik

    2000-01-01

    as young and dynamic. Consequently, the new courses apply IT as a gateway to power engineering. The courses present the students with: (1) a meaningful, easy understandable power engineering problem; (2) a realistic set-up in the laboratory; and (3) a microprocessor system used as a tool to solve...

  9. Evolutionary programming as a platform for in silico metabolic engineering

    Förster Jochen

    2005-12-01

    Full Text Available Abstract Background Through genetic engineering it is possible to introduce targeted genetic changes and hereby engineer the metabolism of microbial cells with the objective to obtain desirable phenotypes. However, owing to the complexity of metabolic networks, both in terms of structure and regulation, it is often difficult to predict the effects of genetic modifications on the resulting phenotype. Recently genome-scale metabolic models have been compiled for several different microorganisms where structural and stoichiometric complexity is inherently accounted for. New algorithms are being developed by using genome-scale metabolic models that enable identification of gene knockout strategies for obtaining improved phenotypes. However, the problem of finding optimal gene deletion strategy is combinatorial and consequently the computational time increases exponentially with the size of the problem, and it is therefore interesting to develop new faster algorithms. Results In this study we report an evolutionary programming based method to rapidly identify gene deletion strategies for optimization of a desired phenotypic objective function. We illustrate the proposed method for two important design parameters in industrial fermentations, one linear and other non-linear, by using a genome-scale model of the yeast Saccharomyces cerevisiae. Potential metabolic engineering targets for improved production of succinic acid, glycerol and vanillin are identified and underlying flux changes for the predicted mutants are discussed. Conclusion We show that evolutionary programming enables solving large gene knockout problems in relatively short computational time. The proposed algorithm also allows the optimization of non-linear objective functions or incorporation of non-linear constraints and additionally provides a family of close to optimal solutions. The identified metabolic engineering strategies suggest that non-intuitive genetic modifications span

  10. Metabolite damage and repair in metabolic engineering design

    Sun, Jiayi; Jeffryes, James G.; Henry, Christopher S.; Bruner, Steven D.; Hanson, Andrew D.

    2017-11-01

    The necessarily sharp focus of metabolic engineering and metabolic synthetic biology on pathways and their fluxes has tended to divert attention from the damaging enzymatic and chemical side-reactions that pathway metabolites can undergo. Although historically overlooked and underappreciated, such metabolite damage reactions are now known to occur throughout metabolism and to generate (formerly enigmatic) peaks detected in metabolomics datasets. It is also now known that metabolite damage is often countered by dedicated repair enzymes that undo or prevent it. Metabolite damage and repair are highly relevant to engineered pathway design: metabolite damage reactions can reduce flux rates and product yields, and repair enzymes can provide robust, host-independent solutions. Herein, after introducing the core principles of metabolite damage and repair, we use case histories to document how damage and repair processes affect efficient operation of engineered pathways - particularly those that are heterologous, non-natural, or cell-free. We then review how metabolite damage reactions can be predicted, how repair reactions can be prospected, and how metabolite damage and repair can be built into genome-scale metabolic models. Lastly, we propose a versatile 'plug and play' set of well-characterized metabolite repair enzymes to solve metabolite damage problems known or likely to occur in metabolic engineering and synthetic biology projects.

  11. Recent applications of synthetic biology tools for yeast metabolic engineering

    Jensen, Michael Krogh; Keasling, Jay

    2015-01-01

    to engineer microbial chemical factories has steadily decreased, improvement is still needed. Through the development of synthetic biology tools for key microbial hosts, it should be possible to further decrease the development times and improve the reliability of the resulting microorganism. Together...... with continuous decreases in price and improvements in DNA synthesis, assembly and sequencing, synthetic biology tools will rationalize time-consuming strain engineering, improve control of metabolic fluxes, and diversify screening assays for cellular metabolism. This review outlines some recently developed...... synthetic biology tools and their application to improve production of chemicals and fuels in yeast. Finally, we provide a perspective for the challenges that lie ahead....

  12. Metabolic Engineering of Chemical Defence Pathways in Plant Disease Control

    Rook, Frederik

    2016-01-01

    on each topic. The chapter reviews the some of the scientific and technical challenges in metabolic engineering and the new possibilities emerging from recent technological developments. It concludes by discussing the outlook for bioengineered chemical defences as part of crop protection strategies, also...... with antimicrobial properties for use in crop protection. It presents an overview of the metabolic engineering efforts made in the area of plant chemical defence. For in-depth information on the characteristics of a specific class of chemical defence compounds, the reader is referred to the specialized reviews...

  13. Metabolic Engineering of Corynebacterium glutamicum for Methanol Metabolism

    Witthoff, Sabrina; Schmitz, Katja; Niedenführ, Sebastian; Nöh, Katharina; Noack, Stephan

    2015-01-01

    Methanol is already an important carbon feedstock in the chemical industry, but it has found only limited application in biotechnological production processes. This can be mostly attributed to the inability of most microbial platform organisms to utilize methanol as a carbon and energy source. With the aim to turn methanol into a suitable feedstock for microbial production processes, we engineered the industrially important but nonmethylotrophic bacterium Corynebacterium glutamicum toward the utilization of methanol as an auxiliary carbon source in a sugar-based medium. Initial oxidation of methanol to formaldehyde was achieved by heterologous expression of a methanol dehydrogenase from Bacillus methanolicus, whereas assimilation of formaldehyde was realized by implementing the two key enzymes of the ribulose monophosphate pathway of Bacillus subtilis: 3-hexulose-6-phosphate synthase and 6-phospho-3-hexuloisomerase. The recombinant C. glutamicum strain showed an average methanol consumption rate of 1.7 ± 0.3 mM/h (mean ± standard deviation) in a glucose-methanol medium, and the culture grew to a higher cell density than in medium without methanol. In addition, [13C]methanol-labeling experiments revealed labeling fractions of 3 to 10% in the m + 1 mass isotopomers of various intracellular metabolites. In the background of a C. glutamicum Δald ΔadhE mutant being strongly impaired in its ability to oxidize formaldehyde to CO2, the m + 1 labeling of these intermediates was increased (8 to 25%), pointing toward higher formaldehyde assimilation capabilities of this strain. The engineered C. glutamicum strains represent a promising starting point for the development of sugar-based biotechnological production processes using methanol as an auxiliary substrate. PMID:25595770

  14. Social Engineering a General Approach

    Valerica GREAVU-SERBAN; Oana SERBAN

    2014-01-01

    Social engineering is considered to be a taboo subject in nowadays society. It involves the use of social skills or to obtain usernames, passwords, credit card data, or to compromise or altering the information and systems of an entity. Social engineering methods are numerous and people using it are extremely ingenious and adaptable. This technique takes advantage of the intrinsic nature of mankind, to manipulate and obtain sensitive information, persuading people into divulge it, using excep...

  15. A systematic approach to engineering ethics education.

    Li, Jessica; Fu, Shengli

    2012-06-01

    Engineering ethics education is a complex field characterized by dynamic topics and diverse students, which results in significant challenges for engineering ethics educators. The purpose of this paper is to introduce a systematic approach to determine what to teach and how to teach in an ethics curriculum. This is a topic that has not been adequately addressed in the engineering ethics literature. This systematic approach provides a method to: (1) develop a context-specific engineering ethics curriculum using the Delphi technique, a process-driven research method; and (2) identify appropriate delivery strategies and instructional strategies using an instructional design model. This approach considers the context-specific needs of different engineering disciplines in ethics education and leverages the collaboration of engineering professors, practicing engineers, engineering graduate students, ethics scholars, and instructional design experts. The proposed approach is most suitable for a department, a discipline/field or a professional society. The approach helps to enhance learning outcomes and to facilitate ethics education curriculum development as part of the regular engineering curriculum.

  16. Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches

    Perrin H. Beatty

    2016-10-01

    Full Text Available A comprehensive understanding of plant metabolism could provide a direct mechanism for improving nitrogen use efficiency (NUE in crops. One of the major barriers to achieving this outcome is our poor understanding of the complex metabolic networks, physiological factors, and signaling mechanisms that affect NUE in agricultural settings. However, an exciting collection of computational and experimental approaches has begun to elucidate whole-plant nitrogen usage and provides an avenue for connecting nitrogen-related phenotypes to genes. Herein, we describe how metabolomics, computational models of metabolism, and flux balance analysis have been harnessed to advance our understanding of plant nitrogen metabolism. We introduce a model describing the complex flow of nitrogen through crops in a real-world agricultural setting and describe how experimental metabolomics data, such as isotope labeling rates and analyses of nutrient uptake, can be used to refine these models. In summary, the metabolomics/computational approach offers an exciting mechanism for understanding NUE that may ultimately lead to more effective crop management and engineered plants with higher yields.

  17. Genetic-Metabolic Coupling for Targeted Metabolic Engineering

    Cardinale, Stefano; Tueros Farfan, Felipe Gonzalo; Sommer, Morten Otto Alexander

    2017-01-01

    pertur-bations, thus limiting their relevance to specific conditions. Here, we address this issue by coupling cell fitness to the production of thiamine diphosphate in Escherichia coli using a synthetic RNA biosensor. We use this strategy to interrogate a library of transposon mutants and elucidate...... the native gene network influencing both cell fitness and thiamine production. Ultimately, we identify effectors of the OxyR-Fur stress response that limit thiamine biosynthesis via alternative regulation of iron storage and Fe-S cluster inclusion in enzymes. This study presents a new approach...

  18. Social Engineering a General Approach

    Valerica GREAVU-SERBAN

    2014-01-01

    Full Text Available Social engineering is considered to be a taboo subject in nowadays society. It involves the use of social skills or to obtain usernames, passwords, credit card data, or to compromise or altering the information and systems of an entity. Social engineering methods are numerous and people using it are extremely ingenious and adaptable. This technique takes advantage of the intrinsic nature of mankind, to manipulate and obtain sensitive information, persuading people into divulge it, using exceptional communication skills. Thus, five models of persuasion were identified, based on: simplicity, interest, incongruity, confidence and empathy, exploiting key factors which predispose people to fall victim to attacks of social engineering such as greed, self-interest, guilt or ignorance. It is well known fact that security is as strong as the weakest link in its chain (individuals therefore, beyond technical measures, staff training is the key to success in defending against such attacks.

  19. Volatile science? Metabolic engineering of terpenoids in plants

    Aharoni, A.; Jongsma, M.A.; Bouwmeester, H.J.

    2005-01-01

    Terpenoids are important for plant survival and also possess biological properties that are beneficial to humans. Here, we describe the state of the art in terpenoid metabolic engineering, showing that significant progress has been made over the past few years. Subcellular targeting of enzymes has

  20. Engineering of aromatic amino acid metabolism in Saccharomyces cerevisiae

    Vuralhan, Z.

    2006-01-01

    Saccharomyces cerevisiae is a popular industrial microorganism. It has since long been used in bread, beer and wine making. More recently it is also being applied for heterologous protein production and as a target organism for metabolic engineering. The work presented in this thesis describes how

  1. Cyanobacterial metabolic engineering for biofuel and chemical production.

    Oliver, Neal J; Rabinovitch-Deere, Christine A; Carroll, Austin L; Nozzi, Nicole E; Case, Anna E; Atsumi, Shota

    2016-12-01

    Rising levels of atmospheric CO 2 are contributing to the global greenhouse effect. Large scale use of atmospheric CO 2 may be a sustainable and renewable means of chemical and liquid fuel production to mitigate global climate change. Photosynthetic organisms are an ideal platform for efficient, natural CO 2 conversion to a broad range of chemicals. Cyanobacteria are especially attractive for these purposes, due to their genetic malleability and relatively fast growth rate. Recent years have yielded a range of work in the metabolic engineering of cyanobacteria and have led to greater knowledge of the host metabolism. Understanding of endogenous and heterologous carbon regulation mechanisms leads to the expansion of productive capacity and chemical variety. This review discusses the recent progress in metabolic engineering of cyanobacteria for biofuel and bulk chemical production since 2014. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Pathway elucidation and metabolic engineering of specialized plant metabolites

    Salomonsen, Bo

    A worldwide need to liberate ourselves from unsustainable petrochemicals has led to numerous metabolic engineering projects, mostly carried out in microbial hosts. Using systems biology for predicting and altering the metabolism of microorganisms towards production of a desired metabolite......, these projects have increased revenues on fermentative production of several biochemicals. The use of systems biology is, however, not limited to microorganisms. Recent advances in biotechnology methods have provided a wealth of data within functional genomics, metabolomics, transcriptomics, proteomics...... and fluxomics for a considerable number of organisms. Unfortunately, transferring the wealth of data to valuable information for metabolic engineering purposes is a non-obvious task. This PhD thesis describes a palate of tools used in generation of cell factories for production of specialized plant metabolites...

  3. Advanced computational approaches to biomedical engineering

    Saha, Punam K; Basu, Subhadip

    2014-01-01

    There has been rapid growth in biomedical engineering in recent decades, given advancements in medical imaging and physiological modelling and sensing systems, coupled with immense growth in computational and network technology, analytic approaches, visualization and virtual-reality, man-machine interaction and automation. Biomedical engineering involves applying engineering principles to the medical and biological sciences and it comprises several topics including biomedicine, medical imaging, physiological modelling and sensing, instrumentation, real-time systems, automation and control, sig

  4. Externally heated valve engine a new approach to piston engines

    Kazimierski, Zbyszko

    2016-01-01

    This book reports on a novel approach for generating mechanical energy from different, external heat sources using the body of a typical piston engine with valves. By presenting simple yet effective numerical models, the authors show how this new approach, which combines existing internal combustion technology with a lubrication system, is able to offer an economic solution to the problem of mechanical energy generation in piston engines. Their results also show that a stable heat generation process can be guaranteed outside of the engine. The book offers a detailed report on physical and numerical models of 4-stroke and 2-stroke versions of the EHVE together with different models of heat exchange, valves and results of their simulations. It also delivers the test results of an engine prototype run in laboratory conditions. By presenting a novel theoretical framework and providing readers with extensive knowledge of both the advantages and challenges of the method, this book is expected to inspire academic re...

  5. An ontological approach to domain engineering

    Falbo, R.A.; Guizzardi, G.; Duarte, K.

    2002-01-01

    Domain engineering aims to support systematic reuse, focusing on modeling common knowledge in a problem domain. Ontologies have also been pointed as holding great promise for software reuse. In this paper, we present ODE (Ontology-based Domain Engineering), an ontological approach for domain

  6. Distributed simulation a model driven engineering approach

    Topçu, Okan; Oğuztüzün, Halit; Yilmaz, Levent

    2016-01-01

    Backed by substantive case studies, the novel approach to software engineering for distributed simulation outlined in this text demonstrates the potent synergies between model-driven techniques, simulation, intelligent agents, and computer systems development.

  7. 2005 Plant Metabolic Engineering Gordon Conference - July 10-15, 2005

    Eleanore T. Wurtzel

    2006-06-30

    The post-genomic era presents new opportunities for manipulating plant chemistry for improvement of plant traits such as disease and stress resistance and nutritional qualities. This conference will provide a setting for developing multidisciplinary collaborations needed to unravel the dynamic complexity of plant metabolic networks and advance basic and applied research in plant metabolic engineering. The conference will integrate recent advances in genomics, with metabolite and gene expression analyses. Research discussions will explore how biosynthetic pathways interact with regard to substrate competition and channeling, plasticity of biosynthetic enzymes, and investigate the localization, structure, and assembly of biosynthetic metabolons in native and nonnative environments. The meeting will develop new perspectives for plant transgenic research with regard to how transgene expression may influence cellular metabolism. Incorporation of spectroscopic approaches for metabolic profiling and flux analysis combined with mathematical modeling will contribute to the development of rational metabolic engineering strategies and lead to the development of new tools to assess temporal and subcellular changes in metabolite pools. The conference will also highlight new technologies for pathway engineering, including use of heterologous systems, directed enzyme evolution, engineering of transcription factors and application of molecular/genetic techniques for controlling biosynthetic pathways.

  8. Corynebacterium glutamicum for Sustainable Bioproduction: From Metabolic Physiology to Systems Metabolic Engineering.

    Becker, Judith; Gießelmann, Gideon; Hoffmann, Sarah Lisa; Wittmann, Christoph

    Since its discovery 60 years ago, Corynebacterium glutamicum has evolved into a workhorse for industrial biotechnology. Traditionally well known for its remarkable capacity to produce amino acids, this Gram-positive soil bacterium, has become a flexible, efficient production platform for various bulk and fine chemicals, materials, and biofuels. The central turnstile of all these achievements is our excellent understanding of its metabolism and physiology. This knowledge base, together with innovative systems metabolic engineering concepts, which integrate systems and synthetic biology into strain engineering, has upgraded C. glutamicum into one of the most successful industrial microorganisms in the world.

  9. Metabolic Engineering of Oleaginous Yeasts for Fatty Alcohol Production

    Wang, Wei; Wei, Hui; Knoshaug, Eric; Van Wychen, Stefanie; Xu, Qi; Himmel, Michael E.; Zhang, Min

    2016-04-25

    To develop pathways for advanced biological upgrading of sugars to hydrocarbons, we are seeking biological approaches to produce high carbon efficiency intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels. In this study, we successfully demonstrated fatty alcohol production by oleaginous yeasts Yarrowia lipolytica and Lipomyces starkeyi by expressing a bacteria-derived fatty acyl-CoA reductase (FAR). Moreover, we find higher extracellular distribution of fatty alcohols produced by FAR-expressing L. starkeyi strain as compared to Y. lipolytica strain, which would benefit the downstream product recovery process. In both oleaginous yeasts, long chain length saturated fatty alcohols were predominant, accounting for more than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Taken together, our work demonstrates that in addition to Y. lipolytica, L. starkeyi can also serve as a platform organism for production of fatty acid-derived biofuels and bioproducts via metabolic engineering. We believe strain and process development both will significantly contribute to our goal of producing scalable and cost-effective fatty alcohols from renewable biomass.

  10. The application of microfluidic-based technologies in the cycle of metabolic engineering

    Xiaoyan Ma

    2016-09-01

    Full Text Available The process of metabolic engineering consists of multiple cycles of design, build, test and learn, which is typically laborious and time-consuming. To increase the efficiency and the rate of success of strain engineering, novel instrumentation must be applied. Microfluidics, the control of liquid flow in microstructures, has enabled flexible, accurate, automatic, and high-throughput manipulation of cells in liquid at picoliter to nanoliter scale. These attributes hold great promise in advancing metabolic engineering in terms of the phases of design, build, test and learn. To promote the application of microfluidic-based technologies in strain improvement, this review addressed the potentials of microfluidics and the related approaches in DNA assembly, transformation, strain screening, genotyping and phenotyping, and highlighted their adaptations for single-cell analysis. As a result, this facilitates in-depth understanding of the metabolic network, which in turn promote efficient optimization in the following cycles of strain engineering. Taken together, microfluidic-based technologies enable on-chip workflow, and could greatly accelerate the turnaround of metabolic engineering.

  11. Thought-evoking approaches in engineering problems

    2014-01-01

    In creating the value-added product in not distant future, it is necessary and inevitable to establish a holistic and though-evoking approach to the engineering problem, which should be at least associated with the inter-disciplinary knowledge and thought processes across the whole engineering spheres. It is furthermore desirable to integrate it with trans-disciplinary aspects ranging from manufacturing culture, through liberal-arts engineering, and industrial sociology.   The thought-evoking approach can be exemplified and typified by representative engineering problems: unveiling essential features in ‘Tangential Force Ratio and Interface Pressure’, prototype development for ‘Bio-mimetic Needle’ and application of ‘Water-jet Machining to Artificial Hip Joint’, product innovation in ‘Heat Sink for Computer’, application of ‘Graph Theory’ to similarity evaluation of production systems, leverage among reciprocity attributes in ‘Industrial and Engineering Designs for Machine Enclosure’,...

  12. Metabolically engineered cells for the production of polyunsaturated fatty acids

    2005-01-01

    The present invention relates to the construction and engineering of cells, more particularly microorganisms for producing PUFAs with four or more double bonds from non-fatty acid substrates through heterologous expression of an oxygen requiring pathway. The invention especially involves...... improvement of the PUFA content in the host organism through fermentation optimization, e.g. decreasing the temperature and/or designing an optimal medium, or through improving the flux towards fatty acids by metabolic engineering, e.g. through over-expression of fatty acid synthases, over-expression of other...

  13. The logics of metabolic regulation in bacteria challenges biosensor-based metabolic engineering

    Matthieu Jules

    2017-12-01

    Full Text Available Synthetic Biology (SB aims at the rational design and engineering of novel biological functions and systems. By facilitating the engineering of living organisms, SB promises to facilitate the development of many new applications for health, biomanufacturing, and the environment. Over the last decade, SB promoted the construction of libraries of components enabling the fine-tuning of genetic circuits expression and the development of novel genome engineering methodologies for many organisms of interest. SB thus opened new perspectives in the field of metabolic engineering, which was until then mainly limited to (overproducing naturally synthesized metabolic compounds. To engineer efficient cell factories, it is key to precisely reroute cellular resources from the central carbon metabolism (CCM to the synthetic circuitry. This task is however difficult as there is still significant lack of knowledge regarding both the function of several metabolic components and the regulation of the CCM fluxes for many industrially important bacteria. Pyruvate is a pivotal metabolite at the heart of the CCM and a key precursor for the synthesis of several commodity compounds and fine chemicals. Numerous bacterial species can also use it as a carbon source when present in the environment but bacterial, pyruvate-specific uptake systems were to be discovered. This is an issue for metabolic engineering as one can imagine to make use of pyruvate transport systems to replenish synthetic metabolic pathways towards the synthesis of chemicals of interest. Here we describe a recent study (MBio 8(5: e00976-17, which identified and characterized a pyruvate transport system in the Gram-positive (G+ve bacterium Bacillus subtilis, a well-established biotechnological workhorse for the production of enzymes, fine chemicals and antibiotics. This study also revealed that the activity of the two-component system (TCS responsible for its induction is retro-inhibited by the level of

  14. Mitigating health risks associated with alcoholic beverages through metabolic engineering.

    Jayakody, Lahiru N; Lane, Stephan; Kim, Heejin; Jin, Yong-Su

    2016-02-01

    Epidemiological studies have established a positive relationship between the occurrence of cancer and consumption of alcoholic beverages. Metabolic engineering of brewing yeast to reduce potential carcinogenic compounds in alcoholic beverage is technically feasible as well as economically promising. This review presents the mechanisms of formation of potentially carcinogenic components in alcoholic beverages, such as formaldehyde, acetaldehyde, ethyl carbamate, acrylamide, and heavy metals, and introduces effective genetic perturbations to minimize the concentrations of these harmful components. As precise and effective genome editing tools for polyploid yeast are now available, we envision that yeast metabolic engineering might open up new research directions for improving brewing yeast in order to ensure product safety as well as to increase overall quality of alcoholic beverages. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Metabolic Engineering of Microorganisms for the Production of Higher Alcohols

    Choi, Yong Jun; Lee, Joungmin; Jang, Yu-Sin

    2014-01-01

    ABSTRACT Due to the increasing concerns about limited fossil resources and environmental problems, there has been much interest in developing biofuels from renewable biomass. Ethanol is currently used as a major biofuel, as it can be easily produced by existing fermentation technology, but it is not the best biofuel due to its low energy density, high vapor pressure, hygroscopy, and incompatibility with current infrastructure. Higher alcohols, including 1-propanol, 1-butanol, isobutanol, 2-methyl-1-butanol, and 3-methyl-1-butanol, which possess fuel properties more similar to those of petroleum-based fuel, have attracted particular interest as alternatives to ethanol. Since microorganisms isolated from nature do not allow production of these alcohols at high enough efficiencies, metabolic engineering has been employed to enhance their production. Here, we review recent advances in metabolic engineering of microorganisms for the production of higher alcohols. PMID:25182323

  16. Two-Scale 13C Metabolic Flux Analysis for Metabolic Engineering.

    Ando, David; Garcia Martin, Hector

    2018-01-01

    Accelerating the Design-Build-Test-Learn (DBTL) cycle in synthetic biology is critical to achieving rapid and facile bioengineering of organisms for the production of, e.g., biofuels and other chemicals. The Learn phase involves using data obtained from the Test phase to inform the next Design phase. As part of the Learn phase, mathematical models of metabolic fluxes give a mechanistic level of comprehension to cellular metabolism, isolating the principle drivers of metabolic behavior from the peripheral ones, and directing future experimental designs and engineering methodologies. Furthermore, the measurement of intracellular metabolic fluxes is specifically noteworthy as providing a rapid and easy-to-understand picture of how carbon and energy flow throughout the cell. Here, we present a detailed guide to performing metabolic flux analysis in the Learn phase of the DBTL cycle, where we show how one can take the isotope labeling data from a 13 C labeling experiment and immediately turn it into a determination of cellular fluxes that points in the direction of genetic engineering strategies that will advance the metabolic engineering process.For our modeling purposes we use the Joint BioEnergy Institute (JBEI) Quantitative Metabolic Modeling (jQMM) library, which provides an open-source, python-based framework for modeling internal metabolic fluxes and making actionable predictions on how to modify cellular metabolism for specific bioengineering goals. It presents a complete toolbox for performing different types of flux analysis such as Flux Balance Analysis, 13 C Metabolic Flux Analysis, and it introduces the capability to use 13 C labeling experimental data to constrain comprehensive genome-scale models through a technique called two-scale 13 C Metabolic Flux Analysis (2S- 13 C MFA) [1]. In addition to several other capabilities, the jQMM is also able to predict the effects of knockouts using the MoMA and ROOM methodologies. The use of the jQMM library is

  17. De Novo Metabolic Engineering and the Promise of Synthetic DNA

    Klein-Marcuschamer, Daniel; Yadav, Vikramaditya G.; Ghaderi, Adel; Stephanopoulos, Gregory N.

    The uncertain price and tight supply of crude oil and the ever-increasing demand for clean energy have prompted heightened attention to the development of sustainable fuel technologies that ensure continued economic development while maintaining stewardship of the environment. In the face of these enormous challenges, biomass has emerged as a viable alternative to petroleum for the production of energy, chemicals, and materials owing to its abundance, inexpensiveness, and carbon-neutrality. Moreover, the immense ease and efficiency of biological systems at converting biomass-derived feedstocks into fuels, chemicals, and materials has generated renewed interest in biotechnology as a replacement for traditional chemical processes. Aided by the ever-expanding repertoire of microbial genetics and plant biotechnology, improved understanding of gene regulation and cellular metabolism, and incessantly accumulating gene and protein data, scientists are now contemplating engineering microbial cell factories to produce fuels, chemical feedstocks, polymers and pharmaceuticals in an economically and environmentally sustainable way. This goal resonates with that of metabolic engineering - the improvement of cellular properties through the intelligent design, rational modification, or directed evolution of biochemical pathways, and arguably, metabolic engineering seems best positioned to achieve the concomittant goals of environmental stewardship and economic prolificity.

  18. Non-photosynthetic plastids as hosts for metabolic engineering.

    Mellor, Silas Busck; Behrendorff, James B Y H; Nielsen, Agnieszka Zygadlo; Jensen, Poul Erik; Pribil, Mathias

    2018-04-13

    Using plants as hosts for production of complex, high-value compounds and therapeutic proteins has gained increasing momentum over the past decade. Recent advances in metabolic engineering techniques using synthetic biology have set the stage for production yields to become economically attractive, but more refined design strategies are required to increase product yields without compromising development and growth of the host system. The ability of plant cells to differentiate into various tissues in combination with a high level of cellular compartmentalization represents so far the most unexploited plant-specific resource. Plant cells contain organelles called plastids that retain their own genome, harbour unique biosynthetic pathways and differentiate into distinct plastid types upon environmental and developmental cues. Chloroplasts, the plastid type hosting the photosynthetic processes in green tissues, have proven to be suitable for high yield protein and bio-compound production. Unfortunately, chloroplast manipulation often affects photosynthetic efficiency and therefore plant fitness. In this respect, plastids of non-photosynthetic tissues, which have focused metabolisms for synthesis and storage of particular classes of compounds, might prove more suitable for engineering the production and storage of non-native metabolites without affecting plant fitness. This review provides the current state of knowledge on the molecular mechanisms involved in plastid differentiation and focuses on non-photosynthetic plastids as alternative biotechnological platforms for metabolic engineering. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  19. Metabolic engineering of microalgal based biofuel production: prospects and challenges

    Chiranjib eBanerjee

    2016-03-01

    Full Text Available The current scenario in renewable energy is focused on development of alternate and sustainable energy sources, amongst which microalgae stands as one of the promising feedstock for biofuel production. It is well known that microalgae generate much larger amounts of biofuels in a shorter time than other sources based on plant seeds. However, the greatest challenge in a transition to algae-based biofuel production is the various other complications involved in microalgal cultivation, its harvesting, concentration, drying and lipid extraction. Several green microalgae accumulate lipids, especially triacylglycerols (TAGs, which are main precursors in the production of lipid. The various aspects on metabolic pathway analysis of an oleaginous microalgae i.e. Chlamydomonas reinhardtii have elucidated some novel metabolically important genes and this enhances the lipid production in this microalgae. Adding to it, various other aspects in metabolic engineering using OptFlux and effectual bioprocess design also gives an interactive snapshot of enhancing lipid production which ultimately improvises the oil yield. This article reviews the current status of microalgal based technologies for biofuel production, bioreactor process design, flux analysis and it also provides various strategies to increase lipids accumulation via metabolic engineering.

  20. Synthetic biology approaches to engineer T cells.

    Wu, Chia-Yung; Rupp, Levi J; Roybal, Kole T; Lim, Wendell A

    2015-08-01

    There is rapidly growing interest in learning how to engineer immune cells, such as T lymphocytes, because of the potential of these engineered cells to be used for therapeutic applications such as the recognition and killing of cancer cells. At the same time, our knowhow and capability to logically engineer cellular behavior is growing rapidly with the development of synthetic biology. Here we describe how synthetic biology approaches are being used to rationally alter the behavior of T cells to optimize them for therapeutic functions. We also describe future developments that will be important in order to construct safe and precise T cell therapeutics. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Comparing Two Approaches for Engineering Education Development

    Edström, Kristina; Kolmos, Anette

    2012-01-01

    During the last decade there have been two dominating models for reforming engineering education: Problem/Project Based Learning (PBL) and the CDIO Initiative. The aim of this paper is to compare the PBL and CDIO approaches to engineering education reform, to identify and explain similarities...... and differences. CDIO and PBL will each be defined and compared in terms of the original need analysis, underlying educational philosophy and the essentials of the respective approaches to engineering education. In these respects we see many similarities. Circumstances that explain differences in history...... approaches have much in common and can be combined, and especially that the practitioners have much to learn from each other’s experiences through a dialogue between the communities. This structured comparison will potentially indicate specifically what an institution experienced in one of the communities...

  2. Synthetic biology and regulatory networks: where metabolic systems biology meets control engineering.

    He, Fei; Murabito, Ettore; Westerhoff, Hans V

    2016-04-01

    Metabolic pathways can be engineered to maximize the synthesis of various products of interest. With the advent of computational systems biology, this endeavour is usually carried out through in silico theoretical studies with the aim to guide and complement further in vitro and in vivo experimental efforts. Clearly, what counts is the result in vivo, not only in terms of maximal productivity but also robustness against environmental perturbations. Engineering an organism towards an increased production flux, however, often compromises that robustness. In this contribution, we review and investigate how various analytical approaches used in metabolic engineering and synthetic biology are related to concepts developed by systems and control engineering. While trade-offs between production optimality and cellular robustness have already been studied diagnostically and statically, the dynamics also matter. Integration of the dynamic design aspects of control engineering with the more diagnostic aspects of metabolic, hierarchical control and regulation analysis is leading to the new, conceptual and operational framework required for the design of robust and productive dynamic pathways. © 2016 The Author(s).

  3. Production of biopharmaceutical proteins by yeast: Advances through metabolic engineering

    Nielsen, Jens

    2013-01-01

    Production of recombinant proteins for use as pharmaceuticals, so-called biopharmaceuticals, is a multi-billion dollar industry. Many different cell factories are used for the production of biopharmaceuticals, but the yeast Saccharomyces cerevisiae is an important cell factory as it is used for p...... production. The involvement of directed metabolic engineering through the integration of tools from genetic engineering, systems biology and mathematical modeling, is also discussed....... by yeast are human serum albumin, hepatitis vaccines and virus like particles used for vaccination against human papillomavirus. Here is given a brief overview of biopharmaceutical production by yeast and it is discussed how the secretory pathway can be engineered to ensure more efficient protein...

  4. Role of glycolytic intermediate in regulation: Improving lycopene production in Escherichia coli by engineering metabolic control

    Farmer, W.R.; Liao, J.C.

    2001-06-01

    Metabolic engineering in the postgenomic era is expected to benefit from a full understanding of the biosynthetic capability of microorganisms as a result of the progress being made in bioinformatics and functional genomics. The immediate advantage of such information is to allow the rational design of novel pathways and the elimination of native reactions that are detrimental or unnecessary for the desired purpose. However, with the ability to manipulate metabolic pathways becoming more effective, metabolic engineering will need to face a new challenge: the reengineering of the regulatory hierarchy that controls gene expression in those pathways. In addition to constructing the genetic composition of a metabolic pathway, they propose that it will become just as important to consider the dynamics of pathways gene expression. It has been widely observed that high-level induction of a recombinant protein or pathway leads to growth retardation and reduced metabolic activity. These phenotypic characteristics result from the fact that the constant demands of production placed upon the cell interfere with its changing requirements for growth. They believe that this common situation in metabolic engineering can be alleviated by designing a dynamic controller that is able to sense the metabolic state of the cell and regulate the expression of the recombinant pathway accordingly. This approach, which is termed metabolic control engineering, involves redesigning the native regulatory circuits and applying them to the recombinant pathway. The general goal of such an effort will be to control the flux to the recombinant pathway adaptively according to the cell's metabolic state. The dynamically controlled recombinant pathway can potentially lead to enhanced production, minimized growth retardation, and reduced toxic by-product formation. The regulation of gene expression in response to the physiological state is also essential to the success of gene therapy. Here they

  5. Engineering design: A cognitive process approach

    Strimel, Greg Joseph

    research objectives of this study. Two independent coders then coded the video/audio recordings and the additional design data using Halfin's (1973) 17 mental processes for technological problem-solving. The results of this study indicated that the participants employed a wide array of mental processes when solving engineering design challenges. However, the findings provide a general analysis of the number of times participants employed each mental process, as well as the amount of time consumed employing the various mental processes through the different stages of the engineering design process. The results indicated many similarities between the students solving the problem, which may highlight voids in current technology and engineering education curricula. Additionally, the findings showed differences between the processes employed by participants that created the most successful solutions and the participants who developed the least effective solutions. Upon comparing and contrasting these processes, recommendations for instructional strategies to enhance a student's capability for solving engineering design problems were developed. The results also indicated that students, when left without teacher intervention, use a simplified and more natural process to solve design challenges than the 12-step engineering design process reported in much of the literature. Lastly, these data indicated that students followed two different approaches to solving the design problem. Some students employed a sequential and logical approach, while others employed a nebulous, solution centered trial-and-error approach to solving the problem. In this study the participants who were more sequential had better performing solutions. Examining these two approaches and the student cognition data enabled the researcher to generate a conceptual engineering design model for the improved teaching and development of engineering design problem solving.

  6. Systems engineering approach to preconceptual design

    1981-01-01

    Steps in the typical pre-conceptual engineering study are: problem identification, identify alternatives, evaluate alternatives, and recommend solution. Three examples of the use of pre-conceptual approach to save money are given; they include the water supply for fire protection in the 200-West Area of Hanford, emergency power for the Purex plant, and new filter for the Purex plant canyon exhaust

  7. Cytochrome P450-mediated metabolic engineering: current progress and future challenges.

    Renault, Hugues; Bassard, Jean-Etienne; Hamberger, Björn; Werck-Reichhart, Danièle

    2014-06-01

    Cytochromes P450 catalyze a broad range of regiospecific, stereospecific and irreversible steps in the biosynthetic routes of plant natural metabolites with important applications in pharmaceutical, cosmetic, fragrance and flavour, or polymer industries. They are consequently essential drivers for the engineered bioproduction of such compounds. Two ground-breaking developments of commercial products driven by the engineering of P450s are the antimalarial drug precursor artemisinic acid and blue roses or carnations. Tedious optimizations were required to generate marketable products. Hurdles encountered in P450 engineering and their potential solutions are summarized here. Together with recent technical developments and novel approaches to metabolic engineering, the lessons from this pioneering work should considerably boost exploitation of the amazing P450 toolkit emerging from accelerated sequencing of plant genomes. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  8. Metabolic network modeling of microbial interactions in natural and engineered environmental systems

    Octavio ePerez-Garcia

    2016-05-01

    Full Text Available We review approaches to characterize metabolic interactions within microbial communities using Stoichiometric Metabolic Network (SMN models for applications in environmental and industrial biotechnology. SMN models are computational tools used to evaluate the metabolic engineering potential of various organisms. They have successfully been applied to design and optimize the microbial production of antibiotics, alcohols and amino acids by single strains. To date however, such models have been rarely applied to analyze and control the metabolism of more complex microbial communities. This is largely attributed to the diversity of microbial community functions, metabolisms and interactions. Here, we firstly review different types of microbial interaction and describe their relevance for natural and engineered environmental processes. Next, we provide a general description of the essential methods of the SMN modeling workflow including the steps of network reconstruction, simulation through Flux Balance Analysis (FBA, experimental data gathering, and model calibration. Then we broadly describe and compare four approaches to model microbial interactions using metabolic networks, i.e. i lumped networks, ii compartment per guild networks, iii bi-level optimization simulations and iv dynamic-SMN methods. These approaches can be used to integrate and analyze diverse microbial physiology, ecology and molecular community data. All of them (except the lumped approach are suitable for incorporating species abundance data but so far they have been used only to model simple communities of two to eight different species. Interactions based on substrate exchange and competition can be directly modeled using the above approaches. However, interactions based on metabolic feedbacks, such as product inhibition and synthropy require extensions to current models, incorporating gene regulation and compounding accumulation mechanisms. SMN models of microbial

  9. 13C Metabolic Flux Analysis for systematic metabolic engineering of S. cerevisiae for overproduction of fatty acids.

    Amit Ghosh

    2016-10-01

    Full Text Available Efficient redirection of microbial metabolism into the abundant production of desired bioproducts remains non-trivial. Here we used flux-based modeling approaches to improve yields of fatty acids in S. cerevisiae. We combined 13C labeling data with comprehensive genome-scale models to shed light onto microbial metabolism and improve metabolic engineering efforts. We concentrated on studying the balance of acetyl-CoA, a precursor metabolite for the biosynthesis of fatty acids. A genome-wide acetyl-CoA balance study showed ATP citrate lyase from Y. lipolytica as a robust source of cytoplasmic acetyl-CoA and malate synthase as a desirable target for down-regulation in terms of acetyl-CoA consumption. These genetic modifications were applied to S. cerevisiae WRY2, a strain that is capable of producing 460 mg L of free fatty acids. With the addition of ATP citrate lyase and down-regulation of malate synthase the engineered strain produced 26 per cent more free fatty acids. Further increases in free fatty acid production of 33 per cent were obtained by knocking out the cytoplasmic glycerol-3-phosphate dehydrogenase, which flux analysis had shown was competing for carbon flux upstream with the carbon flux through the acetyl-CoA production pathway in the cytoplasm. In total, the genetic interventions applied in this work increased fatty acid production by 70 per cent.

  10. Non-photosynthetic plastids as hosts for metabolic engineering

    Mellor, Silas Busck; Behrendorff, James Bruce Yarnton H; Nielsen, Agnieszka Janina Zygadlo

    2018-01-01

    Using plants as hosts for production of complex, high-value compounds and therapeutic proteins has gained increasing momentum over the past decade. Recent advances in metabolic engineering techniques using synthetic biology have set the stage for production yields to become economically attractive......, but more refined design strategies are required to increase product yields without compromising development and growth of the host system. The ability of plant cells to differentiate into various tissues in combination with a high level of cellular compartmentalization represents so far the most...... in green tissues, have proven to be suitable for high yield protein and bio-compound production. Unfortunately, chloroplast manipulation often affects photosynthetic efficiency and therefore plant fitness. In this respect, plastids of non-photosynthetic tissues, which have focused metabolisms for synthesis...

  11. Combining engineering and data-driven approaches

    Fischer, Katharina; De Sanctis, Gianluca; Kohler, Jochen

    2015-01-01

    Two general approaches may be followed for the development of a fire risk model: statistical models based on observed fire losses can support simple cost-benefit studies but are usually not detailed enough for engineering decision-making. Engineering models, on the other hand, require many assump...... to the calibration of a generic fire risk model for single family houses to Swiss insurance data. The example demonstrates that the bias in the risk estimation can be strongly reduced by model calibration.......Two general approaches may be followed for the development of a fire risk model: statistical models based on observed fire losses can support simple cost-benefit studies but are usually not detailed enough for engineering decision-making. Engineering models, on the other hand, require many...... assumptions that may result in a biased risk assessment. In two related papers we show how engineering and data-driven modelling can be combined by developing generic risk models that are calibrated to statistical data on observed fire events. The focus of the present paper is on the calibration procedure...

  12. Production of vanillin by metabolically engineered Escherichia coli.

    Yoon, Sang-Hwal; Li, Cui; Kim, Ju-Eun; Lee, Sook-Hee; Yoon, Ji-Young; Choi, Myung-Suk; Seo, Weon-Taek; Yang, Jae-Kyung; Kim, Jae-Yeon; Kim, Seon-Won

    2005-11-01

    E. coli was metabolically engineered to produce vanillin by expression of the fcs and ech genes from Amycolatopsis sp. encoding feruloyl-CoA synthetase and enoyl-CoA hydratase/aldolase, respectively. Vanillin production was optimized by leaky expression of the genes, under the IPTG-inducible trc promoter, in complex 2YT medium. Supplementation with glucose, fructose, galactose, arabinose or glycerol severely decreased vanillin production. The highest vanillin production of 1.1 g l(-1) was obtained with cultivation for 48 h in 2YT medium with 0.2% (w/v) ferulate, without IPTG and no supplementation of carbon sources.

  13. Synthetic biology for engineering acetyl coenzyme a metabolism in yeast

    Nielsen, Jens

    2014-01-01

    The yeast Saccharomyces cerevisiae is a widely used cell factory for the production of fuels, chemicals, and pharmaceuticals. The use of this cell factory for cost-efficient production of novel fuels and chemicals requires high yields and low by-product production. Many industrially interesting...... chemicals are biosynthesized from acetyl coenzyme A (acetyl-CoA), which serves as a central precursor metabolite in yeast. To ensure high yields in production of these chemicals, it is necessary to engineer the central carbon metabolism so that ethanol production is minimized (or eliminated) and acetyl...

  14. Application of a controllable degron strategy for metabolic engineering

    Knuf, Christoph; Maury, Jerome; Jacobsen, Simo Abdessamad

    2014-01-01

    In numerous cases of metabolic engineering, metabolite pools have to be increased in order to obtain flux into heterologous pathways. A simple tool for this would be the deletion of genes that would practically lead to a block of the natural pathway, so that the carbon can flow into the heterolog...... of intermediates of the mevalonate pathway around 2,3-oxidosqualene, which is the precursor for triterpenoids. Many triterpenoids are pharmaceutically relevant compounds which nowadays need to be extracted from plant material through an intricate and resource consuming process....

  15. Development of biosensors and their application in metabolic engineering

    Zhang, Jie; Jensen, Michael Krogh; Keasling, Jay

    2015-01-01

    and ease of implementation with high-throughput analysis. Here we describe recent progress in biosensor development and their applications in a metabolic engineering context. We also highlight examples of how biosensors can be integrated with synthetic circuits to exert feedback regulation...... for the desired phenotypes. However, methods available for microbial genome diversification far exceed our ability to screen and select for those variants with optimal performance. Genetically encoded biosensors have shown the potential to address this gap, given their ability to respond to small molecule binding...

  16. An engineering design approach to systems biology.

    Janes, Kevin A; Chandran, Preethi L; Ford, Roseanne M; Lazzara, Matthew J; Papin, Jason A; Peirce, Shayn M; Saucerman, Jeffrey J; Lauffenburger, Douglas A

    2017-07-17

    Measuring and modeling the integrated behavior of biomolecular-cellular networks is central to systems biology. Over several decades, systems biology has been shaped by quantitative biologists, physicists, mathematicians, and engineers in different ways. However, the basic and applied versions of systems biology are not typically distinguished, which blurs the separate aspirations of the field and its potential for real-world impact. Here, we articulate an engineering approach to systems biology, which applies educational philosophy, engineering design, and predictive models to solve contemporary problems in an age of biomedical Big Data. A concerted effort to train systems bioengineers will provide a versatile workforce capable of tackling the diverse challenges faced by the biotechnological and pharmaceutical sectors in a modern, information-dense economy.

  17. Metabolic engineering of yeast for lignocellulosic biofuel production.

    Jin, Yong-Su; Cate, Jamie Hd

    2017-12-01

    Production of biofuels from lignocellulosic biomass remains an unsolved challenge in industrial biotechnology. Efforts to use yeast for conversion face the question of which host organism to use, counterbalancing the ease of genetic manipulation with the promise of robust industrial phenotypes. Saccharomyces cerevisiae remains the premier host for metabolic engineering of biofuel pathways, due to its many genetic, systems and synthetic biology tools. Numerous engineering strategies for expanding substrate ranges and diversifying products of S. cerevisiae have been developed. Other yeasts generally lack these tools, yet harbor superior phenotypes that could be exploited in the harsh processes required for lignocellulosic biofuel production. These include thermotolerance, resistance to toxic compounds generated during plant biomass deconstruction, and wider carbon consumption capabilities. Although promising, these yeasts have yet to be widely exploited. By contrast, oleaginous yeasts such as Yarrowia lipolytica capable of producing high titers of lipids are rapidly advancing in terms of the tools available for their metabolic manipulation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Metabolic engineering is key to a sustainable chemical industry.

    Murphy, Annabel C

    2011-08-01

    The depletion of fossil fuel stocks will prohibit their use as the main feedstock of future industrial processes. Biocatalysis is being increasingly used to reduce fossil fuel reliance and to improve the sustainability, efficiency and cost of chemical production. Even with their current small market share, biocatalyzed processes already generate approximately US$50 billion and it has been estimated that they could be used to produce up to 20% of fine chemicals by 2020. Until the advent of molecular biological technologies, the compounds that were readily accessible from renewable biomass were restricted to naturally-occurring metabolites. However, metabolic engineering has considerably broadened the range of compounds now accessible, providing access to compounds that cannot be otherwise reliably sourced, as well as replacing established chemical processes. This review presents the case for continued efforts to promote the adoption of biocatalyzed processes, highlighting successful examples of industrial chemical production from biomass and/or via biocatalyzed processes. A selection of emerging technologies that may further extend the potential and sustainability of biocatalysis are also presented. As the field matures, metabolic engineering will be increasingly crucial in maintaining our quality of life into a future where our current resources and feedstocks cannot be relied upon.

  19. Metabolic engineering of Candida glabrata for diacetyl production.

    Xiang Gao

    Full Text Available In this study, Candida glabrata, an efficient pyruvate-producing strain, was metabolically engineered for the production of the food ingredient diacetyl. A diacetyl biosynthetic pathway was reconstructed based on genetic modifications and medium optimization. The former included (i channeling carbon flux into the diacetyl biosynthetic pathway by amplification of acetolactate synthase, (ii elimination of the branched pathway of α-acetolactate by deleting the ILV5 gene, and (iii restriction of diacetyl degradation by deleting the BDH gene. The resultant strain showed an almost 1∶1 co-production of α-acetolactate and diacetyl (0.95 g L(-1. Furthermore, addition of Fe3+ to the medium enhanced the conversion of α-acetolactate to diacetyl and resulted in a two-fold increase in diacetyl production (2.1 g L(-1. In addition, increased carbon flux was further channeled into diacetyl biosynthetic pathway and a titer of 4.7 g L(-1 of diacetyl was achieved by altering the vitamin level in the flask culture. Thus, this study illustrates that C. glabrata could be tailored as an attractive platform for enhanced biosynthesis of beneficial products from pyruvate by metabolic engineering strategies.

  20. A Nutritional Approach to the Metabolic Syndrome

    Robert H. Lerman

    2011-02-01

    Full Text Available Poor diet and sedentary lifestyle contribute to the development of metabolic syndrome (MetS; addressing both is crucial for its management. A diet featuring the Mediterranean dietary pattern or low glycemic load has been shown to prevent and ameliorate MetS. Plant compounds, including soy protein and phytosterols, have been associated with reduced cardiovascular disease (CVD risk. Recently, phytochemicals from hops and acacia were identified as lipogenic, antiinflammatory compounds that reduced serum insulin and glucose levels in animals. A 12-week, randomized lifestyle intervention study in overweight and obese women with LDL ≥3.37 mmol/L (130 mg/dL compared a Mediterranean-style, low-glycemic-load diet and soy/phytosterol-based medical food to an AHA low-fat diet. The modified Mediterranean diet with medical food was superior in reducing markers of MetS and CVD risk. A subsequent,randomized 12-week study in men and women with MetS and LDL ≥3.37 mmol/L (130 mg/dL showed that supplementation with soy/phytosterol-based medical food plus phytochemicalsenhanced the benefits of a Mediterranean-style low-glycemic-load diet and aerobic exercise. At the completion of the study, 43% of participants receiving medical food and phytochemicalsexhibited net resolution of MetS compared with only 22% of those on diet and exercise alone. A subanalysis of participants at high risk (MetS + LDL ≥4.14 mmol/L [160 mg/dL] indicated minimal benefit from lifestyle change alone but marked benefits with the addition of medical food and phytochemicals. Case studies illustrate long-term benefits of this supplemented lifestyle change program. In conclusion, institution of a phytochemical-enhanced lifestyle intervention promises to be a clinically useful approach in MetS management.

  1. Enterprise architecture approach to mining companies engineering

    Ilin’ Igor

    2017-01-01

    Full Text Available As Russian economy is still largely oriented on commodities production, there are a lot of cities where mining and commodity-oriented enterprises are the backbone of city economy. The mentioned enterprises mostly define the life quality of citizens in such cities, thus there are high requirements for engineering of city-forming enterprises. The paper describes the enterprise architecture approach for management system engineering of the mining enterprises. The paper contains the model of the mining enterprise architecture, the approach to the development and implementation of an integrated management system based on the concept of enterprise architecture and the structure of information systems and information technology infrastructure of the mining enterprise.

  2. Engineering electrical properties of graphene: chemical approaches

    Kim, Yong-Jin; Kim, Yuna; Hong, Byung Hee; Novoselov, Konstantin

    2015-01-01

    To ensure the high performance of graphene-based devices, it is necessary to engineer the electrical properties of graphene with enhanced conductivity, controlled work function, opened or closed bandgaps, etc. This can be performed by various non-covalent chemical approaches, including molecular adsorption, substrate-induced doping, polymerization on graphene, deposition of metallic thin films or nanoparticles, etc. In addition, covalent approaches such as the substitution of carbon atoms with boron or nitrogen and the functionalization with hydrogen or fluorine are useful to tune the bandgaps more efficiently, with better uniformity and stability. In this review, representative examples of chemically engineered graphene and its device applications will be reviewed, and remaining challenges will be discussed. (topical review)

  3. NPP site selection: A systems engineering approach

    Pwani, Henry; Kamanja, Florah; Zolkaffly, Zulfakar; Jung, J. C. [KEPCO International Nuclear Graduate School, Ulsan (Korea, Republic of)

    2012-10-15

    The necessity for improved decision making concerning the siting and licensing of major power facilities has been accelerated in the past decade by the increased environmental consciousness of the public and by the energy crisis. These problems are exceedingly complex due to their multiple objective nature, the many interest groups, the long range time horizons, and the inherent uncertainties of the potential impacts of any decision. Along with the relatively objective economic and engineering concerns, the more subjective factors involving safety, environmental, and social issues are crucial to the problem. The preferences of the general public, as consumers, the utility companies, as builders and operators of power plant facilities, and environmentalists and the government must be accounted for in analyzing power plant siting and licensing issues. We advocate for a systems engineering approach that articulates stake holder's requirements, expert judgements, and a systems decision making approach. The appropriateness and application of systems decision making process is illustrated in this paper.

  4. NPP site selection: A systems engineering approach

    Pwani, Henry; Kamanja, Florah; Zolkaffly, Zulfakar; Jung, J. C.

    2012-01-01

    The necessity for improved decision making concerning the siting and licensing of major power facilities has been accelerated in the past decade by the increased environmental consciousness of the public and by the energy crisis. These problems are exceedingly complex due to their multiple objective nature, the many interest groups, the long range time horizons, and the inherent uncertainties of the potential impacts of any decision. Along with the relatively objective economic and engineering concerns, the more subjective factors involving safety, environmental, and social issues are crucial to the problem. The preferences of the general public, as consumers, the utility companies, as builders and operators of power plant facilities, and environmentalists and the government must be accounted for in analyzing power plant siting and licensing issues. We advocate for a systems engineering approach that articulates stake holder's requirements, expert judgements, and a systems decision making approach. The appropriateness and application of systems decision making process is illustrated in this paper

  5. Metabolic engineering: the ultimate paradigm for continuous pharmaceutical manufacturing.

    Yadav, Vikramaditya G; Stephanopoulos, Gregory

    2014-07-01

    Research and development (R&D) expenditures by pharmaceutical companies doubled over the past decade, yet candidate attrition rates and development times rose markedly during this period. Understandably, companies have begun downsizing their pipelines and diverting investments away from R&D in favor of manufacturing. It is estimated that transitioning to continuous manufacturing could enable companies to compete for a share in emerging markets. Accordingly, the model for continuous manufacturing that has emerged commences with the conversion of late-stage intermediates into the active pharmaceutical ingredient (API) in a series of continuous flow reactors, followed by continuous solid processing to form finished tablets. The use of flow reactions for API synthesis will certainly generate purer products at higher yields in shorter times compared to equivalent batch reactions. However, transitioning from batch to flow configuration simply alleviates transport limitations within the reaction milieu. As the catalogue of reactions used in flow syntheses is a subset of batch-based chemistries, molecules such as natural products will continue to evade drug prospectors. Also, it is uncertain whether flow synthesis can deliver improvements in the atom and energy economies of API production at the scales that would achieve the levels of revenue growth targeted by companies. Instead, it is argued that implementing metabolic engineering for the production of oxidized scaffolds as gateway molecules for flow-based addition of electrophiles is a more effective and scalable strategy for accessing natural product chemical space. This new paradigm for manufacturing, with metabolic engineering as its engine, would also permit rapid optimization of production variables and allow facile scale-up from gram to ton scale to meet material requirements for clinical trials, thus recasting manufacturing as a tool for discovery. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Integrating the protein and metabolic engineering toolkits for next-generation chemical biosynthesis.

    Pirie, Christopher M; De Mey, Marjan; Jones Prather, Kristala L; Ajikumar, Parayil Kumaran

    2013-04-19

    Through microbial engineering, biosynthesis has the potential to produce thousands of chemicals used in everyday life. Metabolic engineering and synthetic biology are fields driven by the manipulation of genes, genetic regulatory systems, and enzymatic pathways for developing highly productive microbial strains. Fundamentally, it is the biochemical characteristics of the enzymes themselves that dictate flux through a biosynthetic pathway toward the product of interest. As metabolic engineers target sophisticated secondary metabolites, there has been little recognition of the reduced catalytic activity and increased substrate/product promiscuity of the corresponding enzymes compared to those of central metabolism. Thus, fine-tuning these enzymatic characteristics through protein engineering is paramount for developing high-productivity microbial strains for secondary metabolites. Here, we describe the importance of protein engineering for advancing metabolic engineering of secondary metabolism pathways. This pathway integrated enzyme optimization can enhance the collective toolkit of microbial engineering to shape the future of chemical manufacturing.

  7. Hydrogen production and metabolic flux analysis of metabolically engineered Escherichia coli strains

    Kim, Seohyoung; Seol, Eunhee; Park, Sunghoon [Department of Chemical and Biochemical Engineering, Pusan National University, Busan 609-735 (Korea); Oh, You-Kwan [Bioenergy Research Center, Korea Institute of Energy Research, Daejeon 305-543 (Korea); Wang, G.Y. [Department of Oceanography, University of Hawaii at Manoa Honolulu, HI 96822 (United States)

    2009-09-15

    Escherichia coli can produce H{sub 2} from glucose via formate hydrogen lyase (FHL). In order to improve the H{sub 2} production rate and yield, metabolically engineered E. coli strains, which included pathway alterations in their H{sub 2} production and central carbon metabolism, were developed and characterized by batch experiments and metabolic flux analysis. Deletion of hycA, a negative regulator for FHL, resulted in twofold increase of FHL activity. Deletion of two uptake hydrogenases (1 (hya) and hydrogenase 2 (hyb)) increased H{sub 2} production yield from 1.20 mol/mol glucose to 1.48 mol/mol glucose. Deletion of lactate dehydrogenase (ldhA) and fumarate reductase (frdAB) further improved the H{sub 2} yield; 1.80 mol/mol glucose under high H{sub 2} pressure or 2.11 mol/mol glucose under reduced H{sub 2} pressure. Several batch experiments at varying concentrations of glucose (2.5-10 g/L) and yeast extract (0.3 or 3.0 g/L) were conducted for the strain containing all these genetic alternations, and their carbon and energy balances were analyzed. The metabolic flux analysis revealed that deletion of ldhA and frdAB directed most of the carbons from glucose to the glycolytic pathway leading to H{sub 2} production by FHL, not to the pentose phosphate pathway. (author)

  8. A POGIL approach to teaching engineering hydrology

    Rutten, M.

    2012-12-01

    This paper presents a case study of the author's experience using Problem Guided Inquiry Learning (POGIL) in an engineering hydrology course. This course is part of an interdisciplinary Water Management program at Bachelor level in the Netherlands. The aims of this approach were to promote constructivism of knowledge, activate critical thinking and reduce math anxiety. POGIL was developed for chemistry education in the United States. To the authors knowledge this is the first application of this approach in Europe. A first trial was done in 2010-2011 and a second trial in 2011-2012 and 55 students participated. The problems that motivated the novel approach, general information on POGIL, its implementation in the course are discussed and the results so far are evaluated.

  9. Cell-free protein synthesis enabled rapid prototyping for metabolic engineering and synthetic biology

    Lihong Jiang

    2018-06-01

    Full Text Available Advances in metabolic engineering and synthetic biology have facilitated the manufacturing of many valuable-added compounds and commodity chemicals using microbial cell factories in the past decade. However, due to complexity of cellular metabolism, the optimization of metabolic pathways for maximal production represents a grand challenge and an unavoidable barrier for metabolic engineering. Recently, cell-free protein synthesis system (CFPS has been emerging as an enabling alternative to address challenges in biomanufacturing. This review summarizes the recent progresses of CFPS in rapid prototyping of biosynthetic pathways and genetic circuits (biosensors to speed up design-build-test (DBT cycles of metabolic engineering and synthetic biology. Keywords: Cell-free protein synthesis, Metabolic pathway optimization, Genetic circuits, Metabolic engineering, Synthetic biology

  10. Polymer reaction engineering, an integrated approach

    Meyer, T.; Keurentjes, J.T.F.; Meyer, T.; Keurentjes, J.T.F.

    2005-01-01

    Summary This chapter contains sections titled: Polymer Materials A Short History of Polymer Reaction Engineering The Position of Polymer Reaction Engineering Toward Integrated Polymer Reaction Engineering The Disciplines in Polymer Reaction Engineering The Future: Product-inspired Polymer Reaction

  11. Document Clustering Approach for Meta Search Engine

    Kumar, Naresh, Dr.

    2017-08-01

    The size of WWW is growing exponentially with ever change in technology. This results in huge amount of information with long list of URLs. Manually it is not possible to visit each page individually. So, if the page ranking algorithms are used properly then user search space can be restricted up to some pages of searched results. But available literatures show that no single search system can provide qualitative results from all the domains. This paper provides solution to this problem by introducing a new meta search engine that determine the relevancy of query corresponding to web page and cluster the results accordingly. The proposed approach reduces the user efforts, improves the quality of results and performance of the meta search engine.

  12. The engineering approach to winter sports

    Cheli, Federico; Maldifassi, Stefano; Melzi, Stefano; Sabbioni, Edoardo

    2016-01-01

    The Engineering Approach to Winter Sports presents the state-of-the-art research in the field of winter sports in a harmonized and comprehensive way for a diverse audience of engineers, equipment and facilities designers, and materials scientists. The book examines the physics and chemistry of snow and ice with particular focus on the interaction (friction) between sports equipment and snow/ice, how it is influenced by environmental factors, such as temperature and pressure, as well as by contaminants and how it can be modified through the use of ski waxes or the microtextures of blades or ski soles. The authors also cover, in turn, the different disciplines in winter sports:  skiing (both alpine and cross country), skating and jumping, bob sledding and skeleton, hockey and curling, with attention given to both equipment design and on the simulation of gesture and  track optimization.

  13. Combination of traditional mutation and metabolic engineering to enhance ansamitocin P-3 production in Actinosynnema pretiosum.

    Du, Zhi-Qiang; Zhang, Yuan; Qian, Zhi-Gang; Xiao, Han; Zhong, Jian-Jiang

    2017-12-01

    Ansamitocin P-3 (AP-3) is a maytansinoid with its most compelling antitumor activity, however, the low production titer of AP-3 greatly restricts its wide commercial application. In this work, a combinatorial approach including random mutation and metabolic engineering was conducted to enhance AP-3 biosynthesis in Actinosynnema pretiosum. First, a mutant strain M was isolated by N-methyl-N'-nitro-N-nitrosoguanidine mutation, which could produce AP-3 almost threefold that of wild type (WT) in 48 deep-well plates. Then, by overexpressing key biosynthetic genes asmUdpg and asm13-17 in the M strain, a further 60% increase of AP-3 production in 250-ml shake flasks was achieved in the engineered strain M-asmUdpg:asm13-17 compared to the M strain, and its maximum AP-3 production reached 582.7 mg/L, which is the highest as ever reported. Both the gene transcription levels and intracellular intermediate concentrations in AP-3 biosynthesis pathway were significantly increased in the M and M-asmUdpg:asm13-17 during fermentation compared to the WT. The good fermentation performance of the engineered strain was also confirmed in a lab-scale bioreactor. This work demonstrated that combination of random mutation and metabolic engineering could promote AP-3 biosynthesis and might be helpful for increasing the production of other industrially important secondary metabolites. © 2017 Wiley Periodicals, Inc.

  14. Biosynthetic Pathway and Metabolic Engineering of Plant Dihydrochalcones.

    Ibdah, Mwafaq; Martens, Stefan; Gang, David R

    2018-03-14

    Dihydrochalcones are plant natural products containing the phenylpropanoid backbone and derived from the plant-specific phenylpropanoid pathway. Dihydrochalcone compounds are important in plant growth and response to stresses and, thus, can have large impacts on agricultural activity. In recent years, these compounds have also received increased attention from the biomedical community for their potential as anticancer treatments and other benefits for human health. However, they are typically produced at relatively low levels in plants. Therefore, an attractive alternative is to express the plant biosynthetic pathway genes in microbial hosts and to engineer the metabolic pathway/host to improve the production of these metabolites. In the present review, we discuss in detail the functions of genes and enzymes involved in the biosynthetic pathway of the dihydrochalcones and the recent strategies and achievements used in the reconstruction of multi-enzyme pathways in microorganisms in efforts to be able to attain higher amounts of desired dihydrochalcones.

  15. Metabolic engineering of Saccharomyces cerevisiae for overproduction of triacylglycerols

    Ferreira, Raphael; Teixeira, Paulo Goncalves; Gossing, Michael

    2018-01-01

    Triacylglycerols (TAGs) are valuable versatile compounds that can be used as metabolites for nutrition and health, as well as feedstocks for biofuel production. Although Saccharomyces cerevisiae is the favored microbial cell factory for industrial production of biochemicals, it does not produce...... large amounts of lipids and TAGs comprise only ~1% of its cell dry weight. Here, we engineered S. cerevisiae to reorient its metabolism for overproduction of TAGs, by regulating lipid droplet associated-proteins involved in TAG synthesis and hydrolysis. We implemented a push-and-pull strategy...... PXA1 led to accumulation of  254 mg∙gCDW−1. The TAG levels achieved here are the highest titer reported in S. cerevisiae, reaching 27.4% of the maximum theoretical yield in minimal medium with 2% glucose. This work shows the potential of using an industrially established and robust yeast species...

  16. Lessons learned from metabolic engineering of cyanogenic glucosides

    Morant, Anne Vinther; Jørgensen, Kirsten; Jørgensen, Bodil

    2007-01-01

    Plants produce a plethora of secondary metabolites which constitute a wealth of potential pharmaceuticals, pro-vitamins, flavours, fragrances, colorants and toxins as well as a source of natural pesticides. Many of these valuable compounds are only synthesized in exotic plant species or in concen......Plants produce a plethora of secondary metabolites which constitute a wealth of potential pharmaceuticals, pro-vitamins, flavours, fragrances, colorants and toxins as well as a source of natural pesticides. Many of these valuable compounds are only synthesized in exotic plant species...... or in concentrations too low to facilitate commercialization. In some cases their presence constitutes a health hazard and renders the crops unsuitable for consumption. Metabolic engineering is a powerful tool to alter and ameliorate the secondary metabolite composition of crop plants and gain new desired traits...

  17. Biobased organic acids production by metabolically engineered microorganisms

    Chen, Yun; Nielsen, Jens

    2016-01-01

    Bio-based production of organic acids via microbial fermentation has been traditionally used in food industry. With the recent desire to develop more sustainable bioprocesses for production of fuels, chemicals and materials, the market for microbial production of organic acids has been further...... expanded as organic acids constitute a key group among top building block chemicals that can be produced from renewable resources. Here we review the current status for production of citric acid and lactic acid, and we highlight the use of modern metabolic engineering technologies to develop high...... performance microbes for production of succinic acid and 3-hydroxypropionic acid. Also, the key limitations and challenges in microbial organic acids production are discussed...

  18. Simple glycolipids of microbes: Chemistry, biological activity and metabolic engineering

    Ahmad Mohammad Abdel-Mawgoud

    2018-03-01

    Full Text Available Glycosylated lipids (GLs are added-value lipid derivatives of great potential. Besides their interesting surface activities that qualify many of them to act as excellent ecological detergents, they have diverse biological activities with promising biomedical and cosmeceutical applications. Glycolipids, especially those of microbial origin, have interesting antimicrobial, anticancer, antiparasitic as well as immunomodulatory activities. Nonetheless, GLs are hardly accessing the market because of their high cost of production. We believe that experience of metabolic engineering (ME of microbial lipids for biofuel production can now be harnessed towards a successful synthesis of microbial GLs for biomedical and other applications. This review presents chemical groups of bacterial and fungal GLs, their biological activities, their general biosynthetic pathways and an insight on ME strategies for their production.

  19. Restructuring cities for sustainability : A metabolism approach

    Schremmer, C.; Stead, D.

    2009-01-01

    The FP7-funded SUME project (Sustainable Urban Metabolism for Europe) is focusing on the way how future urban systems can be designed to be consistently less damaging to the environment and particularly to climate change than in the present. Urban development scenarios linked with an agent-based

  20. In vitro metabolic engineering for the salvage synthesis of NAD(.).

    Honda, Kohsuke; Hara, Naoya; Cheng, Maria; Nakamura, Anna; Mandai, Komako; Okano, Kenji; Ohtake, Hisao

    2016-05-01

    Excellent thermal and operational stabilities of thermophilic enzymes can greatly increase the applicability of biocatalysis in various industrial fields. However, thermophilic enzymes are generally incompatible with thermo-labile substrates, products, and cofactors, since they show the maximal activities at high temperatures. Despite their pivotal roles in a wide range of enzymatic redox reactions, NAD(P)(+) and NAD(P)H exhibit relatively low stabilities at high temperatures, tending to be a major obstacle in the long-term operation of biocatalytic chemical manufacturing with thermophilic enzymes. In this study, we constructed an in vitro artificial metabolic pathway for the salvage synthesis of NAD(+) from its degradation products by the combination of eight thermophilic enzymes. The enzymes were heterologously produced in recombinant Escherichia coli and the heat-treated crude extracts of the recombinant cells were directly used as enzyme solutions. When incubated with experimentally optimized concentrations of the enzymes at 60°C, the NAD(+) concentration could be kept almost constant for 15h. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  1. The approach to engineering tasks composition on knowledge portals

    Novogrudska, Rina; Globa, Larysa; Schill, Alexsander; Romaniuk, Ryszard; Wójcik, Waldemar; Karnakova, Gaini; Kalizhanova, Aliya

    2017-08-01

    The paper presents an approach to engineering tasks composition on engineering knowledge portals. The specific features of engineering tasks are highlighted, their analysis makes the basis for partial engineering tasks integration. The formal algebraic system for engineering tasks composition is proposed, allowing to set the context-independent formal structures for engineering tasks elements' description. The method of engineering tasks composition is developed that allows to integrate partial calculation tasks into general calculation tasks on engineering portals, performed on user request demand. The real world scenario «Calculation of the strength for the power components of magnetic systems» is represented, approving the applicability and efficiency of proposed approach.

  2. Overproduction of Geranylgeraniol by Metabolically Engineered Saccharomyces cerevisiae▿

    Tokuhiro, Kenro; Muramatsu, Masayoshi; Ohto, Chikara; Kawaguchi, Toshiya; Obata, Shusei; Muramoto, Nobuhiko; Hirai, Masana; Takahashi, Haruo; Kondo, Akihiko; Sakuradani, Eiji; Shimizu, Sakayu

    2009-01-01

    (E, E, E)-Geranylgeraniol (GGOH) is a valuable starting material for perfumes and pharmaceutical products. In the yeast Saccharomyces cerevisiae, GGOH is synthesized from the end products of the mevalonate pathway through the sequential reactions of farnesyl diphosphate synthetase (encoded by the ERG20 gene), geranylgeranyl diphosphate synthase (the BTS1 gene), and some endogenous phosphatases. We demonstrated that overexpression of the diacylglycerol diphosphate phosphatase (DPP1) gene could promote GGOH production. We also found that overexpression of a BTS1-DPP1 fusion gene was more efficient for producing GGOH than coexpression of these genes separately. Overexpression of the hydroxymethylglutaryl-coenzyme A reductase (HMG1) gene, which encodes the major rate-limiting enzyme of the mevalonate pathway, resulted in overproduction of squalene (191.9 mg liter−1) rather than GGOH (0.2 mg liter−1) in test tube cultures. Coexpression of the BTS1-DPP1 fusion gene along with the HMG1 gene partially redirected the metabolic flux from squalene to GGOH. Additional expression of a BTS1-ERG20 fusion gene resulted in an almost complete shift of the flux to GGOH production (228.8 mg liter−1 GGOH and 6.5 mg liter−1 squalene). Finally, we constructed a diploid prototrophic strain coexpressing the HMG1, BTS1-DPP1, and BTS1-ERG20 genes from multicopy integration vectors. This strain attained 3.31 g liter−1 GGOH production in a 10-liter jar fermentor with gradual feeding of a mixed glucose and ethanol solution. The use of bifunctional fusion genes such as the BTS1-DPP1 and ERG20-BTS1 genes that code sequential enzymes in the metabolic pathway was an effective method for metabolic engineering. PMID:19592534

  3. Metabolic Engineering and Modeling of Metabolic Pathways to Improve Hydrogen Production by Photosynthetic Bacteria

    Jiao, Y. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Navid, A. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2014-12-19

    traits act as the biocatalysts of the process designed to both enhance the system efficiency of CO2 fixation and the net hydrogen production rate. Additionally we applied metabolic engineering approaches guided by computational modeling for the chosen model microorganisms to enable efficient hydrogen production.

  4. Metabolic engineering of deinococcus radiodurans based on computational analysis and functional genomics

    Edwards, Jeremy, S.

    2005-02-02

    The objective of our work is to develop novel computational tools to analyze the Deinococcus radiodurans DNA repair pathways and the influence of the metabolic flux distribution on DNA repair. These tools will be applied to provide insights for metabolic engineering of strains capable of growing under nutrient poor conditions similar to those found in mixed contaminant sites of interest to the DOE. Over the entire grant period we accomplished all our specific aims and were also able to pursue new directions of research. Below, I will list the major accomplishments over the previous 3 years. (1) Performed Monte Carlo Simulations of RecA Mediated Pairing of Homologous DNA Molecules. (2) Developed a statistical approach to study the gene expression data from D. radiodurans. We have been studying the data from John Batista's. (3) Developed an expression profiling technology to generate very accurate and precise expression data. We followed up on results from John Batista's group using this approach. (4) Developed and put online a database for metabolic reconstructions. (5) We have developed and applied new Monte Carlo algorithms that are optimized for studying biological systems. (6) We developed a flux balance model for the D. radiodurans metabolic network

  5. System engineering approach to GPM retrieval algorithms

    Rose, C. R. (Chris R.); Chandrasekar, V.

    2004-01-01

    System engineering principles and methods are very useful in large-scale complex systems for developing the engineering requirements from end-user needs. Integrating research into system engineering is a challenging task. The proposed Global Precipitation Mission (GPM) satellite will use a dual-wavelength precipitation radar to measure and map global precipitation with unprecedented accuracy, resolution and areal coverage. The satellite vehicle, precipitation radars, retrieval algorithms, and ground validation (GV) functions are all critical subsystems of the overall GPM system and each contributes to the success of the mission. Errors in the radar measurements and models can adversely affect the retrieved output values. Ground validation (GV) systems are intended to provide timely feedback to the satellite and retrieval algorithms based on measured data. These GV sites will consist of radars and DSD measurement systems and also have intrinsic constraints. One of the retrieval algorithms being studied for use with GPM is the dual-wavelength DSD algorithm that does not use the surface reference technique (SRT). The underlying microphysics of precipitation structures and drop-size distributions (DSDs) dictate the types of models and retrieval algorithms that can be used to estimate precipitation. Many types of dual-wavelength algorithms have been studied. Meneghini (2002) analyzed the performance of single-pass dual-wavelength surface-reference-technique (SRT) based algorithms. Mardiana (2003) demonstrated that a dual-wavelength retrieval algorithm could be successfully used without the use of the SRT. It uses an iterative approach based on measured reflectivities at both wavelengths and complex microphysical models to estimate both No and Do at each range bin. More recently, Liao (2004) proposed a solution to the Do ambiguity problem in rain within the dual-wavelength algorithm and showed a possible melting layer model based on stratified spheres. With the No and Do

  6. Metabolic engineering of Escherichia coli for the production of riboflavin

    2014-01-01

    Background Riboflavin (vitamin B2), the precursor of the flavin cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), is used commercially as an animal feed supplement and food colorant. E. coli is a robust host for various genetic manipulations and has been employed for efficient production of biofuels, polymers, amino acids, and bulk chemicals. Thus, the aim of this study was to understand the metabolic capacity of E. coli for the riboflavin production by modification of central metabolism, riboflavin biosynthesis pathway and optimization of the fermentation conditions. Results The basic producer RF01S, in which the riboflavin biosynthesis genes ribABDEC from E. coli were overexpressed under the control of the inducible trc promoter, could accumulate 229.1 mg/L of riboflavin. Further engineering was performed by examining the impact of expression of zwf (encodes glucose 6-phosphate dehydrogenase) and gnd (encodes 6-phosphogluconate dehydrogenase) from Corynebacterium glutamicum and pgl (encodes 6-phosphogluconolactonase) from E. coli on riboflavin production. Deleting pgi (encodes glucose-6-phosphate isomerase) and genes of Entner-Doudoroff (ED) pathway successfully redirected the carbon flux into the oxidative pentose phosphate pathway, and overexpressing the acs (encodes acetyl-CoA synthetase) reduced the acetate accumulation. These modifications increased riboflavin production to 585.2 mg/L. By further modulating the expression of ribF (encodes riboflavin kinase) for reducing the conversion of riboflavin to FMN in RF05S, the final engineering strain RF05S-M40 could produce 1036.1 mg/L riboflavin in LB medium at 37°C. After optimizing the fermentation conditions, strain RF05S-M40 produced 2702.8 mg/L riboflavin in the optimized semi-defined medium, which was a value nearly 12-fold higher than that of RF01S, with a yield of 137.5 mg riboflavin/g glucose. Conclusions The engineered strain RF05S-M40 has the highest yield among all

  7. Metabolic engineering of Escherichia coli for the production of riboflavin.

    Lin, Zhenquan; Xu, Zhibo; Li, Yifan; Wang, Zhiwen; Chen, Tao; Zhao, Xueming

    2014-07-16

    Riboflavin (vitamin B2), the precursor of the flavin cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), is used commercially as an animal feed supplement and food colorant. E. coli is a robust host for various genetic manipulations and has been employed for efficient production of biofuels, polymers, amino acids, and bulk chemicals. Thus, the aim of this study was to understand the metabolic capacity of E. coli for the riboflavin production by modification of central metabolism, riboflavin biosynthesis pathway and optimization of the fermentation conditions. The basic producer RF01S, in which the riboflavin biosynthesis genes ribABDEC from E. coli were overexpressed under the control of the inducible trc promoter, could accumulate 229.1 mg/L of riboflavin. Further engineering was performed by examining the impact of expression of zwf (encodes glucose 6-phosphate dehydrogenase) and gnd (encodes 6-phosphogluconate dehydrogenase) from Corynebacterium glutamicum and pgl (encodes 6-phosphogluconolactonase) from E. coli on riboflavin production. Deleting pgi (encodes glucose-6-phosphate isomerase) and genes of Entner-Doudoroff (ED) pathway successfully redirected the carbon flux into the oxidative pentose phosphate pathway, and overexpressing the acs (encodes acetyl-CoA synthetase) reduced the acetate accumulation. These modifications increased riboflavin production to 585.2 mg/L. By further modulating the expression of ribF (encodes riboflavin kinase) for reducing the conversion of riboflavin to FMN in RF05S, the final engineering strain RF05S-M40 could produce 1036.1 mg/L riboflavin in LB medium at 37°C. After optimizing the fermentation conditions, strain RF05S-M40 produced 2702.8 mg/L riboflavin in the optimized semi-defined medium, which was a value nearly 12-fold higher than that of RF01S, with a yield of 137.5 mg riboflavin/g glucose. The engineered strain RF05S-M40 has the highest yield among all reported riboflavin production

  8. Genomics:GTL Contractor-Grantee Workshop IV and Metabolic Engineering Working Group Inter-Agency Conference on Metabolic Engineering 2006

    Mansfield, Betty Kay [ORNL; Martin, Sheryl A [ORNL

    2006-02-01

    Welcome to the 2006 joint meeting of the fourth Genomics:GTL Contractor-Grantee Workshop and the six Metabolic Engineering Working Group Inter-Agency Conference. The vision and scope of the Genomics:GTL program continue to expand and encompass research and technology issues from diverse scientific disciplines, attracting broad interest and support from researchers at universities, DOE national laboratories, and industry. Metabolic engineering's vision is the targeted and purposeful alteration of metabolic pathways to improve the understanding and use of cellular pathways for chemical transformation, energy transduction, and supramolecular assembly. These two programs have much complementarity in both vision and technological approaches, as reflected in this joint workshop. GLT's challenge to the scientific community remains the further development and use of a broad array of innovative technologies and computational tools to systematically leverage the knowledge and capabilities brought to us by DNA sequencing projects. The goal is to seek a broad and predictive understanding of the functioning and control of complex systems--individual microbes, microbial communities, and plants. GTL's prominent position at the interface of the physical, computational, and biological sciences is both a strength and challenge. Microbes remain GTL's principal biological focus. In the complex 'simplicity' of microbes, they find capabilities needed by DOE and the nation for clean and secure energy, cleanup of environmental contamination, and sequestration of atmospheric carbon dioxide that contributes to global warming. An ongoing challenge for the entire GTL community is to demonstrate that the fundamental science conducted in each of your research projects brings us a step closer to biology-based solutions for these important national energy and environmental needs.

  9. Metabolic engineering of lactic acid bacteria for the production of nutraceuticals

    Hugenholtz, J.; Sybesma, W.; Groot, M.N.; Wisselink, W.; Ladero, V.; Burgess, K.; Sinderen, van D.; Piard, J.C.; Eggink, G.; Smid, E.J.; Savoy, G.; Sesma, F.; Jansen, T.; Hols, P.; Kleerebezem, M.

    2002-01-01

    Lactic acid bacteria display a relatively simple and well-described metabolism where the sugar source is converted mainly to lactic acid. Here we will shortly describe metabolic engineering strategies on the level of sugar metabolism, that lead to either the efficient re-routing of the lactococcal

  10. Metabolic engineering of Corynebacterium glutamicum for fermentative production of chemicals in biorefinery.

    Baritugo, Kei-Anne; Kim, Hee Taek; David, Yokimiko; Choi, Jong-Il; Hong, Soon Ho; Jeong, Ki Jun; Choi, Jong Hyun; Joo, Jeong Chan; Park, Si Jae

    2018-05-01

    Bio-based production of industrially important chemicals provides an eco-friendly alternative to current petrochemical-based processes. Because of the limited supply of fossil fuel reserves, various technologies utilizing microbial host strains for the sustainable production of platform chemicals from renewable biomass have been developed. Corynebacterium glutamicum is a non-pathogenic industrial microbial species traditionally used for L-glutamate and L-lysine production. It is a promising species for industrial production of bio-based chemicals because of its flexible metabolism that allows the utilization of a broad spectrum of carbon sources and the production of various amino acids. Classical breeding, systems, synthetic biology, and metabolic engineering approaches have been used to improve its applications, ranging from traditional amino-acid production to modern biorefinery systems for production of value-added platform chemicals. This review describes recent advances in the development of genetic engineering tools and techniques for the establishment and optimization of metabolic pathways for bio-based production of major C2-C6 platform chemicals using recombinant C. glutamicum.

  11. Metabolic engineering of Agrobacterium sp. ATCC31749 for curdlan production from cellobiose.

    Shin, Hyun-Dong; Liu, Long; Kim, Mi-Kyoung; Park, Yong-Il; Chen, Rachel

    2016-09-01

    Curdlan is a commercial polysaccharide made by fermentation of Agrobacterium sp. Its anticipated expansion to larger volume markets demands improvement in its production efficiency. Metabolic engineering for strain improvement has so far been limited due to the lack of genetic tools. This research aimed to identify strong promoters and to engineer a strain that converts cellobiose efficiently to curdlan. Three strong promoters were identified and were used to install an energy-efficient cellobiose phosphorolysis mechanism in a curdlan-producing strain. The engineered strains were shown with enhanced ability to utilize cellobiose, resulting in a 2.5-fold increase in titer. The availability of metabolically engineered strain capable of producing β-glucan from cellobiose paves the way for its production from cellulose. The identified native promoters from Agrobacterium open up opportunities for further metabolic engineering for improved production of curdlan and other products. The success shown here marks the first such metabolic engineering effort in this microbe.

  12. Expanding the chemical palate of cells by combining systems biology and metabolic engineering.

    Curran, Kathleen A; Alper, Hal S

    2012-07-01

    The field of Metabolic Engineering has recently undergone a transformation that has led to a rapid expansion of the chemical palate of cells. Now, it is conceivable to produce nearly any organic molecule of interest using a cellular host. Significant advances have been made in the production of biofuels, biopolymers and precursors, pharmaceuticals and nutraceuticals, and commodity and specialty chemicals. Much of this rapid expansion in the field has been, in part, due to synergies and advances in the area of systems biology. Specifically, the availability of functional genomics, metabolomics and transcriptomics data has resulted in the potential to produce a wealth of new products, both natural and non-natural, in cellular factories. The sheer amount and diversity of this data however, means that uncovering and unlocking novel chemistries and insights is a non-obvious exercise. To address this issue, a number of computational tools and experimental approaches have been developed to help expedite the design process to create new cellular factories. This review will highlight many of the systems biology enabling technologies that have reduced the design cycle for engineered hosts, highlight major advances in the expanded diversity of products that can be synthesized, and conclude with future prospects in the field of metabolic engineering. Copyright © 2012 Elsevier Inc. All rights reserved.

  13. Expanding beyond canonical metabolism: Interfacing alternative elements, synthetic biology, and metabolic engineering

    Kevin B. Reed

    2018-03-01

    Full Text Available Metabolic engineering offers an exquisite capacity to produce new molecules in a renewable manner. However, most industrial applications have focused on only a small subset of elements from the periodic table, centered around carbon biochemistry. This review aims to illustrate the expanse of chemical elements that can currently (and potentially be integrated into useful products using cellular systems. Specifically, we describe recent advances in expanding the cellular scope to include the halogens, selenium and the metalloids, and a variety of metal incorporations. These examples range from small molecules, heteroatom-linked uncommon elements, and natural products to biomining and nanotechnology applications. Collectively, this review covers the promise of an expanded range of elemental incorporations and the future impacts it may have on biotechnology.

  14. Therapeutic Approaches Using Riboflavin in Mitochondrial Energy Metabolism Disorders.

    Henriques, Bárbara J; Lucas, Tânia G; Gomes, Cláudio M

    2016-01-01

    Riboflavin, or vitamin B2, plays an important role in the cell as biological precursor of FAD and FMN, two important flavin cofactors which are essential for the structure and function of flavoproteins. Riboflavin has been used in therapeutic approaches of various inborn errors of metabolism, notably in metabolic disorders resulting either from defects in proteins involved in riboflavin metabolism and transport or from defects in flavoenzymes. The scope of this review is to provide an updated perspective of clinical cases in which riboflavin was used as a potential therapeutic agent in disorders affecting mitochondrial energy metabolism. In particular, we discuss available mechanistic insights on the role of riboflavin as a pharmacological chaperone for the recovery of misfolded metabolic flavoenzymes.

  15. Dynamic optimal metabolic control theory: a cybernetic approach for modelling of the central nitrogen metabolism of S. cerevisiae

    Riel, van N.A.W.; Giuseppin, M.L.F.; Verrips, C.T.

    2000-01-01

    The theory of dynamic optimal metabolic control (DOMC), as developed by Giuseppin and Van Riel (Metab. Eng., 2000), is applied to model the central nitrogen metabolism (CNM) in Saccharomyces cerevisiae. The CNM represents a typical system encountered in advanced metabolic engineering. The CNM is the

  16. Systematic metabolic engineering of Methylomicrobium alcaliphilum 20Z for 2,3-butanediol production from methane.

    Nguyen, Anh Duc; Hwang, In Yeub; Lee, Ok Kyung; Kim, Donghyuk; Kalyuzhnaya, Marina G; Mariyana, Rina; Hadiyati, Susila; Kim, Min Sik; Lee, Eun Yeol

    2018-04-16

    Methane is considered a next-generation feedstock, and methanotrophic cell-based biorefinery is attractive for production of a variety of high-value compounds from methane. In this work, we have metabolically engineered Methylomicrobium alcaliphilum 20Z for 2,3-butanediol (2,3-BDO) production from methane. The engineered strain 20Z/pBudK.p, harboring the 2,3-BDO synthesis gene cluster (budABC) from Klebsiella pneumoniae, accumulated 2,3-BDO in methane-fed shake flask cultures with a titer of 35.66 mg/L. Expression of the most efficient gene cluster was optimized using selection of promoters, translation initiation rates (TIR), and the combination of 2,3-BDO synthesis genes from different sources. A higher 2,3-BDO titer of 57.7 mg/L was measured in the 20Z/pNBM-Re strain with budA of K. pneumoniae and budB of Bacillus subtilis under the control of the Tac promoter. The genome-scale metabolic network reconstruction of M. alcaliphilum 20Z enabled in silico gene knockout predictions using an evolutionary programming method to couple growth and 2,3-BDO production. The ldh, ack, and mdh genes in M. alcaliphilum 20Z were identified as potential knockout targets. Pursuing these targets, a triple-mutant strain ∆ldh ∆ack ∆mdh was constructed, resulting in a further increase of the 2,3-BDO titer to 68.8 mg/L. The productivity of this optimized strain was then tested in a fed-batch stirred tank bioreactor, where final product concentrations of up to 86.2 mg/L with a yield of 0.0318 g-(2,3-BDO) /g-CH 4 were obtained under O 2 -limited conditions. This study first demonstrates the strategy of in silico simulation-guided metabolic engineering and represents a proof-of-concept for the production of value-added compounds using systematic approaches from engineered methanotrophs. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  17. Metabolic engineering of Dunaliella salina for production of ketocarotenoids.

    Anila, N; Simon, Daris P; Chandrashekar, Arun; Ravishankar, G A; Sarada, R

    2016-03-01

    Dunaliella is a commercially important marine alga producing high amount of β-carotene. The use of Dunaliella as a potential transgenic system for the production of recombinant proteins has been recently recognized. The present study reports for the first time the metabolic engineering of carotenoid biosynthesis in Dunaliella salina for ketocarotenoid production. The pathway modification included the introduction of a bkt gene from H. pluvialis encoding β-carotene ketolase (4,4'β-oxygenase) along with chloroplast targeting for the production of ketocarotenoids. The bkt under the control of Dunaliella Rubisco smaller subunit promoter along with its transit peptide sequence was introduced into the alga through standardized Agrobacterium-mediated transformation procedure. The selected transformants were confirmed using GFP and GUS expression, PCR and southern blot analysis. A notable upregulation of the endogenous hydroxylase level of transformants was observed where the BKT expression was higher in nutrient-limiting conditions. Carotenoid analysis of the transformants through HPLC and MS analysis showed the presence of astaxanthin and canthaxanthin with maximum content of 3.5 and 1.9 µg/g DW, respectively. The present study reports the feasibility of using D. salina for the production of ketocarotenoids including astaxanthin.

  18. Metabolic engineering of carbon overflow metabolism of Bacillus subtilis for improved N-acetyl-glucosamine production.

    Ma, Wenlong; Liu, Yanfeng; Shin, Hyun-Dong; Li, Jianghua; Chen, Jian; Du, Guocheng; Liu, Long

    2018-02-01

    Bacillus subtilis is widely used as cell factories for the production of important industrial biochemicals. Although many studies have demonstrated the effects of organic acidic byproducts, such as acetate, on microbial fermentation, little is known about the effects of blocking the neutral byproduct overflow, such as acetoin, on bioproduction. In this study, we focused on the influences of modulating overflow metabolism on the production of N-acetyl-d-glucosamine (GlcNAc) in engineered B. subtilis. We found that acetoin overflow competes with GlcNAc production, and blocking acetoin overflow increased GlcNAc titer and yield by 1.38- and 1.39-fold, reaching 48.9 g/L and 0.32 g GlcNAc/g glucose, respectively. Further blocking acetate overflow inhibited cell growth and GlcNAc production may be induced by inhibiting glucose uptake. Taken together, our results show that blocking acetoin overflow is a promising strategy for enhancing GlcNAc production. The strategies developed in this work may be useful for engineering strains of B. subtilis for producing other important biochemicals. Copyright © 2017. Published by Elsevier Ltd.

  19. Metabolic engineering of a haploid strain derived from a triploid industrial yeast for producing cellulosic ethanol.

    Kim, Soo Rin; Skerker, Jeffrey M; Kong, In Iok; Kim, Heejin; Maurer, Matthew J; Zhang, Guo-Chang; Peng, Dairong; Wei, Na; Arkin, Adam P; Jin, Yong-Su

    2017-03-01

    Many desired phenotypes for producing cellulosic biofuels are often observed in industrial Saccharomyces cerevisiae strains. However, many industrial yeast strains are polyploid and have low spore viability, making it difficult to use these strains for metabolic engineering applications. We selected the polyploid industrial strain S. cerevisiae ATCC 4124 exhibiting rapid glucose fermentation capability, high ethanol productivity, strong heat and inhibitor tolerance in order to construct an optimal yeast strain for producing cellulosic ethanol. Here, we focused on developing a general approach and high-throughput screening method to isolate stable haploid segregants derived from a polyploid parent, such as triploid ATCC 4124 with a poor spore viability. Specifically, we deleted the HO genes, performed random sporulation, and screened the resulting segregants based on growth rate, mating type, and ploidy. Only one stable haploid derivative (4124-S60) was isolated, while 14 other segregants with a stable mating type were aneuploid. The 4124-S60 strain inherited only a subset of desirable traits present in the parent strain, same as other aneuploids, suggesting that glucose fermentation and specific ethanol productivity are likely to be genetically complex traits and/or they might depend on ploidy. Nonetheless, the 4124-60 strain did inherit the ability to tolerate fermentation inhibitors. When additional genetic perturbations known to improve xylose fermentation were introduced into the 4124-60 strain, the resulting engineered strain (IIK1) was able to ferment a Miscanthus hydrolysate better than a previously engineered laboratory strain (SR8), built by making the same genetic changes. However, the IIK1 strain showed higher glycerol and xylitol yields than the SR8 strain. In order to decrease glycerol and xylitol production, an NADH-dependent acetate reduction pathway was introduced into the IIK1 strain. By consuming 2.4g/L of acetate, the resulting strain (IIK1A

  20. Dynamic Metabolic Model Building Based on the Ensemble Modeling Approach

    Liao, James C. [Univ. of California, Los Angeles, CA (United States)

    2016-10-01

    Ensemble modeling of kinetic systems addresses the challenges of kinetic model construction, with respect to parameter value selection, and still allows for the rich insights possible from kinetic models. This project aimed to show that constructing, implementing, and analyzing such models is a useful tool for the metabolic engineering toolkit, and that they can result in actionable insights from models. Key concepts are developed and deliverable publications and results are presented.

  1. Software quality engineering a practitioner's approach

    Suryn, Witold

    2014-01-01

    Software quality stems from two distinctive, but associated, topics in software engineering: software functional quality and software structural quality. Software Quality Engineering studies the tenets of both of these notions, which focus on the efficiency and value of a design, respectively. The text addresses engineering quality on both the application and system levels with attention to Information Systems and Embedded Systems as well as recent developments. Targeted at graduate engineering students and software quality specialists, the book analyzes the relationship between functionality

  2. Computational design of auxotrophy-dependent microbial biosensors for combinatorial metabolic engineering experiments.

    Tepper, Naama; Shlomi, Tomer

    2011-01-21

    Combinatorial approaches in metabolic engineering work by generating genetic diversity in a microbial population followed by screening for strains with improved phenotypes. One of the most common goals in this field is the generation of a high rate chemical producing strain. A major hurdle with this approach is that many chemicals do not have easy to recognize attributes, making their screening expensive and time consuming. To address this problem, it was previously suggested to use microbial biosensors to facilitate the detection and quantification of chemicals of interest. Here, we present novel computational methods to: (i) rationally design microbial biosensors for chemicals of interest based on substrate auxotrophy that would enable their high-throughput screening; (ii) predict engineering strategies for coupling the synthesis of a chemical of interest with the production of a proxy metabolite for which high-throughput screening is possible via a designed bio-sensor. The biosensor design method is validated based on known genetic modifications in an array of E. coli strains auxotrophic to various amino-acids. Predicted chemical production rates achievable via the biosensor-based approach are shown to potentially improve upon those predicted by current rational strain design approaches. (A Matlab implementation of the biosensor design method is available via http://www.cs.technion.ac.il/~tomersh/tools).

  3. Production of anthocyanins in metabolically engineered microorganisms: Current status and perspectives

    Jian Zha

    2017-12-01

    Full Text Available Microbial production of plant-derived natural products by engineered microorganisms has achieved great success thanks to large extend to metabolic engineering and synthetic biology. Anthocyanins, the water-soluble colored pigments found in terrestrial plants that are responsible for the red, blue and purple coloration of many flowers and fruits, are extensively used in food and cosmetics industry; however, their current supply heavily relies on complex extraction from plant-based materials. A promising alternative is their sustainable production in metabolically engineered microbes. Here, we review the recent progress on anthocyanin biosynthesis in engineered bacteria, with a special focus on the systematic engineering modifications such as selection and engineering of biosynthetic enzymes, engineering of transportation, regulation of UDP-glucose supply, as well as process optimization. These promising engineering strategies will facilitate successful microbial production of anthocyanins in industry in the near future.

  4. Metabolic Engineering of Oleaginous Yeasts for Production of Fuels and Chemicals

    Shuobo Shi

    2017-11-01

    Full Text Available Oleaginous yeasts have been increasingly explored for production of chemicals and fuels via metabolic engineering. Particularly, there is a growing interest in using oleaginous yeasts for the synthesis of lipid-related products due to their high lipogenesis capability, robustness, and ability to utilize a variety of substrates. Most of the metabolic engineering studies in oleaginous yeasts focused on Yarrowia that already has plenty of genetic engineering tools. However, recent advances in systems biology and synthetic biology have provided new strategies and tools to engineer those oleaginous yeasts that have naturally high lipid accumulation but lack genetic tools, such as Rhodosporidium, Trichosporon, and Lipomyces. This review highlights recent accomplishments in metabolic engineering of oleaginous yeasts and recent advances in the development of genetic engineering tools in oleaginous yeasts within the last 3 years.

  5. L-Cysteine Production in Escherichia coli Based on Rational Metabolic Engineering and Modular Strategy.

    Liu, Han; Fang, Guochen; Wu, Hui; Li, Zhimin; Ye, Qin

    2018-05-01

    L-cysteine is an amino acid with important physiological functions and has a wide range of applications in medicine, food, animal feed, and cosmetics industry. In this study, the L-cysteine synthesis in Escherichia coliEscherichia coli is divided into four modules: the transport module, sulfur module, precursor module, and degradation module. The engineered strain LH03 (overexpression of the feedback-insensitive cysE and the exporter ydeD in JM109) accumulated 45.8 mg L -1 of L-cysteine in 48 hr with yield of 0.4% g/g glucose. Further modifications of strains and culture conditions which based on the rational metabolic engineering and modular strategy improved the L-cysteine biosynthesis significantly. The engineered strain LH06 (with additional overexpression of serA, serC, and serB and double mutant of tnaA and sdaA in LH03) produced 620.9 mg L -1 of L-cysteine with yield of 6.0% g/g glucose, which increased the production by 12 times and the yield by 14 times more than those of LH03 in the original condition. In fed-batch fermentation performed in a 5-L reactor, the concentration of L-cysteine achieved 5.1 g L -1 in 32 hr. This work demonstrates that the combination of rational metabolic engineering and module strategy is a promising approach for increasing the L-cysteine production in E. coli. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Metabolic network model guided engineering ethylmalonyl-CoA pathway to improve ascomycin production in Streptomyces hygroscopicus var. ascomyceticus.

    Wang, Junhua; Wang, Cheng; Song, Kejing; Wen, Jianping

    2017-10-03

    Ascomycin is a 23-membered polyketide macrolide with high immunosuppressant and antifungal activity. As the lower production in bio-fermentation, global metabolic analysis is required to further explore its biosynthetic network and determine the key limiting steps for rationally engineering. To achieve this goal, an engineering approach guided by a metabolic network model was implemented to better understand ascomycin biosynthesis and improve its production. The metabolic conservation of Streptomyces species was first investigated by comparing the metabolic enzymes of Streptomyces coelicolor A3(2) with those of 31 Streptomyces strains, the results showed that more than 72% of the examined proteins had high sequence similarity with counterparts in every surveyed strain. And it was found that metabolic reactions are more highly conserved than the enzymes themselves because of its lower diversity of metabolic functions than that of genes. The main source of the observed metabolic differences was from the diversity of secondary metabolism. According to the high conservation of primary metabolic reactions in Streptomyces species, the metabolic network model of Streptomyces hygroscopicus var. ascomyceticus was constructed based on the latest reported metabolic model of S. coelicolor A3(2) and validated experimentally. By coupling with flux balance analysis and using minimization of metabolic adjustment algorithm, potential targets for ascomycin overproduction were predicted. Since several of the preferred targets were highly associated with ethylmalonyl-CoA biosynthesis, two target genes hcd (encoding 3-hydroxybutyryl-CoA dehydrogenase) and ccr (encoding crotonyl-CoA carboxylase/reductase) were selected for overexpression in S. hygroscopicus var. ascomyceticus FS35. Both the mutants HA-Hcd and HA-Ccr showed higher ascomycin titer, which was consistent with the model predictions. Furthermore, the combined effects of the two genes were evaluated and the strain HA

  7. Global Software Engineering: A Software Process Approach

    Richardson, Ita; Casey, Valentine; Burton, John; McCaffery, Fergal

    Our research has shown that many companies are struggling with the successful implementation of global software engineering, due to temporal, cultural and geographical distance, which causes a range of factors to come into play. For example, cultural, project managementproject management and communication difficulties continually cause problems for software engineers and project managers. While the implementation of efficient software processes can be used to improve the quality of the software product, published software process models do not cater explicitly for the recent growth in global software engineering. Our thesis is that global software engineering factors should be included in software process models to ensure their continued usefulness in global organisations. Based on extensive global software engineering research, we have developed a software process, Global Teaming, which includes specific practices and sub-practices. The purpose is to ensure that requirements for successful global software engineering are stipulated so that organisations can ensure successful implementation of global software engineering.

  8. Acetone production with metabolically engineered strains of Acetobacterium woodii.

    Hoffmeister, Sabrina; Gerdom, Marzena; Bengelsdorf, Frank R; Linder, Sonja; Flüchter, Sebastian; Öztürk, Hatice; Blümke, Wilfried; May, Antje; Fischer, Ralf-Jörg; Bahl, Hubert; Dürre, Peter

    2016-07-01

    Expected depletion of oil and fossil resources urges the development of new alternative routes for the production of bulk chemicals and fuels beyond petroleum resources. In this study, the clostridial acetone pathway was used for the formation of acetone in the acetogenic bacterium Acetobacterium woodii. The acetone production operon (APO) containing the genes thlA (encoding thiolase A), ctfA/ctfB (encoding CoA transferase), and adc (encoding acetoacetate decarboxylase) from Clostridium acetobutylicum were cloned under the control of the thlA promoter into four vectors having different replicons for Gram-positives (pIP404, pBP1, pCB102, and pCD6). Stable replication was observed for all constructs. A. woodii [pJIR_actthlA] achieved the maximal acetone concentration under autotrophic conditions (15.2±3.4mM). Promoter sequences of the genes ackA from A. woodii and pta-ack from C. ljungdahlii were determined by primer extension (PEX) and cloned upstream of the APO. The highest acetone production in recombinant A. woodii cells was achieved using the promoters PthlA and Ppta-ack. Batch fermentations using A. woodii [pMTL84151_actthlA] in a bioreactor revealed that acetate concentration had an effect on the acetone production, due to the high Km value of the CoA transferase. In order to establish consistent acetate concentration within the bioreactor and to increase biomass, a continuous fermentation process for A. woodii was developed. Thus, acetone productivity of the strain A. woodii [pMTL84151_actthlA] was increased from 1.2mgL(-1)h(-1) in bottle fermentation to 26.4mgL(-1)h(-1) in continuous gas fermentation. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  9. Hijacking CRISPR-Cas for high-throughput bacterial metabolic engineering: advances and prospects

    Mougiakos, Ioannis; Bosma, Elleke F.; Ganguly, Joyshree

    2018-01-01

    High engineering efficiencies are required for industrial strain development. Due to its user-friendliness and its stringency, CRISPR-Cas-based technologies have strongly increased genome engineering efficiencies in bacteria. This has enabled more rapid metabolic engineering of both the model host...... the range of organisms in which it can be used to create novel production hosts. This review analyses the current status of prokaryotic metabolic engineering towards the production of biotechnologically relevant products, based on the exploitation of different CRISPR-related DNA/RNA endonuclease variants....

  10. Compartmentalized Metabolic Engineering for Artemisinin Biosynthesis and Effective Malaria Treatment by Oral Delivery of Plant Cells.

    Malhotra, Karan; Subramaniyan, Mayavan; Rawat, Khushboo; Kalamuddin, Md; Qureshi, M Irfan; Malhotra, Pawan; Mohmmed, Asif; Cornish, Katrina; Daniell, Henry; Kumar, Shashi

    2016-11-07

    Artemisinin is highly effective against drug-resistant malarial parasites, which affects nearly half of the global population and kills >500 000 people each year. The primary cost of artemisinin is the very expensive process used to extract and purify the drug from Artemisia annua. Elimination of this apparently unnecessary step will make this potent antimalarial drug affordable to the global population living in endemic regions. Here we reported the oral delivery of a non-protein drug artemisinin biosynthesized (∼0.8 mg/g dry weight) at clinically meaningful levels in tobacco by engineering two metabolic pathways targeted to three different cellular compartments (chloroplast, nucleus, and mitochondria). The doubly transgenic lines showed a three-fold enhancement of isopentenyl pyrophosphate, and targeting AACPR, DBR2, and CYP71AV1 to chloroplasts resulted in higher expression and an efficient photo-oxidation of dihydroartemisinic acid to artemisinin. Partially purified extracts from the leaves of transgenic tobacco plants inhibited in vitro growth progression of Plasmodium falciparum-infected red blood cells. Oral feeding of whole intact plant cells bioencapsulating the artemisinin reduced the parasitemia levels in challenged mice in comparison with commercial drug. Such novel synergistic approaches should facilitate low-cost production and delivery of artemisinin and other drugs through metabolic engineering of edible plants. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  11. Production of the sesquiterpenoid (+)-nootkatone by metabolic engineering of Pichia pastoris.

    Wriessnegger, Tamara; Augustin, Peter; Engleder, Matthias; Leitner, Erich; Müller, Monika; Kaluzna, Iwona; Schürmann, Martin; Mink, Daniel; Zellnig, Günther; Schwab, Helmut; Pichler, Harald

    2014-07-01

    The sesquiterpenoid (+)-nootkatone is a highly demanded and highly valued aroma compound naturally found in grapefruit, pummelo or Nootka cypress tree. Extraction of (+)-nootkatone from plant material or its production by chemical synthesis suffers from low yields and the use of environmentally harmful methods, respectively. Lately, major attention has been paid to biotechnological approaches, using cell extracts or whole-cell systems for the production of (+)-nootkatone. In our study, the yeast Pichia pastoris initially was applied as whole-cell biocatalyst for the production of (+)-nootkatone from (+)-valencene, the abundant aroma compound of oranges. Therefore, we generated a strain co-expressing the premnaspirodiene oxygenase of Hyoscyamus muticus (HPO) and the Arabidopsis thaliana cytochrome P450 reductase (CPR) that hydroxylated extracellularly added (+)-valencene. Intracellular production of (+)-valencene by co-expression of valencene synthase from Callitropsis nootkatensis resolved the phase-transfer issues of (+)-valencene. Bi-phasic cultivations of P. pastoris resulted in the production of trans-nootkatol, which was oxidized to (+)-nootkatone by an intrinsic P. pastoris activity. Additional overexpression of a P. pastoris alcohol dehydrogenase and truncated hydroxy-methylglutaryl-CoA reductase (tHmg1p) significantly enhanced the (+)-nootkatone yield to 208mg L(-1) cell culture in bioreactor cultivations. Thus, metabolically engineered yeast P. pastoris represents a valuable, whole-cell system for high-level production of (+)-nootkatone from simple carbon sources. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  12. Metabolic engineering of Ustilago trichophora TZ1 for improved malic acid production

    Thiemo Zambanini

    2017-06-01

    These results open up a wide range of possibilities for further optimization, especially combinatorial metabolic engineering to increase the flux from pyruvate to malic acid and to reduce by-product formation.

  13. Synthetic biology and regulatory networks: where metabolic systems biology meets control engineering

    He, F.; Murabito, E.; Westerhoff, H.V.

    2016-01-01

    Metabolic pathways can be engineered to maximize the synthesis of various products of interest. With the advent of computational systems biology, this endeavour is usually carried out throughin silicotheoretical studies with the aim to guide and complement furtherin vitroandin vivoexperimental

  14. Natural and modified promoters for tailored metabolic engineering of the yeast Saccharomyces cerevisiae

    Hubmann, Georg; Thevelein, Johan M; Nevoigt, Elke

    2014-01-01

    The ease of highly sophisticated genetic manipulations in the yeast Saccharomyces cerevisiae has initiated numerous initiatives towards development of metabolically engineered strains for novel applications beyond its traditional use in brewing, baking, and wine making. In fact, baker's yeast has

  15. Development of Renewable Biofuels Technology by Transcriptomic Analysis and Metabolic Engineering of Diatoms

    Hildebrand, Mark [Univ. of California, San Diego, CA (United States)

    2013-11-18

    There is enormous interest in developing renewable sources of liquid fuels because of depletion of fossil fuel reserves, dependence on foreign sources, and increasing atmospheric CO2 levels. Algae produce neutral lipids that are readily converted into liquid fuels such as biodiesel or JP-8 equivalent, and are attractive sources because they are far more productive than plants (yielding 10 -100’s of time more lipid per land area), and can be grown on non-cultivatable land with non-potable (brackish or salt) water sources. Unicellular algae known as diatoms were the most thoroughly characterized species in the National Renewable Energy Laboratory’s Aquatic Species Program, whose goal was to develop microalgae as renewable fuel sources. Lipid accumulation in microalgae is generally induced by nutrient limitation, which involves a change in environmental conditions. Intrinsic variability in cellular response to environmental changes prevents a high degree of control over the process. Nutrient limitation also inhibits biomass accumulation; therefore a tradeoff between high biomass and lipid production occurs. The goal of this project was to develop metabolic engineering approaches for diatoms to enable induction of lipid accumulation by controllable manipulation of intracellular processes rather than from external environmental conditions, and to manipulate carbon partitioning within the cell between lipid and carbohydrate synthesis to enable both abundant biomass and lipid accumulation. There were two specific objectives for this project; Objective 1:To perform comparative transcriptomic analysis in T. pseudonana and C. cryptica of lipid accumulation resulting from silicon and nitrogen limitation, to identify common and key regulatory steps involved in controlling lipid accumulation and carbon partitioning; and Objective 2: To metabolically engineer the cell to alter carbon partitioning to either trigger lipid induction without the need for nutrient

  16. A bottom-up approach to urban metabolism: the perspective of BRIDGE

    Chrysoulakis, N.; Borrego, C.; San Josè, R.; Grimmond, S. B.; Jones, M. B.; Magliulo, V.; Klostermann, J.; Santamouris, M.

    2011-12-01

    Urban metabolism considers a city as a system and usually distinguishes between energy and material flows as its components. "Metabolic" studies are usually top-down approaches that assess the inputs and outputs of food, water, energy, and pollutants from a city, or that compare the changing metabolic process of several cities. In contrast, bottom-up approaches are based on quantitative estimates of urban metabolism components at local to regional scales. Such approaches consider the urban metabolism as the 3D exchange and transformation of energy and matter between a city and its environment. The city is considered as a system and the physical flows between this system and its environment are quantitatively estimated. The transformation of landscapes from primarily agricultural and forest uses to urbanized landscapes can greatly modify energy and material exchanges and it is, therefore, an important aspect of an urban area. Here we focus on the exchanges and transformation of energy, water, carbon and pollutants. Recent advances in bio-physical sciences have led to new methods and models to estimate local scale energy, water, carbon and pollutant fluxes. However, there is often poor communication of new knowledge and its implications to end-users, such as planners, architects and engineers. The FP7 Project BRIDGE (SustainaBle uRban plannIng Decision support accountinG for urban mEtabolism) aims at bridging this gap and at illustrating the advantages of considering environmental issues in urban planning. BRIDGE does not perform a complete life cycle analysis or calculate whole system urban metabolism, but rather focuses on specific metabolism components (energy, water, carbon and pollutants). Its main goal is the development of a Decision Suport System (DSS) with the potential to select planning actions which better fit the goal of changing the metabolism of urban systems towards sustainability. BRIDGE evaluates how planning alternatives can modify the physical

  17. Engineering Sustainability: A Technical Approach to Sustainability

    Rosen, Marc A.

    2012-01-01

    Sustainability is a critically important goal for human activity and development. Sustainability in the area of engineering is of great importance to any plans for overall sustainability given 1) the pervasiveness of engineering activities in societies, 2) their importance in economic development and living standards, and 3) the significant impacts that engineering processes and systems have had, and continue to have, on the environment. Many factors that need to be considered and appropriate...

  18. Metabolic pathway analysis using a nash equilibrium approach

    Lucia, Angelo; DiMaggio, Peter A.; Alonso-Martinez, Diego

    2018-01-01

    A novel approach to metabolic network analysis using a Nash Equilibrium (NE) formulation is proposed in which enzymes are considered players in a multi-player game. Each player has its own payoff function with the objective of minimizing the Gibbs free energy associated with the biochemical

  19. Toward systems metabolic engineering of Aspergillus and Pichia species for the production of chemicals and biofuels

    Caspeta, Luis; Nielsen, Jens

    2013-01-01

    trends in systems biology of Aspergillus and Pichia species, highlighting the relevance of these developments for systems metabolic engineering of these organisms for the production of hydrolytic enzymes, biofuels and chemicals from biomass. Metabolic engineering is moving from traditional methods...... for the production of hydrolytic enzymes, biofuels and chemicals from biomass. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim....

  20. Metabolomics Approach Reveals Integrated Metabolic Network Associated with Serotonin Deficiency

    Weng, Rui; Shen, Sensen; Tian, Yonglu; Burton, Casey; Xu, Xinyuan; Liu, Yi; Chang, Cuilan; Bai, Yu; Liu, Huwei

    2015-07-01

    Serotonin is an important neurotransmitter that broadly participates in various biological processes. While serotonin deficiency has been associated with multiple pathological conditions such as depression, schizophrenia, Alzheimer’s disease and Parkinson’s disease, the serotonin-dependent mechanisms remain poorly understood. This study therefore aimed to identify novel biomarkers and metabolic pathways perturbed by serotonin deficiency using metabolomics approach in order to gain new metabolic insights into the serotonin deficiency-related molecular mechanisms. Serotonin deficiency was achieved through pharmacological inhibition of tryptophan hydroxylase (Tph) using p-chlorophenylalanine (pCPA) or genetic knockout of the neuronal specific Tph2 isoform. This dual approach improved specificity for the serotonin deficiency-associated biomarkers while minimizing nonspecific effects of pCPA treatment or Tph2 knockout (Tph2-/-). Non-targeted metabolic profiling and a targeted pCPA dose-response study identified 21 biomarkers in the pCPA-treated mice while 17 metabolites in the Tph2-/- mice were found to be significantly altered compared with the control mice. These newly identified biomarkers were associated with amino acid, energy, purine, lipid and gut microflora metabolisms. Oxidative stress was also found to be significantly increased in the serotonin deficient mice. These new biomarkers and the overall metabolic pathways may provide new understanding for the serotonin deficiency-associated mechanisms under multiple pathological states.

  1. An Innovative Approach for online Meta Search Engine Optimization

    Manral, Jai; Hossain, Mohammed Alamgir

    2015-01-01

    This paper presents an approach to identify efficient techniques used in Web Search Engine Optimization (SEO). Understanding SEO factors which can influence page ranking in search engine is significant for webmasters who wish to attract large number of users to their website. Different from previous relevant research, in this study we developed an intelligent Meta search engine which aggregates results from various search engines and ranks them based on several important SEO parameters. The r...

  2. In-silico-driven metabolic engineering of Pseudomonas putida for enhanced production of poly-hydroxyalkanoates

    Poblete-Castro, I.; Binger, D.; Rodrigues, A.; Becker, J.; Martins Dos Santos, V.A.P.; Wittmann, C.

    2013-01-01

    Here, we present systems metabolic engineering driven by in-silico modeling to tailor Pseudomonas putida for synthesis of medium chain length PHAs on glucose. Using physiological properties of the parent wild type as constraints, elementary flux mode analysis of a large-scale model of the metabolism

  3. Metabolic engineering of ethanol production in Thermoanaerobacter mathranii

    Shou Yao

    2010-11-15

    Strain BG1 is a xylanolytic, thermophilic, anaerobic, Gram-positive bacterium originally isolated from an Icelandic hot spring. The strain belongs to the species Thermoanaerobacter mathranii. The strain ferments glucose, xylose, arabinose, galactose and mannose simultaneously and produces ethanol, acetate, lactate, CO{sub 2}, and H2 as fermentation end-products. As a potential ethanol producer from lignocellulosic biomass, tailor-made BG1 strain with the metabolism redirected to produce ethanol is needed. Metabolic engineering of T. mathranii BG1 is therefore necessary to improve ethanol production. Strain BG1 contains four alcohol dehydrogenase (ADH) encoding genes. They are adhA, adhB, bdhA and adhE encoding primary alcohol dehydrogenase, secondary alcohol dehydrogenase, butanol dehydrogenase and bifunctional alcohol/acetaldehyde dehydrogenase, respectively. The presence in an organism of multiple alcohol dehydrogenases with overlapping specificities makes the determination of the specific role of each ADH difficult. Deletion of each individual adh gene in the strain revealed that the adhE deficient mutant strain fails to produce ethanol as the fermentation product. The bifunctional alcohol/acetaldehyde dehydrogenase, AdhE, is therefore proposed responsible for ethanol production in T. mathranii BG1, by catalyzing sequential NADH-dependent reductions of acetyl-CoA to acetaldehyde and then to ethanol under fermentative conditions. Moreover, AdhE was conditionally expressed from a xylose-induced promoter in a recombinant strain (BG1E1) with a concomitant deletion of a lactate dehydrogenase. Over-expression of AdhE in strain BG1E1 with xylose as a substrate facilitates the production of ethanol at an increased yield. With a cofactor-dependent ethanol production pathway in T. mathranii BG1, it may become crucial to regenerate cofactor to increase the ethanol yield. Feeding the cells with a more reduced carbon source, such as mannitol, was shown to increase ethanol

  4. Dehydratase mediated 1-propanol production in metabolically engineered Escherichia coli

    Jain Rachit

    2011-11-01

    Full Text Available Abstract Background With the increasing consumption of fossil fuels, the question of meeting the global energy demand is of great importance in the near future. As an effective solution, production of higher alcohols from renewable sources by microorganisms has been proposed to address both energy crisis and environmental concerns. Higher alcohols contain more than two carbon atoms and have better physiochemical properties than ethanol as fuel substitutes. Results We designed a novel 1-propanol metabolic pathway by expanding the well-known 1,2-propanediol pathway with two more enzymatic steps catalyzed by a 1,2-propanediol dehydratase and an alcohol dehydrogenase. In order to engineer the pathway into E. coli, we evaluated the activities of eight different methylglyoxal synthases which play crucial roles in shunting carbon flux from glycolysis towards 1-propanol biosynthesis, as well as two secondary alcohol dehydrogenases of different origins that reduce both methylglyoxal and hydroxyacetone. It is evident from our results that the most active enzymes are the methylglyoxal synthase from Bacillus subtilis and the secondary alcohol dehydrogenase from Klebsiella pneumoniae, encoded by mgsA and budC respectively. With the expression of these two genes and the E. coli ydjG encoding methylglyoxal reductase, we achieved the production of 1,2-propanediol at 0.8 g/L in shake flask experiments. We then characterized the catalytic efficiency of three different diol dehydratases on 1,2-propanediol and identified the optimal one as the 1,2-propanediol dehydratase from Klebsiella oxytoca, encoded by the operon ppdABC. Co-expressing this enzyme with the above 1,2-propanediol pathway in wild type E. coli resulted in the production of 1-propanol at a titer of 0.25 g/L. Conclusions We have successfully established a new pathway for 1-propanol production by shunting the carbon flux from glycolysis. To our knowledge, it is the first time that this pathway has been

  5. Dynamic gene expression for metabolic engineering of mammalian cells in culture.

    Le, Huong; Vishwanathan, Nandita; Kantardjieff, Anne; Doo, Inseok; Srienc, Michael; Zheng, Xiaolu; Somia, Nikunj; Hu, Wei-Shou

    2013-11-01

    Recombinant mammalian cells are the major hosts for the production of protein therapeutics. In addition to high expression of the product gene, a hyper-producer must also harbor superior phenotypic traits related to metabolism, protein secretion, and growth control. Introduction of genes endowing the relevant hyper-productivity traits is a strategy frequently used to enhance the productivity. Most of such cell engineering efforts have been performed using constitutive expression systems. However, cells respond to various environmental cues and cellular events dynamically according to cellular needs. The use of inducible systems allows for time dependent expression, but requires external manipulation. Ideally, a transgene's expression should be synchronous to the host cell's own rhythm, and at levels appropriate for the objective. To that end, we identified genes with different expression dynamics and intensity ranges using pooled transcriptome data. Their promoters may be used to drive the expression of the transgenes following the desired dynamics. We isolated the promoter of the Thioredoxin-interacting protein (Txnip) gene and demonstrated its capability to drive transgene expression in concert with cell growth. We further employed this Chinese hamster promoter to engineer dynamic expression of the mouse GLUT5 fructose transporter in Chinese hamster ovary (CHO) cells, enabling them to utilize sugar according to cellular needs rather than in excess as typically seen in culture. Thus, less lactate was produced, resulting in a better growth rate, prolonged culture duration, and higher product titer. This approach illustrates a novel concept in metabolic engineering which can potentially be used to achieve dynamic control of cellular behaviors for enhanced process characteristics. © 2013 Published by Elsevier Inc.

  6. The role of metabolic engineering in the production of secondary metabolites

    Nielsen, Jens Bredal

    1998-01-01

    In the production of secondary metabolites yield and productivity are the most important design parameters. The focus is therefore to direct the carbon fluxes towards the product of interest, and this can be obtained through metabolic engineering whereby directed genetic changes are introduced...... into the production strain. In this process it is, however, important to analyze the metabolic network through measurement of the intracellular metabolites and the flux distributions. Besides playing an important role in the optimization of existing processes, metabolic engineering also offers the possibility...

  7. Enzyme and metabolic engineering for the production of novel biopolymers: crossover of biological and chemical processes.

    Matsumoto, Ken'ichiro; Taguchi, Seiichi

    2013-12-01

    The development of synthetic biology has transformed microbes into useful factories for producing valuable polymers and/or their precursors from renewable biomass. Recent progress at the interface of chemistry and biology has enabled the production of a variety of new biopolymers with properties that substantially differ from their petroleum-derived counterparts. This review touches on recent trials and achievements in the field of biopolymer synthesis, including chemo-enzymatically synthesized aliphatic polyesters, wholly biosynthesized lactate-based polyesters, polyhydroxyalkanoates and other unusual bacterially synthesized polyesters. The expanding diversities in structure and the material properties of biopolymers are key for exploring practical applications. The enzyme and metabolic engineering approaches toward this goal are discussed by shedding light on the successful case studies. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Knowledge Engineering Approach to the Geotectonic Discourse

    Pshenichny, Cyril

    2014-05-01

    The intellectual challenge of geotectonics is, and always was, much harder than that of most of the sciences: geotectonics has to say much when there is objectively not too much to say. As the target of study (the genesis of regional and planetary geological structures) is vast and multidisciplinary and is more or less generic for many geological disciplines, its more or less complete description is practically inachievable. Hence, the normal pathway of natural-scientific research - first acquire data, then draw conclusion - unlikely can be the case here. Geotectonics does quite the opposite; its approach is purely abductive: first to suggest a conceptualization (hypothesis) based on some external grounds (either general planetary/cosmic/philosophic/religious considerations, or based on experience gained from research of other structures/regions/planets) and then to acquire data that either support or refute it. In fact, geotectonics defines the context for data acquisition, and hence, the paradigm for the entire body of geology. Being an obvious necessity for a descriptive science, this nevertheless creates a number of threats: • Like any people, scientists like simplicity and unity, and therefore a single geotectonic hypothesis may seem preferable once based on the data available at the moment and oppress other views which may acquire evidence in the future; • As impartial data acquisition is rather a myth than reality even in most of the natural sciences, in a study like geology this process becomes strongly biased by the reigning hypothesis and controlled to supply only supportive evidence; • It becomes collectively agreed that any, or great many, domains of geological knowledge are determined by a geotectonic concept, which is, in turn, offered by a reigning hypothesis (sometimes reclassified as theory) - e.g., exploration geologists must involve the global geotectonic terminology in their technical reports on assessment of mineral or hydrocarbon

  9. Bioethanol a Microbial Biofuel Metabolite; New Insights of Yeasts Metabolic Engineering

    Khaled A. Selim

    2018-03-01

    Full Text Available Scarcity of the non-renewable energy sources, global warming, environmental pollution, and raising the cost of petroleum are the motive for the development of renewable, eco-friendly fuels production with low costs. Bioethanol production is one of the promising materials that can subrogate the petroleum oil, and it is considered recently as a clean liquid fuel or a neutral carbon. Diverse microorganisms such as yeasts and bacteria are able to produce bioethanol on a large scale, which can satisfy our daily needs with cheap and applicable methods. Saccharomyces cerevisiae and Pichia stipitis are two of the pioneer yeasts in ethanol production due to their abilities to produce a high amount of ethanol. The recent focus is directed towards lignocellulosic biomass that contains 30–50% cellulose and 20–40% hemicellulose, and can be transformed into glucose and fundamentally xylose after enzymatic hydrolysis. For this purpose, a number of various approaches have been used to engineer different pathways for improving the bioethanol production with simultaneous fermentation of pentose and hexoses sugars in the yeasts. These approaches include metabolic and flux analysis, modeling and expression analysis, followed by targeted deletions or the overexpression of key genes. In this review, we highlight and discuss the current status of yeasts genetic engineering for enhancing bioethanol production, and the conditions that influence bioethanol production.

  10. [Advances in metabolic engineering for the microbial production of naturally occurring terpenes-limonene and bisabolene: a mini review].

    Pang, Yaru; Hu, Zhihui; Xiao, Dongguang; Yu, Aiqun

    2018-01-25

    Limonene (C₁₀H₁₆) and bisabolene (C₁₅H₂₄) are both naturally occurring terpenes in plants. Depending on the number of C₅ units, limonene and bisabolene are recognized as representative monoterpenes and sesquiterpenes, respectively. Limonene and bisabolene are important pharmaceutical and nutraceutical products used in the prevention and treatment of cancer and many other diseases. In addition, they can be used as starting materials to produce a range of commercially valuable products, such as pharmaceuticals, nutraceuticals, cosmetics, and biofuels. The low abundance or yield of limonene and bisabolene in plants renders their isolation from plant sources non-economically viable. Isolation of limonene and bisabolene from plants also suffers from low efficiency and often requires harsh reaction conditions, prolonged reaction times, and expensive equipment cost. Recently, the rapid developments in metabolic engineering of microbes provide a promising alternative route for producing these plant natural products. Therefore, producing limonene and bisabolene by engineering microbial cells into microbial factories is becoming an attractive alternative approach that can overcome the bottlenecks, making it more sustainable, environmentally friendly and economically competitive. Here, we reviewed the status of metabolic engineering of microbes that produce limonene and bisabolene including microbial hosts, key enzymes, metabolic pathways and engineering of limonene/bisabolene biosynthesis. Furthermore, key challenges and future perspectives were discussed.

  11. A Status Report on the Global Research in Microbial Metabolic Engineering

    Joe, Min Ho; Lim, Sang Yong; Kim, Dong Ho

    2008-09-15

    Biotechnology industry is now a global 'Mega-Trend' and metabolic engineering technology has important role is this area. Therefore, many countries has made efforts in this field to produce top value added bio-products efficiently using microorganisms. It has been applied to increase the production of chemicals that are already produced by the host organism, to produce desired chemical substances from less expensive feedstock, and to generate products that are new to the host organism. Recent experimental advances, the so-called '-omics' technologies, mainly functional genomics, proteomics and metabolomics, have enabled wholesale generation of new genomic, transcriptomic, proteomic, and metabolomic data. This report provides the insights of the integrated view of metabolism generated by metabolic engineering for biotechnological applications of microbial metabolic engineering.

  12. A Status Report on the Global Research in Microbial Metabolic Engineering

    Joe, Min Ho; Lim, Sang Yong; Kim, Dong Ho

    2008-09-01

    Biotechnology industry is now a global 'Mega-Trend' and metabolic engineering technology has important role is this area. Therefore, many countries has made efforts in this field to produce top value added bio-products efficiently using microorganisms. It has been applied to increase the production of chemicals that are already produced by the host organism, to produce desired chemical substances from less expensive feedstock, and to generate products that are new to the host organism. Recent experimental advances, the so-called '-omics' technologies, mainly functional genomics, proteomics and metabolomics, have enabled wholesale generation of new genomic, transcriptomic, proteomic, and metabolomic data. This report provides the insights of the integrated view of metabolism generated by metabolic engineering for biotechnological applications of microbial metabolic engineering

  13. A Status Report on the Global Research in Microbial Metabolic Engineering

    Joe, Min Ho; Lim, Sang Yong; Kim, Dong Ho

    2008-09-15

    Biotechnology industry is now a global 'Mega-Trend' and metabolic engineering technology has important role is this area. Therefore, many countries has made efforts in this field to produce top value added bio-products efficiently using microorganisms. It has been applied to increase the production of chemicals that are already produced by the host organism, to produce desired chemical substances from less expensive feedstock, and to generate products that are new to the host organism. Recent experimental advances, the so-called '-omics' technologies, mainly functional genomics, proteomics and metabolomics, have enabled wholesale generation of new genomic, transcriptomic, proteomic, and metabolomic data. This report provides the insights of the integrated view of metabolism generated by metabolic engineering for biotechnological applications of microbial metabolic engineering.

  14. Investigating cholesterol metabolism and ageing using a systems biology approach.

    Morgan, A E; Mooney, K M; Wilkinson, S J; Pickles, N A; Mc Auley, M T

    2017-08-01

    CVD accounted for 27 % of all deaths in the UK in 2014, and was responsible for 1·7 million hospital admissions in 2013/2014. This condition becomes increasingly prevalent with age, affecting 34·1 and 29·8 % of males and females over 75 years of age respectively in 2011. The dysregulation of cholesterol metabolism with age, often observed as a rise in LDL-cholesterol, has been associated with the pathogenesis of CVD. To compound this problem, it is estimated by 2050, 22 % of the world's population will be over 60 years of age, in culmination with a growing resistance and intolerance to pre-existing cholesterol regulating drugs such as statins. Therefore, it is apparent research into additional therapies for hypercholesterolaemia and CVD prevention is a growing necessity. However, it is also imperative to recognise this complex biological system cannot be studied using a reductionist approach; rather its biological uniqueness necessitates a more integrated methodology, such as that offered by systems biology. In this review, we firstly discuss cholesterol metabolism and how it is affected by diet and the ageing process. Next, we describe therapeutic strategies for hypercholesterolaemia, and finally how the systems biology paradigm can be utilised to investigate how ageing interacts with complex systems such as cholesterol metabolism. We conclude by emphasising the need for nutritionists to work in parallel with the systems biology community, to develop novel approaches to studying cholesterol metabolism and its interaction with ageing.

  15. Review of Microfluidic Photobioreactor Technology for Metabolic Engineering and Synthetic Biology of Cyanobacteria and Microalgae

    Ya-Tang Yang

    2016-10-01

    Full Text Available One goal of metabolic engineering and synthetic biology for cyanobacteria and microalgae is to engineer strains that can optimally produce biofuels and commodity chemicals. However, the current workflow is slow and labor intensive with respect to assembly of genetic parts and characterization of production yields because of the slow growth rates of these organisms. Here, we review recent progress in the microfluidic photobioreactors and identify opportunities and unmet needs in metabolic engineering and synthetic biology. Because of the unprecedented experimental resolution down to the single cell level, long-term real-time monitoring capability, and high throughput with low cost, microfluidic photobioreactor technology will be an indispensible tool to speed up the development process, advance fundamental knowledge, and realize the full potential of metabolic engineering and synthetic biology for cyanobacteria and microalgae.

  16. Design, Optimization and Application of Small Molecule Biosensor in Metabolic Engineering.

    Liu, Yang; Liu, Ye; Wang, Meng

    2017-01-01

    The development of synthetic biology and metabolic engineering has painted a great future for the bio-based economy, including fuels, chemicals, and drugs produced from renewable feedstocks. With the rapid advance of genome-scale modeling, pathway assembling and genome engineering/editing, our ability to design and generate microbial cell factories with various phenotype becomes almost limitless. However, our lack of ability to measure and exert precise control over metabolite concentration related phenotypes becomes a bottleneck in metabolic engineering. Genetically encoded small molecule biosensors, which provide the means to couple metabolite concentration to measurable or actionable outputs, are highly promising solutions to the bottleneck. Here we review recent advances in the design, optimization and application of small molecule biosensor in metabolic engineering, with particular focus on optimization strategies for transcription factor (TF) based biosensors.

  17. Metabollic Engineering of Saccharomyces Cereviae a,omi acid metabolism for production of products of industrial interest

    Chen, Xiao

    -based processes. This study has focused on metabolic engineering of the amino acid metabolism in S. cerevisiae for production of two types of chemicals of industrial interest. The first chemical is δ-(L-α-aminoadipyl)–L-cysteinyl–D-valine (LLD-ACV). ACV belongs to non-ribosomal peptides (NRPs), which......Saccharomyces cerevisiae is widely used in microbial production of chemicals, metabolites and proteins, mainly because genetic manipulation of S. cerevisiae is relatively easy and experiences from its wide application in the existing industrial fermentations directly benefit new S. cerevisiae...

  18. Recent Approaches in Tooth Engineering Research

    Švandová, Eva; Veselá, Barbora; Křivánek, J.; Hampl, A.; Matalová, Eva

    2014-01-01

    Roč. 60, Suppl 1 (2014), s. 21-29 ISSN 0015-5500 R&D Projects: GA ČR GAP304/11/1418; GA MZd(CZ) NT11420 Institutional support: RVO:67985904 Keywords : engineering * tooth * stem cells * culture techniques Subject RIV: EA - Cell Biology Impact factor: 1.000, year: 2014

  19. Greenhouse engineering: New technologies and approaches

    Montero, J.I.; Henten, van E.J.; Son, J.E.; Castilla, N.

    2011-01-01

    Firstly, this article discusses the greenhouse engineering situation in three geographic areas which are relevant in the field of protected cultivation: Northern Asia, The Netherlands and the Mediterranean. For each area, the prevailing greenhouse type and equipment is briefly described. Secondly,

  20. Biomedical Engineering Education: A Conservative Approach

    Niemi, Eugene E., Jr.

    1973-01-01

    Describes the demand for graduates from biomedical engineering programs as being not yet fully able to absorb the supply. Suggests small schools interested in entering the field consider offering their programs at the undergraduate level via a minor or an option. Examples of such options and student projects are included. (CC)

  1. Metabolic engineering of microbial competitive advantage for industrial fermentation processes.

    Shaw, A Joe; Lam, Felix H; Hamilton, Maureen; Consiglio, Andrew; MacEwen, Kyle; Brevnova, Elena E; Greenhagen, Emily; LaTouf, W Greg; South, Colin R; van Dijken, Hans; Stephanopoulos, Gregory

    2016-08-05

    Microbial contamination is an obstacle to widespread production of advanced biofuels and chemicals. Current practices such as process sterilization or antibiotic dosage carry excess costs or encourage the development of antibiotic resistance. We engineered Escherichia coli to assimilate melamine, a xenobiotic compound containing nitrogen. After adaptive laboratory evolution to improve pathway efficiency, the engineered strain rapidly outcompeted a control strain when melamine was supplied as the nitrogen source. We additionally engineered the yeasts Saccharomyces cerevisiae and Yarrowia lipolytica to assimilate nitrogen from cyanamide and phosphorus from potassium phosphite, and they outcompeted contaminating strains in several low-cost feedstocks. Supplying essential growth nutrients through xenobiotic or ecologically rare chemicals provides microbial competitive advantage with minimal external risks, given that engineered biocatalysts only have improved fitness within the customized fermentation environment. Copyright © 2016, American Association for the Advancement of Science.

  2. Design Approaches to Myocardial and Vascular Tissue Engineering.

    Akintewe, Olukemi O; Roberts, Erin G; Rim, Nae-Gyune; Ferguson, Michael A H; Wong, Joyce Y

    2017-06-21

    Engineered tissues represent an increasingly promising therapeutic approach for correcting structural defects and promoting tissue regeneration in cardiovascular diseases. One of the challenges associated with this approach has been the necessity for the replacement tissue to promote sufficient vascularization to maintain functionality after implantation. This review highlights a number of promising prevascularization design approaches for introducing vasculature into engineered tissues. Although we focus on encouraging blood vessel formation within myocardial implants, we also discuss techniques developed for other tissues that could eventually become relevant to engineered cardiac tissues. Because the ultimate solution to engineered tissue vascularization will require collaboration between wide-ranging disciplines such as developmental biology, tissue engineering, and computational modeling, we explore contributions from each field.

  3. Systems Biology as an Integrated Platform for Bioinformatics, Systems Synthetic Biology, and Systems Metabolic Engineering

    Bor-Sen Chen

    2013-10-01

    Full Text Available Systems biology aims at achieving a system-level understanding of living organisms and applying this knowledge to various fields such as synthetic biology, metabolic engineering, and medicine. System-level understanding of living organisms can be derived from insight into: (i system structure and the mechanism of biological networks such as gene regulation, protein interactions, signaling, and metabolic pathways; (ii system dynamics of biological networks, which provides an understanding of stability, robustness, and transduction ability through system identification, and through system analysis methods; (iii system control methods at different levels of biological networks, which provide an understanding of systematic mechanisms to robustly control system states, minimize malfunctions, and provide potential therapeutic targets in disease treatment; (iv systematic design methods for the modification and construction of biological networks with desired behaviors, which provide system design principles and system simulations for synthetic biology designs and systems metabolic engineering. This review describes current developments in systems biology, systems synthetic biology, and systems metabolic engineering for engineering and biology researchers. We also discuss challenges and future prospects for systems biology and the concept of systems biology as an integrated platform for bioinformatics, systems synthetic biology, and systems metabolic engineering.

  4. Systems Biology as an Integrated Platform for Bioinformatics, Systems Synthetic Biology, and Systems Metabolic Engineering

    Chen, Bor-Sen; Wu, Chia-Chou

    2013-01-01

    Systems biology aims at achieving a system-level understanding of living organisms and applying this knowledge to various fields such as synthetic biology, metabolic engineering, and medicine. System-level understanding of living organisms can be derived from insight into: (i) system structure and the mechanism of biological networks such as gene regulation, protein interactions, signaling, and metabolic pathways; (ii) system dynamics of biological networks, which provides an understanding of stability, robustness, and transduction ability through system identification, and through system analysis methods; (iii) system control methods at different levels of biological networks, which provide an understanding of systematic mechanisms to robustly control system states, minimize malfunctions, and provide potential therapeutic targets in disease treatment; (iv) systematic design methods for the modification and construction of biological networks with desired behaviors, which provide system design principles and system simulations for synthetic biology designs and systems metabolic engineering. This review describes current developments in systems biology, systems synthetic biology, and systems metabolic engineering for engineering and biology researchers. We also discuss challenges and future prospects for systems biology and the concept of systems biology as an integrated platform for bioinformatics, systems synthetic biology, and systems metabolic engineering. PMID:24709875

  5. Metabolic engineering to expand the substrate spectrum of Pseudomonas putida toward sucrose.

    Löwe, Hannes; Schmauder, Lukas; Hobmeier, Karina; Kremling, Andreas; Pflüger-Grau, Katharina

    2017-08-01

    Sucrose is an important disaccharide used as a substrate in many industrial applications. It is a major component of molasses, a cheap by-product of the sugar industry. Unfortunately, not all industrially relevant organisms, among them Pseudomonas putida, are capable of metabolizing sucrose. We chose a metabolic engineering approach to circumvent this blockage and equip P. putida with the activities necessary to consume sucrose. Therefore, we constructed a pair of broad-host range mini-transposons (pSST - sucrose splitting transposon), carrying either cscA, encoding an invertase able to split sucrose into glucose and fructose, or additionally cscB, encoding a sucrose permease. Introduction of cscA was sufficient to convey sucrose consumption and the additional presence of cscB had no further effect, though the sucrose permease was built and localized to the membrane. Sucrose was split extracellularly by the activity of the invertase CscA leaking out of the cell. The transposons were also used to confer sucrose consumption to Cupriavidus necator. Interestingly, in this strain, CscB acted as a glucose transporter, such that C. necator also gained the ability to grow on glucose. Thus, the pSST transposons are functional tools to extend the substrate spectrum of Gram-negative bacterial strains toward sucrose. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  6. Harnessing the respiration machinery for high-yield production of chemicals in metabolically engineered Lactococcus lactis

    Liu, Jianming; Wang, Zhihao; Kandasamy, Vijayalakshmi

    2017-01-01

    on metabolically engineered Lactococcus lactis strains to optimize the production of acetoin and (R,R)−2,3-butanediol (R-BDO). In the absence of an external electron acceptor, a surplus of two NADH per acetoin molecule is produced. We found that a fully activated respiration was able to efficiently regenerate NAD......+, and a high titer of 371 mM (32 g/L) of acetoin was obtained with a yield of 82% of the theoretical maximum. Subsequently, we extended the metabolic pathway from acetoin to R-BDO by introducing the butanediol dehydrogenase gene from Bacillus subtilis. Since one mole of NADH is consumed when acetoin...... is converted into R-BDO per mole, only the excess of NADH needs to be oxidized via respiration. Either by fine-tuning the respiration capacity or by using a dual-phase fermentation approach involving a switch from fully respiratory to non-respiratory conditions, we obtained 361 mM (32 g/L) R-BDO with a yield...

  7. Terminal alkenes as versatile chemical reporter groups for metabolic oligosaccharide engineering.

    Späte, Anne-Katrin; Schart, Verena F; Schöllkopf, Sophie; Niederwieser, Andrea; Wittmann, Valentin

    2014-12-08

    The Diels-Alder reaction with inverse electron demand (DAinv reaction) of 1,2,4,5-tetrazines with electron rich or strained alkenes was proven to be a bioorthogonal ligation reaction that proceeds fast and with high yields. An important application of the DAinv reaction is metabolic oligosaccharide engineering (MOE) which allows the visualization of glycoconjugates in living cells. In this approach, a sugar derivative bearing a chemical reporter group is metabolically incorporated into cellular glycoconjugates and subsequently derivatized with a probe by means of a bioorthogonal ligation reaction. Here, we investigated a series of new mannosamine and glucosamine derivatives with carbamate-linked side chains of varying length terminated by alkene groups and their suitability for labeling cell-surface glycans. Kinetic investigations showed that the reactivity of the alkenes in DAinv reactions increases with growing chain length. When applied to MOE, one of the compounds, peracetylated N-butenyloxycarbonylmannosamine, was especially well suited for labeling cell-surface glycans. Obviously, the length of its side chain represents the optimal balance between incorporation efficiency and speed of the labeling reaction. Sialidase treatment of the cells before the bioorthogonal labeling reaction showed that this sugar derivative is attached to the glycans in form of the corresponding sialic acid derivative and not epimerized to another hexosamine derivative to a considerable extent. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Improved Triacylglycerol Production in Acinetobacter baylyi ADP1 by Metabolic Engineering

    Karp Matti

    2011-05-01

    Full Text Available Abstract Background Triacylglycerols are used in various purposes including food applications, cosmetics, oleochemicals and biofuels. Currently the main sources for triacylglycerol are vegetable oils, and microbial triacylglycerol has been suggested as an alternative for these. Due to the low production rates and yields of microbial processes, the role of metabolic engineering has become more significant. As a robust model organism for genetic and metabolic studies, and for the natural capability to produce triacylglycerol, Acinetobacter baylyi ADP1 serves as an excellent organism for modelling the effects of metabolic engineering for energy molecule biosynthesis. Results Beneficial gene deletions regarding triacylglycerol production were screened by computational means exploiting the metabolic model of ADP1. Four deletions, acr1, poxB, dgkA, and a triacylglycerol lipase were chosen to be studied experimentally both separately and concurrently by constructing a knock-out strain (MT with three of the deletions. Improvements in triacylglycerol production were observed: the strain MT produced 5.6 fold more triacylglycerol (mg/g cell dry weight compared to the wild type strain, and the proportion of triacylglycerol in total lipids was increased by 8-fold. Conclusions In silico predictions of beneficial gene deletions were verified experimentally. The chosen single and multiple gene deletions affected beneficially the natural triacylglycerol metabolism of A. baylyi ADP1. This study demonstrates the importance of single gene deletions in triacylglycerol metabolism, and proposes Acinetobacter sp. ADP1 as a model system for bioenergetic studies regarding metabolic engineering.

  9. Engineering microorganisms to increase ethanol production by metabolic redirection

    Deng, Yu; Olson, Daniel G.; van Dijken, Johannes Pieter; Shaw, IV, Arthur J.; Argyros, Aaron; Barrett, Trisha; Caiazza, Nicky; Herring, Christopher D.; Rogers, Stephen R.; Agbogbo, Frank

    2017-10-31

    The present invention provides for the manipulation of carbon flux in a recombinant host cell to increase the formation of desirable products. The invention relates to cellulose-digesting organisms that have been genetically modified to allow the production of ethanol at a high yield by redirecting carbon flux at key steps of central metabolism.

  10. Metabolic engineering toward 1-butanol derivatives in solvent producing clostridia

    Siemerink, M.A.J.

    2010-01-01

    Chapter 1 of this thesis gives an overview about the history of the acetone, butanol and ethanol (ABE) fermentation. The responsible solventogenic clostridia with their central metabolism are briefly discussed. Despite the fact that scientific research on the key organisms of the ABE process has

  11. Helicopter Gas Turbine Engine Performance Analysis : A Multivariable Approach

    Arush, Ilan; Pavel, M.D.

    2017-01-01

    Helicopter performance relies heavily on the available output power of the engine(s) installed. A simplistic single-variable analysis approach is often used within the flight-testing community to reduce raw flight-test data in order to predict the available output power under different atmospheric

  12. The JPL Cost Risk Analysis Approach that Incorporates Engineering Realism

    Harmon, Corey C.; Warfield, Keith R.; Rosenberg, Leigh S.

    2006-01-01

    This paper discusses the JPL Cost Engineering Group (CEG) cost risk analysis approach that accounts for all three types of cost risk. It will also describe the evaluation of historical cost data upon which this method is based. This investigation is essential in developing a method that is rooted in engineering realism and produces credible, dependable results to aid decision makers.

  13. Fumaric acid production in Saccharomyces cerevisiae by in silico aided metabolic engineering.

    Guoqiang Xu

    Full Text Available Fumaric acid (FA is a promising biomass-derived building-block chemical. Bio-based FA production from renewable feedstock is a promising and sustainable alternative to petroleum-based chemical synthesis. Here we report on FA production by direct fermentation using metabolically engineered Saccharomyces cerevisiae with the aid of in silico analysis of a genome-scale metabolic model. First, FUM1 was selected as the target gene on the basis of extensive literature mining. Flux balance analysis (FBA revealed that FUM1 deletion can lead to FA production and slightly lower growth of S. cerevisiae. The engineered S. cerevisiae strain obtained by deleting FUM1 can produce FA up to a concentration of 610±31 mg L(-1 without any apparent change in growth in fed-batch culture. FT-IR and (1H and (13C NMR spectra confirmed that FA was synthesized by the engineered S. cerevisiae strain. FBA identified pyruvate carboxylase as one of the factors limiting higher FA production. When the RoPYC gene was introduced, S. cerevisiae produced 1134±48 mg L(-1 FA. Furthermore, the final engineered S. cerevisiae strain was able to produce 1675±52 mg L(-1 FA in batch culture when the SFC1 gene encoding a succinate-fumarate transporter was introduced. These results demonstrate that the model shows great predictive capability for metabolic engineering. Moreover, FA production in S. cerevisiae can be efficiently developed with the aid of in silico metabolic engineering.

  14. System engineering approach to planning anticancer therapies

    Świerniak, Andrzej; Smieja, Jaroslaw; Puszynski, Krzysztof; Psiuk-Maksymowicz, Krzysztof

    2016-01-01

    This book focuses on the analysis of cancer dynamics and the mathematically based synthesis of anticancer therapy. It summarizes the current state-of-the-art in this field and clarifies common misconceptions about mathematical modeling in cancer. Additionally, it encourages closer cooperation between engineers, physicians and mathematicians by showing the clear benefits of this without stating unrealistic goals. Development of therapy protocols is realized from an engineering point of view, such as the search for a solution to a specific control-optimization problem. Since in the case of cancer patients, consecutive measurements providing information about the current state of the disease are not available, the control laws are derived for an open loop structure. Different forms of therapy are incorporated into the models, from chemotherapy and antiangiogenic therapy to immunotherapy and gene therapy, but the class of models introduced is broad enough to incorporate other forms of therapy as well. The book be...

  15. Agile Service Development: A Rule-Based Method Engineering Approach

    dr. Martijn Zoet; Stijn Hoppenbrouwers; Inge van de Weerd; Johan Versendaal

    2011-01-01

    Agile software development has evolved into an increasingly mature software development approach and has been applied successfully in many software vendors’ development departments. In this position paper, we address the broader agile service development. Based on method engineering principles we

  16. Enterprise Engineering Method supporting Six Sigma Approach

    Jochem, Roland

    2007-01-01

    Enterprise Modeling (EM) is currently in operation either as a technique to represent and understand the structure and behavior of the enterprise, or as a technique to analyze business processes, and in many cases as support technique for business process reengineering. However, EM architectures and methodes for Enterprise Engineering can also used to support new management techniques like SIX SIGMA, because these new techniques need a clear, transparent and integrated definition and descript...

  17. Enhancing gold recovery from electronic waste via lixiviant metabolic engineering in Chromobacterium violaceum

    Tay, Song Buck; Natarajan, Gayathri; Rahim, Muhammad Nadjad bin Abdul; Tan, Hwee Tong; Chung, Maxey Ching Ming; Ting, Yen Peng; Yew, Wen Shan

    2013-01-01

    Conventional leaching (extraction) methods for gold recovery from electronic waste involve the use of strong acids and pose considerable threat to the environment. The alternative use of bioleaching microbes for gold recovery is non-pollutive and relies on the secretion of a lixiviant or (bio)chemical such as cyanide for extraction of gold from electronic waste. However, widespread industrial use of bioleaching microbes has been constrained by the limited cyanogenic capabilities of lixiviant-producing microorganisms such as Chromobacterium violaceum. Here we show the construction of a metabolically-engineered strain of Chromobacterium violaceum that produces more (70%) cyanide lixiviant and recovers more than twice as much gold from electronic waste compared to wild-type bacteria. Comparative proteome analyses suggested the possibility of further enhancement in cyanogenesis through subsequent metabolic engineering. Our results demonstrated the utility of lixiviant metabolic engineering in the construction of enhanced bioleaching microbes for the bioleaching of precious metals from electronic waste. PMID:23868689

  18. Human Metabolic Enzymes Deficiency: A Genetic Mutation Based Approach

    Swati Chaturvedi

    2016-01-01

    Full Text Available One of the extreme challenges in biology is to ameliorate the understanding of the mechanisms which emphasize metabolic enzyme deficiency (MED and how these pretend to have influence on human health. However, it has been manifested that MED could be either inherited as inborn error of metabolism (IEM or acquired, which carries a high risk of interrupted biochemical reactions. Enzyme deficiency results in accumulation of toxic compounds that may disrupt normal organ functions and cause failure in producing crucial biological compounds and other intermediates. The MED related disorders cover widespread clinical presentations and can involve almost any organ system. To sum up the causal factors of almost all the MED-associated disorders, we decided to embark on a less traveled but nonetheless relevant direction, by focusing our attention on associated gene family products, regulation of their expression, genetic mutation, and mutation types. In addition, the review also outlines the clinical presentations as well as diagnostic and therapeutic approaches.

  19. Merged ontology for engineering design: Contrasting empirical and theoretical approaches to develop engineering ontologies

    Ahmed, Saeema; Storga, M

    2009-01-01

    to developing the ontology engineering design integrated taxonomies (EDIT) with a theoretical approach in which concepts and relations are elicited from engineering design theories ontology (DO) The limitations and advantages of each approach are discussed. The research methodology adopted is to map......This paper presents a comparison of two previous and separate efforts to develop an ontology in the engineering design domain, together with an ontology proposal from which ontologies for a specific application may be derived. The research contrasts an empirical, user-centered approach...

  20. Metabolic engineering of Escherichia coli for the production of riboflavin

    Lin, Zhenquan; Xu, Zhibo; Li, Yifan; Wang, Zhiwen; Chen, Tao; Zhao, Xueming

    2014-01-01

    Background Riboflavin (vitamin B2), the precursor of the flavin cofactors flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), is used commercially as an animal feed supplement and food colorant. E. coli is a robust host for various genetic manipulations and has been employed for efficient production of biofuels, polymers, amino acids, and bulk chemicals. Thus, the aim of this study was to understand the metabolic capacity of E. coli for the riboflavin production by modification...

  1. Engineering approach to modeling of piled systems

    Coombs, R.F.; Silva, M.A.G. da

    1980-01-01

    Available methods of analysis of piled systems subjected to dynamic excitation invade areas of mathematics usually beyond the reach of a practising engineer. A simple technique that avoids that conflict is proposed, at least for preliminary studies, and its application, compared with other methods, is shown to be satisfactory. A corrective factor for parameters currently used to represent transmitting boundaries is derived for a finite strip that models an infinite layer. The influence of internal damping on the dynamic stiffness of the layer and on radiation damping is analysed. (Author) [pt

  2. Radio systems engineering a tutorial approach

    Santos, Héctor J De Los; Ponte, Juan

    2015-01-01

    This book is intended for readers who already have knowledge of devices and circuits for radio-frequency (RF) and microwave communication and are ready to study the systems engineering-level aspects of modern radio communications systems. The authors provide a general overview of radio systems with their components, focusing on the analog parts of the system and their non-idealities. Based on the physical functionality of the various building blocks of a modern radio system, block parameters are derived, which allows the examination of their influence on the overall system performance. The dis

  3. Metabolic engineering of yeast for fermentative production of flavonoids

    Rodriguez Prado, Edith Angelica; Strucko, Tomas; Stahlhut, Steen Gustav

    2017-01-01

    Yeast Saccharomyces cerevisiae was engineered for de novo production of six different flavonoids (naringenin, liquiritigenin, kaempferol, resokaempferol, quercetin, and fisetin) directly from glucose, without supplementation of expensive intermediates. This required reconstruction of long...... demonstrates the potential of flavonoid-producing yeast cell factories....

  4. Saccharomyces cerevisiae engineered for xylose metabolism exhibits a respiratory response

    Yong-Su Jin; Jose M. Laplaza; Thomas W. Jeffries

    2004-01-01

    Native strains of Saccharomyces cerevisiae do not assimilate xylose. S. cerevisiae engineered for D-xylose utilization through the heterologous expression of genes for aldose reductase ( XYL1), xylitol dehydrogenase (XYL2), and D-xylulokinase ( XYL3 or XKS1) produce only limited amounts of ethanol in xylose medium. In recombinant S. cerevisiae expressing XYL1, XYL2,...

  5. Systems engineering: A formal approach. Part 1: System concepts

    Vanhee, K. M.

    1993-03-01

    Engineering is the scientific discipline focused on the creation of new artifacts that are supposed to be of some use to our society. Different types of artifacts require different engineering approaches. However, in all these disciplines the development of a new artifact is divided into stages. Three stages can always be recognized: Analysis, Design, and Realization. The book considers only the first two stages of the development process. It focuses on a specific type of artifacts, called discrete dynamic systems. These systems consist of active components of actors that consume and produce passive components or tokens. Three subtypes are studied in more detail: business systems (like a factory or restaurant), information systems (whether automated or not), and automated systems (systems that are controlled by an automated information system). The first subtype is studied by industrial engineers, the last by software engineers and electrical engineers, whereas the second is a battlefield for all three disciplines. The union of these disciplines is called systems engineering.

  6. An algebraic approach to modeling in software engineering

    Loegel, C.J.; Ravishankar, C.V.

    1993-09-01

    Our work couples the formalism of universal algebras with the engineering techniques of mathematical modeling to develop a new approach to the software engineering process. Our purpose in using this combination is twofold. First, abstract data types and their specification using universal algebras can be considered a common point between the practical requirements of software engineering and the formal specification of software systems. Second, mathematical modeling principles provide us with a means for effectively analyzing real-world systems. We first use modeling techniques to analyze a system and then represent the analysis using universal algebras. The rest of the software engineering process exploits properties of universal algebras that preserve the structure of our original model. This paper describes our software engineering process and our experience using it on both research and commercial systems. We need a new approach because current software engineering practices often deliver software that is difficult to develop and maintain. Formal software engineering approaches use universal algebras to describe ''computer science'' objects like abstract data types, but in practice software errors are often caused because ''real-world'' objects are improperly modeled. There is a large semantic gap between the customer's objects and abstract data types. In contrast, mathematical modeling uses engineering techniques to construct valid models for real-world systems, but these models are often implemented in an ad hoc manner. A combination of the best features of both approaches would enable software engineering to formally specify and develop software systems that better model real systems. Software engineering, like mathematical modeling, should concern itself first and foremost with understanding a real system and its behavior under given circumstances, and then with expressing this knowledge in an executable form

  7. A civil engineering approach to ideal MHD

    Jensen, V.O.

    1992-01-01

    It is well known that a magnetic field can be conceived as a medium where an isotropic compressive stress, B 2 /2μ 0 , is superimposed on a tensile stress, B 2 /μ 0 , parallel to the lines of force. When a stationary ideal MHD plasma is present in the magnetic field, the particle pressure adds to the magnetic stresses to form a combined stress tensor. Calculations of plasma equilibria based on this concept are very similar to calculations in civil engineering of static structures based on compressive, tensile, and shear stresses. Therefore the very simple physical pictures known from civil engineering when used in plasma physics provide simple physical understanding and facilitate the physical interpretation of the results. In an earlier paper the concept was used to derive and discuss the equilibrium equations for θ-, Z-, and screw pinches and the Grad-Shafranov shift in a tokamak plasma with circular cross sections of the flux surfaces. Here the concept is used to discuss the virial theorem and to obtain a simple physical interpretation of this theorem. We also reconsider the Grad-Shafranov shift in a tokamak plasma and show that a situation where all flux surfaces have circular cross sections cannot be an exact solution to the ideal MHD equations. (author) 3 refs., 3 figs

  8. Metabolic engineering of strains: from industrial-scale to lab-scale chemical production.

    Sun, Jie; Alper, Hal S

    2015-03-01

    A plethora of successful metabolic engineering case studies have been published over the past several decades. Here, we highlight a collection of microbially produced chemicals using a historical framework, starting with titers ranging from industrial scale (more than 50 g/L), to medium-scale (5-50 g/L), and lab-scale (0-5 g/L). Although engineered Escherichia coli and Saccharomyces cerevisiae emerge as prominent hosts in the literature as a result of well-developed genetic engineering tools, several novel native-producing strains are gaining attention. This review catalogs the current progress of metabolic engineering towards production of compounds such as acids, alcohols, amino acids, natural organic compounds, and others.

  9. Metabolic Engineering for Production of Biorenewable Fuels and Chemicals: Contributions of Synthetic Biology

    Laura R. Jarboe

    2010-01-01

    Full Text Available Production of fuels and chemicals through microbial fermentation of plant material is a desirable alternative to petrochemical-based production. Fermentative production of biorenewable fuels and chemicals requires the engineering of biocatalysts that can quickly and efficiently convert sugars to target products at a cost that is competitive with existing petrochemical-based processes. It is also important that biocatalysts be robust to extreme fermentation conditions, biomass-derived inhibitors, and their target products. Traditional metabolic engineering has made great advances in this area, but synthetic biology has contributed and will continue to contribute to this field, particularly with next-generation biofuels. This work reviews the use of metabolic engineering and synthetic biology in biocatalyst engineering for biorenewable fuels and chemicals production, such as ethanol, butanol, acetate, lactate, succinate, alanine, and xylitol. We also examine the existing challenges in this area and discuss strategies for improving biocatalyst tolerance to chemical inhibitors.

  10. Metabolic engineering of microorganisms for biofuels production: from bugs to synthetic biology to fuels

    Kuk Lee, Sung; Chou, Howard; Ham, Timothy S.; Soon Lee, Taek; Keasling, Jay D.

    2009-12-02

    The ability to generate microorganisms that can produce biofuels similar to petroleum-based transportation fuels would allow the use of existing engines and infrastructure and would save an enormous amount of capital required for replacing the current infrastructure to accommodate biofuels that have properties significantly different from petroleum-based fuels. Several groups have demonstrated the feasibility of manipulating microbes to produce molecules similar to petroleum-derived products, albeit at relatively low productivity (e.g. maximum butanol production is around 20 g/L). For cost-effective production of biofuels, the fuel-producing hosts and pathways must be engineered and optimized. Advances in metabolic engineering and synthetic biology will provide new tools for metabolic engineers to better understand how to rewire the cell in order to create the desired phenotypes for the production of economically viable biofuels.

  11. Metabolic engineering for the microbial production of isoprenoids: Carotenoids and isoprenoid-based biofuels

    Fu-Xing Niu

    2017-09-01

    Full Text Available Isoprenoids are the most abundant and highly diverse group of natural products. Many isoprenoids have been used for pharmaceuticals, nutraceuticals, flavors, cosmetics, food additives and biofuels. Carotenoids and isoprenoid-based biofuels are two classes of important isoprenoids. These isoprenoids have been produced microbially through metabolic engineering and synthetic biology efforts. Herein, we briefly review the engineered biosynthetic pathways in well-characterized microbial systems for the production of carotenoids and several isoprenoid-based biofuels.

  12. Biobased production of alkanes and alkenes through metabolic engineering of microorganisms

    Kang, Min Kyoung; Nielsen, Jens

    2017-01-01

    Advancement in metabolic engineering of microorganisms has enabled bio-based production of a range of chemicals, and such engineered microorganism can be used for sustainable production leading to reduced carbon dioxide emission there. One area that has attained much interest is microbial hydrocarbon biosynthesis, and in particular, alkanes and alkenes are important high-value chemicals as they can be utilized for a broad range of industrial purposes as well as ?drop-in? biofuels. Some microo...

  13. A Systems Engineering Approach to Architecture Development

    Di Pietro, David A.

    2015-01-01

    Architecture development is often conducted prior to system concept design when there is a need to determine the best-value mix of systems that works collectively in specific scenarios and time frames to accomplish a set of mission area objectives. While multiple architecture frameworks exist, they often require use of unique taxonomies and data structures. In contrast, this paper characterizes architecture development using terminology widely understood within the systems engineering community. Using a notional civil space architecture example, it employs a multi-tier framework to describe the enterprise level architecture and illustrates how results of lower tier, mission area architectures integrate into the enterprise architecture. It also presents practices for conducting effective mission area architecture studies, including establishing the trade space, developing functions and metrics, evaluating the ability of potential design solutions to meet the required functions, and expediting study execution through the use of iterative design cycles

  14. Engine Icing Data - An Analytics Approach

    Fitzgerald, Brooke A.; Flegel, Ashlie B.

    2017-01-01

    Engine icing researchers at the NASA Glenn Research Center use the Escort data acquisition system in the Propulsion Systems Laboratory (PSL) to generate and collect a tremendous amount of data every day. Currently these researchers spend countless hours processing and formatting their data, selecting important variables, and plotting relationships between variables, all by hand, generally analyzing data in a spreadsheet-style program (such as Microsoft Excel). Though spreadsheet-style analysis is familiar and intuitive to many, processing data in spreadsheets is often unreproducible and small mistakes are easily overlooked. Spreadsheet-style analysis is also time inefficient. The same formatting, processing, and plotting procedure has to be repeated for every dataset, which leads to researchers performing the same tedious data munging process over and over instead of making discoveries within their data. This paper documents a data analysis tool written in Python hosted in a Jupyter notebook that vastly simplifies the analysis process. From the file path of any folder containing time series datasets, this tool batch loads every dataset in the folder, processes the datasets in parallel, and ingests them into a widget where users can search for and interactively plot subsets of columns in a number of ways with a click of a button, easily and intuitively comparing their data and discovering interesting dynamics. Furthermore, comparing variables across data sets and integrating video data (while extremely difficult with spreadsheet-style programs) is quite simplified in this tool. This tool has also gathered interest outside the engine icing branch, and will be used by researchers across NASA Glenn Research Center. This project exemplifies the enormous benefit of automating data processing, analysis, and visualization, and will help researchers move from raw data to insight in a much smaller time frame.

  15. Engineered proteins with PUF scaffold to manipulate RNA metabolism

    Wang, Yang; Wang, Zefeng; Tanaka Hall, Traci M.

    2013-01-01

    Pumilio/fem-3 mRNA binding factor (FBF) proteins are characterized by a sequence-specific RNA-binding domain. This unique single-stranded RNA recognition module, whose sequence specificity can be reprogrammed, has been fused with functional modules to engineer protein factors with various functions. Here we summarize the advancement in developing RNA regulatory tools and opportunities for the future. PMID:23731364

  16. Discovery of Boolean metabolic networks: integer linear programming based approach.

    Qiu, Yushan; Jiang, Hao; Ching, Wai-Ki; Cheng, Xiaoqing

    2018-04-11

    Traditional drug discovery methods focused on the efficacy of drugs rather than their toxicity. However, toxicity and/or lack of efficacy are produced when unintended targets are affected in metabolic networks. Thus, identification of biological targets which can be manipulated to produce the desired effect with minimum side-effects has become an important and challenging topic. Efficient computational methods are required to identify the drug targets while incurring minimal side-effects. In this paper, we propose a graph-based computational damage model that summarizes the impact of enzymes on compounds in metabolic networks. An efficient method based on Integer Linear Programming formalism is then developed to identify the optimal enzyme-combination so as to minimize the side-effects. The identified target enzymes for known successful drugs are then verified by comparing the results with those in the existing literature. Side-effects reduction plays a crucial role in the study of drug development. A graph-based computational damage model is proposed and the theoretical analysis states the captured problem is NP-completeness. The proposed approaches can therefore contribute to the discovery of drug targets. Our developed software is available at " http://hkumath.hku.hk/~wkc/APBC2018-metabolic-network.zip ".

  17. Metabolic 'engines' of flight drive genome size reduction in birds.

    Wright, Natalie A; Gregory, T Ryan; Witt, Christopher C

    2014-03-22

    The tendency for flying organisms to possess small genomes has been interpreted as evidence of natural selection acting on the physical size of the genome. Nonetheless, the flight-genome link and its mechanistic basis have yet to be well established by comparative studies within a volant clade. Is there a particular functional aspect of flight such as brisk metabolism, lift production or maneuverability that impinges on the physical genome? We measured genome sizes, wing dimensions and heart, flight muscle and body masses from a phylogenetically diverse set of bird species. In phylogenetically controlled analyses, we found that genome size was negatively correlated with relative flight muscle size and heart index (i.e. ratio of heart to body mass), but positively correlated with body mass and wing loading. The proportional masses of the flight muscles and heart were the most important parameters explaining variation in genome size in multivariate models. Hence, the metabolic intensity of powered flight appears to have driven genome size reduction in birds.

  18. Practical approach for the study of metabolic regulation

    D.V. Macedo

    2004-05-01

    Full Text Available First year students in Physical Education must understand metabolic regulation to comprehend thewhole integration of biochemical pathways in attempt to establish the relation with exercise. Thiswhole view is not easy to learn and the task becomes even harder with the lack of time at theend of course, when normally the students think about metabolic integration. Trying to get thestudents attention to this important issue, we developed practical works beginning in the middle ofthe course, in parallel with theory classes. Blood and urine were collected for metabolite analysis ineach practice. The students were divided in groups (10 students and they created the protocols in formthat they only have been guided and directed by the teacher and monitors. The practical activitiesand biochemical analysis were: six 30m sprints with dierent recovery times (blood lactate and meanvelocities, lactate removal from muscle to blood after high intensity exercise (blood lactate, anaerobicthreshold (blood lactate and heart rate, the eect of glycogen depletion after high and moderateintensity exercises (plasma glucose and urea concentrations and low carbohydrate diet vs. normaldiet (plasma glucose and urine ketone bodies. After data collection, discussion and interpretation, thestudents presented orally each work in the same order above. Each presentation had the focus on themetabolic pathways involved in each practice. Group 1: phosphocreatine utilization and resynthesis.Group 2: anaerobic glycolysis, lactate production and removal. Group 3: transition between anaerobicglycolysis and oxidative metabolism. In attempt to promote the integration between muscle and liver-Group 4: protein catabolism after high intensity exercise with low muscular glycogen concentration(transamination, Cori Cycle and gluconeogenesis. Group 5: liver ketogenesis in low carbohydratediet. This sequence was intended to promote the comprehension of integrated metabolism. As a nalactivity, the

  19. Engineering synergy in biotechnology

    Nielsen, Jens; Fussenegger, Martin; Keasling, Jay

    2014-01-01

    In this article, the author focuses on approaches in metabolic engineering and synthetic biology for the creation of efficient cell factories, which can be bused to convert biomass and other feedstocks for the generation of chemicals. Topics discussed include development of restriction enzymes......, engineering plasmids and recyclable markers, production of 1,3-propanediol using a metabolically engineered Escherichia coli and production of isobutanol by using metabolically engineered yeast....

  20. Metabolic engineering in chemolithoautotrophic hosts for the production of fuels and chemicals.

    Nybo, S Eric; Khan, Nymul E; Woolston, Benjamin M; Curtis, Wayne R

    2015-07-01

    The ability of autotrophic organisms to fix CO2 presents an opportunity to utilize this 'greenhouse gas' as an inexpensive substrate for biochemical production. Unlike conventional heterotrophic microorganisms that consume carbohydrates and amino acids, prokaryotic chemolithoautotrophs have evolved the capacity to utilize reduced chemical compounds to fix CO2 and drive metabolic processes. The use of chemolithoautotrophic hosts as production platforms has been renewed by the prospect of metabolically engineered commodity chemicals and fuels. Efforts such as the ARPA-E electrofuels program highlight both the potential and obstacles that chemolithoautotrophic biosynthetic platforms provide. This review surveys the numerous advances that have been made in chemolithoautotrophic metabolic engineering with a focus on hydrogen oxidizing bacteria such as the model chemolithoautotrophic organism (Ralstonia), the purple photosynthetic bacteria (Rhodobacter), and anaerobic acetogens. Two alternative strategies of microbial chassis development are considered: (1) introducing or enhancing autotrophic capabilities (carbon fixation, hydrogen utilization) in model heterotrophic organisms, or (2) improving tools for pathway engineering (transformation methods, promoters, vectors etc.) in native autotrophic organisms. Unique characteristics of autotrophic growth as they relate to bioreactor design and process development are also discussed in the context of challenges and opportunities for genetic manipulation of organisms as production platforms. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  1. Engineering the fatty acid metabolic pathway in Saccharomyces cerevisiae for advanced biofuel production

    Xiaoling Tang

    2015-12-01

    Full Text Available Fatty acid-derived fuels and chemicals have attracted a great deal of attention in recent decades, due to their following properties of high compatibility to gasoline-based fuels and existing infrastructure for their direct utilization, storage and distribution. The yeast Saccharomyces cerevisiae is the ideal biofuel producing candidate, based on the wealth of available genetic information and versatile tools designed to manipulate its metabolic pathways. Engineering the fatty acid metabolic pathways in S. cerevisiae is an effective strategy to increase its fatty acid biosynthesis and provide more pathway precursors for production of targeted products. This review summarizes the recent progress in metabolic engineering of yeast cells for fatty acids and fatty acid derivatives production, including the regulation of acetyl-CoA biosynthesis, NADPH production, fatty acid elongation, and the accumulation of activated precursors of fatty acids for converting enzymes. By introducing specific enzymes in the engineered strains, a powerful platform with a scalable, controllable and economic route for advanced biofuel production has been established. Keywords: Metabolic engineering, Fatty acid biosynthesis, Fatty acid derivatives, Saccharomyces cerevisiae

  2. Improving production of ?-lactam antibiotics by Penicillium chrysogenum : Metabolic engineering based on transcriptome analysis

    Veiga, T.

    2012-01-01

    In Chapters 2-5 of this thesis, the applicability of transcriptome analysis to guide metabolic engineering strategies in P. chrysogenum is explored by investigating four cellular processes that are of potential relevance for industrial production of ?-lactam antibiotics: - Regulation of secondary

  3. Metabolic engineering of free-energy (ATP) conserving reactions in Saccharomyces cerevisiae

    De Kok, S.

    2012-01-01

    Metabolic engineering – the improvement of cellular activities by manipulation of enzymatic, transport and regulatory functions of the cell – has enabled the industrial production of a wide variety of biological molecules from renewable resources. Microbial production of fuels and chemicals thereby

  4. Microalgal bioengineering for sustainable energy development: Recent transgenesis and metabolic engineering strategies.

    Banerjee, Chiranjib; Singh, Puneet Kumar; Shukla, Pratyoosh

    2016-03-01

    Exploring the efficiency of algae to produce remarkable products can be directly benefitted by studying its mechanism at systems level. Recent advents in biotechnology like flux balance analysis (FBA), genomics and in silico proteomics minimize the wet lab exertion. It is understood that FBA predicts the metabolic products, metabolic pathways and alternative pathway to maximize the desired product, and these are key components for microalgae bio-engineering. This review encompasses recent transgenesis techniques and metabolic engineering strategies applied to different microalgae for improving different traits. Further it also throws light on RNAi and riboswitch engineering based methods which may be advantageous for high throughput microalgal research. A valid and optimally designed microalga can be developed where every engineering strategies meet each other successfully and will definitely fulfill the market needs. It is also to be noted that Omics (viz. genetic and metabolic manipulation with bioinformatics) should be integrated to develop a strain which could prove to be a futuristic solution for sustainable development for energy. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. An extensive case study of hairy-root cultures for enhanced secondary-metabolite production through metabolic-pathway engineering.

    Mehrotra, Shakti; Rahman, Laiq Ur; Kukreja, Arun Kumar

    2010-08-23

    An intrinsic improvement is taking place in the methodologies for the development of culture systems with first-rate production of plant-based molecules. The blending of HR (hairy root) cultures with ME (metabolic engineering) approaches offers new insights into, and possibilities for, improving the system productivity for known and/or novel high-value plant-derived active compounds. The introduction and expression of foreign genes in plants results in improvement of cellular activities by manipulating enzymatic, regulatory and transport function of the cell. The rational amendments in the rate-limiting steps of a biosynthetic pathway as well as inactivating the inefficient pathway(s) for by-product formation can be accomplished either through single-step engineering or through the multi-step engineering. The hierarchical control of any metabolic process can lead the engineer to apply the ME ideas and principles to any of the strata, including transcriptional, moving on to translational and enzymatic activity. The HR culture systems offer a remarkable potential for commercial production of a number of low-volume, but high-value, secondary metabolites. Taking HR as a model system, in the present review, we discuss engineering principles and perceptions to exploit secondary-metabolite pathways for the production of important bioactive compounds. We also talk about requisites and possible challenges that occur during ME, with emphasis on examples of various HR systems. Furthermore, it also highlights the utilization of global information obtained from '-omic' platforms in order to explore pathway architecture, structural and functional aspects of important enzymes and genes that can support the design of sets of engineering, resulting in the generation of wide-ranging views of DNA sequence-to-metabolite passageway networking and their control to obtain desired results.

  6. Systems engineering approach towards performance monitoring of emergency diesel generator

    Nurhayati Ramli; Lee, Y.K.

    2013-01-01

    Full-text: Systems engineering is an interdisciplinary approach and means to enable the realization of successful systems. In this study, systems engineering approach towards the performance monitoring of Emergency Diesel Generator (EDG) is presented. Performance monitoring is part and parcel of predictive maintenance where the systems and components conditions can be detected before they result into failures. In an effort to identify the proposal for addressing performance monitoring, the EDG boundary has been defined. Based on the Probabilistic Safety Analysis (PSA) results and industry operating experiences, the most critical component is identified. This paper proposed a systems engineering concept development framework towards EDG performance monitoring. The expected output of this study is that the EDG reliability can be improved by the performance monitoring alternatives through the systems engineering concept development effort. (author)

  7. Using hybrid expert system approaches for engineering applications

    Allen, R. H.; Boarnet, M. G.; Culbert, C. J.; Savely, R. T.

    1987-01-01

    In this paper, the use of hybrid expert system shells and hybrid (i.e., algorithmic and heuristic) approaches for solving engineering problems is reported. Aspects of various engineering problem domains are reviewed for a number of examples with specific applications made to recently developed prototype expert systems. Based on this prototyping experience, critical evaluations of and comparisons between commercially available tools, and some research tools, in the United States and Australia, and their underlying problem-solving paradigms are made. Characteristics of the implementation tool and the engineering domain are compared and practical software engineering issues are discussed with respect to hybrid tools and approaches. Finally, guidelines are offered with the hope that expert system development will be less time consuming, more effective, and more cost-effective than it has been in the past.

  8. Engineering systems reliability, safety, and maintenance an integrated approach

    Dhillon, B S

    2017-01-01

    Today, engineering systems are an important element of the world economy and each year billions of dollars are spent to develop, manufacture, operate, and maintain various types of engineering systems around the globe. Many of these systems are highly sophisticated and contain millions of parts. For example, a Boeing jumbo 747 is made up of approximately 4.5 million parts including fasteners. Needless to say, reliability, safety, and maintenance of systems such as this have become more important than ever before.  Global competition and other factors are forcing manufacturers to produce highly reliable, safe, and maintainable engineering products. Therefore, there is a definite need for the reliability, safety, and maintenance professionals to work closely during design and other phases. Engineering Systems Reliability, Safety, and Maintenance: An Integrated Approach eliminates the need to consult many different and diverse sources in the hunt for the information required to design better engineering syste...

  9. Genome Sequencing of Streptomyces atratus SCSIOZH16 and Activation Production of Nocardamine via Metabolic Engineering

    Yan Li

    2018-06-01

    Full Text Available The Actinomycetes are metabolically flexible microorganisms capable of producing a wide range of interesting compounds, including but by no means limited to, siderophores which have high affinity for ferric iron. In this study, we report the complete genome sequence of marine-derived Streptomyces atratus ZH16 and the activation of an embedded siderophore gene cluster via the application of metabolic engineering methods. The S. atratus ZH16 genome reveals that this strain has the potential to produce 26 categories of natural products (NPs barring the ilamycins. Our activation studies revealed S. atratus SCSIO ZH16 to be a promising source of the production of nocardamine-type (desferrioxamine compounds which are important in treating acute iron intoxication and performing ecological remediation. We conclude that metabolic engineering provides a highly effective strategy by which to discover drug-like compounds and new NPs in the genomic era.

  10. A modular approach to creating large engineered cartilage surfaces.

    Ford, Audrey C; Chui, Wan Fung; Zeng, Anne Y; Nandy, Aditya; Liebenberg, Ellen; Carraro, Carlo; Kazakia, Galateia; Alliston, Tamara; O'Connell, Grace D

    2018-01-23

    Native articular cartilage has limited capacity to repair itself from focal defects or osteoarthritis. Tissue engineering has provided a promising biological treatment strategy that is currently being evaluated in clinical trials. However, current approaches in translating these techniques to developing large engineered tissues remains a significant challenge. In this study, we present a method for developing large-scale engineered cartilage surfaces through modular fabrication. Modular Engineered Tissue Surfaces (METS) uses the well-known, but largely under-utilized self-adhesion properties of de novo tissue to create large scaffolds with nutrient channels. Compressive mechanical properties were evaluated throughout METS specimens, and the tensile mechanical strength of the bonds between attached constructs was evaluated over time. Raman spectroscopy, biochemical assays, and histology were performed to investigate matrix distribution. Results showed that by Day 14, stable connections had formed between the constructs in the METS samples. By Day 21, bonds were robust enough to form a rigid sheet and continued to increase in size and strength over time. Compressive mechanical properties and glycosaminoglycan (GAG) content of METS and individual constructs increased significantly over time. The METS technique builds on established tissue engineering accomplishments of developing constructs with GAG composition and compressive properties approaching native cartilage. This study demonstrated that modular fabrication is a viable technique for creating large-scale engineered cartilage, which can be broadly applied to many tissue engineering applications and construct geometries. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Optimisation of Multilayer Insulation an Engineering Approach

    Chorowski, M; Parente, C; Riddone, G

    2001-01-01

    A mathematical model has been developed to describe the heat flux through multilayer insulation (MLI). The total heat flux between the layers is the result of three distinct heat transfer modes: radiation, residual gas conduction and solid spacer conduction. The model describes the MLI behaviour considering a layer-to-layer approach and is based on an electrical analogy, in which the three heat transfer modes are treated as parallel thermal impedances. The values of each of the transfer mode vary from layer to layer, although the total heat flux remains constant across the whole MLI blanket. The model enables the optimisation of the insulation with regard to different MLI parameters, such as residual gas pressure, number of layers and boundary temperatures. The model has been tested with experimental measurements carried out at CERN and the results revealed to be in a good agreement, especially for insulation vacuum between 10-5 Pa and 10-3 Pa.

  12. Enhancement of Thiamin Content in Arabidopsis thaliana by Metabolic Engineering.

    Dong, Wei; Stockwell, Virginia O; Goyer, Aymeric

    2015-12-01

    Thiamin is an essential nutrient in the human diet. Severe thiamin deficiency leads to beriberi, a lethal disease which is common in developing countries. Thiamin biofortification of staple food crops is a possible strategy to alleviate thiamin deficiency-related diseases. In plants, thiamin plays a role in the response to abiotic and biotic stresses, and data from the literature suggest that boosting thiamin content could increase resistance to stresses. Here, we tested an engineering strategy to increase thiamin content in Arabidopsis. Thiamin is composed of a thiazole ring linked to a pyrimidine ring by a methylene bridge. THI1 and THIC are the first committed steps in the synthesis of the thiazole and pyrimidine moieties, respectively. Arabidopsis plants were transformed with a vector containing the THI1-coding sequence under the control of a constitutive promoter. Total thiamin leaf content in THI1 plants was up approximately 2-fold compared with the wild type. THI1-overexpressing lines were then crossed with pre-existing THIC-overexpressing lines. Resulting THI1 × THIC plants accumulated up to 3.4- and 2.6-fold more total thiamin than wild-type plants in leaf and seeds, respectively. After inoculation with Pseudomonas syringae, THI1 × THIC plants had lower populations than the wild-type control. However, THI1 × THIC plants subjected to various abiotic stresses did not show any visible or biochemical changes compared with the wild type. We discuss the impact of engineering thiamin biosynthesis on the nutritional value of plants and their resistance to biotic and abiotic stresses. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  13. Biobased production of alkanes and alkenes through metabolic engineering of microorganisms

    Kang, Min Kyoung; Nielsen, Jens

    2017-01-01

    Advancement in metabolic engineering of microorganisms has enabled bio-based production of a range of chemicals, and such engineered microorganism can be used for sustainable production leading to reduced carbon dioxide emission there. One area that has attained much interest is microbial...... hydrocarbon biosynthesis, and in particular, alkanes and alkenes are important high-value chemicals as they can be utilized for a broad range of industrial purposes as well as ‘drop-in’ biofuels. Some microorganisms have the ability to biosynthesize alkanes and alkenes naturally, but their production level...... is extremely low. Therefore, there have been various attempts to recruit other microbial cell factories for production of alkanes and alkenes by applying metabolic engineering strategies. Here we review different pathways and involved enzymes for alkane and alkene production and discuss bottlenecks...

  14. Metabolic engineering of Saccharomyces cerevisiae: a key cell factory platform for future biorefineries.

    Hong, Kuk-Ki; Nielsen, Jens

    2012-08-01

    Metabolic engineering is the enabling science of development of efficient cell factories for the production of fuels, chemicals, pharmaceuticals, and food ingredients through microbial fermentations. The yeast Saccharomyces cerevisiae is a key cell factory already used for the production of a wide range of industrial products, and here we review ongoing work, particularly in industry, on using this organism for the production of butanol, which can be used as biofuel, and isoprenoids, which can find a wide range of applications including as pharmaceuticals and as biodiesel. We also look into how engineering of yeast can lead to improved uptake of sugars that are present in biomass hydrolyzates, and hereby allow for utilization of biomass as feedstock in the production of fuels and chemicals employing S. cerevisiae. Finally, we discuss the perspectives of how technologies from systems biology and synthetic biology can be used to advance metabolic engineering of yeast.

  15. Applications of CRISPR/Cas System to Bacterial Metabolic Engineering

    Suhyung Cho

    2018-04-01

    Full Text Available The clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas adaptive immune system has been extensively used for gene editing, including gene deletion, insertion, and replacement in bacterial and eukaryotic cells owing to its simple, rapid, and efficient activities in unprecedented resolution. Furthermore, the CRISPR interference (CRISPRi system including deactivated Cas9 (dCas9 with inactivated endonuclease activity has been further investigated for regulation of the target gene transiently or constitutively, avoiding cell death by disruption of genome. This review discusses the applications of CRISPR/Cas for genome editing in various bacterial systems and their applications. In particular, CRISPR technology has been used for the production of metabolites of high industrial significance, including biochemical, biofuel, and pharmaceutical products/precursors in bacteria. Here, we focus on methods to increase the productivity and yield/titer scan by controlling metabolic flux through individual or combinatorial use of CRISPR/Cas and CRISPRi systems with introduction of synthetic pathway in industrially common bacteria including Escherichia coli. Further, we discuss additional useful applications of the CRISPR/Cas system, including its use in functional genomics.

  16. Comparison of multiple gene assembly methods for metabolic engineering

    Chenfeng Lu; Karen Mansoorabadi; Thomas Jeffries

    2007-01-01

    A universal, rapid DNA assembly method for efficient multigene plasmid construction is important for biological research and for optimizing gene expression in industrial microbes. Three different approaches to achieve this goal were evaluated. These included creating long complementary extensions using a uracil-DNA glycosylase technique, overlap extension polymerase...

  17. Recent advances in rational approaches for enzyme engineering

    Kerstin Steiner

    2012-09-01

    Full Text Available Enzymes are an attractive alternative in the asymmetric syntheses of chiral building blocks. To meet the requirements of industrial biotechnology and to introduce new functionalities, the enzymes need to be optimized by protein engineering. This article specifically reviews rational approaches for enzyme engineering and de novo enzyme design involving structure-based approaches developed in recent years for improvement of the enzymes’ performance, broadened substrate range, and creation of novel functionalities to obtain products with high added value for industrial applications.

  18. Transformation techniques for metabolic engineering of diatoms and haptophytes: current state and prospects.

    Velmurugan, Natarajan; Deka, Deepi

    2018-05-01

    Diatoms and haptophytes represent a key segment of the dominant phytoplankton communities that frequently form massive blooms in the photic zone of the ocean and are considered indicators of global climate changes. Diatoms and haptophytes also play a vital role in the biological carbon fixation in the carbon cycles. Carbon partitioning within diatoms and haptophytes possesses a wide range of chemical compounds and storage materials, such as lipids, carbohydrates, and chlorophyll. Among the marine microorganisms, diatoms and haptophytes have been recognized as promising sources of long- and very long-chain polyunsaturated fatty acids (PUFA). So far, a variety of approaches have been employed for genetic modification in the nuclei of diatoms and haptophytes. Studies on transformation and metabolic engineering in various intracellular genomes, such as chloroplast and mitochondria, are scarce. Particle bombardment, Agrobacterium and PEG-mediated gene transfer, and electroporation have been reported for foreign gene transformation into the diatoms and haptophytes. Antibiotics (G418 and chloramphenicol) and herbicides (zeocin, hygromycin, and norflurazon) have been successfully demonstrated as the best selection markers. Despite the availability of a wide range of molecular tools for foreign gene expression in microalgae, very few promoters (lhcf1, nr, h4, ef2, fcp, and pds) have been reported for diatoms and haptophytes. Therefore, in this review, we first summarize the significant progress that has been achieved in transgene expression in diatoms and haptophytes and highlight the importance and availability of recently developed novel tools that are suitable for transgenic expression in diatoms and haptophytes.

  19. Mechanisms relevant to the enhanced virulence of a dihydroxynaphthalene-melanin metabolically engineered entomopathogen.

    Min-Nan Tseng

    Full Text Available The entomopathogenic fungus Metarhizium anisopliae MA05-169 is a transformant strain that has been metabolically engineered to express dihydroxynaphthalene-melanin biosynthesis genes. In contrast to the wild type strain, the transformant displays a greater resistance to environmental stress and a higher virulence toward target insect host. However, the underlying mechanisms for these characteristics remain unclear; hence experiments were initiated to explore the possible mechanism(s through physiological and molecular approaches. Although both transformant and wild type strains could infect and share the same insect host range, the former germinated faster and produced more appressoria than the latter, both in vivo and in vitro. The transformant showed a significantly shorter median lethal time (LT50 when infecting the diamondback moth (Plutella xylostella and the striped flea beetle (Phyllotreta striolata, than the wild type. Additionally, the transformant was more tolerant to reactive oxygen species (ROS, produced 40-fold more orthosporin and notably overexpressed the transcripts of the pathogenicity-relevant hydrolytic enzymes (chitinase, protease, and phospholipase genes in vivo. In contrast, appressorium turgor pressure and destruxin A content were slightly decreased compared to the wild type. The transformant's high anti-stress tolerance, its high virulence against five important insect pests (cowpea aphid Aphis craccivora, diamondback moth Pl. xylostella, striped flea beetle Ph. striolata, and silverleaf whitefly Bemisia argentifolii and its capacity to colonize the root system are key properties for its potential bio-control field application.

  20. Metabolic Engineering of Yeast to Produce Fatty Acid-derived Biofuels: Bottlenecks and Solutions

    Jiayuan eSheng

    2015-06-01

    Full Text Available Fatty acid-derived biofuels can be a better solution than bioethanol to replace petroleum fuel, since they have similar energy content and combustion properties as current transportation fuels. The environmentally friendly microbial fermentation process has been used to synthesize advanced biofuels from renewable feedstock. Due to their robustness as well as the high tolerance to fermentation inhibitors and phage contamination, yeast strains such as Saccharomyces cerevisiae and Yarrowia lipolytica have attracted tremendous attention in recent studies regarding the production of fatty acid-derived biofuels, including fatty acids, fatty acid ethyl esters, fatty alcohols, and fatty alkanes. However, the native yeast strains cannot produce fatty acids and fatty acid-derived biofuels in large quantities. To this end, we have summarized recent publications in this review on metabolic engineering of yeast strains to improve the production of fatty acid-derived biofuels, identified the bottlenecks that limit the productivity of biofuels, and categorized the appropriate approaches to overcome these obstacles.

  1. High-density biosynthetic fuels: the intersection of heterogeneous catalysis and metabolic engineering.

    Harvey, Benjamin G; Meylemans, Heather A; Gough, Raina V; Quintana, Roxanne L; Garrison, Michael D; Bruno, Thomas J

    2014-05-28

    Biosynthetic valencene, premnaspirodiene, and natural caryophyllene were hydrogenated and evaluated as high performance fuels. The parent sesquiterpenes were then isomerized to complex mixtures of hydrocarbons with the heterogeneous acid catalyst Nafion SAC-13. High density fuels with net heats of combustion ranging from 133-141 000 Btu gal(-1), or up to 13% higher than commercial jet fuel could be generated by this approach. The products of caryophyllene isomerization were primarily tricyclic hydrocarbons which after hydrogenation increased the fuel density by 6%. The isomerization of valencene and premnaspirodiene also generated a variety of sesquiterpenes, but in both cases the dominant product was δ-selinene. Ab initio calculations were conducted to determine the total electronic energies for the reactants and products. In all cases the results were in excellent agreement with the experimental distribution of isomers. The cetane numbers for the sesquiterpane fuels ranged from 20-32 and were highly dependent on the isomer distribution. Specific distillation cuts may have the potential to act as high density diesel fuels, while use of these hydrocarbons as additives to jet fuel will increase the range and/or time of flight of aircraft. In addition to the ability to generate high performance renewable fuels, the powerful combination of metabolic engineering and heterogeneous catalysis will allow for the preparation of a variety of sesquiterpenes with potential for pharmaceutical, flavor, and fragrance applications.

  2. Integrated approach in the treatment of metabolic syndrome

    2014-03-01

    Full Text Available The Goal of this study was to investigate the efficacy of the integrated approach for the treatment of metabolic syndrome (MS aiming to correct all of its components versus standard therapy using clinical outcomes (BMI, waist circumference, blood pressure, lipid levels, assessment of psychological status (Beck Depression Inventory, and quality of life (SF-36. Methods: A total of 60 patients with MS were included in the study. The study group (30 subjects mean age 41.0±11 years, women - 23 (76.7%, men - 7 (23.3% received the complex therapy of MS - pharmacotherapy of obesity (orlistat and insulin resistance (metformin, lipid-lowering therapy (statins or fibrates, antihypertensive therapy. Control group (30 patients mean age 43.4±9.5 years, women - 26 (86.7%, men - 4 (13.3% was treated with statins or fibrates and received antihypertensive therapy when needed. At the inclusion in the study and after 6 months of therapy all patients underwent clinical and laboratory investigation, assessment of depression and quality of life. Results: We found a more significant reduction of all clinical outcomes (body weight, blood pressure, improvement in glucose and lipid metabolism, a significant decrease in the prevalence and severity of the depression, and an improvement in the quality of life in patients of study group compared with standard therapy. Conclusion: Complex treatment of the MS, including pharmacotherapy of obesity (orlistat, Xenical and insulin resistance (metformin, Glucophage is characterized by a greater clinical efficacy compared with standard therapy.

  3. CRISPR/Cas9-coupled recombineering for metabolic engineering of Corynebacterium glutamicum.

    Cho, Jae Sung; Choi, Kyeong Rok; Prabowo, Cindy Pricilia Surya; Shin, Jae Ho; Yang, Dongsoo; Jang, Jaedong; Lee, Sang Yup

    2017-07-01

    Genome engineering of Corynebacterium glutamicum, an important industrial microorganism for amino acids production, currently relies on random mutagenesis and inefficient double crossover events. Here we report a rapid genome engineering strategy to scarlessly knock out one or more genes in C. glutamicum in sequential and iterative manner. Recombinase RecT is used to incorporate synthetic single-stranded oligodeoxyribonucleotides into the genome and CRISPR/Cas9 to counter-select negative mutants. We completed the system by engineering the respective plasmids harboring CRISPR/Cas9 and RecT for efficient curing such that multiple gene targets can be done iteratively and final strains will be free of plasmids. To demonstrate the system, seven different mutants were constructed within two weeks to study the combinatorial deletion effects of three different genes on the production of γ-aminobutyric acid, an industrially relevant chemical of much interest. This genome engineering strategy will expedite metabolic engineering of C. glutamicum. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  4. Synthetic metabolic engineering-a novel, simple technology for designing a chimeric metabolic pathway

    Ye Xiaoting

    2012-09-01

    Full Text Available Abstract Background The integration of biotechnology into chemical manufacturing has been recognized as a key technology to build a sustainable society. However, the practical applications of biocatalytic chemical conversions are often restricted due to their complexities involving the unpredictability of product yield and the troublesome controls in fermentation processes. One of the possible strategies to overcome these limitations is to eliminate the use of living microorganisms and to use only enzymes involved in the metabolic pathway. Use of recombinant mesophiles producing thermophilic enzymes at high temperature results in denaturation of indigenous proteins and elimination of undesired side reactions; consequently, highly selective and stable biocatalytic modules can be readily prepared. By rationally combining those modules together, artificial synthetic pathways specialized for chemical manufacturing could be designed and constructed. Results A chimeric Embden-Meyerhof (EM pathway with balanced consumption and regeneration of ATP and ADP was constructed by using nine recombinant E. coli strains overproducing either one of the seven glycolytic enzymes of Thermus thermophilus, the cofactor-independent phosphoglycerate mutase of Pyrococcus horikoshii, or the non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase of Thermococcus kodakarensis. By coupling this pathway with the Thermus malate/lactate dehydrogenase, a stoichiometric amount of lactate was produced from glucose with an overall ATP turnover number of 31. Conclusions In this study, a novel and simple technology for flexible design of a bespoke metabolic pathway was developed. The concept has been testified via a non-ATP-forming chimeric EM pathway. We designated this technology as “synthetic metabolic engineering”. Our technology is, in principle, applicable to all thermophilic enzymes as long as they can be functionally expressed in the host, and thus would be

  5. Systems engineering approach for future automotive microcontroller solutions; Systems-Engineering-Ansatz zur Entwicklung zukuenftiger Mikrocontroller

    Hilgert, J.; Turski, K.; Vollhardt, S. [NEC Electronics Europe, Duesseldorf (Germany)

    2005-09-01

    In the future, microcontrollers used in automotive applications will have to meet escalating demands from different areas. For this reason, NEC Electronics (Europe) regards the concept of Systems Engineering as the key to handling the development of the complex system vehicle. This article describes how the Systems Engineering approach is applied to the development of new microcontrollers. The example used is the development platform for NEC's upcoming gateway product. (orig.)

  6. Metabolic engineering of oleaginous yeast Yarrowia lipolytica for limonene overproduction.

    Cao, Xuan; Lv, Yu-Bei; Chen, Jun; Imanaka, Tadayuki; Wei, Liu-Jing; Hua, Qiang

    2016-01-01

    Limonene, a monocyclic monoterpene, is known for its using as an important precursor of many flavoring, pharmaceutical, and biodiesel products. Currently, d-limonene has been produced via fractionation from essential oils or as a byproduct of orange juice production, however, considering the increasing need for limonene and a certain amount of pesticides may exist in the limonene obtained from the citrus industry, some other methods should be explored to produce limonene. To construct the limonene synthetic pathway in Yarrowia lipolytica , two genes encoding neryl diphosphate synthase 1 (NDPS1) and limonene synthase (LS) were codon-optimized and heterologously expressed in Y. lipolytica . Furthermore, to maximize limonene production, several genes involved in the MVA pathway were overexpressed, either in different copies of the same gene or in combination. Finally with the optimized pyruvic acid and dodecane concentration in flask culture, a maximum limonene titer and content of 23.56 mg/L and 1.36 mg/g DCW were achieved in the final engineered strain Po1f-LN-051, showing approximately 226-fold increase compared with the initial yield 0.006 mg/g DCW. This is the first report on limonene biosynthesis in oleaginous yeast Y. lipolytica by heterologous expression of codon-optimized tLS and tNDPS1 genes. To our knowledge, the limonene production 23.56 mg/L, is the highest limonene production level reported in yeast. In short, we demonstrate that Y. lipolytica provides a compelling platform for the overproduction of limonene derivatives, and even other monoterpenes.

  7. Ketocarotenoid Production in Soybean Seeds through Metabolic Engineering.

    Emily C Pierce

    Full Text Available The pink or red ketocarotenoids, canthaxanthin and astaxanthin, are used as feed additives in the poultry and aquaculture industries as a source of egg yolk and flesh pigmentation, as farmed animals do not have access to the carotenoid sources of their wild counterparts. Because soybean is already an important component in animal feed, production of these carotenoids in soybean could be a cost-effective means of delivery. In order to characterize the ability of soybean seed to produce carotenoids, soybean cv. Jack was transformed with the crtB gene from Pantoea ananatis, which codes for phytoene synthase, an enzyme which catalyzes the first committed step in the carotenoid pathway. The crtB gene was engineered together in combinations with ketolase genes (crtW from Brevundimonas sp. strain SD212 and bkt1 from Haematococcus pluvialis to produce ketocarotenoids; all genes were placed under the control of seed-specific promoters. HPLC results showed that canthaxanthin is present in the transgenic seeds at levels up to 52 μg/g dry weight. Transgenic seeds also accumulated other compounds in the carotenoid pathway, such as astaxanthin, lutein, β-carotene, phytoene, α-carotene, lycopene, and β-cryptoxanthin, whereas lutein was the only one of these detected in non-transgenic seeds. The accumulation of astaxanthin, which requires a β-carotene hydroxylase in addition to a β-carotene ketolase, in the transgenic seeds suggests that an endogenous soybean enzyme is able to work in combination with the ketolase transgene. Soybean seeds that accumulate ketocarotenoids could potentially be used in animal feed to reduce or eliminate the need for the costly addition of these compounds.

  8. Current progress of targetron technology: development, improvement and application in metabolic engineering.

    Liu, Ya-Jun; Zhang, Jie; Cui, Gu-Zhen; Cui, Qiu

    2015-06-01

    Targetrons are mobile group II introns that can recognize their DNA target sites by base-pairing RNA-DNA interactions with the aid of site-specific binding reverse transcriptases. Targetron technology stands out from recently developed gene targeting methods because of the flexibility, feasibility, and efficiency, and is particularly suitable for the genetic engineering of difficult microorganisms, including cellulolytic bacteria that are considered promising candidates for biomass conversion via consolidated bioprocessing. Along with the development of the thermotargetron method for thermophiles, targetron technology becomes increasingly important for the metabolic engineering of industrial microorganisms aiming at biofuel/chemical production. To summarize the current progress of targetron technology and provide new insights on the use of the technology, this paper reviews the retrohoming mechanisms of both mesophilic and thermophilic targetron methods based on various group II introns, investigates the improvement of targetron tools for high target efficiency and specificity, and discusses the current applications in the metabolic engineering for bacterial producers. Although there are still intellectual property and technical restrictions in targetron applications, we propose that targetron technology will contribute to both biochemistry research and the metabolic engineering for industrial productions. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Perspectives for Cell-homing Approaches to Engineer Dental Pulp.

    Galler, Kerstin M; Widbiller, Matthias

    2017-09-01

    Sufficient proof is available today to demonstrate that dental pulp tissue engineering is possible. The body of evidence was generated mainly on cell transplantation; however, because of several severe problems afflicted with this approach, it might not be feasible for a clinical setting in the near future. More recently, cell homing has been proposed as a viable alternative. We suggest a modification of the tissue engineering paradigm, where resident cells are attracted by endogenous, dentin-derived growth factors that further induce cell proliferation and differentiation and a bioactive scaffold material laden with these growth factors that serves as a template for tissue formation. This article highlights the latest developments regarding scaffold materials, stem cells, and dentin-derived growth factors specifically for a cell-homing approach to engineer dental pulp and summarizes new ideas. Copyright © 2017 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  10. An engineering approach to extending lifespan in C. elegans.

    Dror Sagi

    Full Text Available We have taken an engineering approach to extending the lifespan of Caenorhabditis elegans. Aging stands out as a complex trait, because events that occur in old animals are not under strong natural selection. As a result, lifespan can be lengthened rationally using bioengineering to modulate gene expression or to add exogenous components. Here, we engineered longer lifespan by expressing genes from zebrafish encoding molecular functions not normally present in worms. Additionally, we extended lifespan by increasing the activity of four endogenous worm aging pathways. Next, we used a modular approach to extend lifespan by combining components. Finally, we used cell- and worm-based assays to analyze changes in cell physiology and as a rapid means to evaluate whether multi-component transgenic lines were likely to have extended longevity. Using engineering to add novel functions and to tune endogenous functions provides a new framework for lifespan extension that goes beyond the constraints of the worm genome.

  11. Quantifying the metabolic capabilities of engineered Zymomonas mobilis using linear programming analysis

    Tsantili Ivi C

    2007-03-01

    Full Text Available Abstract Background The need for discovery of alternative, renewable, environmentally friendly energy sources and the development of cost-efficient, "clean" methods for their conversion into higher fuels becomes imperative. Ethanol, whose significance as fuel has dramatically increased in the last decade, can be produced from hexoses and pentoses through microbial fermentation. Importantly, plant biomass, if appropriately and effectively decomposed, is a potential inexpensive and highly renewable source of the hexose and pentose mixture. Recently, the engineered (to also catabolize pentoses anaerobic bacterium Zymomonas mobilis has been widely discussed among the most promising microorganisms for the microbial production of ethanol fuel. However, Z. mobilis genome having been fully sequenced in 2005, there is still a small number of published studies of its in vivo physiology and limited use of the metabolic engineering experimental and computational toolboxes to understand its metabolic pathway interconnectivity and regulation towards the optimization of its hexose and pentose fermentation into ethanol. Results In this paper, we reconstructed the metabolic network of the engineered Z. mobilis to a level that it could be modelled using the metabolic engineering methodologies. We then used linear programming (LP analysis and identified the Z. mobilis metabolic boundaries with respect to various biological objectives, these boundaries being determined only by Z. mobilis network's stoichiometric connectivity. This study revealed the essential for bacterial growth reactions and elucidated the association between the metabolic pathways, especially regarding main product and byproduct formation. More specifically, the study indicated that ethanol and biomass production depend directly on anaerobic respiration stoichiometry and activity. Thus, enhanced understanding and improved means for analyzing anaerobic respiration and redox potential in vivo are

  12. DEVELOPMENT OF MICROORGANISMS FOR CELLULOSE-BIOFUEL CONSOLIDATED BIOPROCESSINGS: METABOLIC ENGINEERS' TRICKS

    Roberto Mazzoli

    2012-10-01

    By starting from the description of natural enzyme systems for plant biomass degradation and natural metabolic pathways for some of the most valuable product (i.e. butanol, ethanol, and hydrogen biosynthesis, this review describes state-of-the-art bottlenecks and solutions for the development of recombinant microbial strains for cellulosic biofuel CBP by metabolic engineering. Complexed cellulases (i.e. cellulosomes benefit from stronger proximity effects and show enhanced synergy on insoluble substrates (i.e. crystalline cellulose with respect to free enzymes. For this reason, special attention was held on strategies involving cellulosome/designer cellulosome-bearing recombinant microorganisms.

  13. Understanding alternative fluxes/effluxes through comparative metabolic pathway analysis of phylum actinobacteria using a simplified approach.

    Verma, Mansi; Lal, Devi; Saxena, Anjali; Anand, Shailly; Kaur, Jasvinder; Kaur, Jaspreet; Lal, Rup

    2013-12-01

    Actinobacteria are known for their diverse metabolism and physiology. Some are dreadful human pathogens whereas some constitute the natural flora for human gut. Therefore, the understanding of metabolic pathways is a key feature for targeting the pathogenic bacteria without disturbing the symbiotic ones. A big challenge faced today is multiple drug resistance by Mycobacterium and other pathogens that utilize alternative fluxes/effluxes. With the availability of genome sequence, it is now feasible to conduct the comparative in silico analysis. Here we present a simplified approach to compare metabolic pathways so that the species specific enzyme may be traced and engineered for future therapeutics. The analyses of four key carbohydrate metabolic pathways, i.e., glycolysis, pyruvate metabolism, tri carboxylic acid cycle and pentose phosphate pathway suggest the presence of alternative fluxes. It was found that the upper pathway of glycolysis was highly variable in the actinobacterial genomes whereas lower glycolytic pathway was highly conserved. Likewise, pentose phosphate pathway was well conserved in contradiction to TCA cycle, which was found to be incomplete in majority of actinobacteria. The clustering based on presence and absence of genes of these metabolic pathways clearly revealed that members of different genera shared identical pathways and, therefore, provided an easy method to identify the metabolic similarities/differences between pathogenic and symbiotic organisms. The analyses could identify isoenzymes and some key enzymes that were found to be missing in some pathogenic actinobacteria. The present work defines a simple approach to explore the effluxes in four metabolic pathways within the phylum actinobacteria. The analysis clearly reflects that actinobacteria exhibit diverse routes for metabolizing substrates. The pathway comparison can help in finding the enzymes that can be used as drug targets for pathogens without effecting symbiotic organisms

  14. Metabolic Engineering toward Sustainable Production of Nylon-6.

    Turk, Stefan C H J; Kloosterman, Wigard P; Ninaber, Dennis K; Kolen, Karin P A M; Knutova, Julia; Suir, Erwin; Schürmann, Martin; Raemakers-Franken, Petronella C; Müller, Monika; de Wildeman, Stefaan M A; Raamsdonk, Leonie M; van der Pol, Ruud; Wu, Liang; Temudo, Margarida F; van der Hoeven, Rob A M; Akeroyd, Michiel; van der Stoel, Roland E; Noorman, Henk J; Bovenberg, Roel A L; Trefzer, Axel C

    2016-01-15

    Nylon-6 is a bulk polymer used for many applications. It consists of the non-natural building block 6-aminocaproic acid, the linear form of caprolactam. Via a retro-synthetic approach, two synthetic pathways were identified for the fermentative production of 6-aminocaproic acid. Both pathways require yet unreported novel biocatalytic steps. We demonstrated proof of these bioconversions by in vitro enzyme assays with a set of selected candidate proteins expressed in Escherichia coli. One of the biosynthetic pathways starts with 2-oxoglutarate and contains bioconversions of the ketoacid elongation pathway known from methanogenic archaea. This pathway was selected for implementation in E. coli and yielded 6-aminocaproic acid at levels up to 160 mg/L in lab-scale batch fermentations. The total amount of 6-aminocaproic acid and related intermediates generated by this pathway exceeded 2 g/L in lab-scale fed-batch fermentations, indicating its potential for further optimization toward large-scale sustainable production of nylon-6.

  15. Recent advances in microbial production of fuels and chemicals using tools and strategies of systems metabolic engineering

    Cho, Changhee; Choi, So Young; Luo, Zi Wei

    2015-01-01

    The advent of various systems metabolic engineering tools and strategies has enabled more sophisticated engineering of microorganisms for the production of industrially useful fuels and chemicals. Advances in systems metabolic engineering have been made in overproducing natural chemicals...... and producing novel non-natural chemicals. In this paper, we review the tools and strategies of systems metabolic engineering employed for the development of microorganisms for the production of various industrially useful chemicals belonging to fuels, building block chemicals, and specialty chemicals......, in particular focusing on those reported in the last three years. It was aimed at providing the current landscape of systems metabolic engineering and suggesting directions to address future challenges towards successfully establishing processes for the bio-based production of fuels and chemicals from renewable...

  16. Recent advances in microbial production of fuels and chemicals using tools and strategies of systems metabolic engineering.

    Cho, Changhee; Choi, So Young; Luo, Zi Wei; Lee, Sang Yup

    2015-11-15

    The advent of various systems metabolic engineering tools and strategies has enabled more sophisticated engineering of microorganisms for the production of industrially useful fuels and chemicals. Advances in systems metabolic engineering have been made in overproducing natural chemicals and producing novel non-natural chemicals. In this paper, we review the tools and strategies of systems metabolic engineering employed for the development of microorganisms for the production of various industrially useful chemicals belonging to fuels, building block chemicals, and specialty chemicals, in particular focusing on those reported in the last three years. It was aimed at providing the current landscape of systems metabolic engineering and suggesting directions to address future challenges towards successfully establishing processes for the bio-based production of fuels and chemicals from renewable resources. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Genome and metabolic engineering in non-conventional yeasts: Current advances and applications

    Ann-Kathrin Löbs

    2017-09-01

    Full Text Available Microbial production of chemicals and proteins from biomass-derived and waste sugar streams is a rapidly growing area of research and development. While the model yeast Saccharomyces cerevisiae is an excellent host for the conversion of glucose to ethanol, production of other chemicals from alternative substrates often requires extensive strain engineering. To avoid complex and intensive engineering of S. cerevisiae, other yeasts are often selected as hosts for bioprocessing based on their natural capacity to produce a desired product: for example, the efficient production and secretion of proteins, lipids, and primary metabolites that have value as commodity chemicals. Even when using yeasts with beneficial native phenotypes, metabolic engineering to increase yield, titer, and production rate is essential. The non-conventional yeasts Kluyveromyces lactis, K. marxianus, Scheffersomyces stipitis, Yarrowia lipolytica, Hansenula polymorpha and Pichia pastoris have been developed as eukaryotic hosts because of their desirable phenotypes, including thermotolerance, assimilation of diverse carbon sources, and high protein secretion. However, advanced metabolic engineering in these yeasts has been limited. This review outlines the challenges of using non-conventional yeasts for strain and pathway engineering, and discusses the developed solutions to these problems and the resulting applications in industrial biotechnology.

  18. Genome and metabolic engineering in non-conventional yeasts: Current advances and applications.

    Löbs, Ann-Kathrin; Schwartz, Cory; Wheeldon, Ian

    2017-09-01

    Microbial production of chemicals and proteins from biomass-derived and waste sugar streams is a rapidly growing area of research and development. While the model yeast Saccharomyces cerevisia e is an excellent host for the conversion of glucose to ethanol, production of other chemicals from alternative substrates often requires extensive strain engineering. To avoid complex and intensive engineering of S. cerevisiae, other yeasts are often selected as hosts for bioprocessing based on their natural capacity to produce a desired product: for example, the efficient production and secretion of proteins, lipids, and primary metabolites that have value as commodity chemicals. Even when using yeasts with beneficial native phenotypes, metabolic engineering to increase yield, titer, and production rate is essential. The non-conventional yeasts Kluyveromyces lactis, K. marxianus, Scheffersomyces stipitis, Yarrowia lipolytica, Hansenula polymorpha and Pichia pastoris have been developed as eukaryotic hosts because of their desirable phenotypes, including thermotolerance, assimilation of diverse carbon sources, and high protein secretion. However, advanced metabolic engineering in these yeasts has been limited. This review outlines the challenges of using non-conventional yeasts for strain and pathway engineering, and discusses the developed solutions to these problems and the resulting applications in industrial biotechnology.

  19. Biofuel production in Escherichia coli. The role of metabolic engineering and synthetic biology

    Clomburg, James M. [Rice Univ., Houston, TX (United States). Dept. of Chemical and Biomolecular Engineering; Gonzalez, Ramon [Rice Univ., Houston, TX (United States). Dept. of Chemical and Biomolecular Engineering; Rice Univ., Houston, TX (United States). Dept. of Bioengineering

    2010-03-15

    The microbial production of biofuels is a promising avenue for the development of viable processes for the generation of fuels from sustainable resources. In order to become cost and energy effective, these processes must utilize organisms that can be optimized to efficiently produce candidate fuels from a variety of feedstocks. Escherichia coli has become a promising host organism for the microbial production of biofuels in part due to the ease at which this organism can be manipulated. Advancements in metabolic engineering and synthetic biology have led to the ability to efficiently engineer E. coli as a biocatalyst for the production of a wide variety of potential biofuels from several biomass constituents. This review focuses on recent efforts devoted to engineering E. coli for the production of biofuels, with emphasis on the key aspects of both the utilization of a variety of substrates as well as the synthesis of several promising biofuels. Strategies for the efficient utilization of carbohydrates, carbohydrate mixtures, and noncarbohydrate carbon sources will be discussed along with engineering efforts for the exploitation of both fermentative and nonfermentative pathways for the production of candidate biofuels such as alcohols and higher carbon biofuels derived from fatty acid and isoprenoid pathways. Continued advancements in metabolic engineering and synthetic biology will help improve not only the titers, yields, and productivities of biofuels discussed herein, but also increase the potential range of compounds that can be produced. (orig.)

  20. Multiplexed genome engineering and genotyping methods applications for synthetic biology and metabolic engineering.

    Wang, Harris H; Church, George M

    2011-01-01

    Engineering at the scale of whole genomes requires fundamentally new molecular biology tools. Recent advances in recombineering using synthetic oligonucleotides enable the rapid generation of mutants at high efficiency and specificity and can be implemented at the genome scale. With these techniques, libraries of mutants can be generated, from which individuals with functionally useful phenotypes can be isolated. Furthermore, populations of cells can be evolved in situ by directed evolution using complex pools of oligonucleotides. Here, we discuss ways to utilize these multiplexed genome engineering methods, with special emphasis on experimental design and implementation. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Metabolic engineering of Saccharomyces cerevisiae for production of germacrene A, a precursor of beta-elemene

    Hu, Yating; Zhou, Yongjin J.; Bao, Jichen

    2017-01-01

    inefficient and suffers from limited natural resources. Here, we engineered a yeast cell factory for the sustainable production of germacrene A, which can be transformed to beta-elemene by a one-step chemical reaction in vitro. Two heterologous germacrene A synthases (GASs) converting farnesyl pyrophosphate...... (FPP) to germacrene A were evaluated in yeast for their ability to produce germacrene A. Thereafter, several metabolic engineering strategies were used to improve the production level. Overexpression of truncated 3-hydroxyl-3-methylglutaryl-CoA reductase and fusion of FPP synthase with GAS, led...

  2. Medicine is not health care, food is health care: plant metabolic engineering, diet and human health.

    Martin, Cathie; Li, Jie

    2017-11-01

    Contents 699 I. 699 II. 700 III. 700 IV. 706 V. 707 VI. 714 714 References 714 SUMMARY: Plants make substantial contributions to our health through our diets, providing macronutrients for energy and growth as well as essential vitamins and phytonutrients that protect us from chronic diseases. Imbalances in our food can lead to deficiency diseases or obesity and associated metabolic disorders, increased risk of cardiovascular diseases and cancer. Nutritional security is now a global challenge which can be addressed, at least in part, through plant metabolic engineering for nutritional improvement of foods that are accessible to and eaten by many. We review the progress that has been made in nutritional enhancement of foods, both improvements through breeding and through biotechnology and the engineering principles on which increased phytonutrient levels are based. We also consider the evidence, where available, that such foods do enhance health and protect against chronic diseases. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  3. Impact of synthetic biology and metabolic engineering on industrial production of fine chemicals.

    Jullesson, David; David, Florian; Pfleger, Brian; Nielsen, Jens

    2015-11-15

    Industrial bio-processes for fine chemical production are increasingly relying on cell factories developed through metabolic engineering and synthetic biology. The use of high throughput techniques and automation for the design of cell factories, and especially platform strains, has played an important role in the transition from laboratory research to industrial production. Model organisms such as Saccharomyces cerevisiae and Escherichia coli remain widely used host strains for industrial production due to their robust and desirable traits. This review describes some of the bio-based fine chemicals that have reached the market, key metabolic engineering tools that have allowed this to happen and some of the companies that are currently utilizing these technologies for developing industrial production processes. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Impact of synthetic biology and metabolic engineering on industrial production of fine chemicals

    Jullesson, David; David, Florian; Pfleger, Brian

    2015-01-01

    Industrial bio-processes for fine chemical production are increasingly relying on cell factories developed through metabolic engineering and synthetic biology. The use of high throughput techniques and automation for the design of cell factories, and especially platform strains, has played...... chemicals that have reached the market, key metabolic engineering tools that have allowed this to happen and some of the companies that are currently utilizing these technologies for developing industrial production processes....... an important role in the transition from laboratory research to industrial production. Model organisms such as Saccharomyces cerevisiae and Escherichia coli remain widely used host strains for industrial production due to their robust and desirable traits. This review describes some of the bio-based fine...

  5. Engineering yeast metabolism for production of terpenoids for use as perfume ingredients, pharmaceuticals and biofuels.

    Zhang, Yueping; Nielsen, Jens; Liu, Zihe

    2017-12-01

    Terpenoids represent a large class of natural products with significant commercial applications. These chemicals are currently mainly obtained through extraction from plants and microbes or through chemical synthesis. However, these sources often face challenges of unsustainability and low productivity. In order to address these issues, Escherichia coli and yeast have been metabolic engineered to produce non-native terpenoids. With recent reports of engineering yeast metabolism to produce several terpenoids at high yields, it has become possible to establish commercial yeast production of terpenoids that find applications as perfume ingredients, pharmaceuticals and advanced biofuels. In this review, we describe the strategies to rewire the yeast pathway for terpenoid biosynthesis. Recent advances will be discussed together with challenges and perspectives of yeast as a cell factory to produce different terpenoids. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. Engineering system dynamics a unified graph-centered approach

    Brown, Forbes T

    2006-01-01

    For today's students, learning to model the dynamics of complex systems is increasingly important across nearly all engineering disciplines. First published in 2001, Forbes T. Brown's Engineering System Dynamics: A Unified Graph-Centered Approach introduced students to a unique and highly successful approach to modeling system dynamics using bond graphs. Updated with nearly one-third new material, this second edition expands this approach to an even broader range of topics. What's New in the Second Edition? In addition to new material, this edition was restructured to build students' competence in traditional linear mathematical methods before they have gone too far into the modeling that still plays a pivotal role. New topics include magnetic circuits and motors including simulation with magnetic hysteresis; extensive new material on the modeling, analysis, and simulation of distributed-parameter systems; kinetic energy in thermodynamic systems; and Lagrangian and Hamiltonian methods. MATLAB(R) figures promi...

  7. Metabolic Engineering of the Actinomycete Amycolatopsis sp. Strain ATCC 39116 towards Enhanced Production of Natural Vanillin.

    Fleige, Christian; Meyer, Florian; Steinbüchel, Alexander

    2016-06-01

    The Gram-positive bacterium Amycolatopsis sp. ATCC 39116 is used for the fermentative production of natural vanillin from ferulic acid on an industrial scale. The strain is known for its outstanding tolerance to this toxic product. In order to improve the productivity of the fermentation process, the strain's metabolism was engineered for higher final concentrations and molar yields. Degradation of vanillin could be decreased by more than 90% through deletion of the vdh gene, which codes for the central vanillin catabolism enzyme, vanillin dehydrogenase. This mutation resulted in improvement of the final concentration of vanillin by more than 2.2 g/liter, with a molar yield of 80.9%. Further improvement was achieved with constitutive expression of the vanillin anabolism genes ech and fcs, coding for the enzymes feruloyl-coenzyme A (CoA) synthetase (fcs) and enoyl-CoA hydratase/aldolase (ech). The transcription of both genes was shown to be induced by ferulic acid, which explains the unwanted adaptation phase in the fermentation process before vanillin was efficiently produced by the wild-type cells. Through the constitutive and enhanced expression of the two genes, the adaptation phase was eliminated and a final vanillin concentration of 19.3 g/liter, with a molar yield of 94.9%, was obtained. Moreover, an even higher final vanillin concentration of 22.3 g/liter was achieved, at the expense of a lower molar yield, by using an improved feeding strategy. This is the highest reported vanillin concentration reached in microbial fermentation processes without extraction of the product. Furthermore, the vanillin was produced almost without by-products, with a molar yield that nearly approached the theoretical maximum. Much effort has been put into optimization of the biotechnological production of natural vanillin. The demand for this compound is growing due to increased consumer concerns regarding chemically produced food additives. Since this compound is toxic to most

  8. Towards Multi-Method Research Approach in Empirical Software Engineering

    Mandić, Vladimir; Markkula, Jouni; Oivo, Markku

    This paper presents results of a literature analysis on Empirical Research Approaches in Software Engineering (SE). The analysis explores reasons why traditional methods, such as statistical hypothesis testing and experiment replication are weakly utilized in the field of SE. It appears that basic assumptions and preconditions of the traditional methods are contradicting the actual situation in the SE. Furthermore, we have identified main issues that should be considered by the researcher when selecting the research approach. In virtue of reasons for weak utilization of traditional methods we propose stronger use of Multi-Method approach with Pragmatism as the philosophical standpoint.

  9. Sundanese ancient manuscripts search engine using probability approach

    Suryani, Mira; Hadi, Setiawan; Paulus, Erick; Nurma Yulita, Intan; Supriatna, Asep K.

    2017-10-01

    Today, Information and Communication Technology (ICT) has become a regular thing for every aspect of live include cultural and heritage aspect. Sundanese ancient manuscripts as Sundanese heritage are in damage condition and also the information that containing on it. So in order to preserve the information in Sundanese ancient manuscripts and make them easier to search, a search engine has been developed. The search engine must has good computing ability. In order to get the best computation in developed search engine, three types of probabilistic approaches: Bayesian Networks Model, Divergence from Randomness with PL2 distribution, and DFR-PL2F as derivative form DFR-PL2 have been compared in this study. The three probabilistic approaches supported by index of documents and three different weighting methods: term occurrence, term frequency, and TF-IDF. The experiment involved 12 Sundanese ancient manuscripts. From 12 manuscripts there are 474 distinct terms. The developed search engine tested by 50 random queries for three types of query. The experiment results showed that for the single query and multiple query, the best searching performance given by the combination of PL2F approach and TF-IDF weighting method. The performance has been evaluated using average time responds with value about 0.08 second and Mean Average Precision (MAP) about 0.33.

  10. A new approach to heart valve tissue engineering

    Kaasi, Andreas; Cestari, Idágene A.; Stolf, Noedir A G.

    2011-01-01

    The 'biomimetic' approach to tissue engineering usually involves the use of a bioreactor mimicking physiological parameters whilst supplying nutrients to the developing tissue. Here we present a new heart valve bioreactor, having as its centrepiece a ventricular assist device (VAD), which exposes...... chamber. Subsequently, applied vacuum to the pneumatic chamber causes the blood chamber to fill. A mechanical heart valve was placed in the VAD's inflow position. The tissue engineered (TE) valve was placed in the outflow position. The VAD was coupled in series with a Windkessel compliance chamber...

  11. A scientific approach to writing for engineers and scientists

    Berger, Robert E

    2014-01-01

    This book is a guide to technical writing, presented in a systematic framework that mirrors the logic associated with the scientific process itself. Other English books merely define concepts and provide rules; this one explains the reasoning behind the rules. Other writing books for scientists and engineers focus primarily on how to gather and organize materials; this one focuses primarily on how to compose a readable sentence. The approach should be satisfying not only to scientists and engineers, but also to anyone that once took a grammar course but can't remember the rules - because there was no exposure to underlying principles.

  12. Metabolic Engineering of the Actinomycete Amycolatopsis sp. Strain ATCC 39116 towards Enhanced Production of Natural Vanillin

    Fleige, Christian; Meyer, Florian; Steinbüchel, Alexander

    2016-01-01

    The Gram-positive bacterium Amycolatopsis sp. ATCC 39116 is used for the fermentative production of natural vanillin from ferulic acid on an industrial scale. The strain is known for its outstanding tolerance to this toxic product. In order to improve the productivity of the fermentation process, the strain's metabolism was engineered for higher final concentrations and molar yields. Degradation of vanillin could be decreased by more than 90% through deletion of the vdh gene, which codes for ...

  13. Increasing galactose consumption by Saccharomyces cerevisiae through metabolic engineering of the GAL gene regulatory network

    Østergaard, Simon; Olsson, Lisbeth; Johnston, M.

    2000-01-01

    Increasing the flux through central carbon metabolism is difficult because of rigidity in regulatory structures, at both the genetic and the enzymatic levels. Here we describe metabolic engineering of a regulatory network to obtain a balanced increase in the activity of all the enzymes in the pat...... media. The improved galactose consumption of the gal mutants did not favor biomass formation, but rather caused excessive respiro-fermentative metabolism, with the ethanol production rate increasing linearly with glycolytic flux....... by eliminating three known negative regulators of the GAL system: Gale, Gal80, and Mig1. This led to a 41% increase in flux through the galactose utilization pathway compared with the wild-type strain. This is of significant interest within the field of biotechnology since galactose is present in many industrial...

  14. Biosynthesis and metabolic engineering of palmitoleate production, an important contributor to human health and sustainable industry.

    Wu, Yongmei; Li, Runzhi; Hildebrand, David F

    2012-10-01

    Palmitoleate (cis-Δ9-16:1) shows numerous health benefits such as increased cell membrane fluidity, reduced inflammation, protection of the cardiovascular system, and inhibition of oncogenesis. Plant oils containing this unusual fatty acid can also be sustainable feedstocks for producing industrially important and high-demand 1-octene. Vegetable oils rich in palmitoleate are the ideal candidates for biodiesel production. Several wild plants are known that can synthesize high levels of palmitoleate in seeds. However, low yields and poor agronomic characteristics of these plants limit their commercialization. Metabolic engineering has been developed to create oilseed crops that accumulate high levels of palmitoleate or other unusual fatty acids, and significant advances have been made recently in this field, particularly using the model plant Arabidopsis as the host. The engineered targets for enhancing palmitoleate synthesis include overexpression of Δ9 desaturase from mammals, yeast, fungi, and plants, down-regulating KASII, coexpression of an ACP-Δ9 desaturase in plastids and CoA-Δ9 desaturase in endoplasmic reticulum (ER), and optimizing the metabolic flux into triacylglycerols (TAGs). This review will mainly describe the recent progress towards producing palmitoleate in transgenic plants by metabolic engineering along with our current understanding of palmitoleate biosynthesis and its regulation, as well as highlighting the bottlenecks that require additional investigation by combining lipidomics, transgenics and other "-omics" tools. A brief review of reported health benefits and non-food uses of palmitoleate will also be covered. Copyright © 2012. Published by Elsevier Ltd.

  15. Validation of RetroPath, a computer-aided design tool for metabolic pathway engineering.

    Fehér, Tamás; Planson, Anne-Gaëlle; Carbonell, Pablo; Fernández-Castané, Alfred; Grigoras, Ioana; Dariy, Ekaterina; Perret, Alain; Faulon, Jean-Loup

    2014-11-01

    Metabolic engineering has succeeded in biosynthesis of numerous commodity or high value compounds. However, the choice of pathways and enzymes used for production was many times made ad hoc, or required expert knowledge of the specific biochemical reactions. In order to rationalize the process of engineering producer strains, we developed the computer-aided design (CAD) tool RetroPath that explores and enumerates metabolic pathways connecting the endogenous metabolites of a chassis cell to the target compound. To experimentally validate our tool, we constructed 12 top-ranked enzyme combinations producing the flavonoid pinocembrin, four of which displayed significant yields. Namely, our tool queried the enzymes found in metabolic databases based on their annotated and predicted activities. Next, it ranked pathways based on the predicted efficiency of the available enzymes, the toxicity of the intermediate metabolites and the calculated maximum product flux. To implement the top-ranking pathway, our procedure narrowed down a list of nine million possible enzyme combinations to 12, a number easily assembled and tested. One round of metabolic network optimization based on RetroPath output further increased pinocembrin titers 17-fold. In total, 12 out of the 13 enzymes tested in this work displayed a relative performance that was in accordance with its predicted score. These results validate the ranking function of our CAD tool, and open the way to its utilization in the biosynthesis of novel compounds. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Learning intervention and the approach to study of engineering undergraduates

    Solomonides, Ian Paul

    The aim of the research was to: investigate the effect of a learning intervention on the Approach to Study of first year engineering degree students. The learning intervention was a local programme of learning to learn' workshops designed and facilitated by the author. The primary aim of these was to develop students' Approaches to Study. Fifty-three first year engineering undergraduates at The Nottingham Trent University participated in the workshops. Approaches to Study were quantified using data obtained from the Revised Approach to Study Inventory (RASI) which was also subjected to a validity and reliability study using local data. Quantitative outcomes were supplemented using a qualitative analysis of essays written by students during the workshops. These were analysed for detail regarding student Approach to Study. It was intended that any findings would inform the local system of Engineering Education, although more general findings also emerged, in particular in relation to the utility of the research instrument. It was concluded that the intervention did not promote the preferential Deep Approach and did not affect Approaches to Study generally as measured by the RASI. This concurred with previous attempts to change student Approaches to Study at the group level. It was also established that subsequent years of the Integrated Engineering degree course are associated with progressively deteriorating Approaches to Study. Students who were exposed to the intervention followed a similar pattern of deteriorating Approaches suggesting that the local course context and its demands had a greater influence over the Approach of students than the intervention did. It was found that academic outcomes were unrelated to the extent to which students took a Deep Approach to the local assessment demands. There appeared therefore to be a mis-match between the Approach students adopted to pass examinations and those that are required for high quality learning outcomes. It is

  17. Nuclear engineering education: A competence based approach to curricula development

    2014-01-01

    Maintaining nuclear competencies in the nuclear industry is a one of the most critical challenges in the near future. With the development of a number of nuclear engineering educational programmes in several States, this publication provides guidance to decision makers in Member States on a competence based approach to curricula development, presenting the established practices and associated requirements for educational programmes in this field. It is a consolidation of best practices that will ensure sustainable, effective nuclear engineering programmes, contributing to the safe, efficient and economic operation of nuclear power plants. The information presented is drawn from a variety of recognized nuclear engineering programmes around the world and contributes to the main areas that are needed to ensure a viable and robust nuclear industry

  18. Parallel science and engineering applications the Charm++ approach

    Kale, Laxmikant V

    2016-01-01

    Developed in the context of science and engineering applications, with each abstraction motivated by and further honed by specific application needs, Charm++ is a production-quality system that runs on almost all parallel computers available. Parallel Science and Engineering Applications: The Charm++ Approach surveys a diverse and scalable collection of science and engineering applications, most of which are used regularly on supercomputers by scientists to further their research. After a brief introduction to Charm++, the book presents several parallel CSE codes written in the Charm++ model, along with their underlying scientific and numerical formulations, explaining their parallelization strategies and parallel performance. These chapters demonstrate the versatility of Charm++ and its utility for a wide variety of applications, including molecular dynamics, cosmology, quantum chemistry, fracture simulations, agent-based simulations, and weather modeling. The book is intended for a wide audience of people i...

  19. Enabling tools for high-throughput detection of metabolites: Metabolic engineering and directed evolution applications.

    Lin, Jyun-Liang; Wagner, James M; Alper, Hal S

    2017-12-01

    Within the Design-Build-Test Cycle for strain engineering, rapid product detection and selection strategies remain challenging and limit overall throughput. Here we summarize a wide variety of modalities that transduce chemical concentrations into easily measured absorbance, luminescence, and fluorescence signals. Specifically, we cover protein-based biosensors (including transcription factors), nucleic acid-based biosensors, coupled enzyme reactions, bioorthogonal chemistry, and fluorescent and chromogenic dyes and substrates as modalities for detection. We focus on the use of these methods for strain engineering and enzyme discovery and conclude with remarks on the current and future state of biosensor development for application in the metabolic engineering field. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Metabolic engineering of Escherichia coli for limonene and perillyl alcohol production.

    Alonso-Gutierrez, Jorge; Chan, Rossana; Batth, Tanveer S; Adams, Paul D; Keasling, Jay D; Petzold, Christopher J; Lee, Taek Soon

    2013-09-01

    Limonene is a valuable monoterpene used in the production of several commodity chemicals and medicinal compounds. Among them, perillyl alcohol (POH) is a promising anti-cancer agent that can be produced by hydroxylation of limonene. We engineered E. coli with a heterologous mevalonate pathway and limonene synthase for production of limonene followed by coupling with a cytochrome P450, which specifically hydroxylates limonene to produce POH. A strain containing all mevalonate pathway genes in a single plasmid produced limonene at titers over 400mg/L from glucose, substantially higher than has been achieved in the past. Incorporation of a cytochrome P450 to hydroxylate limonene yielded approximately 100mg/L of POH. Further metabolic engineering of the pathway and in situ product recovery using anion exchange resins would make this engineered E. coli a potential production platform for any valuable limonene derivative. © 2013 Elsevier Inc. All rights reserved.

  1. Metabolic Engineering

    IAS Admin

    Considering its advantages over the other chemical synthesis routes ... such as an anti-malarial drug (artemesinin), chemicals required as the raw ... Assistant Professor at. Symbiosis ... research interests are in ... Synthetic biology, commodity.

  2. Combining metabolic and process engineering strategies to improve recombinant glycoprotein production and quality.

    Karengera, Eric; Durocher, Yves; De Crescenzo, Gregory; Henry, Olivier

    2017-11-01

    Increasing recombinant protein production while ensuring a high and consistent protein quality remains a challenge in mammalian cell culture process development. In this work, we combined a nutrient substitution approach with a metabolic engineering strategy that improves glucose utilization efficiency. This combination allowed us to tackle both lactate and ammonia accumulation and investigate on potential synergistic effects on protein production and quality. To this end, HEK293 cells overexpressing the pyruvate yeast carboxylase (PYC2) and their parental cells, both stably producing the therapeutic glycoprotein interferon α2b (IFNα2b), were cultured in media deprived of glutamine but containing chosen substitutes. Among the tested substitutes, pyruvate led to the best improvement in growth (integral of viable cell density) for both cell lines in batch cultures, whereas the culture of PYC2 cells without neither glutamine nor any substitute displayed surprisingly enhanced IFNα2b production. The drastic reduction in both lactate and ammonia in the cultures translated into extended high viability conditions and an increase in recombinant protein titer by up to 47% for the parental cells and the PYC2 cells. Product characterization performed by surface plasmon resonance biosensing using Sambucus nigra (SNA) lectin revealed that the increase in yield was however accompanied by a reduction in the degree of sialylation of the product. Supplementing cultures with glycosylation precursors and a cofactor were effective at counterbalancing the lack of glutamine and allowed improvement in IFNα2b quality as evaluated by lectin affinity. Our study provides a strategy to reconcile protein productivity and quality and highlights the advantages of PYC2-overexpressing cells in glutamine-free conditions.

  3. Flow-based approach for holistic factory engineering and design

    Constantinescu, C.; Westkämper, E.

    2010-01-01

    The engineering of future factories requires digital tools along life cycle phases from investment planning to ramp-up. Manufacturers need scientific-based integrated highly dynamic data management systems for the participative and integrated factory planning. The paper presents a new approach for the continuously integrated product design, factory and process planning, through a service-oriented architecture for the implementation of digital factory tools. A first prototype of the digital fa...

  4. ADVANCED APPROACH TO PRODUCTION WORKFLOW COMPOSITION ON ENGINEERING KNOWLEDGE PORTALS

    Novogrudska, Rina; Kot, Tatyana; Globa, Larisa; Schill, Alexander

    2016-01-01

    Background. In the environment of engineering knowledge portals great amount of partial workflows is concentrated. Such workflows are composed into general workflow aiming to perform real complex production task. Characteristics of partial workflows and general workflow structure are not studied enough, that affects the impossibility of general production workflowdynamic composition.Objective. Creating an approach to the general production workflow dynamic composition based on the partial wor...

  5. Extremely Thermophilic Microorganisms as Metabolic Engineering Platforms for Production of Fuels and Industrial Chemicals

    Benjamin M Zeldes

    2015-11-01

    Full Text Available Enzymes from extremely thermophilic microorganisms have been of technological interest for some time because of their ability to catalyze reactions of industrial significance at elevated temperatures. Thermophilic enzymes are now routinely produced in recombinant mesophilic hosts for use as discrete biocatalysts. Genome and metagenome sequence data for extreme thermophiles provide useful information for putative biocatalysts for a wide range of biotransformations, albeit involving at most a few enzymatic steps. However, in the past several years, unprecedented progress has been made in establishing molecular genetics tools for extreme thermophiles to the point that the use of these microorganisms as metabolic engineering platforms has become possible. While in its early days, complex metabolic pathways have been altered or engineered into recombinant extreme thermophiles, such that the production of fuels and chemicals at elevated temperatures has become possible. Not only does this expand the thermal range for industrial biotechnology, it also potentially provides biodiverse options for specific biotransformations unique to these microorganisms. The list of extreme thermophiles growing optimally between 70 and 100°C with genetic toolkits currently available includes archaea and bacteria, aerobes and anaerobes, coming from genera such as Caldicellulosiruptor, Sulfolobus, Thermotoga, Thermococcus and Pyrococcus. These organisms exhibit unusual and potentially useful native metabolic capabilities, including cellulose degradation, metal solubilization, and RuBisCO-free carbon fixation. Those looking to design a thermal bioprocess now have a host of potential candidates to choose from, each with its own advantages and challenges that will influence its appropriateness for specific applications. Here, the issues and opportunities for extremely thermophilic metabolic engineering platforms are considered with an eye towards potential technological

  6. Extremely thermophilic microorganisms as metabolic engineering platforms for production of fuels and industrial chemicals

    Zeldes, Benjamin M.; Keller, Matthew W.; Loder, Andrew J.; Straub, Christopher T.; Adams, Michael W. W.; Kelly, Robert M.

    2015-01-01

    Enzymes from extremely thermophilic microorganisms have been of technological interest for some time because of their ability to catalyze reactions of industrial significance at elevated temperatures. Thermophilic enzymes are now routinely produced in recombinant mesophilic hosts for use as discrete biocatalysts. Genome and metagenome sequence data for extreme thermophiles provide useful information for putative biocatalysts for a wide range of biotransformations, albeit involving at most a few enzymatic steps. However, in the past several years, unprecedented progress has been made in establishing molecular genetics tools for extreme thermophiles to the point that the use of these microorganisms as metabolic engineering platforms has become possible. While in its early days, complex metabolic pathways have been altered or engineered into recombinant extreme thermophiles, such that the production of fuels and chemicals at elevated temperatures has become possible. Not only does this expand the thermal range for industrial biotechnology, it also potentially provides biodiverse options for specific biotransformations unique to these microorganisms. The list of extreme thermophiles growing optimally between 70 and 100°C with genetic toolkits currently available includes archaea and bacteria, aerobes and anaerobes, coming from genera such as Caldicellulosiruptor, Sulfolobus, Thermotoga, Thermococcus, and Pyrococcus. These organisms exhibit unusual and potentially useful native metabolic capabilities, including cellulose degradation, metal solubilization, and RuBisCO-free carbon fixation. Those looking to design a thermal bioprocess now have a host of potential candidates to choose from, each with its own advantages and challenges that will influence its appropriateness for specific applications. Here, the issues and opportunities for extremely thermophilic metabolic engineering platforms are considered with an eye toward potential technological advantages for high

  7. A Predictive Approach to Network Reverse-Engineering

    Wiggins, Chris

    2005-03-01

    A central challenge of systems biology is the ``reverse engineering" of transcriptional networks: inferring which genes exert regulatory control over which other genes. Attempting such inference at the genomic scale has only recently become feasible, via data-intensive biological innovations such as DNA microrrays (``DNA chips") and the sequencing of whole genomes. In this talk we present a predictive approach to network reverse-engineering, in which we integrate DNA chip data and sequence data to build a model of the transcriptional network of the yeast S. cerevisiae capable of predicting the response of genes in unseen experiments. The technique can also be used to extract ``motifs,'' sequence elements which act as binding sites for regulatory proteins. We validate by a number of approaches and present comparison of theoretical prediction vs. experimental data, along with biological interpretations of the resulting model. En route, we will illustrate some basic notions in statistical learning theory (fitting vs. over-fitting; cross- validation; assessing statistical significance), highlighting ways in which physicists can make a unique contribution in data- driven approaches to reverse engineering.

  8. Dynamic optimal control of homeostasis: an integrative system approach for modeling of the central nitrogen metabolism in Saccharomyces cerevisiae.

    van Riel, N A; Giuseppin, M L; Verrips, C T

    2000-01-01

    The theory of dynamic optimal metabolic control (DOMC), as developed by Giuseppin and Van Riel (Metab. Eng., 2000), is applied to model the central nitrogen metabolism (CNM) in Saccharomyces cerevisiae. The CNM represents a typical system encountered in advanced metabolic engineering. The CNM is the source of the cellular amino acids and proteins, including flavors and potentially valuable biomolecules; therefore, it is also of industrial interest. In the DOMC approach the cell is regarded as an optimally controlled system. Given the metabolic genotype, the cell faces a control problem to maintain an optimal flux distribution in a changing environment. The regulation is based on strategies and balances feedback control of homeostasis and feedforward regulation for adaptation. The DOMC approach is an integrative, holistic approach, not based on mechanistic descriptions and (therefore) not biased by the variation present in biochemical and molecular biological data. It is an effective tool to structure the rapidly increasing amount of data on the function of genes and pathways. The DOMC model is used successfully to predict the responses of pulses of ammonia and glutamine to nitrogen-limited continuous cultures of a wild-type strain and a glutamine synthetase-negative mutant. The simulation results are validated with experimental data.

  9. Esophageal tissue engineering: a new approach for esophageal replacement.

    Totonelli, Giorgia; Maghsoudlou, Panagiotis; Fishman, Jonathan M; Orlando, Giuseppe; Ansari, Tahera; Sibbons, Paul; Birchall, Martin A; Pierro, Agostino; Eaton, Simon; De Coppi, Paolo

    2012-12-21

    A number of congenital and acquired disorders require esophageal tissue replacement. Various surgical techniques, such as gastric and colonic interposition, are standards of treatment, but frequently complicated by stenosis and other problems. Regenerative medicine approaches facilitate the use of biological constructs to replace or regenerate normal tissue function. We review the literature of esophageal tissue engineering, discuss its implications, compare the methodologies that have been employed and suggest possible directions for the future. Medline, Embase, the Cochrane Library, National Research Register and ClinicalTrials.gov databases were searched with the following search terms: stem cell and esophagus, esophageal replacement, esophageal tissue engineering, esophageal substitution. Reference lists of papers identified were also examined and experts in this field contacted for further information. All full-text articles in English of all potentially relevant abstracts were reviewed. Tissue engineering has involved acellular scaffolds that were either transplanted with the aim of being repopulated by host cells or seeded prior to transplantation. When acellular scaffolds were used to replace patch and short tubular defects they allowed epithelial and partial muscular migration whereas when employed for long tubular defects the results were poor leading to an increased rate of stenosis and mortality. Stenting has been shown as an effective means to reduce stenotic changes and promote cell migration, whilst omental wrapping to induce vascularization of the construct has an uncertain benefit. Decellularized matrices have been recently suggested as the optimal choice for scaffolds, but smart polymers that will incorporate signalling to promote cell-scaffold interaction may provide a more reproducible and available solution. Results in animal models that have used seeded scaffolds strongly suggest that seeding of both muscle and epithelial cells on scaffolds

  10. Broadening of nuclear engineering programs: An engineering physics approach at Rensselaer

    Malaviya, B.K.

    1990-01-01

    With the maturing of nuclear engineering as an academic discipline and the uncertainty surrounding the nuclear industry, attention is being increasingly turned to ways in which the base of traditional nuclear engineering programs in universities can be broadened to make them more attractively useful to a wider class of potential students and employers while maintaining the strengths in mainstream areas of nuclear technology. An approach that seems to provide a natural evolution combining the existing programmatic strengths, infrastructure, and resources with the trending needs of a broad segment of diversified industries is the development and initiation of an engineering physics degree program as an adjunct to an established nuclear engineering curriculum. In line with these developments, a new comprehensive academic program offering baccalaureate, master's, and doctoral degrees in engineering physics has been developed and formally instituted at Rensselaer Polytechnic Institute (RPI). It provides a valuable opportunity for students to pursue education and research that cuts across traditional disciplinary lines, leading to a wide variety of career opportunities in industry, government, national research and defense laboratories, and academia

  11. OptFlux: an open-source software platform for in silico metabolic engineering.

    Rocha, Isabel; Maia, Paulo; Evangelista, Pedro; Vilaça, Paulo; Soares, Simão; Pinto, José P; Nielsen, Jens; Patil, Kiran R; Ferreira, Eugénio C; Rocha, Miguel

    2010-04-19

    Over the last few years a number of methods have been proposed for the phenotype simulation of microorganisms under different environmental and genetic conditions. These have been used as the basis to support the discovery of successful genetic modifications of the microbial metabolism to address industrial goals. However, the use of these methods has been restricted to bioinformaticians or other expert researchers. The main aim of this work is, therefore, to provide a user-friendly computational tool for Metabolic Engineering applications. OptFlux is an open-source and modular software aimed at being the reference computational application in the field. It is the first tool to incorporate strain optimization tasks, i.e., the identification of Metabolic Engineering targets, using Evolutionary Algorithms/Simulated Annealing metaheuristics or the previously proposed OptKnock algorithm. It also allows the use of stoichiometric metabolic models for (i) phenotype simulation of both wild-type and mutant organisms, using the methods of Flux Balance Analysis, Minimization of Metabolic Adjustment or Regulatory on/off Minimization of Metabolic flux changes, (ii) Metabolic Flux Analysis, computing the admissible flux space given a set of measured fluxes, and (iii) pathway analysis through the calculation of Elementary Flux Modes. OptFlux also contemplates several methods for model simplification and other pre-processing operations aimed at reducing the search space for optimization algorithms. The software supports importing/exporting to several flat file formats and it is compatible with the SBML standard. OptFlux has a visualization module that allows the analysis of the model structure that is compatible with the layout information of Cell Designer, allowing the superimposition of simulation results with the model graph. The OptFlux software is freely available, together with documentation and other resources, thus bridging the gap from research in strain optimization

  12. Metabolic engineering of β-carotene in orange fruit increases its in vivo antioxidant properties.

    Pons, Elsa; Alquézar, Berta; Rodríguez, Ana; Martorell, Patricia; Genovés, Salvador; Ramón, Daniel; Rodrigo, María Jesús; Zacarías, Lorenzo; Peña, Leandro

    2014-01-01

    Orange is a major crop and an important source of health-promoting bioactive compounds. Increasing the levels of specific antioxidants in orange fruit through metabolic engineering could strengthen the fruit's health benefits. In this work, we have afforded enhancing the β-carotene content of orange fruit through blocking by RNA interference the expression of an endogenous β-carotene hydroxylase gene (Csβ-CHX) that is involved in the conversion of β-carotene into xanthophylls. Additionally, we have simultaneously overexpressed a key regulator gene of flowering transition, the FLOWERING LOCUS T from sweet orange (CsFT), in the transgenic juvenile plants, which allowed us to obtain fruit in an extremely short period of time. Silencing the Csβ-CHX gene resulted in oranges with a deep yellow ('golden') phenotype and significant increases (up to 36-fold) in β-carotene content in the pulp. The capacity of β-carotene-enriched oranges for protection against oxidative stress in vivo was assessed using Caenorhabditis elegans as experimental animal model. Golden oranges induced a 20% higher antioxidant effect than the isogenic control. This is the first example of the successful metabolic engineering of the β-carotene content (or the content of any other phytonutrient) in oranges and demonstrates the potential of genetic engineering for the nutritional enhancement of fruit tree crops. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  13. Alternative approaches to providing engineering expertise on shift

    Olson, J.; Schreiber, R.E.; Melber, B.D.

    1984-05-01

    This report represents the conclusions of a project studying the role of engineering expertise on shift in nuclear power plants. Using the present shift technical advisor (STA) position as the base case, several alternatives are analyzed. On-shift alternatives included the STA, the SS (shift supervisor), and the SE (shift engineer). The SE is degreed, experienced, trained, and licensed as a Senior Reactor Operator. Some non-shift alternatives were also studied. These included a cadre of on-call engineers and specialists within continual contact and easy reach of the plant; a technical system of phone and data lines linking the plant with a facility similar to an on-site technical support center; and finally, an SPDS (safety parameter display system) to agument technical upgrading of operator aids presently available. Potential problems considered in the analysis of implementation of these alternatives included job content constraints, problems of crew acceptance, and problems of labor supply and retention. Of the considered alternatives, the SE and SS options appear superior to the current STA approach. The SE approach appears the easiest to implement and the most effective under varied plant conditions. The SE may also serve as liaison to off-site support facilities

  14. Advances in metabolic engineering in the microbial production of fuels and chemicals from C1 gas.

    Humphreys, Christopher M; Minton, Nigel P

    2018-04-01

    The future sustainable production of chemicals and fuels from non-petrochemical sources, while at the same time reducing greenhouse gas (GHG) emissions, represent two of society's greatest challenges. Microbial chassis able to grow on waste carbon monoxide (CO) and carbon dioxide (CO 2 ) can provide solutions to both. Ranging from the anaerobic acetogens, through the aerobic chemoautotrophs to the photoautotrophic cyanobacteria, they are able to convert C1 gases into a range of chemicals and fuels which may be enhanced and extended through appropriate metabolic engineering. The necessary improvements will be facilitated by the increasingly sophisticated gene tools that are beginning to emerge as part of the Synthetic Biology revolution. These tools, in combination with more accurate metabolic and genome scale models, will enable C1 chassis to deliver their full potential. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  15. Metabolic engineering of Escherichia coli for production of mixed-acid fermentation end products

    Andreas Hartmut Förster

    2014-05-01

    Full Text Available Mixed-acid fermentation end products have numerous applications in biotechnology. This is probably the main driving force for the development of multiple strains that are supposed to produce individual end products with high yields. The process of engineering Escherichia coli strains for applied production of ethanol, lactate, succinate, or acetate was initiated several decades ago and is still ongoing. This review follows the path of strain development from the general characteristics of aerobic versus anaerobic metabolism over the regulatory machinery that enables the different metabolic routes. Thereafter, major improvements for broadening the substrate spectrum of Escherichia coli towards cheap carbon sources like molasses or lignocellulose are highlighted before major routes of strain development for the production of ethanol, acetate, lactate and succinate are presented.

  16. Approaches to Low Fuel Regression Rate in Hybrid Rocket Engines

    Dario Pastrone

    2012-01-01

    Full Text Available Hybrid rocket engines are promising propulsion systems which present appealing features such as safety, low cost, and environmental friendliness. On the other hand, certain issues hamper the development hoped for. The present paper discusses approaches addressing improvements to one of the most important among these issues: low fuel regression rate. To highlight the consequence of such an issue and to better understand the concepts proposed, fundamentals are summarized. Two approaches are presented (multiport grain and high mixture ratio which aim at reducing negative effects without enhancing regression rate. Furthermore, fuel material changes and nonconventional geometries of grain and/or injector are presented as methods to increase fuel regression rate. Although most of these approaches are still at the laboratory or concept scale, many of them are promising.

  17. Model-Based Systems Engineering Approach to Managing Mass Margin

    Chung, Seung H.; Bayer, Todd J.; Cole, Bjorn; Cooke, Brian; Dekens, Frank; Delp, Christopher; Lam, Doris

    2012-01-01

    When designing a flight system from concept through implementation, one of the fundamental systems engineering tasks ismanaging the mass margin and a mass equipment list (MEL) of the flight system. While generating a MEL and computing a mass margin is conceptually a trivial task, maintaining consistent and correct MELs and mass margins can be challenging due to the current practices of maintaining duplicate information in various forms, such as diagrams and tables, and in various media, such as files and emails. We have overcome this challenge through a model-based systems engineering (MBSE) approach within which we allow only a single-source-of-truth. In this paper we describe the modeling patternsused to capture the single-source-of-truth and the views that have been developed for the Europa Habitability Mission (EHM) project, a mission concept study, at the Jet Propulsion Laboratory (JPL).

  18. Implementing CDIO Approach in preparing engineers for Space Industry

    Daneykin Yury

    2017-01-01

    Full Text Available The necessity to train highly qualified specialists leads to the development of the trajectory that can allow training specialists for the space industry. Several steps have been undertaken to reach this purpose. First, the University founded the Space Instrument Design Center that promotes a wide range of initiatives in the sphere of educating specialists, retraining specialists, carrying out research and collaborating with profiled enterprises. The University introduced Elite Engineering Education system to attract talented specialist and help them to follow individual trajectory to train unique specialist. The paper discusses the targets necessary for achievement to train specialists. Moreover, the paper presents the compliance of the attempts with the CDIO Approach, which is widely used in leading universities to improve engineering programs.

  19. Multi-hazard approaches to civil infrastructure engineering

    LaFave, James

    2016-01-01

    This collection focuses on the development of novel approaches to address one of the most pressing challenges of civil engineering, namely the mitigation of natural hazards. Numerous engineering books to date have focused on, and illustrate considerable progress toward, mitigation of individual hazards (earthquakes, wind, and so forth.). The current volume addresses concerns related to overall safety, sustainability and resilience of the built environment when subject to multiple hazards: natural disaster events that are concurrent and either correlated (e.g., wind and surge); uncorrelated (e.g., earthquake and flood); cascading (e.g., fire following earthquake); or uncorrelated and occurring at different times (e.g., wind and earthquake). The authors examine a range of specific topics including methodologies for vulnerability assessment of structures, new techniques to reduce the system demands through control systems; instrumentation, monitoring and condition assessment of structures and foundations; new te...

  20. Systems-wide metabolic pathway engineering in Corynebacterium glutamicum for bio-based production of diaminopentane.

    Kind, Stefanie; Jeong, Weol Kyu; Schröder, Hartwig; Wittmann, Christoph

    2010-07-01

    In the present work the Gram-positive bacterium Corynebacterium glutamicum was engineered into an efficient, tailor-made production strain for diaminopentane (cadaverine), a highly attractive building block for bio-based polyamides. The engineering comprised expression of lysine decarboxylase (ldcC) from Escherichia coli, catalyzing the conversion of lysine into diaminopentane, and systems-wide metabolic engineering of central supporting pathways. Substantially re-designing the metabolism yielded superior strains with desirable properties such as (i) the release from unwanted feedback regulation at the level of aspartokinase and pyruvate carboxylase by introducing the point mutations lysC311 and pycA458, (ii) an optimized supply of the key precursor oxaloacetate by amplifying the anaplerotic enzyme, pyruvate carboxylase, and deleting phosphoenolpyruvate carboxykinase which otherwise removes oxaloacetate, (iii) enhanced biosynthetic flux via combined amplification of aspartokinase, dihydrodipicolinate reductase, diaminopimelate dehydrogenase and diaminopimelate decarboxylase, and (iv) attenuated flux into the threonine pathway competing with production by the leaky mutation hom59 in the homoserine dehydrogenase gene. Lysine decarboxylase proved to be a bottleneck for efficient production, since its in vitro activity and in vivo flux were closely correlated. To achieve an optimal strain having only stable genomic modifications, the combination of the strong constitutive C. glutamicum tuf promoter and optimized codon usage allowed efficient genome-based ldcC expression and resulted in a high diaminopentane yield of 200 mmol mol(-1). By supplementing the medium with 1 mgL(-1) pyridoxal, the cofactor of lysine decarboxylase, the yield was increased to 300 mmol mol(-1). In the production strain obtained, lysine secretion was almost completely abolished. Metabolic analysis, however, revealed substantial formation of an as yet unknown by-product. It was identified as an

  1. Synthetic Biology and Metabolic Engineering for Marine Carotenoids: New Opportunities and Future Prospects

    Chonglong Wang

    2014-09-01

    Full Text Available Carotenoids are a class of diverse pigments with important biological roles such as light capture and antioxidative activities. Many novel carotenoids have been isolated from marine organisms to date and have shown various utilizations as nutraceuticals and pharmaceuticals. In this review, we summarize the pathways and enzymes of carotenoid synthesis and discuss various modifications of marine carotenoids. The advances in metabolic engineering and synthetic biology for carotenoid production are also reviewed, in hopes that this review will promote the exploration of marine carotenoid for their utilizations.

  2. OptFlux: an open-source software platform for in silico metabolic engineering

    Rocha, I.; Maia, P.; Evangelista, P.

    2010-01-01

    to address industrial goals. However, the use of these methods has been restricted to bioinformaticians or other expert researchers. The main aim of this work is, therefore, to provide a user-friendly computational tool for Metabolic Engineering applications. Results: OptFlux is an open-source and modular...... available a number of useful tools. Its open-source nature invites contributions by all those interested in making their methods available for the community. Given its plug-in based architecture it can be extended with new functionalities. Currently, several plug-ins are being developed, including network...

  3. Simulation Modeling to Compare High-Throughput, Low-Iteration Optimization Strategies for Metabolic Engineering.

    Heinsch, Stephen C; Das, Siba R; Smanski, Michael J

    2018-01-01

    Increasing the final titer of a multi-gene metabolic pathway can be viewed as a multivariate optimization problem. While numerous multivariate optimization algorithms exist, few are specifically designed to accommodate the constraints posed by genetic engineering workflows. We present a strategy for optimizing expression levels across an arbitrary number of genes that requires few design-build-test iterations. We compare the performance of several optimization algorithms on a series of simulated expression landscapes. We show that optimal experimental design parameters depend on the degree of landscape ruggedness. This work provides a theoretical framework for designing and executing numerical optimization on multi-gene systems.

  4. Synthetic biology and metabolic engineering for marine carotenoids: new opportunities and future prospects.

    Wang, Chonglong; Kim, Jung-Hun; Kim, Seon-Won

    2014-09-17

    Carotenoids are a class of diverse pigments with important biological roles such as light capture and antioxidative activities. Many novel carotenoids have been isolated from marine organisms to date and have shown various utilizations as nutraceuticals and pharmaceuticals. In this review, we summarize the pathways and enzymes of carotenoid synthesis and discuss various modifications of marine carotenoids. The advances in metabolic engineering and synthetic biology for carotenoid production are also reviewed, in hopes that this review will promote the exploration of marine carotenoid for their utilizations.

  5. Synthetic Biology and Metabolic Engineering for Marine Carotenoids: New Opportunities and Future Prospects

    Wang, Chonglong; Kim, Jung-Hun; Kim, Seon-Won

    2014-01-01

    Carotenoids are a class of diverse pigments with important biological roles such as light capture and antioxidative activities. Many novel carotenoids have been isolated from marine organisms to date and have shown various utilizations as nutraceuticals and pharmaceuticals. In this review, we summarize the pathways and enzymes of carotenoid synthesis and discuss various modifications of marine carotenoids. The advances in metabolic engineering and synthetic biology for carotenoid production are also reviewed, in hopes that this review will promote the exploration of marine carotenoid for their utilizations. PMID:25233369

  6. Modeling with a view to target identification in metabolic engineering: a critical evaluation of the available tools.

    Maertens, Jo; Vanrolleghem, Peter A

    2010-01-01

    The state of the art tools for modeling metabolism, typically used in the domain of metabolic engineering, were reviewed. The tools considered are stoichiometric network analysis (elementary modes and extreme pathways), stoichiometric modeling (metabolic flux analysis, flux balance analysis, and carbon modeling), mechanistic and approximative modeling, cybernetic modeling, and multivariate statistics. In the context of metabolic engineering, one should be aware that the usefulness of these tools to optimize microbial metabolism for overproducing a target compound depends predominantly on the characteristic properties of that compound. Because of their shortcomings not all tools are suitable for every kind of optimization; issues like the dependence of the target compound's synthesis on severe (redox) constraints, the characteristics of its formation pathway, and the achievable/desired flux towards the target compound should play a role when choosing the optimization strategy.

  7. Introduction and expression of genes for metabolic engineering applications in Saccharomyces cerevisiae.

    Da Silva, Nancy A; Srikrishnan, Sneha

    2012-03-01

    Metabolic pathway engineering in the yeast Saccharomyces cerevisiae leads to improved production of a wide range of compounds, ranging from ethanol (from biomass) to natural products such as sesquiterpenes. The introduction of multienzyme pathways requires precise control over the level and timing of expression of the associated genes. Gene number and promoter strength/regulation are two critical control points, and multiple studies have focused on modulating these in yeast. This MiniReview focuses on methods for introducing genes and controlling their copy number and on the many promoters (both constitutive and inducible) that have been successfully employed. The advantages and disadvantages of the methods will be presented, and applications to pathway engineering will be highlighted. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  8. Advances in metabolic engineering of yeast Saccharomyces cerevisiae for production of chemicals.

    Borodina, Irina; Nielsen, Jens

    2014-05-01

    Yeast Saccharomyces cerevisiae is an important industrial host for production of enzymes, pharmaceutical and nutraceutical ingredients and recently also commodity chemicals and biofuels. Here, we review the advances in modeling and synthetic biology tools and how these tools can speed up the development of yeast cell factories. We also present an overview of metabolic engineering strategies for developing yeast strains for production of polymer monomers: lactic, succinic, and cis,cis-muconic acids. S. cerevisiae has already firmly established itself as a cell factory in industrial biotechnology and the advances in yeast strain engineering will stimulate development of novel yeast-based processes for chemicals production. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Enhancement of Naringenin Biosynthesis from Tyrosine by Metabolic Engineering of Saccharomyces cerevisiae.

    Lyu, Xiaomei; Ng, Kuan Rei; Lee, Jie Lin; Mark, Rita; Chen, Wei Ning

    2017-08-09

    Flavonoids are an important class of plant polyphenols that possess a variety of health benefits. In this work, S. cerevisiae was metabolically engineered to produce the flavonoid naringenin, using tyrosine as the precursor. Our strategy to improve naringenin production comprised three modules. In module 1, we employed a modified GAL system to overexpress the genes of the naringenin biosynthesis pathway and investigated their synergistic action. In module 2, we simultaneously up-regulated acetyl-CoA production and down-regulated fatty acid biosynthesis in order to increase the precursor supply, malonyl-CoA. In module 3, we engineered the tyrosine biosynthetic pathway to eliminate the feedback inhibition of tyrosine and also down-regulated competing pathways. It was found that modules 1 and 3 played important roles in improving naringenin production. We succeeded in producing up to ∼90 mg/L of naringenin in our final strain, which is a 20-fold increase as compared to the parental strain.

  10. Engineering approach to model and compute electric power markets settlements

    Kumar, J.; Petrov, V.

    2006-01-01

    Back-office accounting settlement activities are an important part of market operations in Independent System Operator (ISO) organizations. A potential way to measure ISO market design correctness is to analyze how well market price signals create incentives or penalties for creating an efficient market to achieve market design goals. Market settlement rules are an important tool for implementing price signals which are fed back to participants via the settlement activities of the ISO. ISO's are currently faced with the challenge of high volumes of data resulting from the increasing size of markets and ever-changing market designs, as well as the growing complexity of wholesale energy settlement business rules. This paper analyzed the problem and presented a practical engineering solution using an approach based on mathematical formulation and modeling of large scale calculations. The paper also presented critical comments on various differences in settlement design approaches to electrical power market design, as well as further areas of development. The paper provided a brief introduction to the wholesale energy market settlement systems and discussed problem formulation. An actual settlement implementation framework and discussion of the results and conclusions were also presented. It was concluded that a proper engineering approach to this domain can yield satisfying results by formalizing wholesale energy settlements. Significant improvements were observed in the initial preparation phase, scoping and effort estimation, implementation and testing. 5 refs., 2 figs

  11. Approaches to Improve Mixing in Compression Ignition Engines

    Boot, M.D.

    2010-04-20

    This thesis presents three approaches to suppress soot emissions in compression ignition (CI) engines. First, a fuel chemistry approach is proposed. A particular class of fuels - cyclic oxygenates - is identified which is capable of significantly reducing engine-out soot emissions. By means of experiments in 'closed' and optical engines, as well as on an industrial burner, two possible mechanisms are identified that could account for the observed reduction in soot: a) an extended ignition delay (ID) and b) a longer flame lift-length (FLoL). Further analysis of the available data suggests that both mechanisms are related to the inherently low reactivity of the fuel class in question. These findings are largely in line with data found in literature. In the second approach, it is attempted to reduce soot by adopting an alternative combustion concept: early direct injection premixed charge compression ignition (EDI PCCI). In this concept, fuel is injected relatively early in the compression stroke instead of conventional, close to top-dead-center (TDC), injection schemes. While the goal of soot reduction can indeed be achieved via this approach, an important drawback must be addressed before this concept can be considered practically viable. Due to the fact that combustion chamber temperature and pressure is relatively low early in the compression stroke, fuel impingement against the cylinder liner (wall-wetting) often occurs. Consequently, high levels of unburned hydrocarbons (UHC), oil dilution and poor efficiency are observed. Several strategies, combining a limited engine modification with dedicated air management and fueling settings, are investigated to tackle this drawback. All of these strategies, and especially their combination, resulted in significantly lower UHC emissions and improved fuel economy. Although UHC emissions are typically a tell-tale sign of wall-wetting, as mentioned earlier, the relation between these two has long been hypothetical

  12. An Event-driven, Value-based, Pull Systems Engineering Scheduling Approach

    2012-03-01

    combining a services approach to systems engineering with a kanban -based scheduling system. It provides the basis for validating the approach with...agent-based simulations. Keywords-systems engineering; systems engineering process; lean; kanban ; process simulation I. INTRODUCTION AND BACKGROUND...approaches [8], [9], we are investigating the use of flow-based pull scheduling techniques ( kanban systems) in a rapid response development

  13. Premigratory fat metabolism in hummingbirds: A rumsfeldian approach

    Raul K. SUAREZ

    2013-06-01

    Full Text Available Hummingbird migration is a remarkable feat, given the small body sizes of migratory species, their high metabolic rates during flight and the long distances traveled using fat to fuel the effort. Equally remarkable is the ability of premigratory hummingbirds in the wild to accumulate fat, synthesized from sugar, at rates as high as 10% of body mass per day. This paper summarizes, using Rumsfeldian terminology, “known knowns” concerning the energetics of hummingbird migration and premigratory fattening. Energy metabolism during hover-feeding on floral nectar is fueled directly by dietary sugar through the pathway recently named the “sugar oxidation cascade”. However, flight without feeding for more than a few minutes requires shifting to fat as a fuel. It is proposed that behavior and metabolic fuel choice are coadapted to maximize the rate of fat deposition during premigratory fattening. The hummingbird liver appears to possess extraordinarily high capacities for fatty acid synthesis. The analysis of “known knowns” leads to identification of “known unknowns”, e.g., the fates of dietary glucose and fructose, the regulation of fat metabolism and metabolic interactions between liver and adipose tissue. The history of science behooves recognition of “unknown unknowns” that, when discovered serendipitously, might shed new light on fundamental mechanisms as well as human pathological conditions [Current Zoology 59 (3: 371–380, 2013].

  14. Process support for Opticam: a concurrent engineering approach

    Czajkowski, Walter C.; Tipps, Joe D., Jr.

    1992-12-01

    Although the principles of concurrent engineering and rapid product cycles are not new concepts in the industrial sector, the optics manufacturing industry has witnessed few technological advances since the 1940's. At present the optics industry maintains outdated stand-alone manufacturing equipment and systems that do little to foster integration or communications. 'Islands of Technology', spawned from the latest offerings of CNC controlled equipment, are generally stand alone systems incapable of supporting communication with other process equipment, not to mention the total business enterprise. This approach increases the cost in design and manufacture of optical systems while negatively impacting competitiveness in the global marketplace.

  15. Sustainable solid waste management a systems engineering approach

    Chang, N

    2015-01-01

    Interactions between human activities and the environment are complicated and often difficult to quantify. In many occasions, judging where the optimal balance should lie among environmental protection, social well-being, economic growth, and technological progress is complex. The use of a systems engineering approach will fill in the gap contributing to how we understand the intricacy by a holistic way and how we generate better sustainable solid waste management practices. This book aims to advance interdisciplinary understanding of intertwined facets between policy and technology relevant to solid waste management issues interrelated to climate change, land use, economic growth, environmental pollution, industrial ecology, and population dynamics.

  16. Energy systems a new approach to engineering thermodynamics

    Gicquel, Renaud

    2011-01-01

    Forewords, About the Author, General introduction, Structure of the book, Objectives, A working tool on many levels, Mind Maps, List of Symbols, Conversion FactorsI First Steps in Engineering Thermodynamics1 A New Educational Paradigm1.1 Introduction1.2 General remarks on the evolution of training specifi cations1.3 Specifi cs of applied thermodynamics teaching1.4 A new educational paradigm1.5 Diapason modules1.6 A three-step progressive approach1.7 Main pedagogic innovations brought by Thermoptim1.8 Digital resources of the Thermoptim-UNIT portal1.9 Comparison with other tools with teaching p

  17. Advanced Imaging Approaches to Characterize Stromal and Metabolic Changes in In Vivo Mammary Tumor Models

    2015-02-01

    Bird , L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, N. Ramanujam, Metabolic mapping of MCF10A human breast cells...1   Award Number: W81XWH-12-1-0025 TITLE: Advanced Imaging Approaches to Characterize Stromal and Metabolic Changes in In Vivo Mammary... Metabolic Changes in In Vivo Mammary Tumor Models 5b. GRANT NUMBER BC112240 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Betty Diamond 5d. PROJECT NUMBER

  18. Production of L-lactic acid from metabolically engineered strain of Enterobacter aerogenes ATCC 29007.

    Thapa, Laxmi Prasad; Lee, Sang Jun; Park, Chulhwan; Kim, Seung Wook

    2017-07-01

    In this study, L-lactic acid production was investigated from metabolically engineered strain of E. aerogenes ATCC 29007. The engineered strain E. aerogenes SUMI01 (Δpta) was generated by the deletion of phosphate acetyltransferase (pta) gene from the chromosome of E. aerogenes ATCC 29007 and deletion was confirmed by colony PCR. Under the optimized fermentation conditions, at 37°C and pH 6 for 84h, the L-lactic acid produced by engineered strain E. aerogenes SUMI01 (Δpta) in flask fermentation using 100g/L mannitol as the carbon source was 40.05g/L as compared to that of the wild type counterpart 20.70g/L. At the end of the batch fermentation in bioreactor the production of L-lactic acid reached to 46.02g/L and yield was 0.41g/g by utilizing 112.32g/L mannitol. This is the first report regarding the production of L-lactic acid from Enterobacter species. We believe that this result may provide valuable guidelines for further engineering Enterobacter strain for the improvement of L-lactic acid production. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Metabolic Network Discovery by Top-Down and Bottom-Up Approaches and Paths for Reconciliation

    Çakır, Tunahan, E-mail: tcakir@gyte.edu.tr [Computational Systems Biology Group, Department of Bioengineering, Gebze Technical University (formerly known as Gebze Institute of Technology), Gebze (Turkey); Khatibipour, Mohammad Jafar [Computational Systems Biology Group, Department of Bioengineering, Gebze Technical University (formerly known as Gebze Institute of Technology), Gebze (Turkey); Department of Chemical Engineering, Gebze Technical University (formerly known as Gebze Institute of Technology), Gebze (Turkey)

    2014-12-03

    The primary focus in the network-centric analysis of cellular metabolism by systems biology approaches is to identify the active metabolic network for the condition of interest. Two major approaches are available for the discovery of the condition-specific metabolic networks. One approach starts from genome-scale metabolic networks, which cover all possible reactions known to occur in the related organism in a condition-independent manner, and applies methods such as the optimization-based Flux-Balance Analysis to elucidate the active network. The other approach starts from the condition-specific metabolome data, and processes the data with statistical or optimization-based methods to extract information content of the data such that the active network is inferred. These approaches, termed bottom-up and top-down, respectively, are currently employed independently. However, considering that both approaches have the same goal, they can both benefit from each other paving the way for the novel integrative analysis methods of metabolome data- and flux-analysis approaches in the post-genomic era. This study reviews the strengths of constraint-based analysis and network inference methods reported in the metabolic systems biology field; then elaborates on the potential paths to reconcile the two approaches to shed better light on how the metabolism functions.

  20. Metabolic Network Discovery by Top-Down and Bottom-Up Approaches and Paths for Reconciliation

    Çakır, Tunahan; Khatibipour, Mohammad Jafar

    2014-01-01

    The primary focus in the network-centric analysis of cellular metabolism by systems biology approaches is to identify the active metabolic network for the condition of interest. Two major approaches are available for the discovery of the condition-specific metabolic networks. One approach starts from genome-scale metabolic networks, which cover all possible reactions known to occur in the related organism in a condition-independent manner, and applies methods such as the optimization-based Flux-Balance Analysis to elucidate the active network. The other approach starts from the condition-specific metabolome data, and processes the data with statistical or optimization-based methods to extract information content of the data such that the active network is inferred. These approaches, termed bottom-up and top-down, respectively, are currently employed independently. However, considering that both approaches have the same goal, they can both benefit from each other paving the way for the novel integrative analysis methods of metabolome data- and flux-analysis approaches in the post-genomic era. This study reviews the strengths of constraint-based analysis and network inference methods reported in the metabolic systems biology field; then elaborates on the potential paths to reconcile the two approaches to shed better light on how the metabolism functions.

  1. Esophageal tissue engineering: A new approach for esophageal replacement

    Giorgia Totonelli; Panagiotis Maghsoudlou; Jonathan M Fishman; Giuseppe Orlando; Tahera Ansari; Paul Sibbons; Martin A Birchall

    2012-01-01

    A number of congenital and acquired disorders require esophageal tissue replacement.Various surgical techniques,such as gastric and colonic interposition,are standards of treatment,but frequently complicated by stenosis and other problems.Regenerative medicine approaches facilitate the use of biological constructs to replace or regenerate normal tissue function.We review the literature of esophageal tissue engineering,discuss its implications,compare the methodologies that have been employed and suggest possible directions for the future.Medline,Embase,the Cochrane Library,National Research Register and ClinicalTrials.gov databases were searched with the following search terms:stem cell and esophagus,esophageal replacement,esophageal tissue engineering,esophageal substitution.Reference lists of papers identified were also examined and experts in this field contacted for further information.All full-text articles in English of all potentially relevant abstracts were reviewed.Tissue engineering has involved acellular scaffolds that were either transplanted with the aim of being repopulated by host cells or seeded prior to transplantation.When acellular scaffolds were used to replace patch and short tubular defects they allowed epithelial and partial muscular migration whereas when employed for long tubular defects the results were poor leading to an increased rate of stenosis and mortality.Stenting has been shown as an effective means to reduce stenotic changes and promote cell migration,whilst omental wrapping to induce vascularization of the construct has an uncertain benefit.Decellularized matrices have been recently suggested as the optimal choice for scaffolds,but smart polymers that will incorporate signalling to promote cell-scaffold interaction may provide a more reproducible and available solution.Results in animal models that have used seeded scaffolds strongly suggest that seeding of both muscle and epithelial cells on scaffolds prior to implantation is a

  2. Biomimetic Materials and Fabrication Approaches for Bone Tissue Engineering.

    Kim, Hwan D; Amirthalingam, Sivashanmugam; Kim, Seunghyun L; Lee, Seunghun S; Rangasamy, Jayakumar; Hwang, Nathaniel S

    2017-12-01

    Various strategies have been explored to overcome critically sized bone defects via bone tissue engineering approaches that incorporate biomimetic scaffolds. Biomimetic scaffolds may provide a novel platform for phenotypically stable tissue formation and stem cell differentiation. In recent years, osteoinductive and inorganic biomimetic scaffold materials have been optimized to offer an osteo-friendly microenvironment for the osteogenic commitment of stem cells. Furthermore, scaffold structures with a microarchitecture design similar to native bone tissue are necessary for successful bone tissue regeneration. For this reason, various methods for fabricating 3D porous structures have been developed. Innovative techniques, such as 3D printing methods, are currently being utilized for optimal host stem cell infiltration, vascularization, nutrient transfer, and stem cell differentiation. In this progress report, biomimetic materials and fabrication approaches that are currently being utilized for biomimetic scaffold design are reviewed. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Molecular basis of glyphosate resistance: Different approaches through protein engineering

    Pollegioni, Loredano; Schonbrunn, Ernst; Siehl, Daniel

    2011-01-01

    Glyphosate (N-phosphonomethyl-glycine) is the most-used herbicide in the world: glyphosate-based formulations exhibit broad-spectrum herbicidal activity with minimal human and environmental toxicity. The extraordinary success of this simple small molecule is mainly due to the high specificity of glyphosate towards the plant enzyme enolpyruvylshikimate-3-phosphate synthase in the shikimate pathway leading to biosynthesis of aromatic amino acids. Starting in 1996, transgenic glyphosate-resistant plants were introduced thus allowing the application of the herbicide to the crop (post-emergence) to remove emerged weeds without crop damage. This review focuses on the evolution of mechanisms of resistance to glyphosate as obtained through natural diversity, the gene shuffling approach to molecular evolution, and a rational, structure-based approach to protein engineering. In addition, we offer rationale for the means by which the modifications made have had their intended effect. PMID:21668647

  4. Mass spectrometry characterisation of fatty acids from metabolically engineered soybean seeds.

    Murad, André M; Vianna, Giovanni R; Machado, Alex M; da Cunha, Nicolau B; Coelho, Cíntia M; Lacerda, Valquiria A M; Coelho, Marly C; Rech, Elibio L

    2014-05-01

    Improving the quality and performance of soybean oil as biodiesel depends on the chemical composition of its fatty acids and requires an increase in monounsaturated acids and a reduction in polyunsaturated acids. Despite its current use as a source of biofuel, soybean oil contains an average of 25 % oleic acid and 13 % palmitic acid, which negatively impacts its oxidative stability and freezing point, causing a high rate of nitrogen oxide emission. Gas chromatography and ion mobility mass spectrometry were conducted on soybean fatty acids from metabolically engineered seed extracts to determine the nature of the structural oleic and palmitic acids. The soybean genes FAD2-1 and FatB were placed under the control of the 35SCaMV constitutive promoter, introduced to soybean embryonic axes by particle bombardment and down-regulated using RNA interference technology. Results indicate that the metabolically engineered plants exhibited a significant increase in oleic acid (up to 94.58 %) and a reduction in palmitic acid (to seed oil content. No structural differences were observed between the fatty acids of the transgenic and non-transgenic oil extracts.

  5. Computational metabolic engineering strategies for growth-coupled biofuel production by Synechocystis

    Kiyan Shabestary

    2016-12-01

    Full Text Available Chemical and fuel production by photosynthetic cyanobacteria is a promising technology but to date has not reached competitive rates and titers. Genome-scale metabolic modeling can reveal limitations in cyanobacteria metabolism and guide genetic engineering strategies to increase chemical production. Here, we used constraint-based modeling and optimization algorithms on a genome-scale model of Synechocystis PCC6803 to find ways to improve productivity of fermentative, fatty-acid, and terpene-derived fuels. OptGene and MOMA were used to find heuristics for knockout strategies that could increase biofuel productivity. OptKnock was used to find a set of knockouts that led to coupling between biofuel and growth. Our results show that high productivity of fermentation or reversed beta-oxidation derived alcohols such as 1-butanol requires elimination of NADH sinks, while terpenes and fatty-acid based fuels require creating imbalances in intracellular ATP and NADPH production and consumption. The FBA-predicted productivities of these fuels are at least 10-fold higher than those reported so far in the literature. We also discuss the physiological and practical feasibility of implementing these knockouts. This work gives insight into how cyanobacteria could be engineered to reach competitive biofuel productivities. Keywords: Cyanobacteria, Modeling, Flux balance analysis, Biofuel, MOMA, OptFlux, OptKnock

  6. Efficient odd straight medium chain free fatty acid production by metabolically engineered Escherichia coli.

    Wu, Hui; San, Ka-Yiu

    2014-11-01

    Free fatty acids (FFAs) can be used as precursors for the production of biofuels or chemicals. Different composition of FFAs will be useful for further modification of the biofuel/biochemical quality. Microbial biosynthesis of even chain FFAs can be achieved by introducing an acyl-acyl carrier protein thioesterase gene into E. coli. In this study, odd straight medium chain FFAs production was investigated by using metabolic engineered E. coli carrying acyl-ACP thioesterase (TE, Ricinus communis), propionyl-CoA synthase (Salmonella enterica), and β-ketoacyl-acyl carrier protein synthase III (four different sources) with supplement of extracellular propionate. By using these metabolically engineered E. coli, significant quantity of C13 and C15 odd straight-chain FFAs could be produced from glucose and propionate. The highest concentration of total odd straight chain FFAs attained was 1205 mg/L by the strain HWK201 (pXZ18, pBHE2), and 85% of the odd straight chain FFAs was C15. However, the highest percentage of odd straight chain FFAs was achieved by the strain HWK201 (pXZ18, pBHE3) of 83.2% at 48 h. This strategy was also applied successfully in strains carrying different TE, such as the medium length acyl-ACP thioesterase gene from Umbellularia californica. C11 and C13 became the major odd straight-chain FFAs. © 2014 Wiley Periodicals, Inc.

  7. Weedy lignocellulosic feedstock and microbial metabolic engineering. Advancing the generation of 'Biofuel'

    Chandel, Anuj K. [Jawaharlal Nehru Technological Univ., Hyderabad (India). Centre of Biotechnology; Singh, Om V. [Pittsburgh Univ., Bradford, PA (United States). Div. of Biological and Health Sciences

    2011-03-15

    Lignocellulosic materials are the most abundant renewable organic resources ({proportional_to}200 billion tons annually) on earth that are readily available for conversion to ethanol and other value-added products, but they have not yet been tapped for the commercial production of fuel ethanol. The lignocellulosic substrates include woody substrates such as hardwood (birch and aspen, etc.) and softwood (spruce and pine, etc.), agro residues (wheat straw, sugarcane bagasse, corn stover, etc.), dedicated energy crops (switch grass, and Miscanthus etc.), weedy materials (Eicchornia crassipes, Lantana camara etc.), and municipal solid waste (food and kitchen waste, etc.). Despite the success achieved in the laboratory, there are limitations to success with lignocellulosic substrates on a commercial scale. The future of lignocellulosics is expected to lie in improvements of plant biomass, metabolic engineering of ethanol, and cellulolytic enzyme-producing microorganisms, fullest exploitation of weed materials, and process integration of the individual steps involved in bioethanol production. Issues related to the chemical composition of various weedy raw substrates for bioethanol formation, including chemical composition-based structural hydrolysis of the substrate, need special attention. This area could be opened up further by exploring genetically modified metabolic engineering routes in weedy materials and in biocatalysts that would make the production of bioethanol more efficient. (orig.)

  8. Transcriptomic Changes in Response to Putrescine Production in Metabolically Engineered Corynebacterium glutamicum

    Zhen Li

    2017-10-01

    Full Text Available Putrescine is widely used in industrial production of bioplastics, pharmaceuticals, agrochemicals, and surfactants. Although engineered Corynebacterium glutamicum has been successfully used to produce high levels of putrescine, the overall cellular physiological and metabolic changes caused by overproduction of putrescine remains unclear. To reveal the transcriptional changes that occur in response to putrescine production in an engineered C. glutamicum strain, a comparative transcriptomic analysis was carried out. Overproduction of putrescine resulted in transcriptional downregulation of genes involved in glycolysis; the TCA cycle, pyruvate degradation, biosynthesis of some amino acids, oxidative phosphorylation; vitamin biosynthesis (thiamine and vitamin 6, metabolism of purine, pyrimidine and sulfur, and ATP-, NAD-, and NADPH-consuming enzymes. The transcriptional levels of genes involved in ornithine biosynthesis and NADPH-forming related enzymes were significantly upregulated in the putrescine producing C. glutamicum strain PUT-ALE. Comparative transcriptomic analysis provided some genetic modification strategies to further improve putrescine production. Repressing ATP- and NADPH-consuming enzyme coding gene expression via CRISPRi enhanced putrescine production.

  9. Transcriptomic Changes in Response to Putrescine Production in Metabolically Engineered Corynebacterium glutamicum

    Li, Zhen; Liu, Jian-Zhong

    2017-01-01

    Putrescine is widely used in industrial production of bioplastics, pharmaceuticals, agrochemicals, and surfactants. Although engineered Corynebacterium glutamicum has been successfully used to produce high levels of putrescine, the overall cellular physiological and metabolic changes caused by overproduction of putrescine remains unclear. To reveal the transcriptional changes that occur in response to putrescine production in an engineered C. glutamicum strain, a comparative transcriptomic analysis was carried out. Overproduction of putrescine resulted in transcriptional downregulation of genes involved in glycolysis; the TCA cycle, pyruvate degradation, biosynthesis of some amino acids, oxidative phosphorylation; vitamin biosynthesis (thiamine and vitamin 6), metabolism of purine, pyrimidine and sulfur, and ATP-, NAD-, and NADPH-consuming enzymes. The transcriptional levels of genes involved in ornithine biosynthesis and NADPH-forming related enzymes were significantly upregulated in the putrescine producing C. glutamicum strain PUT-ALE. Comparative transcriptomic analysis provided some genetic modification strategies to further improve putrescine production. Repressing ATP- and NADPH-consuming enzyme coding gene expression via CRISPRi enhanced putrescine production. PMID:29089930

  10. Metabolic engineering of Escherichia coli for biotechnological production of high-value organic acids and alcohols

    Yu, Chao; Cao, Yujin; Zou, Huibin; Xian, Mo [Chinese Academy of Sciences, Qingdao (China). Key Lab. of Biofuels

    2011-02-15

    Confronted with the gradual and inescapable exhaustion of the earth's fossil energy resources, the bio-based process to produce platform chemicals from renewable carbohydrates is attracting growing interest. Escherichia coli has been chosen as a workhouse for the production of many valuable chemicals due to its clear genetic background, convenient to be genetically modified and good growth properties with low nutrient requirements. Rational strain development of E. coli achieved by metabolic engineering strategies has provided new processes for efficiently biotechnological production of various high-value chemical building blocks. Compared to previous reviews, this review focuses on recent advances in metabolic engineering of the industrial model bacteria E. coli that lead to efficient recombinant biocatalysts for the production of high-value organic acids like succinic acid, lactic acid, 3-hydroxypropanoic acid and glucaric acid as well as alcohols like 1,3-propanediol, xylitol, mannitol, and glycerol with the discussion of the future research in this area. Besides, this review also discusses several platform chemicals, including fumaric acid, aspartic acid, glutamic acid, sorbitol, itaconic acid, and 2,5-furan dicarboxylic acid, which have not been produced by E. coli until now. (orig.)

  11. Clinical approach to inherited metabolic disorders in neonates

    Saudubray, J. M.; Narcy, C.; Lyonnet, L.; Bonnefont, J. P.; Poll The, B. T.; Munnich, A.

    1990-01-01

    Most inborn errors of intermediary metabolism presenting in the neonatal period fall schematically into three clinical categories: (1) those which lead to a neurological distress 'intoxication type' with a symptom-free interval, vomiting, comas, hypertonia, abnormal movements and frequent humoral

  12. Some approaches to system reliability improvement in engineering design

    Shen, Kecheng.

    1990-01-01

    In this thesis some approaches to system reliability improvement in engineering design are studied. In particular, the thesis aims at developing alternative methodologies for ranking of component importance which are more related to the design practice and which are more useful in system synthesis than the existing ones. It also aims at developing component reliability models by means of stress-strength interference which will enable both component reliability prediction and design for reliability. A new methodology for ranking of component importance is first developed based on the notion of the increase of the expected system yield. This methodology allows for incorporation of different improvement actions at the component level such as parallel redundancy, standby redundancy, burn-in, minimal repair and perfect replacement. For each of these improvement actions, the increase of system reliability is studied and used as the component importance measure. A possible connection between the commonly known models of component lifetimes and the stress-strength interference models is suggested. Under some general conditions the relationship between component failure rate and the stress and strength distribution characteristics is studied. A heuristic approach for obtaining bounds on failure probability through stress-strength interference is also presented. A case study and a worked example are presented, which illustrate and verify the developed importance measures and their applications in the analytical as well as synthetical work of engineering design. (author)

  13. Metabolic engineering of cyanobacteria for photosynthetic 3-hydroxypropionic acid production from CO2 using Synechococcus elongatus PCC 7942.

    Lan, Ethan I; Chuang, Derrick S; Shen, Claire R; Lee, Annabel M; Ro, Soo Y; Liao, James C

    2015-09-01

    Photosynthetic conversion of CO2 to chemicals using cyanobacteria is an attractive approach for direct recycling of CO2 to useful products. 3-Hydroxypropionic acid (3 HP) is a valuable chemical for the synthesis of polymers and serves as a precursor to many other chemicals such as acrylic acid. 3 HP is naturally produced through glycerol metabolism. However, cyanobacteria do not possess pathways for synthesizing glycerol and converting glycerol to 3 HP. Furthermore, the latter pathway requires coenzyme B12, or an oxygen sensitive, coenzyme B12-independent enzyme. These characteristics present major challenges for production of 3 HP using cyanobacteria. To overcome such difficulties, we constructed two alternative pathways in Synechococcus elongatus PCC 7942: a malonyl-CoA dependent pathway and a β-alanine dependent pathway. Expression of the malonyl-CoA dependent pathway genes (malonyl-CoA reductase and malonate semialdehyde reductase) enabled S. elongatus to synthesize 3 HP to a final titer of 665 mg/L. β-Alanine dependent pathway expressing S. elongatus produced 3H P to final titer of 186 mg/L. These results demonstrated the feasibility of converting CO2 into 3 HP using cyanobacteria. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  14. APPROACHING CARBOHYDRATES AND ITS METABOLISM: AN EXPERIENCE FOR EDUCATIONAL GAMES

    Sandro Henrique Dias Ribeiro

    2016-11-01

    Full Text Available INTRODUCTION: The study of carbohydrates, its metabolism and many other fields of biochemistry are often understood by students as a junction of chemical structures and reactions of difficult compression. However, Biochemistry should no longer be seen as an abstruse field, but a way to know the human body and its components, including molecular, structural and functional aspects. Therefore, some alternatives are being evaluated in order to assist and improve the dissemination of knowledge among them highlights are the educational games. OBJECTIVES: The aim of this work is the production of two educational games able to include the contents of carbohydrates and its metabolism in higher education. MATERIALS AND METHODS: The educational games produced were made from available materials and low cost. The games were tested in courses of Biological Sciences and Biotechnology of the Federal University of Uberlândia and the response of the students towards the activities was analyzed. The application, had the presence of trained students to instruct on the activity and correcting. DISCUSSION AND RESULTS: "What is the Carbohydrate?" and "Mastering the metabolism" are two educational games covering the content of structure and function carbohydrates and basal metabolism, respectively. "What is the Carbohydrate?" consists in unravel amid several options the carbohydrate in the hands of the others players. For this, several questions with two possible answers, “yes” or “not”, are accepted each round, and if the player find difficulty in formulating questions, there are cards tips. “Mastering the metabolism” consists in a combination of cards that simulate pieces of a domino that must be mounted following the metabolic pathway of carbohydrates, and as the game progresses, the main points of regulation of the pathway will be accompanied by surprise questions. The games showed great acceptance by students. CONCLUSION: “What is the Carbohydrate

  15. Metabolic and process engineering for biodesulfurization in Gram-negative bacteria.

    Martínez, I; El-Said Mohamed, M; Santos, V E; García, J L; García-Ochoa, F; Díaz, E

    2017-11-20

    Microbial desulfurization or biodesulfurization (BDS) is an attractive low-cost and environmentally friendly complementary technology to the hydrotreating chemical process based on the potential of certain bacteria to specifically remove sulfur from S-heterocyclic compounds of crude fuels that are recalcitrant to the chemical treatments. The 4S or Dsz sulfur specific pathway for dibenzothiophene (DBT) and alkyl-substituted DBTs, widely used as model S-heterocyclic compounds, has been extensively studied at the physiological, biochemical and genetic levels mainly in Gram-positive bacteria. Nevertheless, several Gram-negative bacteria have been also used in BDS because they are endowed with some properties, e.g., broad metabolic versatility and easy genetic and genomic manipulation, that make them suitable chassis for systems metabolic engineering strategies. A high number of recombinant bacteria, many of which are Pseudomonas strains, have been constructed to overcome the major bottlenecks of the desulfurization process, i.e., expression of the dsz operon, activity of the Dsz enzymes, retro-inhibition of the Dsz pathway, availability of reducing power, uptake-secretion of substrate and intermediates, tolerance to organic solvents and metals, and other host-specific limitations. However, to attain a BDS process with industrial applicability, it is necessary to apply all the knowledge and advances achieved at the genetic and metabolic levels to the process engineering level, i.e., kinetic modelling, scale-up of biphasic systems, enhancing mass transfer rates, biocatalyst separation, etc. The production of high-added value products derived from the organosulfur material present in oil can be regarded also as an economically viable process that has barely begun to be explored. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Metabolic engineering of Corynebacterium glutamicum to produce GDP-L-fucose from glucose and mannose.

    Chin, Young-Wook; Park, Jin-Byung; Park, Yong-Cheol; Kim, Kyoung Heon; Seo, Jin-Ho

    2013-06-01

    Wild-type Corynebacterium glutamicum was metabolically engineered to convert glucose and mannose into guanosine 5'-diphosphate (GDP)-L-fucose, a precursor of fucosyl-oligosaccharides, which are involved in various biological and pathological functions. This was done by introducing the gmd and wcaG genes of Escherichia coli encoding GDP-D-mannose-4,6-dehydratase and GDP-4-keto-6-deoxy-D-mannose-3,5-epimerase-4-reductase, respectively, which are known as key enzymes in the production of GDP-L-fucose from GDP-D-mannose. Coexpression of the genes allowed the recombinant C. glutamicum cells to produce GDP-L-fucose in a minimal medium containing glucose and mannose as carbon sources. The specific product formation rate was much higher during growth on mannose than on glucose. In addition, the specific product formation rate was further increased by coexpressing the endogenous phosphomanno-mutase gene (manB) and GTP-mannose-1-phosphate guanylyl-transferase gene (manC), which are involved in the conversion of mannose-6-phosphate into GDP-D-mannose. However, the overexpression of manA encoding mannose-6-phosphate isomerase, catalyzing interconversion of mannose-6-phosphate and fructose-6-phosphate showed a negative effect on formation of the target product. Overall, coexpression of gmd, wcaG, manB and manC in C. glutamicum enabled production of GDP-L-fucose at the specific rate of 0.11 mg g cell(-1) h(-1). The specific GDP-L-fucose content reached 5.5 mg g cell(-1), which is a 2.4-fold higher than that of the recombinant E. coli overexpressing gmd, wcaG, manB and manC under comparable conditions. Well-established metabolic engineering tools may permit optimization of the carbon and cofactor metabolisms of C. glutamicum to further improve their production capacity.

  17. Metabolic engineering of riboflavin production in Ashbya gossypii through pathway optimization.

    Ledesma-Amaro, Rodrigo; Serrano-Amatriain, Cristina; Jiménez, Alberto; Revuelta, José Luis

    2015-10-14

    The industrial production of riboflavin mostly relies on the microbial fermentation of flavinogenic microorganisms and Ashbya gossypii is the main industrial producer of the vitamin. Accordingly, bioengineering strategies aimed at increasing riboflavin production in A. gossypii are highly valuable for industry. We analyze the contribution of all the RIB genes to the production of riboflavin in A. gossypii. Two important metabolic rate-limiting steps that limit the overproduction of riboflavin have been found: first, low mRNA levels of the RIB genes hindered the overproduction of riboflavin; second, the competition of the AMP branch for purinogenic precursors also represents a limitation for riboflavin overproduction. Thus, overexpression of the RIB genes resulted in a significant increase in riboflavin yield. Moreover, both the inactivation and the underexpression of the ADE12 gene, which controls the first step of the AMP branch, also proved to have a positive effect on riboflavin production. Accordingly, a strain that combines both the overexpression of the RIB genes and the underexpression of the ADE12 gene was engineered. This strain produced 523 mg/L of riboflavin (5.4-fold higher than the wild-type), which is the highest titer of riboflavin obtained by metabolic engineering in A. gossypii so far. Riboflavin production in A. gossypii is limited by a low transcription activity of the RIB genes. Flux limitation towards AMP provides committed substrate GTP for riboflavin overproduction without detrimental effects on biomass formation. A multiple-engineered Ashbya strain that produces up to 523 mg/L of riboflavin was generated.

  18. Metabolic engineering of the pentose phosphate pathway for enhanced limonene production in the cyanobacterium Synechocysti s sp. PCC 6803.

    Lin, Po-Cheng; Saha, Rajib; Zhang, Fuzhong; Pakrasi, Himadri B

    2017-12-13

    Isoprenoids are diverse natural compounds, which have various applications as pharmaceuticals, fragrances, and solvents. The low yield of isoprenoids in plants makes them difficult for cost-effective production, and chemical synthesis of complex isoprenoids is impractical. Microbial production of isoprenoids has been considered as a promising approach to increase the yield. In this study, we engineered the model cyanobacterium Synechocystis sp. PCC 6803 for sustainable production of a commercially valuable isoprenoid, limonene. Limonene synthases from the plants Mentha spicata and Citrus limon were expressed in cyanobacteria for limonene production. Production of limonene was two-fold higher with limonene synthase from M. spicata than that from C. limon. To enhance isoprenoid production, computational strain design was conducted by applying the OptForce strain design algorithm on Synechocystis 6803. Based on the metabolic interventions suggested by this algorithm, genes (ribose 5-phosphate isomerase and ribulose 5-phosphate 3-epimerase) in the pentose phosphate pathway were overexpressed, and a geranyl diphosphate synthase from the plant Abies grandis was expressed to optimize the limonene biosynthetic pathway. The optimized strain produced 6.7 mg/L of limonene, a 2.3-fold improvement in productivity. Thus, this study presents a feasible strategy to engineer cyanobacteria for photosynthetic production of isoprenoids.

  19. Metabolism

    ... Are More Common in People With Type 1 Diabetes Metabolic Syndrome Your Child's Weight Healthy Eating Endocrine System Blood Test: Basic Metabolic Panel (BMP) Activity: Endocrine System Growth Disorders Diabetes Center Thyroid Disorders Your Endocrine System Movie: Endocrine ...

  20. Engineering nanomaterials with a combined electrochemical and molecular biomimetic approach

    Dai, Haixia

    Biocomposite materials, such as bones, teeth, and shells, are created using mild aqueous solution-based processes near room temperature. Proteins add flexibility to these processes by facilitating the nucleation, growth, and ordering of specific inorganic materials into hierarchical structures. We aim to develop a biomimetic strategy for engineering technologically relevant inorganic materials with controlled compositions and structures, as Nature does, using proteins to orchestrate material formation and assembly. This approach involves three basic steps: (i) preparation of inorganic substrates compatible with combinatorial polypeptide screening; (ii) identification of inorganic-binding polypeptides and their engineering into inorganic-binding proteins; and (iii) protein-mediated inorganic nucleation and organization. Cuprous oxide (Cu2O), a p-type semiconductor, has been used to demonstrate all three steps. Zinc oxide (ZnO), an n-type semiconductor, has been used to show the generality of selected steps. Step (i), preparation of high quality inorganic substrates to select inorganic-binding polypeptides, was accomplished using electrochemical microfabrication to grow and pattern Cu2O and ZnO. Raman spectroscopy and x-ray photoelectron spectroscopy were used to verify phase purity and compositional stability of these surfaces during polypeptide screening. Step (ii), accomplished in collaboration with personnel in Prof Baneyx' lab at the University of Washington, involved incubating the inorganic substrates with the FliTrx(TM) random peptide library to identify cysteine-constrained dodecapeptides that bind the targeted inorganic. Insertion of a Cu2O-binding dodecapeptide into the DNA-binding protein TraI endowed the engineered TraI with strong affinity for Cu2O (Kd ≈ 10 -8 M). Finally, step (iii) involved nonequilibrium synthesis and organization of Cu2O nanoparticles, taking advantage of the inorganic and DNA recognition properties of the engineered TraI. The

  1. APPROACHING CARBOHYDRATES AND ITS METABOLISM: AN EXPERIENCE FOR EDUCATIONAL GAMES

    Ribeiro, Sandro Henrique Dias; Mestanza, Paulo Enrique Cuevas; Freitas, Vitor de; Chagas, Letícia de Fátima; Espindola, Foued Salmen; Rodrigues, Renata Santos; Tudini, Kelly Aparecida Geraldo Yoneyama; Ávila, Veridiana Melo Rodrigues

    2016-01-01

    INTRODUCTION: The study of carbohydrates, its metabolism and many other fields of biochemistry are often understood by students as a junction of chemical structures and reactions of difficult compression. However, Biochemistry should no longer be seen as an abstruse field, but a way to know the human body and its components, including molecular, structural and functional aspects. Therefore, some alternatives are being evaluated in order to assist and improve the dissemination of knowledge amo...

  2. Metabolic engineering of Escherichia coli: a sustainable industrial platform for bio-based chemical production.

    Chen, Xianzhong; Zhou, Li; Tian, Kangming; Kumar, Ashwani; Singh, Suren; Prior, Bernard A; Wang, Zhengxiang

    2013-12-01

    In order to decrease carbon emissions and negative environmental impacts of various pollutants, more bulk and/or fine chemicals are produced by bioprocesses, replacing the traditional energy and fossil based intensive route. The Gram-negative rod-shaped bacterium, Escherichia coli has been studied extensively on a fundamental and applied level and has become a predominant host microorganism for industrial applications. Furthermore, metabolic engineering of E. coli for the enhanced biochemical production has been significantly promoted by the integrated use of recent developments in systems biology, synthetic biology and evolutionary engineering. In this review, we focus on recent efforts devoted to the use of genetically engineered E. coli as a sustainable platform for the production of industrially important biochemicals such as biofuels, organic acids, amino acids, sugar alcohols and biopolymers. In addition, representative secondary metabolites produced by E. coli will be systematically discussed and the successful strategies for strain improvements will be highlighted. Moreover, this review presents guidelines for future developments in the bio-based chemical production using E. coli as an industrial platform. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. Selection Finder (SelFi: A computational metabolic engineering tool to enable directed evolution of enzymes

    Neda Hassanpour

    2017-06-01

    Full Text Available Directed evolution of enzymes consists of an iterative process of creating mutant libraries and choosing desired phenotypes through screening or selection until the enzymatic activity reaches a desired goal. The biggest challenge in directed enzyme evolution is identifying high-throughput screens or selections to isolate the variant(s with the desired property. We present in this paper a computational metabolic engineering framework, Selection Finder (SelFi, to construct a selection pathway from a desired enzymatic product to a cellular host and to couple the pathway with cell survival. We applied SelFi to construct selection pathways for four enzymes and their desired enzymatic products xylitol, D-ribulose-1,5-bisphosphate, methanol, and aniline. Two of the selection pathways identified by SelFi were previously experimentally validated for engineering Xylose Reductase and RuBisCO. Importantly, SelFi advances directed evolution of enzymes as there is currently no known generalized strategies or computational techniques for identifying high-throughput selections for engineering enzymes.

  4. Glycerol positive promoters for tailored metabolic engineering of the yeast Saccharomyces cerevisiae.

    Ho, Ping-Wei; Klein, Mathias; Futschik, Matthias; Nevoigt, Elke

    2018-05-01

    Glycerol offers several advantages as a substrate for biotechnological applications. An important step toward using the popular production host Saccharomyces cerevisiae for glycerol-based bioprocesses has been the fact that in recent studies commonly used S. cerevisiae strains were engineered to grow in synthetic medium containing glycerol as the sole carbon source. For metabolic engineering projects of S. cerevisiae growing on glycerol, characterized promoters are missing. In the current study, we used transcriptome analysis and a yECitrine-based fluorescence reporter assay to select and characterize 25 useful promoters. The promoters of the genes ALD4 and ADH2 showed 4.2-fold and 3-fold higher activities compared to the well-known strong TEF1 promoter. Moreover, the collection contains promoters with graded activities in synthetic glycerol medium and different degrees of glucose repression. To demonstrate the general applicability of the promoter collection, we successfully used a subset of the characterized promoters with graded activities in order to optimize growth on glycerol in an engineered derivative of CEN.PK, in which glycerol catabolism exclusively occurs via a non-native DHA pathway.

  5. Phytohormones and their metabolic engineering for abiotic stress tolerance in crop plants

    Shabir H. Wani

    2016-06-01

    Full Text Available Abiotic stresses including drought, salinity, heat, cold, flooding, and ultraviolet radiation causes crop losses worldwide. In recent times, preventing these crop losses and producing more food and feed to meet the demands of ever-increasing human populations have gained unprecedented importance. However, the proportion of agricultural lands facing multiple abiotic stresses is expected only to rise under a changing global climate fueled by anthropogenic activities. Identifying the mechanisms developed and deployed by plants to counteract abiotic stresses and maintain their growth and survival under harsh conditions thus holds great significance. Recent investigations have shown that phytohormones, including the classical auxins, cytokinins, ethylene, and gibberellins, and newer members including brassinosteroids, jasmonates, and strigolactones may prove to be important metabolic engineering targets for producing abiotic stress-tolerant crop plants. In this review, we summarize and critically assess the roles that phytohormones play in plant growth and development and abiotic stress tolerance, besides their engineering for conferring abiotic stress tolerance in transgenic crops. We also describe recent successes in identifying the roles of phytohormones under stressful conditions. We conclude by describing the recent progress and future prospects including limitations and challenges of phytohormone engineering for inducing abiotic stress tolerance in crop plants.

  6. Cerebellar involvement in metabolic disorders: a pattern-recognition approach

    Steinlin, M.; Boltshauser, E.; Blaser, S.

    1998-01-01

    Inborn errors of metabolism can affect the cerebellum during development, maturation and later during life. We have established criteria for pattern recognition of cerebellar abnormalities in metabolic disorders. The abnormalities can be divided into four major groups: cerebellar hypoplasia (CH), hyperplasia, cerebellar atrophy (CA), cerebellar white matter abnormalities (WMA) or swelling, and involvement of the dentate nuclei (DN) or cerebellar cortex. CH can be an isolated typical finding, as in adenylsuccinase deficiency, but is also occasionally seen in many other disorders. Differentiation from CH and CA is often difficult, as in carbohydrate deficient glycoprotein syndrome or 2-l-hydroxyglutaric acidaemia. In cases of atrophy the relationship of cerebellar to cerebral atrophy is important. WMA may be diffuse or patchy, frequently predominantly around the DN. Severe swelling of white matter is present during metabolic crisis in maple syrup urine disease. The DN can be affected by metabolite deposition, necrosis, calcification or demyelination. Involvement of cerebellar cortex is seen in infantile neuroaxonal dystrophy. Changes in DN and cerebellar cortex are rather typical and therefore most helpful; additional features should be sought as they are useful in narrowing down the differential diagnosis. (orig.)

  7. Fatigue in engineering structures. A three fold analysis approach

    Malik, Afzaal M.; Qureshi, Ejaz M.; Dar, Naeem Ullah; Khan, Iqbal

    2007-01-01

    The integrity in most of the engineering structures in influenced by the presence of cracks or crack like defects. These structures fail, even catastrophically if a crack greater than a critically safe size exist. Although most of the optimal designed structures are initially free from critical cracks, sub-critical cracks can lead to failures under cyclic loadings, called fatigue crack growth. It is nearly impractical to prevent sub-critical crack growth in engineering structures particularly in crack sensitive structures like most of the structures in nuclear, aerospace and aeronautical domains. However, it is essential to predict the fatigue crack growth for these structures to preclude the in service failures causing loss of assets. The present research presents an automatic procedure for the prediction of fatigue crack growth in three dimensional engineering structures and the key data for the fracture mechanics based design: the stress intensity factors. Three fold analysis procedures are adopted to investigate the effects of repetitive (cyclic) loadings on the fatigue life of different geometries of aluminum alloy 2219-O. A general purpose Finite Element (FE) Code ANSYS-8.0 is used to predict/estimate the fatigue life of the geometries. Computer codes utilizing the Green's Function are developed to calculate the stress intensity factors. Another code based on superposition technique presented by Shivakumara and Foreman is developed to calculate the fatigue crack growth rate, fatigue life (No. of loading cycles are developed to validate the results and finally full scale laboratory tests are conducted for the comparison of the results. The results showing a close co-relation between the different techniques employed gives the promising feature of the analysis approach for the future work. (author)

  8. Engineered nanomaterial-mediated changes in the metabolism of terrestrial plants

    Hatami, Mehrnaz, E-mail: m-hatami@araku.ac.ir [Department of Medicinal Plants, Faculty of Agriculture and Natural Resources, Arak University, 38156-8-8349 Arak (Iran, Islamic Republic of); Kariman, Khalil [School of Earth and Environment M004, The University of Western Australia, Crawley, WA 6009 (Australia); Ghorbanpour, Mansour, E-mail: m-ghorbanpour@araku.ac.ir [Department of Medicinal Plants, Faculty of Agriculture and Natural Resources, Arak University, 38156-8-8349 Arak (Iran, Islamic Republic of)

    2016-11-15

    Engineered nanomaterials (ENMs) possess remarkable physicochemical characteristics suitable for different applications in medicine, pharmaceuticals, biotechnology, energy, cosmetics and electronics. Because of their ultrafine size and high surface reactivity, ENMs can enter plant cells and interact with intracellular structures and metabolic pathways which may produce toxicity or promote plant growth and development by diverse mechanisms. Depending on their type and concentration, ENMs can have positive or negative effects on photosynthesis, photochemical fluorescence and quantum yield as well as photosynthetic pigments status of the plants. Some studies have shown that ENMs can improve photosynthetic efficiency via increasing chlorophyll content and light absorption and also broadening the spectrum of captured light, suggesting that photosynthesis can be nano-engineered for harnessing more solar energy. Both up- and down-regulation of primary metabolites such as proteins and carbohydrates have been observed following exposure of plants to various ENMs. The potential capacity of ENMs for changing the rate of primary metabolites lies in their close relationship with activation and biosynthesis of the key enzymes. Several classes of secondary metabolites such as phenolics, flavonoids, and alkaloids have been shown to be induced (mostly accompanied by stress-related factors) in plants exposed to different ENMs, highlighting their great potential as elicitors to enhance both quantity and quality of biologically active secondary metabolites. Considering reports on both positive and negative effects of ENMs on plant metabolism, in-depth studies are warranted to figure out the most appropriate ENMs (type, size and optimal concentration) in order to achieve the desirable effect on specific metabolites in a given plant species. In this review, we summarize the studies performed on the impacts of ENMs on biosynthesis of plant primary and secondary metabolites and mention the

  9. Engineered nanomaterial-mediated changes in the metabolism of terrestrial plants

    Hatami, Mehrnaz; Kariman, Khalil; Ghorbanpour, Mansour

    2016-01-01

    Engineered nanomaterials (ENMs) possess remarkable physicochemical characteristics suitable for different applications in medicine, pharmaceuticals, biotechnology, energy, cosmetics and electronics. Because of their ultrafine size and high surface reactivity, ENMs can enter plant cells and interact with intracellular structures and metabolic pathways which may produce toxicity or promote plant growth and development by diverse mechanisms. Depending on their type and concentration, ENMs can have positive or negative effects on photosynthesis, photochemical fluorescence and quantum yield as well as photosynthetic pigments status of the plants. Some studies have shown that ENMs can improve photosynthetic efficiency via increasing chlorophyll content and light absorption and also broadening the spectrum of captured light, suggesting that photosynthesis can be nano-engineered for harnessing more solar energy. Both up- and down-regulation of primary metabolites such as proteins and carbohydrates have been observed following exposure of plants to various ENMs. The potential capacity of ENMs for changing the rate of primary metabolites lies in their close relationship with activation and biosynthesis of the key enzymes. Several classes of secondary metabolites such as phenolics, flavonoids, and alkaloids have been shown to be induced (mostly accompanied by stress-related factors) in plants exposed to different ENMs, highlighting their great potential as elicitors to enhance both quantity and quality of biologically active secondary metabolites. Considering reports on both positive and negative effects of ENMs on plant metabolism, in-depth studies are warranted to figure out the most appropriate ENMs (type, size and optimal concentration) in order to achieve the desirable effect on specific metabolites in a given plant species. In this review, we summarize the studies performed on the impacts of ENMs on biosynthesis of plant primary and secondary metabolites and mention the

  10. Emerging nanotechnology approaches in tissue engineering for peripheral nerve regeneration.

    Cunha, Carla; Panseri, Silvia; Antonini, Stefania

    2011-02-01

    Effective nerve regeneration and functional recovery subsequent to peripheral nerve injury is still a clinical challenge. Autologous nerve graft transplantation is a feasible treatment in several clinical cases, but it is limited by donor site morbidity and insufficient donor tissue, impairing complete functional recovery. Tissue engineering has introduced innovative approaches to promote and guide peripheral nerve regeneration by using biomimetic conduits creating favorable microenvironments for nervous ingrowth, but despite the development of a plethora of nerve prostheses, few approaches have as yet entered the clinic. Promising strategies using nanotechnology have recently been proposed, such as the use of scaffolds with functionalized cell-binding domains, the use of guidance channels with cell-scale internally oriented fibers, and the possibility of sustained release of neurotrophic factors. This review addresses the fabrication, advantages, drawbacks, and results achieved by the most recent nanotechnology approaches in view of future solutions for peripheral nerve repair. Peripheral nerve repair strategies are very limited despite numerous advances on the field of neurosciences and regenerative medicine. This review discusses nanotechnology based strategies including scaffolds with functionalized cell binding domains, the use of guidance channels, and the potential use of sustained release neurotropic factors. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. Complex approach in telecommunication engineering education: develop engineering skills by a team project

    Scripcariu Luminița

    2017-01-01

    Full Text Available This paper provides an overview of the educational process of telecommunication engineering students by presenting the preparation of a team project focused on information security. Our educational approach combines basic knowledge such as mathematics with specialized engineering notions and various skills. The project theme is to design, implement and test an encryption algorithm. Students are provided with online courses, specific software programs and Internet access. They have to choose an encryption algorithm, to study its details and to write the script of the encryption algorithm in MATLAB program. The algorithm is implemented in C/C++ programming language and tested. Finally, a concurrent team tries to break the algorithm by finding the decryption key. It is an interactive approach which combines various education methods including gaming concepts. The covered topics provide students professional outcomes such as knowledge and use of specific mathematical tools and software environments (C/C ++ programming languages, MATLAB, abilities to design, develop, implement and test software algorithms. The project also provides transversal outcomes such as ability to team work, skills of computer use and information technology and capability to take responsibilities. Creativity is also encouraged by extending the algorithm to other encryption key lengths than the usual ones.

  12. Approaches to nontraditional delivery of nuclear engineering education

    Malaviya, B.K.

    1991-01-01

    At Rensselaer Polytechnic Institute, the faculty of the nuclear engineering and engineering physics department have, over the years, been involved in a variety of such approaches in response to the changing needs of nuclear industry personnel. A number of different types of short course and workshop programs have been developed and implemented both on and off campus in such areas as basic nuclear technology, reactor design computer codes and applications, nuclear power plant design and maintenance, reactor operations, health physics, modern developments in boiling heat transfer and two-phase flow, and probabilistic risk assessment. Customized coursed tailored to meet the particular needs of personnel in specialized areas can also be offered on specific industrial site locations, generally resulting in substantial savings of time as well as costs associated with tuition, travel, lodging. The Rensselaer Satellite Video Program (RSVP) brings the latest technological aids to the nontraditional delivery of courses and provides the facilities and opportunities for off-campus students and professional personnel to participate in regular academic programs and courses without leaving their industrial sites

  13. Synthetic biology approaches to engineering the nitrogen symbiosis in cereals.

    Rogers, Christian; Oldroyd, Giles E D

    2014-05-01

    Nitrogen is abundant in the earth's atmosphere but, unlike carbon, cannot be directly assimilated by plants. The limitation this places on plant productivity has been circumvented in contemporary agriculture through the production and application of chemical fertilizers. The chemical reduction of nitrogen for this purpose consumes large amounts of energy and the reactive nitrogen released into the environment as a result of fertilizer application leads to greenhouse gas emissions, as well as widespread eutrophication of aquatic ecosystems. The environmental impacts are intensified by injudicious use of fertilizers in many parts of the world. Simultaneously, limitations in the production and supply of chemical fertilizers in other regions are leading to low agricultural productivity and malnutrition. Nitrogen can be directly fixed from the atmosphere by some bacteria and Archaea, which possess the enzyme nitrogenase. Some plant species, most notably legumes, have evolved close symbiotic associations with nitrogen-fixing bacteria. Engineering cereal crops with the capability to fix their own nitrogen could one day address the problems created by the over- and under-use of nitrogen fertilizers in agriculture. This could be achieved either by expression of a functional nitrogenase enzyme in the cells of the cereal crop or through transferring the capability to form a symbiotic association with nitrogen-fixing bacteria. While potentially transformative, these biotechnological approaches are challenging; however, with recent advances in synthetic biology they are viable long-term goals. This review discusses the possibility of these biotechnological solutions to the nitrogen problem, focusing on engineering the nitrogen symbiosis in cereals.

  14. A Systems Engineering Approach to Quality Assurance for Aerospace Testing

    Shepherd, Christena C.

    2015-01-01

    On the surface, it appears that AS91001 has little to say about how to apply a Quality Management System (QMS) to major aerospace test programs (or even smaller ones). It also appears that there is little in the quality engineering Body of Knowledge (BOK)2 that applies to testing, unless it is nondestructive examination (NDE), or some type of lab or bench testing associated with the manufacturing process. However, if one examines: a) how the systems engineering (SE) processes are implemented throughout a test program; and b) how these SE processes can be mapped to the requirements of AS9100, a number of areas for involvement of the quality professional are revealed. What often happens is that quality assurance during a test program is limited to inspections of the test article; what could be considered a manufacturing al fresco approach. This limits the quality professional and is a disservice to the programs and projects, since there are a number of ways that quality can enhance critical processes, and support efforts to improve risk reduction, efficiency and effectiveness.

  15. Energetics of glucose metabolism: a phenomenological approach to metabolic network modeling.

    Diederichs, Frank

    2010-08-12

    A new formalism to describe metabolic fluxes as well as membrane transport processes was developed. The new flux equations are comparable to other phenomenological laws. Michaelis-Menten like expressions, as well as flux equations of nonequilibrium thermodynamics, can be regarded as special cases of these new equations. For metabolic network modeling, variable conductances and driving forces are required to enable pathway control and to allow a rapid response to perturbations. When applied to oxidative phosphorylation, results of simulations show that whole oxidative phosphorylation cannot be described as a two-flux-system according to nonequilibrium thermodynamics, although all coupled reactions per se fulfill the equations of this theory. Simulations show that activation of ATP-coupled load reactions plus glucose oxidation is brought about by an increase of only two different conductances: a [Ca(2+)] dependent increase of cytosolic load conductances, and an increase of phosphofructokinase conductance by [AMP], which in turn becomes increased through [ADP] generation by those load reactions. In ventricular myocytes, this feedback mechanism is sufficient to increase cellular power output and O(2) consumption several fold, without any appreciable impairment of energetic parameters. Glucose oxidation proceeds near maximal power output, since transformed input and output conductances are nearly equal, yielding an efficiency of about 0.5. This conductance matching is fulfilled also by glucose oxidation of β-cells. But, as a price for the metabolic mechanism of glucose recognition, β-cells have only a limited capability to increase their power output.

  16. Integrating biocompatible chemistry and manipulating cofactor partitioning in metabolically engineeredLactococcus lactisfor fermentative production of (3S)-acetoin

    Liu, Jianming; Solem, Christian; Jensen, Peter Ruhdal

    2016-01-01

    Biocompatible chemistry (BC), i.e. non-enzymatic chemical reactions compatible with living organisms, is increasingly used in conjunction with metabolically engineered microorganisms for producing compounds that do not usually occur naturally. Here we report production of one such compound, (3S......)-acetoin, a valuable precursor for chiral synthesis, using a metabolically engineered Lactococcus lactis strain growing under respiratory conditions with ferric iron serving as a BC component. The strain used has all competing product pathways inactivated, and an appropriate cofactor balance is achieved by fine...

  17. Personalized Education Approaches for Chemical Engineering and Relevant Majors

    Zhao Feng-qing

    2016-01-01

    Full Text Available Personalized education has drawn increasing attention in universities these years. With the purpose of improving the studentss’ comprehensive ability and developing teaching strategies to ensure students’ education is tailored to their needs, we proposed Three-Stage Approach (TSA to enhance personalized education for chemical engineering and relevant majors: professional tutorial system--equipping with professional guidance teachers for freshman students to guide their learning activities and provide professional guidance; open experimental project--setting up open experimental projects for sophomore and junior students to choose freely; individualized education module--setting up 10 different individualized education modules for senior students to select. After years of practice, the personalized education model is improved day by day and proved effective and fruitful.

  18. Altered Levels of Aroma and Volatiles by Metabolic Engineering of Shikimate Pathway Genes in Tomato Fruits

    Vered Tzin

    2015-06-01

    Full Text Available The tomato (Solanum lycopersicum fruit is an excellent source of antioxidants, dietary fibers, minerals and vitamins and therefore has been referred to as a “functional food”. Ripe tomato fruits produce a large number of specialized metabolites including volatile organic compounds. These volatiles serve as key components of the tomato fruit flavor, participate in plant pathogen and herbivore defense, and are used to attract seed dispersers. A major class of specialized metabolites is derived from the shikimate pathway followed by aromatic amino acid biosynthesis of phenylalanine, tyrosine and tryptophan. We attempted to modify tomato fruit flavor by overexpressing key regulatory genes in the shikimate pathway. Bacterial genes encoding feedback-insensitive variants of 3-Deoxy-D-Arabino-Heptulosonate 7-Phosphate Synthase (DAHPS; AroG209-9 and bi-functional Chorismate Mutase/Prephenate Dehydratase (CM/PDT; PheA12 were expressed under the control of a fruit-specific promoter. We crossed these transgenes to generate tomato plants expressing both the AroG209 and PheA12 genes. Overexpression of the AroG209-9 gene had a dramatic effect on the overall metabolic profile of the fruit, including enhanced levels of multiple volatile and non-volatile metabolites. In contrast, the PheA12 overexpression line exhibited minor metabolic effects compared to the wild type fruit. Co-expression of both the AroG209-9 and PheA12 genes in tomato resulted overall in a similar metabolic effect to that of expressing only the AroG209-9 gene. However, the aroma ranking attributes of the tomato fruits from PheA12//AroG209-9 were unique and different from those of the lines expressing a single gene, suggesting a contribution of the PheA12 gene to the overall metabolic profile. We suggest that expression of bacterial genes encoding feedback-insensitive enzymes of the shikimate pathway in tomato fruits provides a useful metabolic engineering tool for the modification of

  19. Enhancing Carbon Fixation by Metabolic Engineering: A Model System of Complex Network Modulation

    Dr. Gregory Stephanopoulos

    2008-04-10

    In the first two years of this research we focused on the development of a DNA microarray for transcriptional studies in the photosynthetic organism Synechocystis and the elucidation of the metabolic pathway for biopolymer synthesis in this organism. In addition we also advanced the molecular biological tools for metabolic engineering of biopolymer synthesis in Synechocystis and initiated a series of physiological studies for the elucidation of the carbon fixing pathways and basic central carbon metabolism of these organisms. During the last two-year period we focused our attention on the continuation and completion of the last task, namely, the development of tools for basic investigations of the physiology of these cells through, primarily, the determination of their metabolic fluxes. The reason for this decision lies in the importance of fluxes as key indicators of physiology and the high level of information content they carry in terms of identifying rate limiting steps in a metabolic pathway. While flux determination is a well-advanced subject for heterotrophic organisms, for the case of autotrophic bacteria, like Synechocystis, some special challenges had to be overcome. These challenges stem mostly from the fact that if one uses {sup 13}C labeled CO{sub 2} for flux determination, the {sup 13}C label will mark, at steady state, all carbon atoms of all cellular metabolites, thus eliminating the necessary differentiation required for flux determination. This peculiarity of autotrophic organisms makes it imperative to carry out flux determination under transient conditions, something that had not been accomplished before. We are pleased to report that we have solved this problem and we are now able to determine fluxes in photosynthetic organisms from stable isotope labeling experiments followed by measurements of label enrichment in cellular metabolites using Gas Chromatography-Mass Spectrometry. We have conducted extensive simulations to test the method and

  20. Metabolic engineering of plant monoterpenes, sesquiterpenes and diterpenes--current status and future opportunities.

    Lange, B Markus; Ahkami, Amirhossein

    2013-02-01

    Terpenoids (a.k.a. isoprenoids) represent the most diverse class of natural products found in plants, with tens of thousands of reported structures. Plant-derived terpenoids have a multitude of pharmaceutical and industrial applications, but the natural resources for their extraction are often limited and, in many cases, synthetic routes are not commercially viable. Some of the most valuable terpenoids are not accumulated in model plants or crops, and genetic resources for breeding of terpenoid natural product traits are thus poorly developed. At present, metabolic engineering, either in the native producer or a heterologous host, is the only realistic alternative to improve yield and accessibility. In this review article, we will evaluate the state of the art of modulating the biosynthetic pathways for the production of mono-, sesqui- and diterpenes in plants. © 2012 The Authors Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  1. Systems metabolic engineering as an enabling technology in accomplishing sustainable development goals.

    Yang, Dongsoo; Cho, Jae Sung; Choi, Kyeong Rok; Kim, Hyun Uk; Lee, Sang Yup

    2017-09-01

    With pressing issues arising in recent years, the United Nations proposed 17 Sustainable Development Goals (SDGs) as an agenda urging international cooperations for sustainable development. In this perspective, we examine the roles of systems metabolic engineering (SysME) and its contribution to improving the quality of life and protecting our environment, presenting how this field of study offers resolutions to the SDGs with relevant examples. We conclude with offering our opinion on the current state of SysME and the direction it should move forward in the generations to come, explicitly focusing on addressing the SDGs. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  2. Advances in metabolic engineering of yeast Saccharomyces cerevisiae for production of chemicals

    Borodina, Irina; Nielsen, Jens

    2014-01-01

    Yeast Saccharomyces cerevisiae is an important industrial host for production of enzymes, pharmaceutical and nutraceutical ingredients and recently also commodity chemicals and biofuels. Here, we review the advances in modeling and synthetic biology tools and how these tools can speed up the deve......Yeast Saccharomyces cerevisiae is an important industrial host for production of enzymes, pharmaceutical and nutraceutical ingredients and recently also commodity chemicals and biofuels. Here, we review the advances in modeling and synthetic biology tools and how these tools can speed up...... the development of yeast cell factories. We also present an overview of metabolic engineering strategies for developing yeast strains for production of polymer monomers: lactic, succinic, and cis,cis-muconic acids. S. cerevisiae has already firmly established itself as a cell factory in industrial biotechnology...

  3. Prognostic relevance of metabolic approach in patients with heart failure.

    Di Napoli, P; Barsotti, A

    2009-01-01

    Progressions in acute cardiac care have improved survival after acute myocardial infarction, but in contraposition with this, there has been an increase in mortality because of heart failure. For this reason congestive heart failure is an increasingly widespread, costly and deadly disease, frequently named as epidemic of the XXI century. Despite advancement in modern treatment, mortality rate in heart failure patients remains high. In these patients more importance was attributed in the management of the left ventricle dysfunction. In fact, the heart failure patients have still a poor prognosis due to the ineluctable progression of contractile dysfunction and ventricular remodeling. The classical management of left ventricle dysfunction includes the pharmacological treatment with beta-blockers, ACE-inhibitors and aldosterone antagonists, and various surgical or electrophysiological interventions. Emerging evidence suggests that myocardium dysfunction is also due to substrate metabolism alterations. In particular, there is evidence that, in the failing heart, shifting metabolism away from a preference for fatty acids towards more carbohydrate oxidation could recover contractile function. Trimetazidine has been shown to improve symptoms and ventricular function and to have a beneficial effect on the inflammatory profile and endothelial function in these patients. Recently, it has been suggested that trimetazidine could also reduce ventricular remodeling, slowing down the progression of pump failure, and improve prognosis. These results suggest that trimetazidine is a useful adjunct to our current armamentarium for the treatment of heart failure patients.

  4. Understanding Fatty Acid Metabolism through an Active Learning Approach

    Fardilha, M.; Schrader, M.; da Cruz e Silva, O. A. B.; da Cruz e Silva, E. F.

    2010-01-01

    A multi-method active learning approach (MALA) was implemented in the Medical Biochemistry teaching unit of the Biomedical Sciences degree at the University of Aveiro, using problem-based learning as the main learning approach. In this type of learning strategy, students are involved beyond the mere exercise of being taught by listening. Less…

  5. Metabolic engineering of β-oxidation in Penicillium chrysogenum for improved semi-synthetic cephalosporin biosynthesis.

    Veiga, Tânia; Gombert, Andreas K; Landes, Nils; Verhoeven, Maarten D; Kiel, Jan A K W; Krikken, Arjen M; Nijland, Jeroen G; Touw, Hesselien; Luttik, Marijke A H; van der Toorn, John C; Driessen, Arnold J M; Bovenberg, Roel A L; van den Berg, Marco A; van der Klei, Ida J; Pronk, Jack T; Daran, Jean-Marc

    2012-07-01

    Industrial production of semi-synthetic cephalosporins by Penicillium chrysogenum requires supplementation of the growth media with the side-chain precursor adipic acid. In glucose-limited chemostat cultures of P. chrysogenum, up to 88% of the consumed adipic acid was not recovered in cephalosporin-related products, but used as an additional carbon and energy source for growth. This low efficiency of side-chain precursor incorporation provides an economic incentive for studying and engineering the metabolism of adipic acid in P. chrysogenum. Chemostat-based transcriptome analysis in the presence and absence of adipic acid confirmed that adipic acid metabolism in this fungus occurs via β-oxidation. A set of 52 adipate-responsive genes included six putative genes for acyl-CoA oxidases and dehydrogenases, enzymes responsible for the first step of β-oxidation. Subcellular localization of the differentially expressed acyl-CoA oxidases and dehydrogenases revealed that the oxidases were exclusively targeted to peroxisomes, while the dehydrogenases were found either in peroxisomes or in mitochondria. Deletion of the genes encoding the peroxisomal acyl-CoA oxidase Pc20g01800 and the mitochondrial acyl-CoA dehydrogenase Pc20g07920 resulted in a 1.6- and 3.7-fold increase in the production of the semi-synthetic cephalosporin intermediate adipoyl-6-APA, respectively. The deletion strains also showed reduced adipate consumption compared to the reference strain, indicating that engineering of the first step of β-oxidation successfully redirected a larger fraction of adipic acid towards cephalosporin biosynthesis. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Engineering metabolic pathways in Amycolatopsis japonicum for the optimization of the precursor supply for heterologous brasilicardin congeners production

    Paul N. Schwarz

    2018-03-01

    Full Text Available The isoprenoid brasilicardin A is a promising immunosuppressant compound with a unique mode of action, high potency and reduced toxicity compared to today's standard drugs. However, production of brasilicardin has been hampered since the producer strain Nocardia terpenica IFM0406 synthesizes brasilicardin in only low amounts and is a biosafety level 2 organism. Previously, we were able to heterologously express the brasilicardin gene cluster in the nocardioform actinomycete Amycolatopsis japonicum. Four brasilicardin congeners, intermediates of the BraA biosynthesis, were produced. Since chemical synthesis of the brasilicardin core structure has remained elusive we intended to produce high amounts of the brasilicardin backbone for semi synthesis and derivatization. Therefore, we used a metabolic engineering approach to increase heterologous production of brasilicardin in A. japonicum. Simultaneous heterologous expression of genes encoding the MVA pathway and expression of diterpenoid specific prenyltransferases were used to increase the provision of the isoprenoid precursor isopentenyl diphosphate (IPP and to channel the precursor into the direction of diterpenoid biosynthesis. Both approaches contributed to an elevated heterologous production of the brasilicardin backbone, which can now be used as a starting point for semi synthesis of new brasilicardin congeners with better properties.

  7. Metabolic engineering of Pseudomonas fluorescens for the production of vanillin from ferulic acid.

    Di Gioia, Diana; Luziatelli, Francesca; Negroni, Andrea; Ficca, Anna Grazia; Fava, Fabio; Ruzzi, Maurizio

    2011-12-20

    Vanillin is one of the most important flavors in the food industry and there is great interest in its production through biotechnological processes starting from natural substrates such as ferulic acid. Among bacteria, recombinant Escherichia coli strains are the most efficient vanillin producers, whereas Pseudomonas spp. strains, although possessing a broader metabolic versatility, rapidly metabolize various phenolic compounds including vanillin. In order to develop a robust Pseudomonas strain that can produce vanillin in high yields and at high productivity, the vanillin dehydrogenase (vdh)-encoding gene of Pseudomonas fluorescens BF13 strain was inactivated via targeted mutagenesis. The results demonstrated that engineered derivatives of strain BF13 accumulate vanillin if inactivation of vdh is associated with concurrent expression of structural genes for feruloyl-CoA synthetase (fcs) and hydratase/aldolase (ech) from a low-copy plasmid. The conversion of ferulic acid to vanillin was enhanced by optimization of growth conditions, growth phase and parameters of the bioconversion process. The developed strain produced up to 8.41 mM vanillin, which is the highest final titer of vanillin produced by a Pseudomonas strain to date and opens new perspectives in the use of bacterial biocatalysts for biotechnological production of vanillin from agro-industrial wastes which contain ferulic acid. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. Metabolic engineering of the phenylpropanoid pathway enhances the antioxidant capacity of Saussurea involucrata.

    Jian Qiu

    Full Text Available The rare wild species of snow lotus Saussurea involucrata is a commonly used medicinal herb with great pharmacological value for human health, resulting from its uniquely high level of phenylpropanoid compound production. To gain information on the phenylpropanid biosynthetic pathway genes in this critically important medicinal plant, global transcriptome sequencing was performed. It revealed that the phenylpropanoid pathway genes were well represented in S. involucrata. In addition, we introduced two key phenylpropanoid pathway inducing transcription factors (PAP1 and Lc into this medicinal plant. Transgenic S. involucrata co-expressing PAP1 and Lc exhibited purple pigments due to a massive accumulation of anthocyanins. The over-expression of PAP1 and Lc largely activated most of the phenylpropanoid pathway genes, and increased accumulation of several phenylpropanoid compounds significantly, including chlorogenic acid, syringin, cyanrine and rutin. Both ABTS (2,2'-azinobis-3-ethylbenzotiazo-line-6-sulfonic acid and FRAP (ferric reducing anti-oxidant power assays revealed that the antioxidant capacity of transgenic S. involucrata lines was greatly enhanced over controls. In addition to providing a deeper understanding of the molecular basis of phenylpropanoid metabolism, our results potentially enable an alternation of bioactive compound production in S. involucrata through metabolic engineering.

  9. Genome-scale modeling enables metabolic engineering of Saccharomyces cerevisiae for succinic acid production.

    Agren, Rasmus; Otero, José Manuel; Nielsen, Jens

    2013-07-01

    In this work, we describe the application of a genome-scale metabolic model and flux balance analysis for the prediction of succinic acid overproduction strategies in Saccharomyces cerevisiae. The top three single gene deletion strategies, Δmdh1, Δoac1, and Δdic1, were tested using knock-out strains cultivated anaerobically on glucose, coupled with physiological and DNA microarray characterization. While Δmdh1 and Δoac1 strains failed to produce succinate, Δdic1 produced 0.02 C-mol/C-mol glucose, in close agreement with model predictions (0.03 C-mol/C-mol glucose). Transcriptional profiling suggests that succinate formation is coupled to mitochondrial redox balancing, and more specifically, reductive TCA cycle activity. While far from industrial titers, this proof-of-concept suggests that in silico predictions coupled with experimental validation can be used to identify novel and non-intuitive metabolic engineering strategies.

  10. Unraveling and engineering the production of 23,24-bisnorcholenic steroids in sterol metabolism.

    Xu, Li-Qin; Liu, Yong-Jun; Yao, Kang; Liu, Hao-Hao; Tao, Xin-Yi; Wang, Feng-Qing; Wei, Dong-Zhi

    2016-02-22

    The catabolism of sterols in mycobacteria is highly important due to its close relevance in the pathogenesis of pathogenic strains and the biotechnological applications of nonpathogenic strains for steroid synthesis. However, some key metabolic steps remain unknown. In this study, the hsd4A gene from Mycobacterium neoaurum ATCC 25795 was investigated. The encoded protein, Hsd4A, was characterized as a dual-function enzyme, with both 17β-hydroxysteroid dehydrogenase and β-hydroxyacyl-CoA dehydrogenase activities in vitro. Using a kshAs-null strain of M. neoaurum ATCC 25795 (NwIB-XII) as a model, Hsd4A was further confirmed to exert dual-function in sterol catabolism in vivo. The deletion of hsd4A in NwIB-XII resulted in the production of 23,24-bisnorcholenic steroids (HBCs), indicating that hsd4A plays a key role in sterol side-chain degradation. Therefore, two competing pathways, the AD and HBC pathways, were proposed for the side-chain degradation. The proposed HBC pathway has great value in illustrating the production mechanism of HBCs in sterol catabolism and in developing HBCs producing strains for industrial application via metabolic engineering. Through the combined modification of hsd4A and other genes, three HBCs producing strains were constructed that resulted in promising productivities of 0.127, 0.109 and 0.074 g/l/h, respectively.

  11. Monitoring Bone Tissue Engineered (BTE) Constructs Based on the Shifting Metabolism of Differentiating Stem Cells.

    Simmons, Aaron D; Sikavitsas, Vassilios I

    2018-01-01

    Ever-increasing demand for bone grafts necessitates the realization of clinical implementation of bone tissue engineered constructs. The predominant hurdle to implementation remains to be securing FDA approval, based on the lack of viable methods for the rigorous monitoring of said constructs. The study presented herein details a method for such monitoring based on the shifting metabolism of mesenchymal stem cells (MSCs) as they differentiate into osteoblasts. To that end, rat MSCs seeded on 85% porous spunbonded poly(L-lactic acid) scaffolds were cultured in flow perfusion bioreactors with baseline or osteoinductive media, and levels of key physio-metabolic markers (oxygen, glucose, osteoprotegerin, and osteocalcin) were monitored throughout culture. Comparison of these non-destructively obtained values and current standard destructive analyses demonstrated key trends useful for the concurrent real-time monitoring of construct cellularity and maturation. Principle among these is the elucidation of the ratio of the rates of oxygen uptake to glucose consumption as a powerful quality marker. This ratio, supported on a physiological basis, has been shown herein to be reliable in the determination of both construct maturation (defined as osteoblastic differentiation and accompanying mineralization) and construct cellularity. Supplementary monitoring of OPG and OCN are shown to provide further validation of such metrics.

  12. Metabonomics Approach to Assessing the Metabolism Variation and Endoexogenous Metabolic Interaction of Ginsenosides in Cold Stress Rats.

    Zhang, Zhihao; Wang, Xiaoyan; Wang, Jingcheng; Jia, Zhiying; Liu, Yumin; Xie, Xie; Wang, Chongchong; Jia, Wei

    2016-06-03

    Metabolic profiling technology, a massive information provider, has promoted the understanding of the metabolism of multicomponent medicines and its interactions with endogenous metabolites, which was previously a challenge in clarification. In this study, an untargeted GC/MS-based approach was employed to investigate the urinary metabolite profile in rats with oral administration of ginsenosides and the control group. Significant changes of urinary metabolites contents were observed in the total ginsenosides group, revealing the impact of ginsenosides as indicated by the up- or down-regulation of several pathways involving neurotransmitter-related metabolites, tricarboxylic acid (TCA) cycle, fatty acids β-oxidation, and intestinal microflora metabolites. Meanwhile, a targeted UPLC-QQQ/MS-based metabonomic approach was developed to investigate the changes of urinary ginsenoside metabolites during the process of acute cold stress. Metabolic analysis indicated that upstream ginsenosides (rg1, re, and rf) increased significantly, whereas downstream ginsenosides (ck, ppd, and ppt) decreased correspondingly after cold exposure. Finally, the relationships between ginsenosides and significantly changed metabolites were investigated by correlation analysis.

  13. Improving polyglucan production in cyanobacteria and microalgae via cultivation design and metabolic engineering.

    Aikawa, Shimpei; Ho, Shih-Hsin; Nakanishi, Akihito; Chang, Jo-Shu; Hasunuma, Tomohisa; Kondo, Akihiko

    2015-06-01

    Photosynthetic microorganisms, such as cyanobacteria and microalgae, are currently being investigated as alternative biomass resources for bioethanol production, owing to their benefits, including high-photosynthetic activity and whole-year cultivation without utilization of arable land. Polyglucans comprise the major carbohydrate content of these organisms. Polyglucans can be utilized as a carbon source for microbial fermentation. Although polyglucan production has so far been promoted by nutrient limitation, it must be further enhanced to accommodate market demand. This review focuses on the recent progress in the production of α-polyglucans such asglycogen and starch in cyanobacteria and green microalgae via cultivation design, including modifying the nutrient supply and replacing the growth medium. The control and manipulation of polyglucan metabolism necessitates the elucidation of the polyglucan production mechanism. We reviewed gene expression and metabolite accumulation profiles of cyanobacteria and green microalgae during nutrient limitation-stimulated α-polyglucan accumulation. We also focus on the enhancement in cyanobacterial glycogen production via the genetic engineering of glycolysis, CO2 concentration mechanism, and photosynthetic light-harvesting protein based on the polyglucan accumulation mechanism. The combined strategies of cultivation design and genetic engineering should be considered for further enhancement of polyglucan productivity for bioethanol production. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Production of 3-hydroxypropionic acid from glucose and xylose by metabolically engineered Saccharomyces cerevisiae

    Kanchana R. Kildegaard

    2015-12-01

    Full Text Available Biomass, the most abundant carbon source on the planet, may in the future become the primary feedstock for production of fuels and chemicals, replacing fossil feedstocks. This will, however, require development of cell factories that can convert both C6 and C5 sugars present in lignocellulosic biomass into the products of interest. We engineered Saccharomyces cerevisiae for production of 3-hydroxypropionic acid (3HP, a potential building block for acrylates, from glucose and xylose. We introduced the 3HP biosynthetic pathways via malonyl-CoA or β-alanine intermediates into a xylose-consuming yeast. Using controlled fed-batch cultivation, we obtained 7.37±0.17 g 3HP L−1 in 120 hours with an overall yield of 29±1% Cmol 3HP Cmol−1 xylose. This study is the first demonstration of the potential of using S. cerevisiae for production of 3HP from the biomass sugar xylose. Keywords: Metabolic engineering, Biorefineries, 3-hydroxypropionic acid, Saccharomyces cerevisiae, Xylose utilization

  15. Dedicated Industrial Oilseed Crops as Metabolic Engineering Platforms for Sustainable Industrial Feedstock Production.

    Zhu, Li-Hua; Krens, Frans; Smith, Mark A; Li, Xueyuan; Qi, Weicong; van Loo, Eibertus N; Iven, Tim; Feussner, Ivo; Nazarenus, Tara J; Huai, Dongxin; Taylor, David C; Zhou, Xue-Rong; Green, Allan G; Shockey, Jay; Klasson, K Thomas; Mullen, Robert T; Huang, Bangquan; Dyer, John M; Cahoon, Edgar B

    2016-02-26

    Feedstocks for industrial applications ranging from polymers to lubricants are largely derived from petroleum, a non-renewable resource. Vegetable oils with fatty acid structures and storage forms tailored for specific industrial uses offer renewable and potentially sustainable sources of petrochemical-type functionalities. A wide array of industrial vegetable oils can be generated through biotechnology, but will likely require non-commodity oilseed platforms dedicated to specialty oil production for commercial acceptance. Here we show the feasibility of three Brassicaceae oilseeds crambe, camelina, and carinata, none of which are widely cultivated for food use, as hosts for complex metabolic engineering of wax esters for lubricant applications. Lines producing wax esters >20% of total seed oil were generated for each crop and further improved for high temperature oxidative stability by down-regulation of fatty acid polyunsaturation. Field cultivation of optimized wax ester-producing crambe demonstrated commercial utility of these engineered crops and a path for sustainable production of other industrial oils in dedicated specialty oilseeds.

  16. Metabolic engineering of Corynebacterium glutamicum aimed at alternative carbon sources and new products

    Volker Fritz Wendisch

    2012-10-01

    Full Text Available Corynebacterium glutamicum is well known as the amino acid-producing workhorse of fermentation industry, being used for multi-million-ton scale production of glutamate and lysine for more than 60 years. However, it is only recently that extensive research has focused on engineering it beyond the scope of amino acids. Meanwhile, a variety of corynebacterial strains allows access to alternative carbon sources and/or allows production of a wide range of industrially relevant compounds. Some of these efforts set new standards in terms of titers and productivities achieved whereas others represent a proof-of-principle. These achievements manifest the position of C. glutamicum as an important industrial microorganism with capabilities far beyond the traditional amino acid production. In this review we focus on the state of the art of metabolic engineering of C. glutamicum for utilization of alternative carbon sources, (e.g. coming from wastes and unprocessed sources, and construction of C. glutamicum strains for production of new products such as diamines, organic acids and alcohols.

  17. Metabolic transcription analysis of engineered Escherichia coli strains that overproduce L-phenylalanine

    Gosset Guillermo

    2007-09-01

    Full Text Available Abstract Background The rational design of L-phenylalanine (L-Phe overproducing microorganisms has been successfully achieved by combining different genetic strategies such as inactivation of the phosphoenolpyruvate: phosphotransferase transport system (PTS and overexpression of key genes (DAHP synthase, transketolase and chorismate mutase-prephenate dehydratase, reaching yields of 0.33 (g-Phe/g-Glc, which correspond to 60% of theoretical maximum. Although genetic modifications introduced into the cell for the generation of overproducing organisms are specifically targeted to a particular pathway, these can trigger unexpected transcriptional responses of several genes. In the current work, metabolic transcription analysis (MTA of both L-Phe overproducing and non-engineered strains using Real-Time PCR was performed, allowing the detection of transcriptional responses to PTS deletion and plasmid presence of genes related to central carbon metabolism. This MTA included 86 genes encoding enzymes of glycolysis, gluconeogenesis, pentoses phosphate, tricarboxylic acid cycle, fermentative and aromatic amino acid pathways. In addition, 30 genes encoding regulatory proteins and transporters for aromatic compounds and carbohydrates were also analyzed. Results MTA revealed that a set of genes encoding carbohydrate transporters (galP, mglB, gluconeogenic (ppsA, pckA and fermentative enzymes (ldhA were significantly induced, while some others were down-regulated such as ppc, pflB, pta and ackA, as a consequence of PTS inactivation. One of the most relevant findings was the coordinated up-regulation of several genes that are exclusively gluconeogenic (fbp, ppsA, pckA, maeB, sfcA, and glyoxylate shunt in the best PTS- L-Phe overproducing strain (PB12-ev2. Furthermore, it was noticeable that most of the TCA genes showed a strong up-regulation in the presence of multicopy plasmids by an unknown mechanism. A group of genes exhibited transcriptional responses to

  18. The catchment based approach using catchment system engineering

    Jonczyk, Jennine; Quinn, Paul; Barber, Nicholas; Wilkinson, Mark

    2015-04-01

    The catchment based approach (CaBa) has been championed as a potential mechanism for delivery of environmental directives such as the Water Framework Directive in the UK. However, since its launch in 2013, there has been only limited progress towards achieving sustainable, holistic management, with only a few of examples of good practice ( e.g. from the Tyne Rivers trust). Common issues with developing catchment plans over a national scale include limited data and resources to identify issues and source of those issues, how to systematically identify suitable locations for measures or suites of measures that will have the biggest downstream impact and how to overcome barriers for implementing solutions. Catchment System Engineering (CSE) is an interventionist approach to altering the catchment scale runoff regime through the manipulation of hydrological flow pathways throughout the catchment. A significant component of the runoff generation can be managed by targeting hydrological flow pathways at source, such as overland flow, field drain and ditch function, greatly reducing erosive soil losses. Coupled with management of farm nutrients at source, many runoff attenuation features or measures can be co-located to achieve benefits for water quality and biodiversity. A catchment, community-led mitigation measures plan using the CSE approach will be presented from a catchment in Northumberland, Northern England that demonstrate a generic framework for identification of multi-purpose features that slow, store and filter runoff at strategic locations in the landscape. Measures include within-field barriers, edge of field traps and within-ditch measures. Progress on the implementation of measures will be reported alongside potential impacts on the runoff regime at both local and catchment scale and costs.

  19. Sustainable Development in Engineering Education: A Pedagogical Approach

    Ahrens, A.; Zascerinska, J.

    2012-01-01

    Engineering education is facing a challenge of the development of student engineers' social responsibility in the context of sustainable development. The aim of the research is to analyze efficiency of engineering curriculum in the context of sustainable development underpinning elaboration of pedagogical guidelines on the development of students'…

  20. A Novel Approach to Physiology Education for Biomedical Engineering Students

    DiCecco, J.; Wu, J.; Kuwasawa, K.; Sun, Y.

    2007-01-01

    It is challenging for biomedical engineering programs to incorporate an indepth study of the systemic interdependence of cells, tissues, and organs into the rigorous mathematical curriculum that is the cornerstone of engineering education. To be sure, many biomedical engineering programs require their students to enroll in anatomy and physiology…

  1. A review of metabolic and enzymatic engineering strategies for designing and optimizing performance of microbial cell factories

    Amanda K. Fisher

    2014-08-01

    Full Text Available Microbial cell factories (MCFs are of considerable interest to convert low value renewable substrates to biofuels and high value chemicals. This review highlights the progress of computational models for the rational design of an MCF to produce a target bio-commodity. In particular, the rational design of an MCF involves: (i product selection, (ii de novo biosynthetic pathway identification (i.e., rational, heterologous, or artificial, (iii MCF chassis selection, (iv enzyme engineering of promiscuity to enable the formation of new products, and (v metabolic engineering to ensure optimal use of the pathway by the MCF host. Computational tools such as (i de novo biosynthetic pathway builders, (ii docking, (iii molecular dynamics (MD and steered MD (SMD, and (iv genome-scale metabolic flux modeling all play critical roles in the rational design of an MCF. Genome-scale metabolic flux models are of considerable use to the design process since they can reveal metabolic capabilities of MCF hosts. These can be used for host selection as well as optimizing precursors and cofactors of artificial de novo biosynthetic pathways. In addition, recent advances in genome-scale modeling have enabled the derivation of metabolic engineering strategies, which can be implemented using the genomic tools reviewed here as well.

  2. Metabolic engineering of a diazotrophic bacterium improves ammonium release and biofertilization of plants and microalgae.

    Ambrosio, Rafael; Ortiz-Marquez, Juan Cesar Federico; Curatti, Leonardo

    2017-03-01

    The biological nitrogen fixation carried out by some Bacteria and Archaea is one of the most attractive alternatives to synthetic nitrogen fertilizers. However, with the exception of the symbiotic rhizobia-legumes system, progress towards a more extensive realization of this goal has been slow. In this study we manipulated the endogenous regulation of both nitrogen fixation and assimilation in the aerobic bacterium Azotobacter vinelandii. Substituting an exogenously inducible promoter for the native promoter of glutamine synthetase produced conditional lethal mutant strains unable to grow diazotrophically in the absence of the inducer. This mutant phenotype could be reverted in a double mutant strain bearing a deletion in the nifL gene that resulted in constitutive expression of nif genes and increased production of ammonium. Under GS non-inducing conditions both the single and the double mutant strains consistently released very high levels of ammonium (>20mM) into the growth medium. The double mutant strain grew and excreted high levels of ammonium under a wider range of concentrations of the inducer than the single mutant strain. Induced mutant cells could be loaded with glutamine synthetase at different levels, which resulted in different patterns of extracellular ammonium accumulation afterwards. Inoculation of the engineered bacteria into a microalgal culture in the absence of sources of C and N other than N 2 and CO 2 from the air, resulted in a strong proliferation of microalgae that was suppressed upon addition of the inducer. Both single and double mutant strains also promoted growth of cucumber plants in the absence of added N-fertilizer, while this property was only marginal in the parental strain. This study provides a simple synthetic genetic circuit that might inspire engineering of optimized inoculants that efficiently channel N 2 from the air into crops. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All

  3. An engineering thermodynamic approach to select the electromagnetic wave effective on cell growth.

    Lucia, Umberto; Grisolia, Giulia; Ponzetto, Antonio; Silvagno, Francesca

    2017-09-21

    To date, the choice of the characteristics of the extremely low-frequency electromagnetic field beneficial in proliferative disorders is still empirical. In order to make the ELF interaction selective, we applied the thermodynamic and biochemical principles to the analysis of the thermo-chemical output generated by the cell in the environment. The theoretical approach applied an engineering bio-thermodynamic approach recently developed in order to obtain a physical-mathematical model that calculated the frequency of the field able to maximize the mean entropy changes as a function of cellular parameters. The combined biochemical approach envisioned the changes of entropy as a metabolic shift leading to a reduction of cell growth. The proliferation of six human cancer cell lines was evaluated as the output signal able to confirm the correctness of the mathematical model. By considering the cell as a reactive system able to respond to the unbalancing external stimuli, for the first time we could calculate and validate the frequencies of the field specifically effective on distinct cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Metabolism

    ... lin), which signals cells to increase their anabolic activities. Metabolism is a complicated chemical process, so it's not ... how those enzymes or hormones work. When the metabolism of body chemicals is ... Hyperthyroidism (pronounced: hi-per-THIGH-roy-dih-zum). Hyperthyroidism ...

  5. A Systems Engineering Approach to Electro-Mechanical Actuator Diagnostic and Prognostic Development

    National Aeronautics and Space Administration — The authors have formulated a Comprehensive Systems Engineering approach to Electro-Mechanical Actuator (EMA) Prognostics and Health Management (PHM) system...

  6. Assembly and multiple gene expression of thermophilic enzymes in Escherichia coli for in vitro metabolic engineering.

    Ninh, Pham Huynh; Honda, Kohsuke; Sakai, Takaaki; Okano, Kenji; Ohtake, Hisao

    2015-01-01

    In vitro reconstitution of an artificial metabolic pathway is an emerging approach for the biocatalytic production of industrial chemicals. However, several enzymes have to be separately prepared (and purified) for the construction of an in vitro metabolic pathway, thereby limiting the practical applicability of this approach. In this study, genes encoding the nine thermophilic enzymes involved in a non-ATP-forming chimeric glycolytic pathway were assembled in an artificial operon and co-expressed in a single recombinant Escherichia coli strain. Gene expression levels of the thermophilic enzymes were controlled by their sequential order in the artificial operon. The specific activities of the recombinant enzymes in the cell-free extract of the multiple-gene-expression E. coli were 5.0-1,370 times higher than those in an enzyme cocktail prepared from a mixture of single-gene-expression strains, in each of which a single one of the nine thermophilic enzymes was overproduced. Heat treatment of a crude extract of the multiple-gene-expression cells led to the denaturation of indigenous proteins and one-step preparation of an in vitro synthetic pathway comprising only a limited number of thermotolerant enzymes. Coupling this in vitro pathway with other thermophilic enzymes including the H2 O-forming NADH oxidase or the malate/lactate dehydrogenase facilitated one-pot conversion of glucose to pyruvate or lactate, respectively. © 2014 Wiley Periodicals, Inc.

  7. Development of a screening approach for exploring cell factory potential through metabolic flux analysis and physiology

    Knudsen, Peter Boldsen; Nielsen, Kristian Fog; Thykær, Jette

    2012-01-01

    The recent developments within the field of metabolic engineering have significantly increased the speed by which fungal recombinant strains are being constructed, pushing focus towards physiological characterisation and analysis. This raises demand for a tool for diligent analysis of the recombi......The recent developments within the field of metabolic engineering have significantly increased the speed by which fungal recombinant strains are being constructed, pushing focus towards physiological characterisation and analysis. This raises demand for a tool for diligent analysis...... of the validation, already well described fungal strains were selected and tested using the described method and the developed method was subsequently used to test recombinant fungal strains producing the model polyketide 6-methylsalicylic acid. Diligent application of this strategy significantly reduces the cost...

  8. Metabolism of chlorofluorocarbons and polybrominated compounds by Pseudomonas putida G786(pHG-2) via an engineered metabolic pathway.

    Hur, H G; Sadowsky, M J; Wackett, L P

    1994-01-01

    The recombinant bacterium Pseudomonas putida G786(pHG-2) metabolizes pentachloroethane to glyoxylate and carbon dioxide, using cytochrome P-450CAM and toluene dioxygenase to catalyze consecutive reductive and oxidative dehalogenation reactions (L.P. Wackett, M.J. Sadowsky, L.N. Newman, H.-G. Hur, and S. Li, Nature [London] 368:627-629, 1994). The present study investigated metabolism of brominated and chlorofluorocarbon compounds by the recombinant strain. Under anaerobic conditions, P. putid...

  9. Partitioning the Metabolic Cost of Human Running: A Task-by-Task Approach

    Arellano, Christopher J.; Kram, Rodger

    2014-01-01

    Compared with other species, humans can be very tractable and thus an ideal “model system” for investigating the metabolic cost of locomotion. Here, we review the biomechanical basis for the metabolic cost of running. Running has been historically modeled as a simple spring-mass system whereby the leg acts as a linear spring, storing, and returning elastic potential energy during stance. However, if running can be modeled as a simple spring-mass system with the underlying assumption of perfect elastic energy storage and return, why does running incur a metabolic cost at all? In 1980, Taylor et al. proposed the “cost of generating force” hypothesis, which was based on the idea that elastic structures allow the muscles to transform metabolic energy into force, and not necessarily mechanical work. In 1990, Kram and Taylor then provided a more explicit and quantitative explanation by demonstrating that the rate of metabolic energy consumption is proportional to body weight and inversely proportional to the time of foot-ground contact for a variety of animals ranging in size and running speed. With a focus on humans, Kram and his colleagues then adopted a task-by-task approach and initially found that the metabolic cost of running could be “individually” partitioned into body weight support (74%), propulsion (37%), and leg-swing (20%). Summing all these biomechanical tasks leads to a paradoxical overestimation of 131%. To further elucidate the possible interactions between these tasks, later studies quantified the reductions in metabolic cost in response to synergistic combinations of body weight support, aiding horizontal forces, and leg-swing-assist forces. This synergistic approach revealed that the interactive nature of body weight support and forward propulsion comprises ∼80% of the net metabolic cost of running. The task of leg-swing at most comprises ∼7% of the net metabolic cost of running and is independent of body weight support and forward

  10. ECSIN's methodological approach for hazard evaluation of engineered nanomaterials

    Bregoli, Lisa; Benetti, Federico; Venturini, Marco; Sabbioni, Enrico

    2013-04-01

    The increasing production volumes and commercialization of engineered nanomaterials (ENM), together with data on their higher biological reactivity when compared to bulk counterpart and ability to cross biological barriers, have caused concerns about their potential impacts on the health and safety of both humans and the environment. A multidisciplinary component of the scientific community has been called to evaluate the real risks associated with the use of products containing ENM, and is today in the process of developing specific definitions and testing strategies for nanomaterials. At ECSIN we are developing an integrated multidisciplinary methodological approach for the evaluation of the biological effects of ENM on the environment and human health. While our testing strategy agrees with the most widely advanced line of work at the European level, the choice of methods and optimization of protocols is made with an extended treatment of details. Our attention to the methodological and technical details is based on the acknowledgment that the innovative characteristics of matter at the nano-size range may influence the existing testing methods in a partially unpredictable manner, an aspect which is frequently recognized at the discussion level but oftentimes disregarded at the laboratory bench level. This work outlines the most important steps of our testing approach. In particular, each step will be briefly discussed in terms of potential technical and methodological pitfalls that we have encountered, and which are often ignored in nanotoxicology research. The final aim is to draw attention to the need of preliminary studies in developing reliable tests, a crucial aspect to confirm the suitability of the chosen analytical and toxicological methods to be used for the specific tested nanoparticle, and to express the idea that in nanotoxicology,"devil is in the detail".

  11. ECSIN's methodological approach for hazard evaluation of engineered nanomaterials

    Bregoli, Lisa; Benetti, Federico; Venturini, Marco; Sabbioni, Enrico

    2013-01-01

    The increasing production volumes and commercialization of engineered nanomaterials (ENM), together with data on their higher biological reactivity when compared to bulk counterpart and ability to cross biological barriers, have caused concerns about their potential impacts on the health and safety of both humans and the environment. A multidisciplinary component of the scientific community has been called to evaluate the real risks associated with the use of products containing ENM, and is today in the process of developing specific definitions and testing strategies for nanomaterials. At ECSIN we are developing an integrated multidisciplinary methodological approach for the evaluation of the biological effects of ENM on the environment and human health. While our testing strategy agrees with the most widely advanced line of work at the European level, the choice of methods and optimization of protocols is made with an extended treatment of details. Our attention to the methodological and technical details is based on the acknowledgment that the innovative characteristics of matter at the nano-size range may influence the existing testing methods in a partially unpredictable manner, an aspect which is frequently recognized at the discussion level but oftentimes disregarded at the laboratory bench level. This work outlines the most important steps of our testing approach. In particular, each step will be briefly discussed in terms of potential technical and methodological pitfalls that we have encountered, and which are often ignored in nanotoxicology research. The final aim is to draw attention to the need of preliminary studies in developing reliable tests, a crucial aspect to confirm the suitability of the chosen analytical and toxicological methods to be used for the specific tested nanoparticle, and to express the idea that in nanotoxicology,'devil is in the detail'.

  12. A unified approach to failure assessment of engineering structures

    Harrison, R.P.

    1977-01-01

    A codified procedure for the failure assessment of engineering structures is presented which has as its basis the two criteria approach of Dowling and Townley (Int. J. Press. Vessels and Piping; 3:77 (1975)) and the Bilby, Cottrell and Swinden (Proc. R. Soc.; A272:304 (1963)) and Dugdale (J. Mech. Phys. Sol.; 8:100 (1960)) model of yielding ahead of a crack tip. The procedure consists of independently assessing the risk of failure (a) under linear elastic conditions only and (b) under plastic collapse conditions only. These two limiting criteria are then plotted as a co-ordinate point on a Failure Assessment Diagram. From this a measure of the degree of safety of the structure can be obtained. As examples, several of the HSST vessel tests are used to indicate the simplicity and versatility of the procedure. It is shown how maximum allowable pressures or defect sizes can be obtained and how safety factors can be readily incorporated on any of the parameters used in the assessment. It is also demonstrated how helpful the procedure is in designing not only working structures, but also structures that are to be used for testing. (author)

  13. A TOTAL QUALITY MANAGEMENT APPROACH TO CIVIL ENGINEERING CONSULTING

    G.A.L. Ambrose

    2012-01-01

    Full Text Available

    ENGLISH ABSTRACT: The article describes the development of a framework for the implementation of a Total Quality Management (TQM approach in the civil consulting industry through the five phases of the typical civil engineering project. For any project to be successful the project team members must take collective and individual responsibility for their role in the project implementation. The organisational goal is “to make money now and in the future”. To achieve this goal the organization must provide value to its clientele. The clients are searching for the silver bullet; double the performance at half the price. TQM provides a means to achieve this goal.

    AFRIKAANSE OPSOMMING: Die artikel beskryf die ontwikkeling van 'n raamwerk vir die implementering van 'n gehaltebestuurstelsel vir die siviele raadgewende bedryf soos dit deelneem aan die vyf fases van 'n tipiese siviele ingenieursprojek. Vir die suksesvolle afhandeling van 'n projek is dit noodsaaklik dat alle projekspanlede gesamentlik en afsonderlik verantwoordelikheid moet aanvaar vir hulle eie betrokkenheid by die projekimplementering. Die organisasie se oogmerk is om “nou en in die toekoms geld te maak”. Ten einde hierdie oogmerk te bereik moet die organisasie waarde vir sy kliënte bied. 'n Gehaltebestuurstelsel voorsien die gereedskap om hierdie doelwit te bereik.

  14. Defining Resilience and Vulnerability Based on Ontology Engineering Approach

    Kumazawa, T.; Matsui, T.; Endo, A.

    2014-12-01

    It is necessary to reflect the concepts of resilience and vulnerability into the assessment framework of "Human-Environmental Security", but it is also in difficulty to identify the linkage between both concepts because of the difference of the academic community which has discussed each concept. The authors have been developing the ontology which deals with the sustainability of the social-ecological systems (SESs). Resilience and vulnerability are also the concepts in the target world which this ontology covers. Based on this point, this paper aims at explicating the semantic relationship between the concepts of resilience and vulnerability based on ontology engineering approach. For this purpose, we first examine the definitions of resilience and vulnerability which the existing literatures proposed. Second, we incorporate the definitions in the ontology dealing with sustainability of SESs. Finally, we focus on the "Water-Energy-Food Nexus Index" to assess Human-Environmental Security, and clarify how the concepts of resilience and vulnerability are linked semantically through the concepts included in these index items.

  15. A co-metabolic approach to groundwater remediation

    Palumbo, A.V.; Boerman, P.A.; Herbes, S.E.; White, D.C.; Strandberg, G.W.; Donaldson, T.L.; Lucero, A.J.; Jennings, H.L.; Phelps, T.J.; White, D.C.

    1991-01-01

    In support of the US Department of Energy's (DOE) Integrated Demonstration (Cleanup of Organics in Soils and Groundwater at Non-arid Sites) at the Savannah River Site (SRS), Oak Ridge National Laboratory (ORNL) and the University of Tennessee (UT) are involved in demonstrations of the use of methanotrophs in bioreactors for remediation of contaminated groundwater. In preparation for a field demonstration at ORNL's K-25 Site in Oak Ridge, Tennessee, ORNL is conducting batch experiments, is operating a number of bench-scale bioreactors, has designed pretreatment systems, and has modified a field-scale bioreactor provided by the Air Force Engineering and Services Center for use at the site. UT is operating benchscale bioreactors with the goal of determining the stability of a trichloroethylene-degrading methanotrophic consortia during shifts in operating conditions (e.g. pH, nutrient inputs, and contaminant mixtures). These activities are all aimed at providing the knowledge base necessary for successful treatment of contaminated groundwater at the SRS and K-25 sites as well as other DOE sites

  16. A co-metabolic approach to groundwater remediation

    Palumbo, A.V.; Boerman, P.A.; Strandberg, G.W.; Donaldson, T.L.; Jennings, H.L.; Lucero, A.J.; Herbes, S.E.; Phelps, T.J.; White, D.C.

    1991-01-01

    In support of the US Department of Energy's (DOE) Integrated Demonstration (Cleanup of Organics in Soils and Groundwater at Non-arid Sites) at the Savannah River Site (SRS), Oak Ridge National Laboratory (ORNL) and the University of Tennessee (UT) are involved in demonstrations of the use of methanotrophs in bioreactors for remediation of contaminated groundwater. In preparation for a field demonstration at ORNL's K-25 Site in Oak Ridge, Tennessee, ORNL is conducting batch experiments, is operating a number of bench-scale bioreactors, has designed pretreatment systems, and has modified a field-scale bioreactor provided by the Air Force Engineering and Services Center for use at the site. UT is operating bench-scale bioreactors with the goal of determining the stability of a trichloroethylene-degrading methanotrophic consortia during shifts in operating conditions (e.g. pH, nutrient inputs, and contaminant mixtures). These activities are all aimed at providing the knowledge base necessary for successful treatment of contaminated groundwater at the SRS and K-25 sites as well as other DOE sites. 18 refs., 1 fig. , 1 tab

  17. Metabolism of chlorofluorocarbons and polybrominated compounds by Pseudomonas putida G786(pHG-2) via an engineered metabolic pathway.

    Hur, H G; Sadowsky, M J; Wackett, L P

    1994-11-01

    The recombinant bacterium Pseudomonas putida G786(pHG-2) metabolizes pentachloroethane to glyoxylate and carbon dioxide, using cytochrome P-450CAM and toluene dioxygenase to catalyze consecutive reductive and oxidative dehalogenation reactions (L.P. Wackett, M.J. Sadowsky, L.N. Newman, H.-G. Hur, and S. Li, Nature [London] 368:627-629, 1994). The present study investigated metabolism of brominated and chlorofluorocarbon compounds by the recombinant strain. Under anaerobic conditions, P. putida G786(pHG-2) reduced 1,1,2,2-tetrabromoethane, 1,2-dibromo-1,2-dichloroethane, and 1,1,1,2-tetrachloro-2,2-difluoroethane to products bearing fewer halogen substituents. Under aerobic conditions, P. putida G786(pHG-2) oxidized cis- and trans-1,2-dibromoethenes, 1,1-dichloro-2,2-difluoroethene, and 1,2-dichloro-1-fluoroethene. Several compounds were metabolized by sequential reductive and oxidative reactions via the constructed metabolic pathway. For example, 1,1,2,2-tetrabromoethane was reduced by cytochrome P-450CAM to 1,2-dibromoethenes, which were subsequently oxidized by toluene dioxygenase. The same pathway metabolized 1,1,1,2-tetrachloro-2,2-difluoroethane to oxalic acid as one of the final products. The results obtained in this study indicate that P. putida G786(pHG-2) metabolizes polyfluorinated, chlorinated, and brominated compounds and further demonstrates the value of using a knowledge of catabolic enzymes and recombinant DNA technology to construct useful metabolic pathways.

  18. Climate-resilient agroforestry: physiological responses to climate change and engineering of crassulacean acid metabolism (CAM) as a mitigation strategy.

    Borland, Anne M; Wullschleger, Stan D; Weston, David J; Hartwell, James; Tuskan, Gerald A; Yang, Xiaohan; Cushman, John C

    2015-09-01

    Global climate change threatens the sustainability of agriculture and agroforestry worldwide through increased heat, drought, surface evaporation and associated soil drying. Exposure of crops and forests to warmer and drier environments will increase leaf:air water vapour-pressure deficits (VPD), and will result in increased drought susceptibility and reduced productivity, not only in arid regions but also in tropical regions with seasonal dry periods. Fast-growing, short-rotation forestry (SRF) bioenergy crops such as poplar (Populus spp.) and willow (Salix spp.) are particularly susceptible to hydraulic failure following drought stress due to their isohydric nature and relatively high stomatal conductance. One approach to sustaining plant productivity is to improve water-use efficiency (WUE) by engineering crassulacean acid metabolism (CAM) into C3 crops. CAM improves WUE by shifting stomatal opening and primary CO2 uptake and fixation to the night-time when leaf:air VPD is low. CAM members of the tree genus Clusia exemplify the compatibility of CAM performance within tree species and highlight CAM as a mechanism to conserve water and maintain carbon uptake during drought conditions. The introduction of bioengineered CAM into SRF bioenergy trees is a potentially viable path to sustaining agroforestry production systems in the face of a globally changing climate. © 2014 John Wiley & Sons Ltd.

  19. Systems approach to managing educational quality in the engineering classroom

    Grygoryev, Kostyantyn

    Today's competitive environment in post-secondary education requires universities to demonstrate the quality of their programs in order to attract financing, and student and academic talent. Despite significant efforts devoted to improving the quality of higher education, systematic, continuous performance measurement and management still have not reached the level where educational outputs and outcomes are actually produced---the classroom. An engineering classroom is a complex environment in which educational inputs are transformed by educational processes into educational outputs and outcomes. By treating a classroom as a system, one can apply tools such as Structural Equation Modeling, Statistical Process Control, and System Dynamics in order to discover cause-and-effect relationships among the classroom variables, control the classroom processes, and evaluate the effect of changes to the course organization, content, and delivery, on educational processes and outcomes. Quality improvement is best achieved through the continuous, systematic application of efforts and resources. Improving classroom processes and outcomes is an iterative process that starts with identifying opportunities for improvement, designing the action plan, implementing the changes, and evaluating their effects. Once the desired objectives are achieved, the quality improvement cycle may start again. The goal of this research was to improve the educational processes and outcomes in an undergraduate engineering management course taught at the University of Alberta. The author was involved with the course, first, as a teaching assistant, and, then, as a primary instructor. The data collected from the course over four years were used to create, first, a static and, then, a dynamic model of a classroom system. By using model output and qualitative feedback from students, changes to the course organization and content were introduced. These changes led to a lower perceived course workload and

  20. Developing a New Industrial Engineering Curriculum Using a Systems Engineering Approach

    Buyurgan, Nebil; Kiassat, Corey

    2017-01-01

    This paper reports on the development of an engineering curriculum for a new industrial engineering programme at a medium-sized private university in the northeast United States. A systems engineering process has been followed to design and develop the new curriculum. Considering the programme curriculum as a system, first the stakeholders have…

  1. Design of an ectoine-responsive AraC mutant and its application in metabolic engineering of ectoine biosynthesis.

    Chen, Wei; Zhang, Shan; Jiang, Peixia; Yao, Jun; He, Yongzhi; Chen, Lincai; Gui, Xiwu; Dong, Zhiyang; Tang, Shuang-Yan

    2015-07-01

    Advanced high-throughput screening methods for small molecules may have important applications in the metabolic engineering of the biosynthetic pathways of these molecules. Ectoine is an excellent osmoprotectant that has been widely used in cosmetics. In this study, the Escherichia coli regulatory protein AraC was engineered to recognize ectoine as its non-natural effector and to activate transcription upon ectoine binding. As an endogenous reporter of ectoine, the mutated AraC protein was successfully incorporated into high-throughput screening of ectoine hyper-producing strains. The ectoine biosynthetic cluster from Halomonas elongata was cloned into E. coli. By engineering the rate-limiting enzyme L-2,4-diaminobutyric acid (DABA) aminotransferase (EctB), ectoine production and the specific activity of the EctB mutant were increased. Thus, these results demonstrated the effectiveness of engineering regulatory proteins into sensitive and rapid screening tools for small molecules and highlighted the importance and efficacy of directed evolution strategies applied to the engineering of genetic components for yield improvement in the biosynthesis of small molecules. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  2. 13C Metabolic Flux Analysis for Systematic Metabolic Engineering of S. cerevisiae for Overproduction of Fatty Acids

    Ghosh, Amit; Ando, David; Gin, Jennifer

    2016-01-01

    Efficient redirection of microbial metabolism into the abundant production of desired bioproducts remains non-trivial. Here, we used flux-based modeling approaches to improve yields of fatty acids in Saccharomyces cerevisiae. We combined 13C labeling data with comprehensive genome-scale models...

  3. Deconstructing Black Swans: An Introductory Approach to Inherited Metabolic Disorders in the Neonate.

    Mew, Nicholas Ah; Viall, Sarah; Kirmse, Brian; Chapman, Kimberly A

    2015-08-01

    Inherited metabolic disorders (IMDs) are individually rare but collectively common disorders that frequently require rapid or urgent therapy. This article provides a generalized approach to IMDs, as well as some investigations and safe therapies that may be initiated pending the metabolic consult. An overview of the research supporting management strategies is provided. In addition, the newborn metabolic screen is reviewed. Caring for infants with IMDs can seem difficult because each of the types is rarely seen; however, collectively the management can be seen as similar. When an IMD is suspected, a metabolic specialist should be consulted for expert advice regarding appropriate laboratory investigations and management. Because rapid intervention of IMDs before the onset of symptoms may prevent future irreversible sequelae, each abnormal newborn screen must be addressed promptly. Management can be difficult. Research in this area is limited and can be difficult without multisite coordination since sample sizes of any significance are difficult to achieve.

  4. New transposon tools tailored for metabolic engineering of Gram-negative microbial cell factories

    Esteban eMartínez-García

    2014-10-01

    Full Text Available Re-programming microorganisms to modify their existing functions and/or to bestow bacteria with entirely new-to-Nature tasks have largely relied so far on specialized molecular biology tools. Such endeavors are not only relevant in the burgeoning metabolic engineering arena, but also instrumental to explore the functioning of complex regulatory networks from a fundamental point of view. À la carte modification of bacterial genomes thus calls for novel tools to make genetic manipulations easier. We propose the use of a series of new broad-host-range mini-Tn5 vectors, termed pBAMDs, for the delivery of gene(s into the chromosome of Gram-negative bacteria and for generating saturated mutagenesis libraries in gene function studies. These delivery vectors endow the user with the possibility of easy cloning and subsequent insertion of functional cargoes with three different antibiotic resistance markers (kanamycin, streptomycin, and gentamicin. After validating the pBAMD vectors in the environmental bacterium Pseudomonas putida KT2440, their use was also illustrated by inserting the entire poly(3-hydroxybutyrate (PHB synthesis pathway from Cupriavidus necator in the chromosome of a phosphotransacetylase mutant of Escherichia coli. PHB is a completely biodegradable polyester with a number of industrial applications that make it attractive as a potential replacement of oil-based plastics. The non-selective nature of chromosomal insertions of the biosynthetic genes was evidenced by a large landscape of PHB synthesis levels in independent clones. One clone was selected and further characterized as a microbial cell factory for PHB accumulation, and it achieved polymer accumulation levels comparable to those of a plasmid-bearing recombinant. Taken together, our results demonstrate that the new mini-Tn5 vectors can be used to confer interesting phenotypes in Gram-negative bacteria that would be very difficult to engineer through direct manipulation of the

  5. Systems metabolic engineering of Corynebacterium glutamicum for production of the chemical chaperone ectoine.

    Becker, Judith; Schäfer, Rudolf; Kohlstedt, Michael; Harder, Björn J; Borchert, Nicole S; Stöveken, Nadine; Bremer, Erhard; Wittmann, Christoph

    2013-11-15

    The stabilizing and function-preserving effects of ectoines have attracted considerable biotechnological interest up to industrial scale processes for their production. These rely on the release of ectoines from high-salinity-cultivated microbial producer cells upon an osmotic down-shock in rather complex processor configurations. There is growing interest in uncoupling the production of ectoines from the typical conditions required for their synthesis, and instead design strains that naturally release ectoines into the medium without the need for osmotic changes, since the use of high-salinity media in the fermentation process imposes notable constraints on the costs, design, and durability of fermenter systems. Here, we used a Corynebacterium glutamicum strain as a cellular chassis to establish a microbial cell factory for the biotechnological production of ectoines. The implementation of a mutant aspartokinase enzyme ensured efficient supply of L-aspartate-beta-semialdehyde, the precursor for ectoine biosynthesis. We further engineered the genome of the basic C. glutamicum strain by integrating a codon-optimized synthetic ectABCD gene cluster under expressional control of the strong and constitutive C. glutamicum tuf promoter. The resulting recombinant strain produced ectoine and excreted it into the medium; however, lysine was still found as a by-product. Subsequent inactivation of the L-lysine exporter prevented the undesired excretion of lysine while ectoine was still exported. Using the streamlined cell factory, a fed-batch process was established that allowed the production of ectoine with an overall productivity of 6.7 g L(-1) day(-1) under growth conditions that did not rely on the use of high-salinity media. The present study describes the construction of a stable microbial cell factory for recombinant production of ectoine. We successfully applied metabolic engineering strategies to optimize its synthetic production in the industrial workhorse C

  6. Metabolic engineering of Chinese hamster ovary cells: towards a bioengineered heparin.

    Baik, Jong Youn; Gasimli, Leyla; Yang, Bo; Datta, Payel; Zhang, Fuming; Glass, Charles A; Esko, Jeffrey D; Linhardt, Robert J; Sharfstein, Susan T

    2012-03-01

    Heparin is the most widely used pharmaceutical to control blood coagulation in modern medicine. A health crisis that took place in 2008 led to a demand for production of heparin from non-animal sources. Chinese hamster ovary (CHO) cells, commonly used mammalian host cells for production of foreign pharmaceutical proteins in the biopharmaceutical industry, are capable of producing heparan sulfate (HS), a related polysaccharide naturally. Since heparin and HS share the same biosynthetic pathway, we hypothesized that heparin could be produced in CHO cells by metabolic engineering. Based on the expression of endogenous enzymes in the HS/heparin pathways of CHO-S cells, human N-deacetylase/N-sulfotransferase (NDST2) and mouse heparan sulfate 3-O-sulfotransferase 1 (Hs3st1) genes were transfected sequentially into CHO host cells growing in suspension culture. Transfectants were screened using quantitative RT-PCR and Western blotting. Out of 120 clones expressing NDST2 and Hs3st1, 2 clones, Dual-3 and Dual-29, were selected for further analysis. An antithrombin III (ATIII) binding assay using flow cytometry, designed to recognize a key sugar structure characteristic of heparin, indicated that Hs3st1 transfection was capable of increasing ATIII binding. An anti-factor Xa assay, which affords a measure of anticoagulant activity, showed a significant increase in activity in the dual-expressing cell lines. Disaccharide analysis of the engineered HS showed a substantial increase in N-sulfo groups, but did not show a pattern consistent with pharmacological heparin, suggesting that further balancing the expression of transgenes with the expression levels of endogenous enzymes involved in HS/heparin biosynthesis might be necessary. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. New Transposon Tools Tailored for Metabolic Engineering of Gram-Negative Microbial Cell Factories

    Martínez-García, Esteban; Aparicio, Tomás; Lorenzo, Víctor de; Nikel, Pablo I.

    2014-01-01

    Re-programming microorganisms to modify their existing functions and/or to bestow bacteria with entirely new-to-Nature tasks have largely relied so far on specialized molecular biology tools. Such endeavors are not only relevant in the burgeoning metabolic engineering arena but also instrumental to explore the functioning of complex regulatory networks from a fundamental point of view. À la carte modification of bacterial genomes thus calls for novel tools to make genetic manipulations easier. We propose the use of a series of new broad-host-range mini-Tn5-vectors, termed pBAMDs, for the delivery of gene(s) into the chromosome of Gram-negative bacteria and for generating saturated mutagenesis libraries in gene function studies. These delivery vectors endow the user with the possibility of easy cloning and subsequent insertion of functional cargoes with three different antibiotic-resistance markers (kanamycin, streptomycin, and gentamicin). After validating the pBAMD vectors in the environmental bacterium Pseudomonas putida KT2440, their use was also illustrated by inserting the entire poly(3-hydroxybutyrate) (PHB) synthesis pathway from Cupriavidus necator in the chromosome of a phosphotransacetylase mutant of Escherichia coli. PHB is a completely biodegradable polyester with a number of industrial applications that make it attractive as a potential replacement of oil-based plastics. The non-selective nature of chromosomal insertions of the biosynthetic genes was evidenced by a large landscape of PHB synthesis levels in independent clones. One clone was selected and further characterized as a microbial cell factory for PHB accumulation, and it achieved polymer accumulation levels comparable to those of a plasmid-bearing recombinant. Taken together, our results demonstrate that the new mini-Tn5-vectors can be used to confer interesting phenotypes in Gram-negative bacteria that would be very difficult to engineer through direct manipulation of the structural genes.

  8. Turnstiles and bifurcators: the disequilibrium converting engines that put metabolism on the road.

    Branscomb, Elbert; Russell, Michael J

    2013-02-01

    The Submarine Hydrothermal Alkaline Spring Theory for the emergence of life holds that it is the ordered delivery of hydrogen and methane in alkaline hydrothermal solutions at a spontaneously precipitated inorganic osmotic and catalytic membrane to the carbon dioxide and other electron acceptors in the earliest acidulous cool ocean that, through these gradients, drove life into being. That such interactions between hydrothermal fuels and potential oxidants have so far not been accomplished in the lab is because some steps along the necessary metabolic pathways are endergonic and must therefore be driven by being coupled to thermodynamically larger exergonic processes. But coupling of this kind is far from automatic and it is not enough to merely sum the ΔGs of two supposedly coupled reactions and show their combined thermodynamic viability. An exergonic reaction will not drive an endergonic one unless 'forced' to do so by being tied to it mechanistically via an organized "engine" of "Free Energy Conversion" (FEC). Here we discuss the thermodynamics of FEC and advance proposals regarding the nature and roles of the FEC devices that could, in principle, have arisen spontaneously in the alkaline hydrothermal context and have forced the onset of a protometabolism. The key challenge is to divine what these initial engines of life were in physicochemical terms and as part of that, what structures provided the first "turnstile-like" mechanisms needed to couple the partner processes in free energy conversion; in particular to couple the dissipation of geochemically given gradients to, say, the reduction of CO(2) to formate and the generation of a pyrophosphate disequilibrium. This article is part of a Special Issue entitled: The evolutionary aspects of bioenergetic systems. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. Camelina sativa: An ideal platform for the metabolic engineering and field production of industrial lipids.

    Bansal, Sunil; Durrett, Timothy P

    2016-01-01

    Triacylglycerols (TAG) containing modified fatty acids with functionality beyond those found in commercially grown oil seed crops can be used as feedstocks for biofuels and bio-based materials. Over the years, advances have been made in transgenically engineering the production of various modified fatty acids in the model plant Arabidopsis thaliana. However, the inability to produce large quantities of transgenic seed has limited the functional testing of the modified oil. In contrast, the emerging oil seed crop Camelina sativa possesses important agronomic traits that recommend it as an ideal production platform for biofuels and industrial feedstocks. Camelina possesses low water and fertilizer requirements and is capable of yields comparable to other oil seed crops, particularly under stress conditions. Importantly, its relatively short growing season enables it to be grown as part of a double cropping system. In addition to these valuable agronomic features, Camelina is amenable to rapid metabolic engineering. The development of a simple and effective transformation method, combined with the availability of abundant transcriptomic and genomic data, has allowed the generation of transgenic Camelina lines capable of synthesizing high levels of unusual lipids. In some cases these levels have surpassed what was achieved in Arabidopsis. Further, the ability to use Camelina as a crop production system has allowed for the large scale growth of transgenic oil seed crops, enabling subsequent physical property testing. The application of new techniques such as genome editing will further increase the suitability of Camelina as an ideal platform for the production of biofuels and bio-materials. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  10. New Transposon Tools Tailored for Metabolic Engineering of Gram-Negative Microbial Cell Factories

    Martínez-García, Esteban; Aparicio, Tomás; Lorenzo, Víctor de; Nikel, Pablo I., E-mail: pablo.nikel@cnb.csic.es [Systems and Synthetic Biology Program, Centro Nacional de Biotecnología (CNB-CSIC), Madrid (Spain)

    2014-10-28

    Re-programming microorganisms to modify their existing functions and/or to bestow bacteria with entirely new-to-Nature tasks have largely relied so far on specialized molecular biology tools. Such endeavors are not only relevant in the burgeoning metabolic engineering arena but also instrumental to explore the functioning of complex regulatory networks from a fundamental point of view. À la carte modification of bacterial genomes thus calls for novel tools to make genetic manipulations easier. We propose the use of a series of new broad-host-range mini-Tn5-vectors, termed pBAMDs, for the delivery of gene(s) into the chromosome of Gram-negative bacteria and for generating saturated mutagenesis libraries in gene function studies. These delivery vectors endow the user with the possibility of easy cloning and subsequent insertion of functional cargoes with three different antibiotic-resistance markers (kanamycin, streptomycin, and gentamicin). After validating the pBAMD vectors in the environmental bacterium Pseudomonas putida KT2440, their use was also illustrated by inserting the entire poly(3-hydroxybutyrate) (PHB) synthesis pathway from Cupriavidus necator in the chromosome of a phosphotransacetylase mutant of Escherichia coli. PHB is a completely biodegradable polyester with a number of industrial applications that make it attractive as a potential replacement of oil-based plastics. The non-selective nature of chromosomal insertions of the biosynthetic genes was evidenced by a large landscape of PHB synthesis levels in independent clones. One clone was selected and further characterized as a microbial cell factory for PHB accumulation, and it achieved polymer accumulation levels comparable to those of a plasmid-bearing recombinant. Taken together, our results demonstrate that the new mini-Tn5-vectors can be used to confer interesting phenotypes in Gram-negative bacteria that would be very difficult to engineer through direct manipulation of the structural genes.

  11. Ensemble Modeling for Robustness Analysis in engineering non-native metabolic pathways.

    Lee, Yun; Lafontaine Rivera, Jimmy G; Liao, James C

    2014-09-01

    Metabolic pathways in cells must be sufficiently robust to tolerate fluctuations in expression levels and changes in environmental conditions. Perturbations in expression levels may lead to system failure due to the disappearance of a stable steady state. Increasing evidence has suggested that biological networks have evolved such that they are intrinsically robust in their network structure. In this article, we presented Ensemble Modeling for Robustness Analysis (EMRA), which combines a continuation method with the Ensemble Modeling approach, for investigating the robustness issue of non-native pathways. EMRA investigates a large ensemble of reference models with different parameters, and determines the effects of parameter drifting until a bifurcation point, beyond which a stable steady state disappears and system failure occurs. A pathway is considered to have high bifurcational robustness if the probability of system failure is low in the ensemble. To demonstrate the utility of EMRA, we investigate the bifurcational robustness of two synthetic central metabolic pathways that achieve carbon conservation: non-oxidative glycolysis and reverse glyoxylate cycle. With EMRA, we determined the probability of system failure of each design and demonstrated that alternative designs of these pathways indeed display varying degrees of bifurcational robustness. Furthermore, we demonstrated that target selection for flux improvement should consider the trade-offs between robustness and performance. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Spatial separation of photosynthesis and ethanol production by cell type-specific metabolic engineering of filamentous cyanobacteria.

    Ehira, Shigeki; Takeuchi, Takuto; Higo, Akiyoshi

    2018-02-01

    Cyanobacteria, which perform oxygenic photosynthesis, have drawn attention as hosts for the direct production of biofuels and commodity chemicals from CO 2 and H 2 O using light energy. Although cyanobacteria capable of producing diverse chemicals have been generated by metabolic engineering, anaerobic non-photosynthetic culture conditions are often necessary for their production. In this study, we conducted cell type-specific metabolic engineering of the filamentous cyanobacterium Anabaena sp. PCC 7120, which forms a terminally differentiated cell called a heterocyst with a semi-regular spacing of 10-15 cells. Because heterocysts are specialized cells for nitrogen fixation, the intracellular oxygen level of heterocysts is maintained very low even when adjacent cells perform oxygenic photosynthesis. Pyruvate decarboxylase of Zymomonas mobilis and alcohol dehydrogenase of Synechocystis sp. PCC 6803 were exclusively expressed in heterocysts. Ethanol production was concomitant with nitrogen fixation in genetically engineered Anabaena sp. PCC 7120. Engineering of carbon metabolism in heterocysts improved ethanol production, and strain ET14, with an extra copy of the invB gene expressed from a heterocyst-specific promoter, produced 130.9 mg L -1 of ethanol after 9 days. ET14 produced 1681.9 mg L -1 of ethanol by increasing the CO 2 supply. Ethanol production per heterocyst cell was approximately threefold higher than that per cell of unicellular cyanobacterium. This study demonstrates the potential of heterocysts for anaerobic production of biofuels and commodity chemicals under oxygenic photosynthetic conditions.

  13. An Alternative Educational Approach for an Inorganic Chemistry Laboratory Course in Industrial and Chemical Engineering

    Garces, Andres; Sanchez-Barba, Luis Fernando

    2011-01-01

    We describe an alternative educational approach for an inorganic chemistry laboratory module named "Experimentation in Chemistry", which is included in Industrial Engineering and Chemical Engineering courses. The main aims of the new approach were to reduce the high levels of failure and dropout on the module and to make the content match the…

  14. An Integrated Neuroscience and Engineering Approach to Classifying Human Brain-States

    2015-12-22

    AFRL-AFOSR-VA-TR-2016-0037 An Integrated Neuroscience and Engineering Approach to Classifying Human Brain-States Adrian Lee UNIVERSITY OF WASHINGTON...to 14-09-2015 4. TITLE AND SUBTITLE An Integrated Neuroscience and Engineering Approach to Classifying Human Brain- States 5a.  CONTRACT NUMBER 5b...specific cognitive states remains elusive, owing perhaps to limited crosstalk between the fields of neuroscience and engineering. Here, we report a

  15. N-Glycosylation optimization of recombinant antibodies in CHO cell through process and metabolic engineering

    Fan, Yuzhou

    , analysis, control and optimization of N-glycosylation were thoroughly reviewed. In particular, how to control and optimize N-glycosylation in CHO cells was exclusively studied. The main focus of this PhD project is to find effective approaches of modulating N-glycosylation of CHO-derived recombinant...... galactose as feed additives, changing process parameters such as seeding density and cultivation duration are all demonstrated to be effective. The causal explanation of their impact on glycosylation can be various, including product, metabolism, proteome and physiology-associated mechanism. In the middle...... part of the thesis, both literature reviews and experimental applications were provided to demonstrate how to use omics data and implement systems biology to understand biological activities, especially N-glycosylation in CHO cells. In the last part of the thesis, the second strategy that apply genetic...

  16. Arctic Ice Management: an integrated approach to climate engineering

    Desch, S. J.; Hartnett, H. E.; Groppi, C. E.; Romaniello, S. J.

    2017-12-01

    The warming climate is having the most rapid and pronounced effects in the high Arctic. The loss of Arctic sea ice is not only changing the physical oceanography of the Arctic Ocean and its coastlines; it is also promoting new conversations about the dangers and benefits for trade, transportation, and industry in the Arctic. The rate of decrease of summer sea ice in the Arctic is currently -300 km3 yr-1, a rate that will lead to complete loss of end-summer sea ice as soon as 2030. Preventing the strong positive feedbacks and increased warming due to sea ice albedo loss must be an important component of climate mitigation strategies. Here, we explore a direct engineering approach we call Arctic Ice Management (AIM) to reduce the loss of Arctic sea ice. We predict that pumping seawater onto the ice surface during the Arctic winter using wind-powered pumps can thicken sea ice by up to 1 m per year, reversing the current loss rates and prolonging the time until the Arctic Ocean is ice-free. Thickening sea ice would not change CO2 levels, which are the underlying cause of ice loss, but it would prevent some of the strongest feedbacks and would buy time to develop the tools and governance systems necessary to achieve carbon-neutrality. We advocate exploration of AIM as a mitigation strategy employed in parallel with CO2 reduction efforts. The opportunity and risk profiles of AIM differ from other geoengineering proposals. While similar in principle to solar radiation management, AIM may present fewer large-scale environmental risks. AIM is separate from greenhouse gas emission reduction or sequestration, but might help prevent accelerated release of methane from thawing permafrost. Further, AIM might be usefully employed at regional and local scales to preserve Arctic ecosystems and possibly reduce the effects of ice-loss induced coastal erosion. Through presentation of the AIM concept, we hope to spark new conversations between scientists, stakeholders, and decision

  17. Accelerating cocaine metabolism as an approach to the treatment of cocaine abuse and toxicity

    Schindler, Charles W; Goldberg, Steven R

    2012-01-01

    One pharmacokinetic approach to the treatment of cocaine abuse and toxicity involves the development of compounds that can be safely administered to humans and that accelerate the metabolism of cocaine to inactive components. Catalytic antibodies have been developed and shown to accelerate cocaine metabolism, but their catalytic efficiency for cocaine is relatively low. Mutations of human butyrylcholinesterase and a bacterial cocaine esterase found in the soil of coca plants have also been developed. These compounds accelerate cocaine metabolism and antagonize the behavioral and toxic effects of cocaine in animal models. Of these two approaches, the human butyrylcholinesterase mutants show the most immediate promise as they would not be expected to evoke an immune response in humans. PMID:22300096

  18. Metabolic engineering pathways for rare sugars biosynthesis, physiological functionalities, and applications-a review.

    Bilal, Muhammad; Iqbal, Hafiz M N; Hu, Hongbo; Wang, Wei; Zhang, Xuehong

    2017-06-29

    Biomolecules like rare sugars and their derivatives are referred to as monosaccharides particularly uncommon in nature. Remarkably, many of them have various known physiological functions and biotechnological applications in cosmetics, nutrition, and pharmaceutical industries. Also, they can be exploited as starting materials for synthesizing fascinating natural bioproducts with significant biological activities. Regrettably, most of the rare sugars are quite expensive, and their synthetic chemical routes are both limited and economically unfeasible due to expensive raw materials. On the other hand, their production by enzymatic means often suffers from low space-time yields and high catalyst costs due to hasty enzyme denaturation/degradation. In this context, biosynthesis of rare sugars with industrial importance is receiving renowned scientific attention, across the globe. Moreover, the utilization of renewable resources as energy sources via microbial fermentation or microbial metabolic engineering has appeared a new tool. This article presents a comprehensive review of physiological functions and biotechnological applications of rare ketohexoses and aldohexoses, including D-psicose, D-tagatose, L-tagatose, D-sorbose, L-fructose, D-allose, L-glucose, D-gulose, L-talose, L-galactose, and L-fucose. Novel in-vivo recombination pathways based on aldolase and phosphatase for the biosynthesis of rare sugars, particularly D-psicose and D-sorbose using robust microbial strains are also deliberated.

  19. Genetic and metabolic engineering for microbial production of poly-γ-glutamic acid.

    Cao, Mingfeng; Feng, Jun; Sirisansaneeyakul, Sarote; Song, Cunjiang; Chisti, Yusuf

    2018-05-28

    Poly-γ-glutamic acid (γ-PGA) is a natural biopolymer of glutamic acid. The repeating units of γ-PGA may be derived exclusively from d-glutamic acid, or l-glutamic acid, or both. The monomer units are linked by amide bonds between the α-amino group and the γ-carboxylic acid group. γ-PGA is biodegradable, edible and water-soluble. It has numerous existing and emerging applications in processing of foods, medicines and cosmetics. This review focuses on microbial production of γ-PGA via genetically and metabolically engineered recombinant bacteria. Strategies for improving production of γ-PGA include modification of its biosynthesis pathway, enhancing the production of its precursor (glutamic acid), and preventing loss of the precursor to competing byproducts. These and other strategies are discussed. Heterologous synthesis of γ-PGA in industrial bacterial hosts that do not naturally produce γ-PGA is discussed. Emerging trends and the challenges affecting the production of γ-PGA are reviewed. Copyright © 2018. Published by Elsevier Inc.

  20. Enhancing GDP-fucose production in recombinant Escherichia coli by metabolic pathway engineering.

    Zhai, Yafei; Han, Donglei; Pan, Ying; Wang, Shuaishuai; Fang, Junqiang; Wang, Peng; Liu, Xian-wei

    2015-02-01

    Guanosine 5'-diphosphate (GDP)-fucose is the indispensible donor substrate for fucosyltransferase-catalyzed synthesis of fucose-containing biomolecules, which have been found involving in various biological functions. In this work, the salvage pathway for GDP-fucose biosynthesis from Bacterioides fragilis was introduced into Escherichia coli. Besides, the biosynthesis of guanosine 5'-triphosphate (GTP), an essential substrate for GDP-fucose biosynthesis, was enhanced via overexpression of enzymes involved in the salvage pathway of GTP biosynthesis. The production capacities of metabolically engineered strains bearing different combinations of recombinant enzymes were compared. The shake flask fermentation of the strain expressing Fkp, Gpt, Gmk and Ndk obtained the maximum GDP-fucose content of 4.6 ± 0.22 μmol/g (dry cell mass), which is 4.2 fold that of the strain only expressing Fkp. Through fed-batch fermentation, the GDP-fucose content further rose to 6.6 ± 0.14 μmol/g (dry cell mass). In addition to a better productivity than previous fermentation processes based on the de novo pathway for GDP-fucose biosynthesis, the established schemes in this work also have the advantage to be a potential avenue to GDP-fucose analogs encompassing chemical modification on the fucose residue. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Metabolic engineering of Pseudomonas putida KT2440 for the production of para-hydroxy benzoic acid

    Shiqin Yu

    2016-11-01

    Full Text Available para-hydroxy benzoic acid (PHBA is the key component for preparing parabens, a common preservatives in food, drugs and personal care products, as well as high performance bioplastics such as liquid crystal polymers (LCP. Pseudomonas putida KT2440 was engineered to produce PHBA from glucose via the shikimate pathway intermediate chorismate. To obtain the PHBA production strain, chorismate lyase UbiC from Escherichia coli and a feedback resistant 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase encoded by gene aroGD146N were overexpressed individually and simultaneously. In addition, genes related to product degradation (pobA or competing for the precursor chorismate (pheA and trpE were deleted from the genome. To further improve PHBA production, the glucose metabolism repressor hexR was knocked out in order to increase erythrose-4- phosphate and NAPH supply. The best strain achieved a maximum titre of 1.73 g L-1 and a carbon yield of 18.1 % (C-mol C-mol-1 in a non-optimized fed-batch fermentation. This is to date the highest PHBA concentration produced by P. putida using a chorismate lyase.

  2. Analysis and design of a genetic circuit for dynamic metabolic engineering.

    Anesiadis, Nikolaos; Kobayashi, Hideki; Cluett, William R; Mahadevan, Radhakrishnan

    2013-08-16

    Recent advances in synthetic biology have equipped us with new tools for bioprocess optimization at the genetic level. Previously, we have presented an integrated in silico design for the dynamic control of gene expression based on a density-sensing unit and a genetic toggle switch. In the present paper, analysis of a serine-producing Escherichia coli mutant shows that an instantaneous ON-OFF switch leads to a maximum theoretical productivity improvement of 29.6% compared to the mutant. To further the design, global sensitivity analysis is applied here to a mathematical model of serine production in E. coli coupled with a genetic circuit. The model of the quorum sensing and the toggle switch involves 13 parameters of which 3 are identified as having a significant effect on serine concentration. Simulations conducted in this reduced parameter space further identified the optimal ranges for these 3 key parameters to achieve productivity values close to the maximum theoretical values. This analysis can now be used to guide the experimental implementation of a dynamic metabolic engineering strategy and reduce the time required to design the genetic circuit components.

  3. Production of ethanol from thin stillage by metabolically engineered Escherichia coli.

    Gonzalez, Ramon; Campbell, Paul; Wong, Matthew

    2010-03-01

    Thin stillage is a by-product generated in large amounts during the production of ethanol that is rich in carbon sources like glycerol, glucose and maltose. Unfortunately, the fermentation of thin stillage results in a mixture of organic acids and ethanol and minimum utilization of glycerol, the latter a compound that can represent up to 80% of the available substrates in this stream. We report here the efficient production of ethanol from thin stillage by a metabolically engineered strain of Escherichia coli. Simultaneous utilization of glycerol and sugars was achieved by overexpressing either the fermentative or the respiratory glycerol-utilization pathway. However, amplification of the fermentative pathway (encoded by gldA and dhaKLM) led to more efficient consumption of glycerol and promoted the synthesis of reduced products, including ethanol. A previously constructed strain, EH05, containing mutations that prevented the accumulation of competing by-products (i.e. lactate, acetate, and succinate) and overexpressing the fermentative pathway for glycerol utilization [i.e. strain EH05 (pZSKLMgldA)], efficiently converted thin stillage supplemented with only mineral salts to ethanol at yields close to 85% of the theoretical maximum. Ethanol accounted for about 90% (w/w) of the product mixture. These results, along with the comparable performance of strain EH05 (pZSKLMgldA) in 0.5 and 5 l fermenters, indicate a great potential for the adoption of this process by the biofuels industry.

  4. Lifestyle and metabolic approaches to maximizing erectile and vascular health.

    Meldrum, D R; Gambone, J C; Morris, M A; Esposito, K; Giugliano, D; Ignarro, L J

    2012-01-01

    Oxidative stress and inflammation, which disrupt nitric oxide (NO) production directly or by causing resistance to insulin, are central determinants of vascular diseases including ED. Decreased vascular NO has been linked to abdominal obesity, smoking and high intakes of fat and sugar, which all cause oxidative stress. Men with ED have decreased vascular NO and circulating and cellular antioxidants. Oxidative stress and inflammatory markers are increased in men with ED, and all increase with age. Exercise increases vascular NO, and more frequent erections are correlated with decreased ED, both in part due to stimulation of endothelial NO production by shear stress. Exercise and weight loss increase insulin sensitivity and endothelial NO production. Potent antioxidants or high doses of weaker antioxidants increase vascular NO and improve vascular and erectile function. Antioxidants may be particularly important in men with ED who smoke, are obese or have diabetes. Omega-3 fatty acids reduce inflammatory markers, decrease cardiac death and increase endothelial NO production, and are therefore critical for men with ED who are under age 60 years, and/or have diabetes, hypertension or coronary artery disease, who are at increased risk of serious or even fatal cardiac events. Phosphodiesterase inhibitors have recently been shown to improve antioxidant status and NO production and allow more frequent and sustained penile exercise. Some angiotensin II receptor blockers decrease oxidative stress and improve vascular and erectile function and are therefore preferred choices for lowering blood pressure in men with ED. Lifestyle modifications, including physical and penile-specific exercise, weight loss, omega-3 and folic acid supplements, reduced intakes of fat and sugar, and improved antioxidant status through diet and/or supplements should be integrated into any comprehensive approach to maximizing erectile function, resulting in greater overall success and patient

  5. Metabolic Interaction between Urea Cycle and Citric Acid Cycle Shunt: A Guided Approach

    Pesi, Rossana; Balestri, Francesco; Ipata, Piero L.

    2018-01-01

    This article is a guided pedagogical approach, devoted to postgraduate students specializing in biochemistry, aimed at presenting all single reactions and overall equations leading to the metabolic interaction between ureagenesis and citric acid cycle to be incorporated into a two-three lecture series about the interaction of urea cycle with other…

  6. An Industrial Engineering Approach to Cost Containment of Pharmacy Education.

    Duncan, Wendy; Bottenberg, Michelle; Chase, Marilea; Chesnut, Renae; Clarke, Cheryl; Schott, Kathryn; Torry, Ronald; Welty, Tim

    2015-11-25

    A 2-semester project explored employing teams of fourth-year industrial engineering students to optimize some of our academic management processes. Results included significant cost savings and increases in efficiency, effectiveness, and student and faculty satisfaction. While we did not adopt all of the students' recommendations, we did learn some important lessons. For example, an initial investment of time in developing a mutually clear understanding of the problems, constraints, and goals maximizes the value of industrial engineering analysis and recommendations. Overall, industrial engineering was a valuable tool for optimizing certain academic management processes.

  7. Engineering the just war: examination of an approach to teaching engineering ethics.

    Haws, David R

    2006-04-01

    The efficiency of engineering applied to civilian projects sometimes threatens to run away with the social agenda, but in military applications, engineering often adds a devastating sleekness to the inevitable destruction of life. The relative crudeness of terrorism (e.g., 9/11) leaves a stark after-image, which belies the comparative insignificance of random (as opposed to orchestrated) belligerence. Just as engineering dwarfs the bricolage of vernacular design 'moving us past the appreciation of brush-strokes, so to speak' the scale of engineered destruction makes it difficult to focus on the charred remains of individual lives. Engineers need to guard against the inappropriate military subsumption of their effort. Fortunately, the ethics of warfare has been an ongoing topic of discussion for millennia. This paper will examine the university core class I've developed (The Moral Dimensions of Technology) to meet accreditation requirements in engineering ethics, and the discussion with engineering and non-engineering students focused by the life of electrical engineer Vannevar Bush, with selected readings in moral philosophy from the Dao de Jing, Lao Tze, Cicero, Aurelius Augustinus, Kant, Annette Baier, Peter Singer, Elizabeth Anscombe, Philippa Foot, and Judith Thomson.

  8. Expert System Approach For Generating And Evaluating Engine Design Alternatives

    Shen, Stewart N. T.; Chew, Meng-Sang; Issa, Ghassan F.

    1989-03-01

    Artificial intelligence is becoming an increasingly important subject of study for computer scientists, engineering designers, as well as professionals in other fields. Even though AI technology is a relatively new discipline, many of its concepts have already found practical applications. Expert systems, in particular, have made significant contributions to technologies in such fields as business, medicine, engineering design, chemistry, and particle physics. This paper describes an expert system developed to aid the mechanical designer with the preliminary design of variable-stroke internal-combustion engines. The expert system accomplished its task by generating and evaluating a large number of design alternatives represented in the form of graphs. Through the application of structural and design rules directly to the graphs, optimal and near optimal preliminary design configurations of engines are deduced.

  9. Biological augmentation and tissue engineering approaches in meniscus surgery.

    Moran, Cathal J; Busilacchi, Alberto; Lee, Cassandra A; Athanasiou, Kyriacos A; Verdonk, Peter C

    2015-05-01

    The purpose of this review was to evaluate the role of biological augmentation and tissue engineering strategies in meniscus surgery. Although clinical (human), preclinical (animal), and in vitro tissue engineering studies are included here, we have placed additional focus on addressing preclinical and clinical studies reported during the 5-year period used in this review in a systematic fashion while also providing a summary review of some important in vitro tissue engineering findings in the field over the past decade. A search was performed on PubMed for original works published from 2009 to March 31, 2014 using the term "meniscus" with all the following terms: "scaffolds," "constructs," "cells," "growth factors," "implant," "tissue engineering," and "regenerative medicine." Inclusion criteria were the following: English-language articles and original clinical, preclinical (in vivo), and in vitro studies of tissue engineering and regenerative medicine application in knee meniscus lesions published from 2009 to March 31, 2014. Three clinical studies and 18 preclinical studies were identified along with 68 tissue engineering in vitro studies. These reports show the increasing promise of biological augmentation and tissue engineering strategies in meniscus surgery. The role of stem cell and growth factor therapy appears to be particularly useful. A review of in vitro tissue engineering studies found a large number of scaffold types to be of promise for meniscus replacement. Limitations include a relatively low number of clinical or preclinical in vivo studies, in addition to the fact there is as yet no report in the literature of a tissue-engineered meniscus construct used clinically. Neither does the literature provide clarity on the optimal meniscus scaffold type or biological augmentation with which meniscus repair or replacement would be best addressed in the future. There is increasing focus on the role of mechanobiology and biomechanical and

  10. A resilience engineering approach to assess major accident risks

    Hollnagel, E.

    2013-01-01

    This chapter describes how the principles of Resilience Engineering can be used to make a risk assessment of an Integrated Operations (IO) scenario. It refers to the case study provided in Chapter 12.......This chapter describes how the principles of Resilience Engineering can be used to make a risk assessment of an Integrated Operations (IO) scenario. It refers to the case study provided in Chapter 12....

  11. Hybrid Engine Powered City Car: Fuzzy Controlled Approach

    Rahman, Ataur; Mohiuddin, AKM; Hawlader, MNA; Ihsan, Sany

    2017-03-01

    This study describes a fuzzy controlled hybrid engine powered car. The car is powered by the lithium ion battery capacity of 1000 Wh is charged by the 50 cc hybrid engine and power regenerative mode. The engine is operated with lean mixture at 3000 rpm to charge the battery. The regenerative mode that connects with the engine generates electrical power of 500-600 W for the deceleration of car from 90 km/h to 20 km/h. The regenerated electrical power has been used to power the air-conditioning system and to meet the other electrical power. The battery power only used to propel the car. The regenerative power also found charging the battery for longer operation about 40 minutes and more. The design flexibility of this vehicle starts with whole-vehicle integration based on radical light weighting, drag reduction, and accessory efficiency. The energy efficient hybrid engine cut carbon dioxide (CO2) and nitrogen oxides (N2O) emission about 70-80% as the loads on the crankshaft such as cam-follower and its associated rotating components are replaced by electromagnetic systems, and the flywheel, alternator and starter motor are replaced by a motor generator. The vehicle was tested and found that it was able to travel 70 km/litre with the power of hybrid engine.

  12. Engineering spinal fusion: evaluating ceramic materials for cell based tissue engineered approaches

    Wilson, C.E.

    2011-01-01

    The principal aim of this thesis was to advance the development of tissue engineered posterolateral spinal fusion by investigating the potential of calcium phosphate ceramic materials to support cell based tissue engineered bone formation. This was accomplished by developing several novel model

  13. Terpene metabolic engineering via nuclear or chloroplast genomes profoundly and globally impacts off-target pathways through metabolite signalling.

    Pasoreck, Elise K; Su, Jin; Silverman, Ian M; Gosai, Sager J; Gregory, Brian D; Yuan, Joshua S; Daniell, Henry

    2016-09-01

    The impact of metabolic engineering on nontarget pathways and outcomes of metabolic engineering from different genomes are poorly understood questions. Therefore, squalene biosynthesis genes FARNESYL DIPHOSPHATE SYNTHASE (FPS) and SQUALENE SYNTHASE (SQS) were engineered via the Nicotiana tabacum chloroplast (C), nuclear (N) or both (CN) genomes to promote squalene biosynthesis. SQS levels were ~4300-fold higher in C and CN lines than in N, but all accumulated ~150-fold higher squalene due to substrate or storage limitations. Abnormal leaf and flower phenotypes, including lower pollen production and reduced fertility, were observed regardless of the compartment or level of transgene expression. Substantial changes in metabolomes of all lines were observed: levels of 65-120 unrelated metabolites, including the toxic alkaloid nicotine, changed by as much as 32-fold. Profound effects of transgenesis on nontarget gene expression included changes in the abundance of 19 076 transcripts by up to 2000-fold in CN; 7784 transcripts by up to 1400-fold in N; and 5224 transcripts by as much as 2200-fold in C. Transporter-related transcripts were induced, and cell cycle-associated transcripts were disproportionally repressed in all three lines. Transcriptome changes were validated by qRT-PCR. The mechanism underlying these large changes likely involves metabolite-mediated anterograde and/or retrograde signalling irrespective of the level of transgene expression or end product, due to imbalance of metabolic pools, offering new insight into both anticipated and unanticipated consequences of metabolic engineering. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  14. A Novel Evolutionary Engineering Design Approach for Mixed-Domain Systems

    Fan, Zhun; Hu, J.; Seo, K.

    2004-01-01

    This paper presents an approach to engineering design of mixed-domain dynamic systems. The approach aims at system-level design and has two key features: first, it generates engineering designs that satisfy predefined specifications in an automatic manner; second, it can design systems belonging ...... often encountered in evolutionary computation, a HFC (Hierarchical Fair Competition) model is adopted in this work. Examples of an analog filter design and a MEM filter design illustrate the application of the approach....

  15. Reducing Diesel Engine Emission Using Reactivity Controlled Approach

    Osama Hasib Ghazal

    2018-01-01

    Full Text Available Several automobile manufacturers are interested in investigating of dual fuel internal combustion engines, due to high efficiencand low emissions. Many alternative fuels have been used in dual fuel mode for IC engine, such as methane, hydrogen, and natural gas. In the present study, a reactivity controlled compression ignition (RCCI engine using gasoline/diesel (G/D dual fuel has been investigated. The effectof mixing gasoline with diesel fuel on combustion characteristic, engine performance and emissions has been studied. The gasoline was injected in the engine intake port, to produce a homogeneous mixture with air. The diesel fuel was injected directly to the combustion chamber during compression stroke to initiate the combustion process. A direct injection compression ignition engine has been built and simulated using ANSYS Forte professional code. The gasoline amount in the simulation varied from (50%-80% by volume. The diesel fuel was injected to the cylinder in two stages. The model has been validated and calibrated for neat diesel fuel using available data from the literature. The results show that the heat release rate and the cylinder pressure increased when the amount of added gasoline is between 50%-60% volume of the total injected fuels, compared to the neat diesel fuel. Further addition of gasoline will have a contrary effect. In addition, the combustion duration is extended drastically when the gasoline ratio is higher than 60% which results in an incomplete combustion. The NO emission decreased drastically as the gasoline ratio increased. Moreover, addition of gasoline to the mixture increased the engine power, thermal efficienc and combustion efficienc compared to neat diesel fuel.

  16. The "parallel pathway": a novel nutritional and metabolic approach to cancer patients.

    Muscaritoli, Maurizio; Molfino, Alessio; Gioia, Gianfranco; Laviano, Alessandro; Rossi Fanelli, Filippo

    2011-04-01

    Cancer-associated malnutrition results from a deadly combination of anorexia, which leads to reduced food intake, and derangements of host metabolism inducing body weight loss, and hindering its reversal with nutrient supplementation. Cancer patients often experience both anorexia and weight loss, contributing to the onset of the clinical feature named as anorexia-cachexia syndrome. This condition has a negative impact upon patients' nutritional status. The pathogenesis of the anorexia-cachexia syndrome is multifactorial, and is related to: tumour-derived factors, host-derived factors inducing metabolic derangements, and side effects of anticancer therapies. In addition, the lack of awareness of cancer patients' nutritional issues and status by many oncologists, frequently results in progressive weight loss going undiagnosed until it becomes severe. The critical involvement of host inflammatory response in the development of weight loss, and, in particular, lean body mass depletion, limits the response to the provision of standard nutrition support. A novel nutritional and metabolic approach, named "parallel pathway", has been devised that may help maintain or improve nutritional status, and prevent or delay the onset of cancer cachexia. Such an approach may improve tolerance to aggressive anticancer therapies, and ameliorate the functional capacity and quality of life even in advanced disease stages. The "parallel pathway" implies a multiprofessional and multimodal approach aimed at ensuring early, appropriate and continuous nutritional and metabolic support to cancer patients in any phase of their cancer journey.

  17. Construction of expression vectors for metabolic engineering of the vanillin-producing actinomycete Amycolatopsis sp. ATCC 39116.

    Fleige, Christian; Steinbüchel, Alexander

    2014-01-01

    Amycolatopsis sp. ATCC 39116 is able to synthesize the important flavoring agent vanillin from cheap natural substrates. The bacterium is therefore of great interest for the industry and used for the fermentative production of vanillin. In order to improve the production of natural vanillin with Amycolatopsis sp. ATCC 39116, the strain has been genetically engineered to optimize the metabolic flux towards the desired product. Extensive metabolic engineering was hitherto hampered, due to the lack of genetic tools like functional promoters and expression vectors. In this study, we report the establishment of a plasmid-based gene expression system for Amycolatopsis sp. ATCC 39116 that allows a further manipulation of the genotype. Four new Escherichia coli-Amycolatopsis shuttle vectors harboring different promoter elements were constructed, and the functionality of these regulatory elements was proven by the expression of the reporter gene gusA, encoding a β-glucuronidase. Glucuronidase activity was detected in all plasmid-harboring strains, and remarkable differences in the expression strength of the reporter gene depending on the used promoter were observed. The new expression vectors will promote the further genetic engineering of Amycolatopsis sp. ATCC 39116 to get insight into the metabolic network and to improve the strain for a more efficient industrial use.

  18. Vanillin production using metabolically engineered Escherichia coli under non-growing conditions.

    Barghini, Paolo; Di Gioia, Diana; Fava, Fabio; Ruzzi, Maurizio

    2007-04-16

    Vanillin is one of the most important aromatic flavour compounds used in the food and cosmetic industries. Natural vanillin is extracted from vanilla beans and is relatively expensive. Moreover, the consumer demand for natural vanillin highly exceeds the amount of vanillin extracted by plant sources. This has led to the investigation of other routes to obtain this flavour such as the biotechnological production from ferulic acid. Studies concerning the use of engineered recombinant Escherichia coli cells as biocatalysts for vanillin production are described in the literature, but yield optimization and biotransformation conditions have not been investigated in details. Effect of plasmid copy number in metabolic engineering of E. coli for the synthesis of vanillin has been evaluated by the use of genes encoding feruloyl-CoA synthetase and feruloyl hydratase/aldolase from Pseudomonas fluorescens BF13. The higher vanillin production yield was obtained using resting cells of E. coli strain JM109 harbouring a low-copy number vector and a promoter exhibiting a low activity to drive the expression of the catabolic genes. Optimization of the bioconversion of ferulic acid to vanillin was accomplished by a response surface methodology. The experimental conditions that allowed us to obtain high values for response functions were 3.3 mM ferulic acid and 4.5 g/L of biomass, with a yield of 70.6% and specific productivity of 5.9 micromoles/g x min after 3 hours of incubation. The final concentration of vanillin in the medium was increased up to 3.5 mM after a 6-hour incubation by sequential spiking of 1.1 mM ferulic acid. The resting cells could be reused up to four times maintaining the production yield levels over 50%, thus increasing three times the vanillin obtained per gram of biomass. Ferulic acid can be efficiently converted to vanillin, without accumulation of undesirable vanillin reduction/oxidation products, using E. coli JM109 cells expressing genes from the ferulic

  19. Vanillin production using metabolically engineered Escherichia coli under non-growing conditions

    Fava Fabio

    2007-04-01

    Full Text Available Abstract Background Vanillin is one of the most important aromatic flavour compounds used in the food and cosmetic industries. Natural vanillin is extracted from vanilla beans and is relatively expensive. Moreover, the consumer demand for natural vanillin highly exceeds the amount of vanillin extracted by plant sources. This has led to the investigation of other routes to obtain this flavour such as the biotechnological production from ferulic acid. Studies concerning the use of engineered recombinant Escherichia coli cells as biocatalysts for vanillin production are described in the literature, but yield optimization and biotransformation conditions have not been investigated in details. Results Effect of plasmid copy number in metabolic engineering of E. coli for the synthesis of vanillin has been evaluated by the use of genes encoding feruloyl-CoA synthetase and feruloyl hydratase/aldolase from Pseudomonas fluorescens BF13. The higher vanillin production yield was obtained using resting cells of E. coli strain JM109 harbouring a low-copy number vector and a promoter exhibiting a low activity to drive the expression of the catabolic genes. Optimization of the bioconversion of ferulic acid to vanillin was accomplished by a response surface methodology. The experimental conditions that allowed us to obtain high values for response functions were 3.3 mM ferulic acid and 4.5 g/L of biomass, with a yield of 70.6% and specific productivity of 5.9 μmoles/g × min after 3 hours of incubation. The final concentration of vanillin in the medium was increased up to 3.5 mM after a 6-hour incubation by sequential spiking of 1.1 mM ferulic acid. The resting cells could be reused up to four times maintaining the production yield levels over 50%, thus increasing three times the vanillin obtained per gram of biomass. Conclusion Ferulic acid can be efficiently converted to vanillin, without accumulation of undesirable vanillin reduction/oxidation products

  20. Metabolic adaptation of a human pathogen during chronic infections - a systems biology approach

    Thøgersen, Juliane Charlotte

    modeling to uncover how human pathogens adapt to the human host. Pseudomonas aeruginosa infections in cystic fibrosis patients are used as a model system for under-­‐ standing these adaptation processes. The exploratory systems biology approach facilitates identification of important phenotypes...... by classical molecular biology approaches where genes and reactions typically are investigated in a one to one relationship. This thesis is an example of how mathematical approaches and modeling can facilitate new biologi-­‐ cal understanding and provide new surprising ideas to important biological processes....... and metabolic pathways that are necessary or related to establishment of chronic infections. Archetypal analysis showed to be successful in extracting relevant phenotypes from global gene expression da-­‐ ta. Furthermore, genome-­‐scale metabolic modeling showed to be useful in connecting the genotype...