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Sample records for plant metabolic engineering

  1. Key applications of plant metabolic engineering.

    Directory of Open Access Journals (Sweden)

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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. Engineering central metabolism - a grand challenge for plant biologists.

    Science.gov (United States)

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

    2016-12-22

    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 using fast-to-transform model plants as test beds. This approach can be realised by coupling new molecular tools to incorporate multiple transgenes in nuclear and plastid genomes with computational modelling to design the engineering strategy and to understand the metabolic phenotype of the engineered organism. We also envisage that mutagenesis could be used to fine-tune the balance between the endogenous metabolic network and the introduced enzymes. Finally, we emphasise the importance of considering the plant as a whole system and not isolated organs: the greatest increase in crop productivity will be achieved if both source and sink metabolism are engineered.

  4. Metabolic engineering of plants using a disarmed potyvirus vector

    OpenAIRE

    Majer, Eszter

    2016-01-01

    [EN] Plant viruses are obligate intracellular parasites which were used to develop recombinant plant virus vectors to express heterologous proteins and to modify endogenous metabolic pathways of natural products in plants. The main limitation of many plant virus-based systems is the difficulty to co-express various heterologous proteins in the same cell with proper subcellular localization, which is a crucial question in metabolic engineering. This work provides a solution to overcome this pr...

  5. Plant Metabolic Modeling: Achieving New Insight into Metabolism and Metabolic Engineering

    Science.gov (United States)

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

    2014-01-01

    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. PMID:25344492

  6. Metabolic engineering for the production of plant isoquinoline alkaloids.

    Science.gov (United States)

    Diamond, Andrew; Desgagné-Penix, Isabel

    2016-06-01

    Several plant isoquinoline alkaloids (PIAs) possess powerful pharmaceutical and biotechnological properties. Thus, PIA metabolism and its fascinating molecules, including morphine, colchicine and galanthamine, have attracted the attention of both the industry and researchers involved in plant science, biochemistry, chemical bioengineering and medicine. Currently, access and availability of high-value PIAs [commercialized (e.g. galanthamine) or not (e.g. narciclasine)] is limited by low concentration in nature, lack of cultivation or geographic access, seasonal production and risk of overharvesting wild plant species. Nevertheless, most commercial PIAs are still extracted from plant sources. Efforts to improve the production of PIA have largely been impaired by the lack of knowledge on PIA metabolism. With the development and integration of next-generation sequencing technologies, high-throughput proteomics and metabolomics analyses and bioinformatics, systems biology was used to unravel metabolic pathways allowing the use of metabolic engineering and synthetic biology approaches to increase production of valuable PIAs. Metabolic engineering provides opportunity to overcome issues related to restricted availability, diversification and productivity of plant alkaloids. Engineered plant, plant cells and microbial cell cultures can act as biofactories by offering their metabolic machinery for the purpose of optimizing the conditions and increasing the productivity of a specific alkaloid. In this article, is presented an update on the production of PIA in engineered plant, plant cell cultures and heterologous micro-organisms.

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

    DEFF Research Database (Denmark)

    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...

  8. Metabolic engineering of micronutrients in crop plants.

    Science.gov (United States)

    Blancquaert, Dieter; De Steur, Hans; Gellynck, Xavier; Van Der Straeten, Dominique

    2017-02-01

    Micronutrient deficiency is a widespread phenomenon, most prevalent in developing countries. Being causally linked to the occurrence of a range of diseases, it affects billions of people worldwide. Enhancing the content of micronutrients in crop products through biotechnology is a promising technique to fight micronutrient malnutrition worldwide. Micronutrient fortification of food products has been implemented in a number of Western countries, but remains inaccessible for poor rural populations in a major part of the developing world. Moreover, evidence of the negative impacts of this practice on human health, at least for some vitamins, is accumulating. Biofortification of crop plants-the enhancement of vitamins and minerals through plant biotechnology-is a promising alternative or complement in the battle against micronutrient deficiencies. Owing to a growing knowledge about vitamin metabolism, as well as mineral uptake and reallocation in plants, it is today possible to enhance micronutrient levels in crop plants, offering a sustainable solution to populations with a suboptimal micronutrient intake. © 2016 New York Academy of Sciences.

  9. Engineering Plant One-Carbon Metabolism

    Energy Technology Data Exchange (ETDEWEB)

    David Rhodes

    2005-02-09

    Primary and secondary metabolism intersect in the one-carbon (C1) area. Primary metabolism supplies most of the C1 units and competes with secondary metabolism for their use. This competition is potentially severe because secondary products such as lignin, alkaloids, and glycine betaine (GlyBet) require massive amounts of C1 units. Towards the goal of understanding how C1 metabolism is regulated at the metabolic and gene levels so as to successfully engineer C1 supply to match demand, we have: (1) cloned complete suites of C1 genes from maize and tobacco, and incorporated them into DNA arrays; (2) prepared antisense constructs and mutants engineered with alterations in C1 unit supply and demand; and (3) have quantified the impacts of these alterations on gene expression (using DNA arrays), and on metabolic fluxes (by combining isotope labeling, MS, NMR and computer modeling). Metabolic flux analysis and modeling in tobacco engineered for GlyBet synthesis by expressing choline oxidizing enzymes in either the chloroplast or cytosol, has shown that the choline biosynthesis network is rigid, and tends to resist large changes in C1 demand. A major constraint on engineering enhanced flux to GlyBet in tobacco is a low capacity of choline transport across the chloroplast envelope. Maize and sorghum mutants defective in GlyBet synthesis show greatly reduced flux of C1 units into choline in comparison to GlyBet-accumulating wildtypes, but this is not associated with altered expression of any of the C1 genes. Control of C1 flux to choline in tobacco, maize and sorghum appears to reside primarily at the level of N-methylation of phosphoethanolamine. A candidate signal for the control of this flux is the pool size of phosphocholine which down-regulates and feedback inhibits phosphoethanolamine N-methyltransferase. Methionine S-methyltransferase (MMT) catalyzes the synthesis of S-methylmethionine (SMM) from methionine (Met) and S-adenosylmethionine (AdoMet). SMM can be

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

    DEFF Research Database (Denmark)

    Rook, Frederik

    2016-01-01

    Plants produce a wide variety of specialized (or secondary) metabolites that function as chemical defence compounds and provide protection against microbial pathogens or herbivores. This chapter focuses on the metabolic engineering of biosynthetic pathways for plant chemical defence compounds...... 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...

  11. Engineering Plant One-Carbon Metabolism

    Energy Technology Data Exchange (ETDEWEB)

    David Rhodes

    2005-02-09

    Primary and secondary metabolism intersect in the one-carbon (C1) area. Primary metabolism supplies most of the C1 units and competes with secondary metabolism for their use. This competition is potentially severe because secondary products such as lignin, alkaloids, and glycine betaine (GlyBet) require massive amounts of C1 units. Towards the goal of understanding how C1 metabolism is regulated at the metabolic and gene levels so as to successfully engineer C1 supply to match demand, we have: (1) cloned complete suites of C1 genes from maize and tobacco, and incorporated them into DNA arrays; (2) prepared antisense constructs and mutants engineered with alterations in C1 unit supply and demand; and (3) have quantified the impacts of these alterations on gene expression (using DNA arrays), and on metabolic fluxes (by combining isotope labeling, MS, NMR and computer modeling). Metabolic flux analysis and modeling in tobacco engineered for GlyBet synthesis by expressing choline oxidizing enzymes in either the chloroplast or cytosol, has shown that the choline biosynthesis network is rigid, and tends to resist large changes in C1 demand. A major constraint on engineering enhanced flux to GlyBet in tobacco is a low capacity of choline transport across the chloroplast envelope. Maize and sorghum mutants defective in GlyBet synthesis show greatly reduced flux of C1 units into choline in comparison to GlyBet-accumulating wildtypes, but this is not associated with altered expression of any of the C1 genes. Control of C1 flux to choline in tobacco, maize and sorghum appears to reside primarily at the level of N-methylation of phosphoethanolamine. A candidate signal for the control of this flux is the pool size of phosphocholine which down-regulates and feedback inhibits phosphoethanolamine N-methyltransferase. Methionine S-methyltransferase (MMT) catalyzes the synthesis of S-methylmethionine (SMM) from methionine (Met) and S-adenosylmethionine (AdoMet). SMM can be

  12. Volatile science? Metabolic engineering of terpenoids in plants.

    Science.gov (United States)

    Aharoni, Asaph; Jongsma, Maarten A; Bouwmeester, Harro J

    2005-12-01

    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 demonstrated that terpenoid precursors in subcellular compartments are not as strictly separated as previously thought and that multistep pathway engineering is feasible, even across cell compartments. These engineered plants show that insect behavior is influenced by terpenoids. In the future, we expect rapid progress in the engineering of terpenoid production in plants. In addition to commercial applications, such transgenic plants should increase our understanding of the biological relevance of these volatile secondary metabolites.

  13. Volatile science? Metabolic engineering of terpenoids in plants

    NARCIS (Netherlands)

    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 d

  14. Metabolic Engineering of Tropane Alkaloid Biosynthesis in Plants

    Institute of Scientific and Technical Information of China (English)

    Lei ZHANG; Guo-Yin KAI; Bei-Bei LU; Han-Ming ZHANG; Ke-Xuan TANG; Ji-Hong JIANG; Wan-Sheng CHEN

    2005-01-01

    Over the past decade, the evolving commercial importance of so-called plant secondary metabolites has resulted in a great interest in secondary metabolism and, particularly, in the possibilities to enhance the yield of fine metabolites by means of genetic engineering. Plant alkaloids, which constitute one of the largest groups of natural products, provide many pharmacologically active compounds. Several genes in the tropane alkaloids biosynthesis pathways have been cloned, making the metabolic engineering of these alkaloids possible. The content of the target chemical scopolamine could be significantly increased by various approaches, such as introducing genes encoding the key biosynthetic enzymes or genes encoding regulatory proteins to overcome the specific rate-limiting steps. In addition, antisense genes have been used to block competitive pathways. These investigations have opened up new, promising perspectives for increased production in plants or plant cell culture. Recent achievements have been made in the metabolic engineering of plant tropane alkaloids and some new powerful strategies are reviewed in the present paper.

  15. Metabolic engineering of monoterpene biosynthesis in plants

    NARCIS (Netherlands)

    Lücker, J.

    2002-01-01

    Monoterpenes are a large group of compounds that belong to the terpenoid family of natural compounds in plants. They are small, volatile, lipophilic substances of which around one thousand different structures have been identified. Monoterpenes are

  16. Plant metabolic engineering strategies for the production of pharmaceutical terpenoids

    Directory of Open Access Journals (Sweden)

    Xu Lu

    2016-11-01

    Full Text Available Pharmaceutical terpenoids belong to the most diverse class of natural products. They have significant curative effects on a variety of diseases, such as cancer, cardiovascular diseases, malaria and Alzheimer’s disease. Nowadays, elicitors, including biotic and abiotic elicitors, are often used to activate the pathway of secondary metabolism and enhance the production of target terpenoids. Based on Agrobacterium-mediated genetic transformation, several plant metabolic engineering strategies hold great promise to regulate the biosynthesis of pharmaceutical terpenoids. Overexpressing terpenoids biosynthesis pathway genes in homologous and ectopic plants is an effective strategy to enhance the yield of pharmaceutical terpenoids. Another strategy is to suppress the expression of competitive metabolic pathways. In addition, global regulation which includes regulating the relative transcription factors, endogenous phytohormones and primary metabolism could also markedly increase their yield. All these strategies offer great opportunities to enhance the supply of scarce terpenoids drugs, reduce the price of expensive drugs and improve people's standards of living.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  19. Pathway elucidation and metabolic engineering of specialized plant metabolites

    DEFF Research Database (Denmark)

    Salomonsen, Bo

    , 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......, namely the plant defense compounds camalexin and glucosinolates. The thesis shows methodologies for elucidation of biosynthesis pathways and describes how to transfer obtained knowledge of metabolic pathways to other organisms through establishment of a synthetic biology platform. Through pathway...

  20. Natural plant genetic engineer Agrobacterium rhizogenes: role of T-DNA in plant secondary metabolism.

    Science.gov (United States)

    Chandra, Sheela

    2012-03-01

    Agrobacterium rhizogenes is a natural plant genetic engineer. It is a gram-negative soil bacterium that induces hairy root formation. Success has been obtained in exploring the molecular mechanisms of transferred DNA (T-DNA) transfer, interaction with host plant proteins, plant defense signaling and integration to plant genome for successful plant genetic transformation. T-DNA and corresponding expression of rol genes alter morphology and plant host secondary metabolism. During transformation, there is a differential loss of a few T-DNA genes. Loss of a few ORFs drastically affect the growth and morphological patterns of hairy roots, expression pattern of biosynthetic pathway genes and accumulation of specific secondary metabolites.

  1. Knowledge-driven approaches for engineering complex metabolic pathways in plants.

    Science.gov (United States)

    Farré, Gemma; Twyman, Richard M; Christou, Paul; Capell, Teresa; Zhu, Changfu

    2015-04-01

    Plant metabolic pathways are complex and often feature multiple levels of regulation. Until recently, metabolic engineering in plants relied on the laborious testing of ad hoc modifications to achieve desirable changes in the metabolic profile. However, technological advances in data mining, modeling, multigene engineering and genome editing are now taking away much of the guesswork by allowing the impact of modifications to be predicted more accurately. In this review we discuss recent developments in knowledge-based metabolic engineering strategies, that is the gathering and mining of genomic, transcriptomic, proteomic and metabolomic data to generate models of metabolic pathways that help to define and refine optimal intervention strategies.

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

    Science.gov (United States)

    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.

  3. Plant cell, tissue and organ culture: the most flexible foundations for plant metabolic engineering applications.

    Science.gov (United States)

    Ogita, Shinjiro

    2015-05-01

    Significant advances in plant cell, tissue and organ culture (PCTOC) have been made in the last five decades. PCTOC is now thought to be the underlying technique for understanding general or specific biological functions of the plant kingdom, and it is one of the most flexible foundations for morphological, physiological and molecular biological applications of plants. Furthermore, the recent advances in the field of information technology (IT) have enabled access to a large amount of information regarding all aspects of plant biology. For example, sequencing information is stored in mega repositories such as the National Center for Biotechnology Information (NCBI), which can be easily accessed by researchers worldwide. To date, the PCTOC and IT combination strategy for regulation of target plant metabolism and the utilization of bioactive plant metabolites for commercial purposes is essential. In this review, the advantages and the limitations of these methodologies, especially regarding the production of bioactive plant secondary metabolites and metabolic engineering in target plants are discussed mainly from the phenotypic view point.

  4. 2005 Plant Metabolic Engineering Gordon Conference - July 10-15, 2005

    Energy Technology Data Exchange (ETDEWEB)

    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.

  5. Flow cytometric methods to investigate culture heterogeneities for plant metabolic engineering.

    Science.gov (United States)

    Gaurav, Vishal; Kolewe, Martin E; Roberts, Susan C

    2010-01-01

    Plant cell cultures provide an important method for production and supply of a variety of natural products, where conditions can be easily controlled, manipulated, and optimized. Development and optimization of plant cell culture processes require both bioprocess engineering and metabolic engineering approaches. Cultures are generally highly heterogeneous, with significant variability amongst cells in terms of growth, metabolism, and productivity of key metabolites. Taxus cultures produce the important anti-cancer agent Taxol((R)) (i.e., paclitaxel) and have demonstrated significant variability amongst cell populations in culture with regard to paclitaxel accumulation, cell cycle participation, and protein synthesis. To fully understand the link between cellular metabolism and culture behavior and to enable targeted metabolic engineering approaches, cultures need to be studied at a single cell level. This chapter describes the application of plant cell flow cytometric techniques to investigate culture heterogeneity at the single cell level, in order to optimize culture performance through targeted metabolic engineering. Flow cytometric analytical methods are described to study Taxus single cells, protoplasts, and nuclei suspensions with respect to secondary metabolite accumulation, DNA content, cell size, and complexity. Reproducible methods to isolate these single particle suspensions from aggregated Taxus cultures are discussed. Methods to stain both fixed and live cells for a variety of biological markers are provided to enable characterization of cell phenotypes. Fluorescence-activated cell sorting (FACS) methods are also presented to facilitate isolation of certain plant cell culture populations for both analysis and propagation of superior cell lines for use in bioprocesses.

  6. Metabolic engineering of chloroplasts for artemisinic acid biosynthesis and impact on plant growth

    Indian Academy of Sciences (India)

    Bhawna Saxena; Mayavan Subramaniyan; Karan Malhotra; Neel Sarovar Bhavesh; Shobha Devi Potlakayala; Shashi Kumar

    2014-03-01

    Chloroplasts offer high-level transgene expression and transgene containment due to maternal inheritance, and are ideal hosts for biopharmaceutical biosynthesis via multigene engineering. To exploit these advantages, we have expressed 12 enzymes in chloroplasts for the biosynthesis of artemisinic acid (precursor of artemisinin, antimalarial drug) in an alternative plant system. Integration of transgenes into the tobacco chloroplast genome via homologous recombination was confirmed by molecular analysis, and biosynthesis of artemisinic acid in plant leaf tissues was detected with the help of 13C NMR and ESI-mass spectrometry. The excess metabolic flux of isopentenyl pyrophosphate generated by an engineered mevalonate pathway was diverted for the biosynthesis of artemisinic acid. However, expression of megatransgenes impacted the growth of the transplastomic plantlets. By combining two exogenous pathways, artemisinic acid was produced in transplastomic plants, which can be improved further using better metabolic engineering strategies for commercially viable yield of desirable isoprenoid products.

  7. Advances in the Plant Isoprenoid Biosynthesis Pathway and Its Metabolic Engineering

    Institute of Scientific and Technical Information of China (English)

    Yan LIU; Hong WANG; He-Chun YE; Guo-Feng LI

    2005-01-01

    Although the cytosolic isoprenoid biosynthetic pathway, mavolonate pathway, in plants has been known for many years, a new plastidial 1-deoxyxylulose-5-phosphate (DXP) pathway was identified in the past few years and its related intermediates, enzymes, and genes have been characterized quite recently.With a deep insight into the biosynthetic pathway of isoprenoids, investigations into the metabolic engineering of isoprenoid biosynthesis have started to prosper. In the present article, recent advances in the discoveries and regulatory roles of new genes and enzymes in the plastidial isoprenoid biosynthesis path way are reviewed and examples of the metabolic engineering of cytosolic and plastidial isoprenoids biosnthesis are discussed.

  8. Multi-scale engineering of plant cell cultures for promotion of specialized metabolism.

    Science.gov (United States)

    Wilson, Sarah A; Cummings, Elizabeth M; Roberts, Susan C

    2014-10-01

    To establish plant culture systems for product synthesis, a multi-scale engineering approach is necessary. At the intracellular level, the influx of 'omics' data has necessitated development of new methods to properly annotate and establish useful metabolic models that can be applied to elucidate unknown steps in specialized metabolite biosynthesis, define effective metabolic engineering strategies and increase enzyme diversity available for synthetic biology platforms. On an intercellular level, the presence of aggregates in culture leads to distinct metabolic sub-populations. Recent advances in flow cytometric analyses and mass spectrometry imaging allow for resolution of metabolites on the single cell level, providing an increased understanding of culture heterogeneity. Finally, extracellular engineering can be used to enhance culture performance through media manipulation, co-culture with bacteria, the use of exogenous elicitors or modulation of shear stress. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. When plants produce not enough or at all: metabolic engineering of flavonoids in microbial hosts

    Directory of Open Access Journals (Sweden)

    Emmanouil Antonios Trantas

    2015-01-01

    Full Text Available Flavonoid metabolism and its fascinating molecules that are natural products in plants, have attracted the attention of industry and researchers involved in plant science, nutrition, bio/chemistry, chemical bioengineering, pharmacy, medicine, etc., since flavonoids were found to be directly or indirectly connected to health. Subsequently, in the last few years flavonoids became top stories in pharmaceutical industry, which is continually seeking for novel ways to produce safe and efficient drugs. Microbial cell cultures can act as workhorse bio-factories by offering their metabolic machinery for the benefit of optimizing the conditions and increasing the productivity of a selective flavonoid. Furthermore, metabolic engineering methodology came to reinforce what nature does best by tuning inadequacies and dead-ends of a metabolic pathway. Combinatorial biosynthesis techniques led to discovery of novel ways to produce plant natural and even unnatural flavonoids, while on top of that metabolic engineering gave the opportunity to industry to invest in synthetic biology to overcome restricted diversification and productivity issues existing so far in synthetic chemistry protocols. In this review, we present an update on rationalized approaches for the production of natural or unnatural flavonoids through biotechnology, analyzing the significance of combinatorial biosynthesis of agricultural/ pharmaceutical compounds produced in heterologous organisms. We also quote strategies and achievements thrived so far in the area of synthetic biology, with emphasis on metabolic engineering targeting the cellular optimization of microorganisms and plants producing flavonoids, stressing the advances in flux dynamic control and optimization. The involvement of the rapidly increasing numbers of assembled genomes that contribute to the gene- or pathway- mining to identify gene(s responsible for producing species-specific secondary metabolites is finally considered.

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

    Science.gov (United States)

    Martin, Cathie; Li, Jie

    2017-08-10

    Contents I. II. III. IV. V. VI. References 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.

  11. Strategic patent analysis in plant biotechnology: terpenoid indole alkaloid metabolic engineering as a case study.

    Science.gov (United States)

    Miralpeix, Bruna; Sabalza, Maite; Twyman, Richard M; Capell, Teresa; Christou, Paul

    2014-02-01

    The do-it-yourself patent search is a useful alternative to professional patent analysis particularly in the context of publicly funded projects where funds for IP activities may be limited. As a case study, we analysed patents related to the engineering of terpenoid indole alkaloid (TIA) metabolism in plants. We developed a focused search strategy to remove redundancy and reduce the workload without missing important and relevant patents. This resulted in the identification of approximately 50 key patents associated with TIA metabolic engineering in plants, which could form the basis of a more detailed freedom-to-operate analysis. The structural elements of this search strategy could easily be transferred to other contexts, making it a useful generic model for publicly funded research projects. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  12. Metabolic engineering by plastid transformation as a strategy to modulate isoprenoid yield in plants.

    Science.gov (United States)

    Hasunuma, Tomohisa; Kondo, Akihiko; Miyake, Chikahiro

    2010-01-01

    Plants synthesize a large number of isoprenoid compounds that have diverse structures and functions. All isoprenoids are synthesized through consecutive condensation of five-carbon precursors, isopentenyl diphosphate (IPP) and its allyl isomer dimethylallyl diphosphate (DMAPP). With recent success in the cloning of genes that encode the enzymes of isoprenoid biosynthesis, genetic engineering strategies for the improvement of plant isoprenoid metabolism have emerged. Plastid transformation technology offers attractive features in plant genetic engineering. It has many advantages over nuclear genome transformation: high-level foreign protein expression, no need for a transit peptide, absence of gene silencing, and convenient transgene stacking in operons. We demonstrated that this technology is a remarkable tool for the production of isoprenoids in plants through metabolic engineering. The expression of bacterial genes encoding CrtW (beta-carotene ketolase) and CrtZ (beta-carotene hydroxylase) or cyanobacterial genes encoding DXR (1-deoxy-D-xylulose-5-phosphate reductoisomerase) in the plastid genome leads to alteration in isoprenoid content of tobacco leaves.

  13. Bioengineering of plant (tri)terpenoids: from metabolic engineering of plants to synthetic biology in vivo and in vitro.

    Science.gov (United States)

    Moses, Tessa; Pollier, Jacob; Thevelein, Johan M; Goossens, Alain

    2013-10-01

    Terpenoids constitute a large and diverse class of natural products that serve many functions in nature. Most of the tens of thousands of the discovered terpenoids are synthesized by plants, where they function as primary metabolites involved in growth and development, or as secondary metabolites that optimize the interaction between the plant and its environment. Several plant terpenoids are economically important molecules that serve many applications as pharmaceuticals, pesticides, etc. Major challenges for the commercialization of plant-derived terpenoids include their low production levels in planta and the continuous demand of industry for novel molecules with new or superior biological activities. Here, we highlight several synthetic biology methods to enhance and diversify the production of plant terpenoids, with a foresight towards triterpenoid engineering, the least engineered class of bioactive terpenoids. Increased or cheaper production of valuable triterpenoids may be obtained by 'classic' metabolic engineering of plants or by heterologous production of the compounds in other plants or microbes. Novel triterpenoid structures can be generated through combinatorial biosynthesis or directed enzyme evolution approaches. In its ultimate form, synthetic biology may lead to the production of large amounts of plant triterpenoids in in vitro systems or custom-designed artificial biological systems.

  14. Metabolic engineering of plant oils and waxes for use as industrial feedstocks.

    Science.gov (United States)

    Vanhercke, Thomas; Wood, Craig C; Stymne, Sten; Singh, Surinder P; Green, Allan G

    2013-02-01

    Society has come to rely heavily on mineral oil for both energy and petrochemical needs. Plant lipids are uniquely suited to serve as a renewable source of high-value fatty acids for use as chemical feedstocks and as a substitute for current petrochemicals. Despite the broad variety of acyl structures encountered in nature and the cloning of many genes involved in their biosynthesis, attempts at engineering economic levels of specialty industrial fatty acids in major oilseed crops have so far met with only limited success. Much of the progress has been hampered by an incomplete knowledge of the fatty acid biosynthesis and accumulation pathways. This review covers new insights based on metabolic flux and reverse engineering studies that have changed our view of plant oil synthesis from a mostly linear process to instead an intricate network with acyl fluxes differing between plant species. These insights are leading to new strategies for high-level production of industrial fatty acids and waxes. Furthermore, progress in increasing the levels of oil and wax structures in storage and vegetative tissues has the potential to yield novel lipid production platforms. The challenge and opportunity for the next decade will be to marry these technologies when engineering current and new crops for the sustainable production of oil and wax feedstocks. © 2012 CSIRO Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  15. Phytohormones and their metabolic engineering for abiotic stress tolerance in crop plants

    Institute of Scientific and Technical Information of China (English)

    Shabir H. Wani; Vinay Kumar; Varsha Shriram; Saroj Kumar Sah

    2016-01-01

    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.

  16. Phytohormones and their metabolic engineering for abiotic stress tolerance in crop plants

    Institute of Scientific and Technical Information of China (English)

    Shabir H.Wani; Vinay Kumar; Varsha Shriram; Saroj Kumar Sah

    2016-01-01

    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.

  17. Phytohormones and their metabolic engineering for abiotic stress tolerance in crop plants

    Directory of Open Access Journals (Sweden)

    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.

  18. Sinapate esters in brassicaceous plants: biochemistry, molecular biology, evolution and metabolic engineering.

    Science.gov (United States)

    Milkowski, Carsten; Strack, Dieter

    2010-06-01

    Brassicaceous plants are characterized by a pronounced metabolic flux toward sinapate, produced by the shikimate/phenylpropanoid pathway, which is converted into a broad spectrum of O-ester conjugates. The abundant sinapate esters in Brassica napus and Arabidopsis thaliana reflect a well-known metabolic network, including UDP-glucose:sinapate glucosyltransferase (SGT), sinapoylglucose:choline sinapoyltransferase (SCT), sinapoylglucose:L-malate sinapoyltransferase (SMT) and sinapoylcholine (sinapine) esterase (SCE). 1-O-Sinapoylglucose, produced by SGT during seed development, is converted to sinapine by SCT and hydrolyzed by SCE in germinating seeds. The released sinapate feeds via sinapoylglucose into the biosynthesis of sinapoylmalate in the seedlings catalyzed by SMT. Sinapoylmalate is involved in protecting the leaves against the deleterious effects of UV-B radiation. Sinapine might function as storage vehicle for ready supply of choline for phosphatidylcholine biosynthesis in young seedlings. The antinutritive character of sinapine and related sinapate esters hamper the use of the valuable seed protein of the oilseed crop B. napus for animal feed and human nutrition. Due to limited variation in seed sinapine content within the assortment of B. napus cultivars, low sinapine lines cannot be generated by conventional breeding giving rise to genetic engineering of sinapate ester metabolism as a promising means. In this article we review the progress made throughout the last decade in identification of genes involved in sinapate ester metabolism and characterization of the encoded enzymes. Based on gene structures and enzyme recruitment, evolution of sinapate ester metabolism is discussed. Strategies of targeted metabolic engineering, designed to generate low-sinapate ester lines of B. napus, are evaluated.

  19. Engineering Triterpene and Methylated Triterpene Production in Plants Provides Biochemical and Physiological Insights into Terpene Metabolism.

    Science.gov (United States)

    Jiang, Zuodong; Kempinski, Chase; Bush, Caroline J; Nybo, S Eric; Chappell, Joe

    2016-02-01

    Linear, branch-chained triterpenes, including squalene (C30), botryococcene (C30), and their methylated derivatives (C31-C37), generated by the green alga Botryococcus braunii race B have received significant attention because of their utility as chemical and biofuel feedstocks. However, the slow growth habit of B. braunii makes it impractical as a production system. In this study, we evaluated the potential of generating high levels of botryococcene in tobacco (Nicotiana tabacum) plants by diverting carbon flux from the cytosolic mevalonate pathway or the plastidic methylerythritol phosphate pathway by the targeted overexpression of an avian farnesyl diphosphate synthase along with two versions of botryococcene synthases. Up to 544 µg g(-1) fresh weight of botryococcene was achieved when this metabolism was directed to the chloroplasts, which is approximately 90 times greater than that accumulating in plants engineered for cytosolic production. To test if methylated triterpenes could be produced in tobacco, we also engineered triterpene methyltransferases (TMTs) from B. braunii into wild-type plants and transgenic lines selected for high-level triterpene accumulation. Up to 91% of the total triterpene contents could be converted to methylated forms (C31 and C32) by cotargeting the TMTs and triterpene biosynthesis to the chloroplasts, whereas only 4% to 14% of total triterpenes were methylated when this metabolism was directed to the cytoplasm. When the TMTs were overexpressed in the cytoplasm of wild-type plants, up to 72% of the total squalene was methylated, and total triterpene (C30+C31+C32) content was elevated 7-fold. Altogether, these results point to innate mechanisms controlling metabolite fluxes, including a homeostatic role for squalene.

  20. Plastoquinone and Ubiquinone in Plants: Biosynthesis, Physiological Function and Metabolic Engineering

    Science.gov (United States)

    Liu, Miaomiao; Lu, Shanfa

    2016-01-01

    Plastoquinone (PQ) and ubiquinone (UQ) are two important prenylquinones, functioning as electron transporters in the electron transport chain of oxygenic photosynthesis and the aerobic respiratory chain, respectively, and play indispensable roles in plant growth and development through participating in the biosynthesis and metabolism of important chemical compounds, acting as antioxidants, being involved in plant response to stress, and regulating gene expression and cell signal transduction. UQ, particularly UQ10, has also been widely used in people’s life. It is effective in treating cardiovascular diseases, chronic gingivitis and periodontitis, and shows favorable impact on cancer treatment and human reproductive health. PQ and UQ are made up of an active benzoquinone ring attached to a polyisoprenoid side chain. Biosynthesis of PQ and UQ is very complicated with more than thirty five enzymes involved. Their synthetic pathways can be generally divided into two stages. The first stage leads to the biosynthesis of precursors of benzene quinone ring and prenyl side chain. The benzene quinone ring for UQ is synthesized from tyrosine or phenylalanine, whereas the ring for PQ is derived from tyrosine. The prenyl side chains of PQ and UQ are derived from glyceraldehyde 3-phosphate and pyruvate through the 2-C-methyl-D-erythritol 4-phosphate pathway and/or acetyl-CoA and acetoacetyl-CoA through the mevalonate pathway. The second stage includes the condensation of ring and side chain and subsequent modification. Homogentisate solanesyltransferase, 4-hydroxybenzoate polyprenyl diphosphate transferase and a series of benzene quinone ring modification enzymes are involved in this stage. PQ exists in plants, while UQ widely presents in plants, animals and microbes. Many enzymes and their encoding genes involved in PQ and UQ biosynthesis have been intensively studied recently. Metabolic engineering of UQ10 in plants, such as rice and tobacco, has also been tested. In this

  1. Plastoquinone and ubiquinone in plants: biosynthesis, physiological function and metabolic engineering

    Directory of Open Access Journals (Sweden)

    Miaomiao Liu

    2016-12-01

    Full Text Available Plastoquinone (PQ and ubiquinone (UQ are two important prenylquinones, functioning as electron transporters in the electron transport chain of oxygenic photosynthesis and the aerobic respiratory chain, respectively, and play indispensable roles in plant growth and development through participating in the biosynthesis and metabolism of important chemical compounds, acting as antioxidants, being involved in plant response to stress, and regulating gene expression and cell signal transduction. UQ, particularly UQ10, has also been widely used in people’s life. It is effective in treating cardiovascular diseases, chronic gingivitis and periodontitis, and shows favorable impact on cancer treatment and human reproductive health. PQ and UQ are made up of an active benzoquinone ring attached to a polyisoprenoid side chain. Biosynthesis of PQ and UQ is very complicated with more than thirty five enzymes involved. Their synthetic pathways can be generally divided into two stages. The first stage leads to the biosynthesis of precursors of benzene quinone ring and prenyl side chain. The benzene quinone ring for UQ is synthesized from tyrosine or phenylalanine, whereas the ring for PQ is derived from tyrosine. The prenyl side chains of PQ and UQ are derived from glyceraldehyde 3-phosphate and pyruvate through the 2-C-methyl-D-erythritol 4-phosphate pathway and/or acetyl-CoA and acetoacetyl-CoA through the mevalonate pathway. The second stage includes the condensation of ring and side chain and subsequent modification. Homogentisate solanesyltransferase, 4-hydroxybenzoate polyprenyl diphosphate transferase and a series of benzene quinone ring modification enzymes are involved in this stage. PQ exists in plants, while UQ widely presents in plants, animals and microbes. Many enzymes and their encoding genes involved in PQ and UQ biosynthesis have been intensively studied recently. Metabolic engineering of UQ10 in plants, such as rice and tobacco, has also been

  2. Metabolic Engineering X Conference

    Energy Technology Data Exchange (ETDEWEB)

    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.

  3. Biosynthesis of plant-specific stilbene polyketides in metabolically engineered Escherichia coli

    Directory of Open Access Journals (Sweden)

    Schmidt-Dannert Claudia

    2006-03-01

    -coumaric acid and caffeic acid, can be efficiently converted to stilbene compounds by recombinant E. coli cells expressing plant biosynthetic genes. Optimization of precursor conversion and cyclization of the bulky ferulic acid precursor by host metabolic engineering and protein engineering may afford the synthesis of even more structurally diverse stilbene compounds.

  4. Metabolic engineering of a diazotrophic bacterium improves ammonium release and biofertilization of plants and microalgae.

    Science.gov (United States)

    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 N2 and CO2 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 N2 from the air into crops. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All

  5. Engineering Cellular Metabolism

    DEFF Research Database (Denmark)

    Nielsen, Jens; Keasling, Jay

    2016-01-01

    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....

  6. Metabolic engineering to enhance the value of plants as green factories.

    Science.gov (United States)

    Yuan, Ling; Grotewold, Erich

    2015-01-01

    The promise of plants to serve as the green factories of the future is ever increasing. Plants have been used traditionally for construction, energy, food and feed. Bioactive compounds primarily derived from specialized plant metabolism continue to serve as important scaffold molecules for pharmaceutical drug production. Yet, the past few years have witnessed a growing interest on plants as the ultimate harvesters of carbon and energy from the sun, providing carbohydrate and lipid biofuels that would contribute to balancing atmospheric carbon. How can the metabolic output from plants be increased even further, and what are the bottlenecks? Here, we present what we perceive to be the main opportunities and challenges associated with increasing the efficiency of plants as chemical factories. We offer some perspectives on when it makes sense to use plants as production systems because the amount of biomass needed makes any other system unfeasible. However, there are other instances in which plants serve as great sources of biological catalysts, yet are not necessarily the best-suited systems for production. We also present emerging opportunities for manipulating plant genomes to make plant synthetic biology a reality.

  7. Recent advances in the metabolic engineering of lignan biosynthesis pathways for the production of transgenic plant-based foods and supplements.

    Science.gov (United States)

    Satake, Honoo; Ono, Eiichiro; Murata, Jun

    2013-12-04

    Plant physiological, epidemiological, and food science studies have shed light on lignans as healthy diets for the reduction of the risk of lifestyle-related noncommunicable diseases and, thus, the demand for lignans has been rapidly increasing. However, the low efficiency and instability of lignan production via extraction from plant resources remain to be resolved, indicating the requirement for the development of new procedures for lignan production. The metabolic engineering of lignan-biosynthesizing plants is expected to be most promising for efficient, sustainable, and stable lignan production. This is supported by the recent verification of biosynthetic pathways of major dietary lignans and the exploration of lignan production via metabolic engineering using transiently gene-transfected or transgenic plants. The aim of this review is to present an overview of the biosynthetic pathways, biological activities, and metabolic engineering of lignans and also perspectives in metabolic engineering-based lignan production using transgenic plants for practical application.

  8. Engineering Mycorrhizal Symbioses to Alter Plant Metabolism and Improve Crop Health

    Science.gov (United States)

    French, Katherine E.

    2017-01-01

    Creating sustainable bioeconomies for the 21st century relies on optimizing the use of biological resources to improve agricultural productivity and create new products. Arbuscular mycorrhizae (phylum Glomeromycota) form symbiotic relationships with over 80% of vascular plants. In return for carbon, these fungi improve plant health and tolerance to environmental stress. This symbiosis is over 400 million years old and there are currently over 200 known arbuscular mycorrhizae, with dozens of new species described annually. Metagenomic sequencing of native soil communities, from species-rich meadows to mangroves, suggests biologically diverse habitats support a variety of mycorrhizal species with potential agricultural, medical, and biotechnological applications. This review looks at the effect of mycorrhizae on plant metabolism and how we can harness this symbiosis to improve crop health. I will first describe the mechanisms that underlie this symbiosis and what physiological, metabolic, and environmental factors trigger these plant-fungal relationships. These include mycorrhizal manipulation of host genetic expression, host mitochondrial and plastid proliferation, and increased production of terpenoids and jasmonic acid by the host plant. I will then discuss the effects of mycorrhizae on plant root and foliar secondary metabolism. I subsequently outline how mycorrhizae induce three key benefits in crops: defense against pathogen and herbivore attack, drought resistance, and heavy metal tolerance. I conclude with an overview of current efforts to harness mycorrhizal diversity to improve crop health through customized inoculum. I argue future research should embrace synthetic biology to create mycorrhizal chasses with improved symbiotic abilities and potentially novel functions to improve plant health. As the effects of climate change and anthropogenic disturbance increase, the global diversity of arbuscular mycorrhizal fungi should be monitored and protected to

  9. Engineering Mycorrhizal Symbioses to Alter Plant Metabolism and Improve Crop Health

    Directory of Open Access Journals (Sweden)

    Katherine E. French

    2017-07-01

    Full Text Available Creating sustainable bioeconomies for the 21st century relies on optimizing the use of biological resources to improve agricultural productivity and create new products. Arbuscular mycorrhizae (phylum Glomeromycota form symbiotic relationships with over 80% of vascular plants. In return for carbon, these fungi improve plant health and tolerance to environmental stress. This symbiosis is over 400 million years old and there are currently over 200 known arbuscular mycorrhizae, with dozens of new species described annually. Metagenomic sequencing of native soil communities, from species-rich meadows to mangroves, suggests biologically diverse habitats support a variety of mycorrhizal species with potential agricultural, medical, and biotechnological applications. This review looks at the effect of mycorrhizae on plant metabolism and how we can harness this symbiosis to improve crop health. I will first describe the mechanisms that underlie this symbiosis and what physiological, metabolic, and environmental factors trigger these plant-fungal relationships. These include mycorrhizal manipulation of host genetic expression, host mitochondrial and plastid proliferation, and increased production of terpenoids and jasmonic acid by the host plant. I will then discuss the effects of mycorrhizae on plant root and foliar secondary metabolism. I subsequently outline how mycorrhizae induce three key benefits in crops: defense against pathogen and herbivore attack, drought resistance, and heavy metal tolerance. I conclude with an overview of current efforts to harness mycorrhizal diversity to improve crop health through customized inoculum. I argue future research should embrace synthetic biology to create mycorrhizal chasses with improved symbiotic abilities and potentially novel functions to improve plant health. As the effects of climate change and anthropogenic disturbance increase, the global diversity of arbuscular mycorrhizal fungi should be monitored

  10. Metabolic Engineering VII Conference

    Energy Technology Data Exchange (ETDEWEB)

    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.

  11. Genetic and metabolic engineering

    OpenAIRE

    Yang,Yea-Tyng; Bennett, George N.; San, Ka-yiu

    1998-01-01

    Recent advances in molecular biology techniques, analytical methods and mathematical tools have led to a growing interest in using metabolic engineering to redirect metabolic fluxes for industrial and medical purposes. Metabolic engineering is referred to as the directed improvement of cellular properties through the modification of specific biochemical reactions or the introduction of new ones, with the use of recombinant DNA technology (Stephanopoulos, 1999). This multidisciplinary field dr...

  12. Engineering Cellular Metabolism.

    Science.gov (United States)

    Nielsen, Jens; Keasling, Jay D

    2016-03-10

    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, and pharmaceuticals. However, making cells into efficient factories is challenging because cells have evolved robust metabolic networks with hard-wired, tightly regulated lines of communication between molecular pathways that resist efforts to divert resources. Here, we will review the current status and challenges 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.

  13. Metabolic Engineering of Synechocystis sp. PCC 6803 for Production of the Plant Diterpenoid Manoyl Oxide.

    Science.gov (United States)

    Englund, Elias; Andersen-Ranberg, Johan; Miao, Rui; Hamberger, Björn; Lindberg, Pia

    2015-12-18

    Forskolin is a high value diterpenoid with a broad range of pharmaceutical applications, naturally found in root bark of the plant Coleus forskohlii. Because of its complex molecular structure, chemical synthesis of forskolin is not commercially attractive. Hence, the labor and resource intensive extraction and purification from C. forskohlii plants remains the current source of the compound. We have engineered the unicellular cyanobacterium Synechocystis sp. PCC 6803 to produce the forskolin precursor 13R-manoyl oxide (13R-MO), paving the way for light driven biotechnological production of this high value compound. In the course of this work, a new series of integrative vectors for use in Synechocystis was developed and used to create stable lines expressing chromosomally integrated CfTPS2 and CfTPS3, the enzymes responsible for the formation of 13R-MO in C. forskohlii. The engineered strains yielded production titers of up to 0.24 mg g(-1) DCW 13R-MO. To increase the yield, 13R-MO producing strains were further engineered by introduction of selected enzymes from C. forskohlii, improving the titer to 0.45 mg g(-1) DCW. This work forms a basis for further development of production of complex plant diterpenoids in cyanobacteria.

  14. Recent developments in systems biology and metabolic engineering of plant microbe interactions

    Directory of Open Access Journals (Sweden)

    Vishal Kumar

    2016-09-01

    Full Text Available Microorganisms play a crucial role in the sustainability of the various ecosystems. The characterization of various interactions between microorganisms and other biotic factors is a necessary footstep to understand the association and functions of microbial communities. Among the different microbial interactions in an ecosystem, plant-microbe interaction plays an important role to balance the ecosystem. The present review explores plant microbe interactions using gene editing and system biology tools towards the comprehension in improvement of plant traits. Further, system biology tools like FBA, OptKnock and constrain based modeling helps in understanding such interactions as a whole. In addition, various gene editing tools have been summarized and a strategy has been hypothesized for the development of disease free plants. Furthermore, we have tried to summarize the predictions through data retrieved from various types of sources such as high throughput sequencing data (e.g. single nucleotide polymorphism (SNP detection, RNA-seq, proteomics and metabolic models have been reconstructed from such sequences for species communities. It is well known fact that systems biology approaches and modeling of biological networks will enable us to learn the insight of such network and will also help further in understanding these interactions.

  15. Recent Developments in Systems Biology and Metabolic Engineering of Plant-Microbe Interactions.

    Science.gov (United States)

    Kumar, Vishal; Baweja, Mehak; Singh, Puneet K; Shukla, Pratyoosh

    2016-01-01

    Microorganisms play a crucial role in the sustainability of the various ecosystems. The characterization of various interactions between microorganisms and other biotic factors is a necessary footstep to understand the association and functions of microbial communities. Among the different microbial interactions in an ecosystem, plant-microbe interaction plays an important role to balance the ecosystem. The present review explores plant-microbe interactions using gene editing and system biology tools toward the comprehension in improvement of plant traits. Further, system biology tools like FBA (flux balance analysis), OptKnock, and constraint-based modeling helps in understanding such interactions as a whole. In addition, various gene editing tools have been summarized and a strategy has been hypothesized for the development of disease free plants. Furthermore, we have tried to summarize the predictions through data retrieved from various types of sources such as high throughput sequencing data (e.g., single nucleotide polymorphism detection, RNA-seq, proteomics) and metabolic models have been reconstructed from such sequences for species communities. It is well known fact that systems biology approaches and modeling of biological networks will enable us to learn the insight of such network and will also help further in understanding these interactions.

  16. Metabolic Engineering of Yeast and Plants for the Production of the Biologically Active Hydroxystilbene, Resveratrol

    Directory of Open Access Journals (Sweden)

    Philippe Jeandet

    2012-01-01

    Full Text Available Resveratrol, a stilbenic compound deriving from the phenyalanine/polymalonate route, being stilbene synthase the last and key enzyme of this pathway, recently has become the focus of a number of studies in medicine and plant physiology. Increased demand for this molecule for nutraceutical, cosmetic and possibly pharmaceutic uses, makes its production a necessity. In this context, the use of biotechnology through recombinant microorganisms and plants is particularly promising. Interesting results can indeed arise from the potential of genetically modified microorganisms as an alternative mechanism for producing resveratrol. Strategies used to tailoring yeast as they do not possess the genes that encode for the resveratrol pathway, will be described. On the other hand, most interest has centered in recent years, on STS gene transfer experiments from various origins to the genome of numerous plants. This work also presents a comprehensive review on plant molecular engineering with the STS gene, resulting in disease resistance against microorganisms and the enhancement of the antioxidant activities of several fruits in transgenic lines.

  17. Metabolic Engineering of Yeast and Plants for the Production of the Biologically Active Hydroxystilbene, Resveratrol

    Science.gov (United States)

    Jeandet, Philippe; Delaunois, Bertrand; Aziz, Aziz; Donnez, David; Vasserot, Yann; Cordelier, Sylvain; Courot, Eric

    2012-01-01

    Resveratrol, a stilbenic compound deriving from the phenyalanine/polymalonate route, being stilbene synthase the last and key enzyme of this pathway, recently has become the focus of a number of studies in medicine and plant physiology. Increased demand for this molecule for nutraceutical, cosmetic and possibly pharmaceutic uses, makes its production a necessity. In this context, the use of biotechnology through recombinant microorganisms and plants is particularly promising. Interesting results can indeed arise from the potential of genetically modified microorganisms as an alternative mechanism for producing resveratrol. Strategies used to tailoring yeast as they do not possess the genes that encode for the resveratrol pathway, will be described. On the other hand, most interest has centered in recent years, on STS gene transfer experiments from various origins to the genome of numerous plants. This work also presents a comprehensive review on plant molecular engineering with the STS gene, resulting in disease resistance against microorganisms and the enhancement of the antioxidant activities of several fruits in transgenic lines. PMID:22654481

  18. Analytics for Metabolic Engineering.

    Science.gov (United States)

    Petzold, Christopher J; Chan, Leanne Jade G; Nhan, Melissa; Adams, Paul D

    2015-01-01

    Realizing the promise of metabolic engineering has been slowed by challenges related to moving beyond proof-of-concept examples to robust and economically viable systems. Key to advancing metabolic engineering beyond trial-and-error research is access to parts with well-defined performance metrics that can be readily applied in vastly different contexts with predictable effects. As the field now stands, research depends greatly on analytical tools that assay target molecules, transcripts, proteins, and metabolites across different hosts and pathways. Screening technologies yield specific information for many thousands of strain variants, while deep omics analysis provides a systems-level view of the cell factory. Efforts focused on a combination of these analyses yield quantitative information of dynamic processes between parts and the host chassis that drive the next engineering steps. Overall, the data generated from these types of assays aid better decision-making at the design and strain construction stages to speed progress in metabolic engineering research.

  19. Collaborative Research: Metabolic Engineering of E. coli Sugar-Utilization Regulatory Systems for the Consumption of Plant Biomass Sugars.

    Energy Technology Data Exchange (ETDEWEB)

    Ramon Gonzalez (PI); J. V. Shanks (Co-PI); K-Y. San (Co-PI).

    2006-03-31

    The overall objective of this project is to metabolically engineer the E. coli sugar-utilization regulatory systems (SURS) to utilize sugar mixtures obtained from plant biomass. Of particular relevance is the implementation of a metabolic engineering cycle aided by functional genomics and systems biology tools. Our findings will help in the establishment of a platform for the efficient production of fuels and chemicals from lignocellulosic sugars. Our research has improved the understanding of the role of SURS in regulating sugar utilization and several other cellular functions. For example, we discovered that Mlc, a global regulatory protein, regulates the utilization of xylose and demonstrated the existence of an important link between catabolite repression and respiratory/fermentative metabolism. The study of SURS mutants also revealed a connection between flagellar biosynthesis and catabolite repression. Several tools were also developed as part of this project. A novel tool (Elementary Network Decomposition, END) to help elucidate the network topology of regulatory systems was developed and its utility as a discovery tool was demonstrated by applying it to the SURS in E. coli. A novel method (and software) to estimate metabolic fluxes that uses labeling experiments and eliminates reliance on extracellular fluxes was also developed. Although not initially considered in the scope of this project, we have developed a novel and superior method for optimization of HPLC separation and applied it to the simultaneous quantification of different functionalities (sugars, organic acids, ethanol, etc.) present in our fermentation samples. Currently under development is a genetic network driven metabolic flux analysis framework to integrate transcriptional and flux data.

  20. Engineering Triterpene and Methylated Triterpene Production in Plants Provides Biochemical and Physiological Insights into Terpene Metabolism1[OPEN

    Science.gov (United States)

    Jiang, Zuodong; Kempinski, Chase; Bush, Caroline J.; Nybo, S. Eric; Chappell, Joe

    2016-01-01

    Linear, branch-chained triterpenes, including squalene (C30), botryococcene (C30), and their methylated derivatives (C31–C37), generated by the green alga Botryococcus braunii race B have received significant attention because of their utility as chemical and biofuel feedstocks. However, the slow growth habit of B. braunii makes it impractical as a production system. In this study, we evaluated the potential of generating high levels of botryococcene in tobacco (Nicotiana tabacum) plants by diverting carbon flux from the cytosolic mevalonate pathway or the plastidic methylerythritol phosphate pathway by the targeted overexpression of an avian farnesyl diphosphate synthase along with two versions of botryococcene synthases. Up to 544 µg g−1 fresh weight of botryococcene was achieved when this metabolism was directed to the chloroplasts, which is approximately 90 times greater than that accumulating in plants engineered for cytosolic production. To test if methylated triterpenes could be produced in tobacco, we also engineered triterpene methyltransferases (TMTs) from B. braunii into wild-type plants and transgenic lines selected for high-level triterpene accumulation. Up to 91% of the total triterpene contents could be converted to methylated forms (C31 and C32) by cotargeting the TMTs and triterpene biosynthesis to the chloroplasts, whereas only 4% to 14% of total triterpenes were methylated when this metabolism was directed to the cytoplasm. When the TMTs were overexpressed in the cytoplasm of wild-type plants, up to 72% of the total squalene was methylated, and total triterpene (C30+C31+C32) content was elevated 7-fold. Altogether, these results point to innate mechanisms controlling metabolite fluxes, including a homeostatic role for squalene. PMID:26603654

  1. Metabolic engineering of biomass for high energy density: oilseed-like triacylglycerol yields from plant leaves.

    Science.gov (United States)

    Vanhercke, Thomas; El Tahchy, Anna; Liu, Qing; Zhou, Xue-Rong; Shrestha, Pushkar; Divi, Uday K; Ral, Jean-Philippe; Mansour, Maged P; Nichols, Peter D; James, Christopher N; Horn, Patrick J; Chapman, Kent D; Beaudoin, Frederic; Ruiz-López, Noemi; Larkin, Philip J; de Feyter, Robert C; Singh, Surinder P; Petrie, James R

    2014-02-01

    High biomass crops have recently attracted significant attention as an alternative platform for the renewable production of high energy storage lipids such as triacylglycerol (TAG). While TAG typically accumulates in seeds as storage compounds fuelling subsequent germination, levels in vegetative tissues are generally low. Here, we report the accumulation of more than 15% TAG (17.7% total lipids) by dry weight in Nicotiana tabacum (tobacco) leaves by the co-expression of three genes involved in different aspects of TAG production without severely impacting plant development. These yields far exceed the levels found in wild-type leaf tissue as well as previously reported engineered TAG yields in vegetative tissues of Arabidopsis thaliana and N. tabacum. When translated to a high biomass crop, the current levels would translate to an oil yield per hectare that exceeds those of most cultivated oilseed crops. Confocal fluorescence microscopy and mass spectrometry imaging confirmed the accumulation of TAG within leaf mesophyll cells. In addition, we explored the applicability of several existing oil-processing methods using fresh leaf tissue. Our results demonstrate the technical feasibility of a vegetative plant oil production platform and provide for a step change in the bioenergy landscape, opening new prospects for sustainable food, high energy forage, biofuel and biomaterial applications.

  2. Analytics for metabolic engineering

    Directory of Open Access Journals (Sweden)

    Christopher J Petzold

    2015-09-01

    Full Text Available Realizing the promise of metabolic engineering has been slowed by challenges related to moving beyond proof-of-concept examples to robust and economically viable systems. Key to advancing metabolic engineering beyond trial-and-error research is access to parts with well-defined performance metrics that can be readily applied in vastly different contexts with predictable effects. As the field now stands, research depends greatly on analytical tools that assay target molecules, transcripts, proteins, and metabolites across different hosts and pathways. Screening technologies yield specific information for many thousands of strain variants while deep omics analysis provide a systems-level view of the cell factory. Efforts focused on a combination of these analyses yield quantitative information of dynamic processes between parts and the host chassis that drive the next engineering steps. Overall, the data generated from these types of assays aid better decision-making at the design and strain construction stages to speed progress in metabolic engineering research.

  3. Analytics for Metabolic Engineering

    Science.gov (United States)

    Petzold, Christopher J.; Chan, Leanne Jade G.; Nhan, Melissa; Adams, Paul D.

    2015-01-01

    Realizing the promise of metabolic engineering has been slowed by challenges related to moving beyond proof-of-concept examples to robust and economically viable systems. Key to advancing metabolic engineering beyond trial-and-error research is access to parts with well-defined performance metrics that can be readily applied in vastly different contexts with predictable effects. As the field now stands, research depends greatly on analytical tools that assay target molecules, transcripts, proteins, and metabolites across different hosts and pathways. Screening technologies yield specific information for many thousands of strain variants, while deep omics analysis provides a systems-level view of the cell factory. Efforts focused on a combination of these analyses yield quantitative information of dynamic processes between parts and the host chassis that drive the next engineering steps. Overall, the data generated from these types of assays aid better decision-making at the design and strain construction stages to speed progress in metabolic engineering research. PMID:26442249

  4. Metabolic Engineering Strategies for the Optimization of Medicinal and Aromatic Plants : Expectations and Realities

    NARCIS (Netherlands)

    Kayser, O.; Baricevic, D; Novak, J; Pank, F

    2010-01-01

    In recent years classic genetic and molecular biology strategies (Bioballistics, Agrobacterium tumefaciens transformation, recombinant enzymes) for production of natural compounds or even breeding of medicinal and aromatic plants have expanded and improved productivity of plant-derived fine chemical

  5. Metabolic engineering strategies for the optimization of medicinal and aromatic plants : realities and expectations

    NARCIS (Netherlands)

    Hendrawati, O.; Woerdenbag, H. J.; Hille, J.; Kayser, O.

    2010-01-01

    In recent years, strategies and techniques for the production of natural compounds (plant derived fine chemicals) and/or the breeding of medicinal and aromatic plants has expanded. Efficient production of high value natural products with medicinal and cosmetic purpose (e.g. essential oils, paclitaxe

  6. Genome scale engineering techniques for metabolic engineering.

    Science.gov (United States)

    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.

  7. Metabolic Engineering of Yeast and Plants for the Production of the Biologically Active Hydroxystilbene, Resveratrol

    OpenAIRE

    Philippe Jeandet; Bertrand Delaunois; Aziz Aziz; David Donnez; Yann Vasserot; Sylvain Cordelier; Eric Courot

    2012-01-01

    Resveratrol, a stilbenic compound deriving from the phenyalanine/polymalonate route, being stilbene synthase the last and key enzyme of this pathway, recently has become the focus of a number of studies in medicine and plant physiology. Increased demand for this molecule for nutraceutical, cosmetic and possibly pharmaceutic uses, makes its production a necessity. In this context, the use of biotechnology through recombinant microorganisms and plants is particularly promising. Interesting resu...

  8. Metabolic engineering plant seeds with fish oil-like levels of DHA.

    Directory of Open Access Journals (Sweden)

    James R Petrie

    Full Text Available BACKGROUND: Omega-3 long-chain (≥C(20 polyunsaturated fatty acids (ω3 LC-PUFA have critical roles in human health and development with studies indicating that deficiencies in these fatty acids can increase the risk or severity of cardiovascular and inflammatory diseases in particular. These fatty acids are predominantly sourced from fish and algal oils, but it is widely recognised that there is an urgent need for an alternative and sustainable source of EPA and DHA. Since the earliest demonstrations of ω3 LC-PUFA engineering there has been good progress in engineering the C(20 EPA with seed fatty acid levels similar to that observed in bulk fish oil (∼18%, although undesirable ω6 PUFA levels have also remained high. METHODOLOGY/PRINCIPAL FINDINGS: The transgenic seed production of the particularly important C(22 DHA has been problematic with many attempts resulting in the accumulation of EPA/DPA, but only a few percent of DHA. This study describes the production of up to 15% of the C(22 fatty acid DHA in Arabidopsis thaliana seed oil with a high ω3/ω6 ratio. This was achieved using a transgenic pathway to increase the C(18 ALA which was then converted to DHA by a microalgal Δ6-desaturase pathway. CONCLUSIONS/SIGNIFICANCE: The amount of DHA described in this study exceeds the 12% level at which DHA is generally found in bulk fish oil. This is a breakthrough in the development of sustainable alternative sources of DHA as this technology should be applicable in oilseed crops. One hectare of a Brassica napus crop containing 12% DHA in seed oil would produce as much DHA as approximately 10,000 fish.

  9. The modification of plant oil composition via metabolic engineering--better nutrition by design.

    Science.gov (United States)

    Haslam, Richard P; Ruiz-Lopez, Noemi; Eastmond, Peter; Moloney, Maurice; Sayanova, Olga; Napier, Johnathan A

    2013-02-01

    This article will focus on the modification of plant seed oils to enhance their nutritional composition. Such modifications will include C18 Δ6-desaturated fatty acids such as γ-linolenic and stearidonic acid, omega-6 long-chain polyunsaturated fatty acids such as arachidonic acid, as well as the omega-3 long-chain polyunsaturated fatty acids (often named 'fish oils') such as eicosapentaenoic acid and docosahexaenoic acid. We will consider how new technologies (such as synthetic biology, next-generation sequencing and lipidomics) can help speed up and direct the development of desired traits in transgenic oilseeds. We will also discuss how manipulating triacylglycerol structure can further enhance the nutritional value of 'designer' oils. We will also consider how advances in model systems have translated into crops and the potential end-users for such novel oils (e.g. aquaculture, animal feed, human nutrition).

  10. Metabolic engineering of Escherichia coli for the synthesis of the plant polyphenol pinosylvin.

    Science.gov (United States)

    van Summeren-Wesenhagen, Philana Veronica; Marienhagen, Jan

    2015-02-01

    Plant polyphenols are of great interest for drug discovery and drug development since many of these compounds have health-promoting activities as treatments against various diseases, such as diabetes, cancer, or heart diseases. However, the limited availability of polyphenols represents a major obstacle to clinical applications that must be overcome. In comparison to the quantities of these compounds obtained by isolation from natural sources or costly chemical synthesis, the microbial production of these compounds could provide sufficient quantities from inexpensive substrates. In this work, we describe the development of an Escherichia coli platform strain for the production of pinosylvin, a stilbene found in the heartwood of pine trees which could aid in the treatment of various cancers and cardiovascular diseases. Initially, several configurations of the three-step biosynthetic pathway to pinosylvin were constructed from a set of two different enzymes for each enzymatic step. After optimization of gene expression and evaluation of different construct environments, low pinosylvin concentrations up to 3 mg/liter could be detected. Analysis of the precursor supply and a comparative analysis of the intracellular pools of pathway intermediates and product identified the limited malonyl coenzyme A (malonyl-CoA) availability and low stilbene synthase activity in the heterologous host to be the main bottlenecks during pinosylvin production. Addition of cerulenin for increasing intracellular malonyl-CoA pools and the in vivo evolution of the stilbene synthase from Pinus strobus for improved activity in E. coli proved to be the keys to elevated product titers. These measures allowed product titers of 70 mg/liter pinosylvin from glucose, which could be further increased to 91 mg/liter by the addition of l-phenylalanine.

  11. Metabolic engineering of glycine betaine synthesis: plant betaine aldehyde dehydrogenases lacking typical transit peptides are targeted to tobacco chloroplasts where they confer betaine aldehyde resistance.

    Science.gov (United States)

    Rathinasabapathi, B; McCue, K F; Gage, D A; Hanson, A D

    1994-01-01

    Certain higher plants synthesize and accumulate glycine betaine, a compound with osmoprotectant properties. Biosynthesis of glycine betaine proceeds via the pathway choline-->betaine aldehyde-->glycine betaine. Plants such as tobacco (Nicotiana tabacum L.) which do not accumulate glycine betaine lack the enzymes catalyzing both reactions. As a step towards engineering glycine betaine accumulation into a non-accumulator, spinach and sugar beet complementary-DNA sequences encoding the second enzyme of glycine-betaine synthesis (betaine aldehyde dehydrogenase, BADH, EC 1.2.1.8) were expressed in tobacco. Despite the absence of a typical transit peptide, BADH was targeted to the chloroplast in leaves of transgenic plants. Levels of extractable BADH were comparable to those in spinach and sugar beet, and the molecular weight, isoenzyme profile and Km for betaine aldehyde of the BADH enzymes from transgenic plants were the same as for native spinach or sugar beet BADH. Transgenic plants converted supplied betaine aldehyde to glycine betaine at high rates, demonstrating that they were able to transport betaine aldehyde across both the plasma membrane and the chloroplast envelope. The glycine betaine produced in this way was not further metabolized and reached concentrations similar to those in plants which accumulate glycine betaine naturally. Betaine aldehyde was toxic to non-transformed tobacco tissues whereas transgenic tissues were resistant due to detoxification of betaine aldehyde to glycine betaine. Betaine aldehyded ehydrogenase is therefore of interest as a potential selectable marker, as well as in the metabolic engineering of osmoprotectant biosynthesis.

  12. Safe genetically engineered plants

    Science.gov (United States)

    Rosellini, D.; Veronesi, F.

    2007-10-01

    The application of genetic engineering to plants has provided genetically modified plants (GMPs, or transgenic plants) that are cultivated worldwide on increasing areas. The most widespread GMPs are herbicide-resistant soybean and canola and insect-resistant corn and cotton. New GMPs that produce vaccines, pharmaceutical or industrial proteins, and fortified food are approaching the market. The techniques employed to introduce foreign genes into plants allow a quite good degree of predictability of the results, and their genome is minimally modified. However, some aspects of GMPs have raised concern: (a) control of the insertion site of the introduced DNA sequences into the plant genome and of its mutagenic effect; (b) presence of selectable marker genes conferring resistance to an antibiotic or an herbicide, linked to the useful gene; (c) insertion of undesired bacterial plasmid sequences; and (d) gene flow from transgenic plants to non-transgenic crops or wild plants. In response to public concerns, genetic engineering techniques are continuously being improved. Techniques to direct foreign gene integration into chosen genomic sites, to avoid the use of selectable genes or to remove them from the cultivated plants, to reduce the transfer of undesired bacterial sequences, and make use of alternative, safer selectable genes, are all fields of active research. In our laboratory, some of these new techniques are applied to alfalfa, an important forage plant. These emerging methods for plant genetic engineering are briefly reviewed in this work.

  13. Plant plastid engineering.

    Science.gov (United States)

    Wani, Shabir H; Haider, Nadia; Kumar, Hitesh; Singh, N B

    2010-11-01

    Genetic material in plants is distributed into nucleus, plastids and mitochondria. Plastid has a central role of carrying out photosynthesis in plant cells. Plastid transformation is becoming more popular and an alternative to nuclear gene transformation because of various advantages like high protein levels, the feasibility of expressing multiple proteins from polycistronic mRNAs, and gene containment through the lack of pollen transmission. Recently, much progress in plastid engineering has been made. In addition to model plant tobacco, many transplastomic crop plants have been generated which possess higher resistance to biotic and abiotic stresses and molecular pharming. In this mini review, we will discuss the features of the plastid DNA and advantages of plastid transformation. We will also present some examples of transplastomic plants developed so far through plastid engineering, and the various applications of plastid transformation.

  14. Engineering triterpene metabolism in tobacco.

    Science.gov (United States)

    Wu, Shuiqin; Jiang, Zuodong; Kempinski, Chase; Eric Nybo, S; Husodo, Satrio; Williams, Robert; Chappell, Joe

    2012-09-01

    Terpenes comprise a distinct class of natural products that serve a diverse range of physiological functions, provide for interactions between plants and their environment and represent a resource for many kinds of practical applications. To better appreciate the importance of terpenes to overall growth and development, and to create a production capacity for specific terpenes of industrial interest, we have pioneered the development of strategies for diverting carbon flow from the native terpene biosynthetic pathways operating in the cytosol and plastid compartments of tobacco for the generation of specific classes of terpenes. In the current work, we demonstrate how difficult it is to divert the 5-carbon intermediates DMAPP and IPP from the mevalonate pathway operating in the cytoplasm for triterpene biosynthesis, yet diversion of the same intermediates from the methylerythritol phosphate pathway operating in the plastid compartment leads to the accumulation of very high levels of the triterpene squalene. This was assessed by the co-expression of an avian farnesyl diphosphate synthase and yeast squalene synthase genes targeting metabolism in the cytoplasm or chloroplast. We also evaluated the possibility of directing this metabolism to the secretory trichomes of tobacco by comparing the effects of trichome-specific gene promoters to strong, constitutive viral promoters. Surprisingly, when transgene expression was directed to trichomes, high-level squalene accumulation was observed, but overall plant growth and physiology were reduced up to 80 % of the non-transgenic controls. Our results support the notion that the biosynthesis of a desired terpene can be dramatically improved by directing that metabolism to a non-native cellular compartment, thus avoiding regulatory mechanisms that might attenuate carbon flux within an engineered pathway.

  15. Metabolic engineering of plant cells for biotransformation of hesperedin into neohesperidin, a substrate for production of the low-calorie sweetener and flavor enhancer NHDC.

    Science.gov (United States)

    Frydman, Ahuva; Weisshaus, Oori; Huhman, David V; Sumner, Lloyd W; Bar-Peled, Maor; Lewinsohn, Efraim; Fluhr, Robert; Gressel, Jonathan; Eyal, Yoram

    2005-12-14

    Neohesperidin dihydrochalcone (NHDC) is a seminatural, safe, low-calorie sweetener, bitterness blocker, and flavor enhancer with unique properties and applications for the food, beverage, pharmaceutical, and animal feed industries. Current production is limited by the availability of the substrate neohesperidin, a flavonoid that accumulates to significant levels only in the inedible bitter citrus species. We propose a process to convert hesperidin, a tasteless flavonoid extracted from orange peels that are abundant byproducts of the vast orange juice industry, into neohesperidin using metabolic engineering and biotransformation via three steps: (i) extraction of hesperidin from orange peels, (ii) hydrolysis of sugar moieties, and (iii) biotransformation of hesperidin hydrolysis products into neohesperidin. We overcame the current technological bottleneck in biotransformation of hesperidin hydrolysis products into neohesperidin using metabolically engineered plant cell cultures expressing a recombinant flavanone-7-O-glucoside-2-O-rhamnosyltransferase. A small-scale production experiment established the feasibility of the proposed process.

  16. Metabolic engineering of terpene biosynthesis in plants using a trichome-specific transcription factor MsYABBY5 from spearmint (Mentha spicata).

    Science.gov (United States)

    Wang, Qian; Reddy, Vaishnavi Amarr; Panicker, Deepa; Mao, Hui-Zhu; Kumar, Nadimuthu; Rajan, Chakravarthy; Venkatesh, Prasanna Nori; Chua, Nam-Hai; Sarojam, Rajani

    2016-07-01

    In many aromatic plants including spearmint (Mentha spicata), the sites of secondary metabolite production are tiny specialized structures called peltate glandular trichomes (PGT). Having high commercial values, these secondary metabolites are exploited largely as flavours, fragrances and pharmaceuticals. But, knowledge about transcription factors (TFs) that regulate secondary metabolism in PGT remains elusive. Understanding the role of TFs in secondary metabolism pathway will aid in metabolic engineering for increased yield of secondary metabolites and also the development of new production techniques for valuable metabolites. Here, we isolated and functionally characterized a novel MsYABBY5 gene that is preferentially expressed in PGT of spearmint. We generated transgenic plants in which MsYABBY5 was either overexpressed or silenced using RNA interference (RNAi). Analysis of the transgenic lines showed that the reduced expression of MsYABBY5 led to increased levels of terpenes and that overexpression decreased terpene levels. Additionally, ectopic expression of MsYABBY5 in Ocimum basilicum and Nicotiana sylvestris decreased secondary metabolite production in them, suggesting that the encoded transcription factor is probably a repressor of secondary metabolism. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  17. Computer-aided design for metabolic engineering.

    Science.gov (United States)

    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.

  18. Engineering of plant chromosomes.

    Science.gov (United States)

    Mette, Michael Florian; Houben, Andreas

    2015-02-01

    Engineered minimal chromosomes with sufficient mitotic and meiotic stability have an enormous potential as vectors for stacking multiple genes required for complex traits in plant biotechnology. Proof of principle for essential steps in chromosome engineering such as truncation of chromosomes by T-DNA-mediated telomere seeding and de novo formation of centromeres by cenH3 fusion protein tethering has been recently obtained. In order to generate robust protocols for application in plant biotechnology, these steps need to be combined and supplemented with additional methods such as site-specific recombination for the directed transfer of multiple genes of interest on the minichromosomes. At the same time, the development of these methods allows new insight into basic aspects of plant chromosome functions such as how centromeres assure proper distribution of chromosomes to daughter cells or how telomeres serve to cap the chromosome ends to prevent shortening of ends over DNA replication cycles and chromosome end fusion.

  19. Essences in Metabolic Engineering of Lignan Biosynthesis

    Directory of Open Access Journals (Sweden)

    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.

  20. Synthetic biology and metabolic engineering.

    Science.gov (United States)

    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.

  1. Metabolic Engineering of Plant-derived (E)-β-farnesene Synthase Genes for a Novel Type of Aphid-resistant Genetically Modified Crop Plants

    Institute of Scientific and Technical Information of China (English)

    Xiu-Dao Yu; John Pickett; You-Zhi Ma; Toby Bruce; Johnathan Napier; Huw D.Jones; Lan-Qin Xia

    2012-01-01

    Aphids are major agricultural pests that cause significant yield losses of crop plants each year.Excessive dependence on insecticides for long-term aphid control is undesirable because of the development of insecticide resistance,the potential negative effects on non-target organisms and environmental pollution.Transgenic crops engineered for resistance to aphids via a non-toxic mode of action could be an efficient alternative strategy.(E)-β-Farnesene (EβF) synthases catalyze the formation of EβF,which for many pest aphids is the main component of the alarm pheromone involved in the chemical communication within these species.EβF can also be synthesized by certain plants but is then normally contaminated with inhibitory compounds.Engineering of crop plants capable of synthesizing and emitting EβF could cause repulsion of aphids and also the attraction of natural enemies that use EβF as a foraging cue,thus minimizing aphid infestation.In this review,the effects of aphids on host plants,plants' defenses against aphid herbivory and the recruitment of natural enemies for aphid control in an agricultural setting are briefly introduced.Furthermore,the plant-derived EβF synthase genes cloned to date along with their potential roles in generating novel aphid resistance via genetically modified approaches are discussed.

  2. Production and metabolic engineering of terpenoid indole alkaloids in cell cultures of the medicinal plant Catharanthus roseus (L.) G. Don (Madagascar periwinkle).

    Science.gov (United States)

    Zhou, Mei-Liang; Shao, Ji-Rong; Tang, Yi-Xiong

    2009-04-01

    The Madagascar periwinkle [Catharanthus roseus (L.) G. Don] is a plant species known for its production of TIAs (terpenoid indole alkaloids), many of which are pharmaceutically important. Ajmalicine and serpentine are prescribed for the treatment of hypertension, whereas the bisindoles vinblastine, vincristine and 3',4'-anhydrovinblastine are used for their antineoplastic activity in the treatment of many cancers. However, TIAs are produced in small yields in C. roseus, which make them expensive. Cell and metabolic engineering has focused on increasing flux through the TIA pathway by various means, including optimization of medium composition, elicitation, construction of noval culture systems and introduction of genes encoding specific metabolic enzymes into the C. roseus genome. The present review will attempt to present the state-of-the-art of research in this area and provide an update on the cell and metabolic engineering of TIAs in C. roseus. We hope that this will contribute to a better understanding of the ways in which TIA production can be achieved in different C. roseus culture systems.

  3. Engineered minichromosomes in plants.

    Science.gov (United States)

    Birchler, James A

    2015-02-01

    Engineered minichromosomes have been produced in several plant species via telomere-mediated chromosomal truncation. This approach bypasses the complications of the epigenetic nature of centromere function in plants, which has to date precluded the production of minichromosomes by the re-introduction of centromere sequences to a plant cell. Genes to be added to a cleaved chromosome are joined together with telomere repeats on one side. When these constructs are introduced into plant cells, the genes are ligated to the broken chromosomes but the telomere repeats will catalyze the formation of a telomere on the other end cutting the chromosome at that point. Telomere-mediated chromosomal truncation is sufficiently efficient that very small chromosomes can be generated consisting of basically the endogenous centromere and the added transgenes. The added transgenes provide a platform onto which it should be possible to assemble a synthetic chromosome to specification. Combining engineered minichromosomes with doubled haploid breeding should greatly expedite the transfer of transgenes to new lines and to test the interaction of transgenes in new background genotypes. Potential basic and applied applications of synthetic chromosomes are discussed.

  4. Engineered plant virus resistance.

    Science.gov (United States)

    Galvez, Leny C; Banerjee, Joydeep; Pinar, Hasan; Mitra, Amitava

    2014-11-01

    Virus diseases are among the key limiting factors that cause significant yield loss and continuously threaten crop production. Resistant cultivars coupled with pesticide application are commonly used to circumvent these threats. One of the limitations of the reliance on resistant cultivars is the inevitable breakdown of resistance due to the multitude of variable virus populations. Similarly, chemical applications to control virus transmitting insect vectors are costly to the farmers, cause adverse health and environmental consequences, and often result in the emergence of resistant vector strains. Thus, exploiting strategies that provide durable and broad-spectrum resistance over diverse environments are of paramount importance. The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Genetic engineering offers various options for introducing transgenic virus resistance into crop plants to provide a wide range of resistance to viral pathogens. This review examines the current strategies of developing virus resistant transgenic plants.

  5. An engineered pathway for glyoxylate metabolism in tobacco plants aimed to avoid the release of ammonia in photorespiration

    Directory of Open Access Journals (Sweden)

    Carvalho Josirley de FC

    2011-11-01

    Full Text Available Abstract Background The photorespiratory nitrogen cycle in C3 plants involves an extensive diversion of carbon and nitrogen away from the direct pathways of assimilation. The liberated ammonia is re-assimilated, but up to 25% of the carbon may be released into the atmosphere as CO2. Because of the loss of CO2 and high energy costs, there has been considerable interest in attempts to decrease the flux through the cycle in C3 plants. Transgenic tobacco plants were generated that contained the genes gcl and hyi from E. coli encoding glyoxylate carboligase (EC 4.1.1.47 and hydroxypyruvate isomerase (EC 5.3.1.22 respectively, targeted to the peroxisomes. It was presumed that the two enzymes could work together and compete with the aminotransferases that convert glyoxylate to glycine, thus avoiding ammonia production in the photorespiratory nitrogen cycle. Results When grown in ambient air, but not in elevated CO2, the transgenic tobacco lines had a distinctive phenotype of necrotic lesions on the leaves. Three of the six lines chosen for a detailed study contained single copies of the gcl gene, two contained single copies of both the gcl and hyi genes and one line contained multiple copies of both gcl and hyi genes. The gcl protein was detected in the five transgenic lines containing single copies of the gcl gene but hyi protein was not detected in any of the transgenic lines. The content of soluble amino acids including glycine and serine, was generally increased in the transgenic lines growing in air, when compared to the wild type. The content of soluble sugars, glucose, fructose and sucrose in the shoot was decreased in transgenic lines growing in air, consistent with decreased carbon assimilation. Conclusions Tobacco plants have been generated that produce bacterial glyoxylate carboligase but not hydroxypyruvate isomerase. The transgenic plants exhibit a stress response when exposed to air, suggesting that some glyoxylate is diverted away from

  6. Metabolic engineering of aroma components in fruits.

    Science.gov (United States)

    Aragüez, Irene; Valpuesta, Victoriano

    2013-10-01

    Plants have the ability to produce a diversity of volatile metabolites, which attract pollinators and seed dispersers and strengthen plant defense responses. Selection by plant breeders of traits such as rapid growth and yield leads, in many cases, to the loss of flavor and aroma quality in crops. How the aroma can be improved without affecting other fruit attributes is a major unsolved issue. Significant advances in metabolic engineering directed at improving the set of volatiles that the fruits emit has been aided by the characterization of enzymes involved in the biosynthesis of flavor and aroma compounds in some fruits. However, before this technology can be successfully applied to modulate the production of volatiles in different crops, further basic research is needed on the mechanisms that lead to the production of these compounds in plants. Here we review the biosynthesis and function of volatile compounds in plants, and the attempts that have been made to manipulate fruit aroma biosynthesis by metabolic engineering. In addition, we discuss the possibilities that molecular breeding offers for aroma enhancement and the implications of the latest advances in biotechnological modification of fruit flavor and aroma.

  7. Metabolic engineering in methanotrophic bacteria

    Energy Technology Data Exchange (ETDEWEB)

    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.

  8. Carotenoid metabolism in plants

    Science.gov (United States)

    Carotenoids are mostly C40 terpenoids, a class of hydrocarbons that participate in various biological processes in plants, such as photosynthesis, photomorphogenesis, photoprotection, and development. Carotenoids also serve as precursors for two plant hormones and a diverse set of apocarotenoids. Th...

  9. Cellulose metabolism in plants.

    Science.gov (United States)

    Hayashi, Takahisa; Yoshida, Kouki; Park, Yong Woo; Konishi, Teruko; Baba, Kei'ichi

    2005-01-01

    Many bacterial genomes contain a cellulose synthase operon together with a cellulase gene, indicating that cellulase is required for cellulose biosynthesis. In higher plants, there is evidence that cell growth is enhanced by the overexpression of cellulase and prevented by its suppression. Cellulase overexpression could modify cell walls not only by trimming off the paracrystalline sites of cellulose microfibrils, but also by releasing xyloglucan tethers between the microfibrils. Mutants for membrane-anchored cellulase (Korrigan) also show a typical phenotype of prevention of cellulose biosynthesis in tissues. All plant cellulases belong to family 9, which endohydrolyzes cellulose, but are not strong enough to cause the bulk degradation of cellulose microfibrils in a plant body. It is hypothesized that cellulase participates primarily in repairing or arranging cellulose microfibrils during cellulose biosynthesis in plants. A scheme for the roles of plant cellulose and cellulases is proposed.

  10. 2011 Plant Lipids: Structure, Metabolism, & Function Gordon Research Conference

    Energy Technology Data Exchange (ETDEWEB)

    Christopher Benning

    2011-02-04

    This is the second Gordon Research Conference on 'Plant Lipids: Structure, Metabolism & Function'. It covers current topics in lipid structure, metabolism and function in eukaryotic photosynthetic organisms including seed plants, algae, mosses and ferns. Work in photosynthetic bacteria is considered as well as it serves the understanding of specific aspects of lipid metabolism in plants. Breakthroughs are discussed in research on plant lipids as diverse as glycerolipids, sphingolipids, lipids of the cell surface, isoprenoids, fatty acids and their derivatives. The program covers nine concepts at the forefront of research under which afore mentioned plant lipid classes are discussed. The goal is to integrate areas such as lipid signaling, basic lipid metabolism, membrane function, lipid analysis, and lipid engineering to achieve a high level of stimulating interaction among diverse researchers with interests in plant lipids. One Emphasis is on the dynamics and regulation of lipid metabolism during plant cell development and in response to environmental factors.

  11. Progress in Metabolic Engineering of Saccharomyces cerevisiae

    OpenAIRE

    Nevoigt, Elke

    2008-01-01

    Summary: The traditional use of the yeast Saccharomyces cerevisiae in alcoholic fermentation has, over time, resulted in substantial accumulated knowledge concerning genetics, physiology, and biochemistry as well as genetic engineering and fermentation technologies. S. cerevisiae has become a platform organism for developing metabolic engineering strategies, methods, and tools. The current review discusses the relevance of several engineering strategies, such as rational and inverse metabolic...

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

    Science.gov (United States)

    Lai, Martin C; Lan, Ethan I

    2015-10-27

    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 diversity of bioproducts synthesized by engineered cyanobacteria, the metabolic pathways used, and the current engineering strategies used for increasing their titers.

  13. Engineering disease resistance in plants

    NARCIS (Netherlands)

    Custers, J.H.H.V.

    2007-01-01

    The genetic engineering of plants for increased pathogen resistance has engaged researchers and companies for decades. Until now, thenumberof crops with genetically engineered disease resistance traits which have entered the market are limited to products displaying virus an

  14. Systems metabolic engineering for chemicals and materials.

    Science.gov (United States)

    Lee, Jeong Wook; Kim, Tae Yong; Jang, Yu-Sin; Choi, Sol; Lee, Sang Yup

    2011-08-01

    Metabolic engineering has contributed significantly to the enhanced production of various value-added and commodity chemicals and materials from renewable resources in the past two decades. Recently, metabolic engineering has been upgraded to the systems level (thus, systems metabolic engineering) by the integrated use of global technologies of systems biology, fine design capabilities of synthetic biology, and rational-random mutagenesis through evolutionary engineering. By systems metabolic engineering, production of natural and unnatural chemicals and materials can be better optimized in a multiplexed way on a genome scale, with reduced time and effort. Here, we review the recent trends in systems metabolic engineering for the production of chemicals and materials by presenting general strategies and showcasing representative examples.

  15. Development and analysis of a highly flexible multi-gene expression system for metabolic engineering in Arabidopsis seeds and other plant tissues.

    Science.gov (United States)

    Shockey, Jay; Mason, Catherine; Gilbert, Matthew; Cao, Heping; Li, Xiangjun; Cahoon, Edgar; Dyer, John

    2015-09-01

    Production of novel value-added compounds in transgenic crops has become an increasingly viable approach in recent years. However, in many cases, product yield still falls short of the levels necessary for optimal profitability. Determination of the limiting factors is thus of supreme importance for the long-term viability of this approach. A significant challenge to most metabolic engineering projects is the need for strong coordinated co-expression of multiple transgenes. Strong constitutive promoters have been well-characterized during the >30 years since plant transformation techniques were developed. However, organ- or tissue-specific promoters are poorly characterized in many cases. Oilseeds are one such example. Reports spanning at least 20 years have described the use of certain seed-specific promoters to drive expression of individual transgenes. Multi-gene engineering strategies are often hampered by sub-optimal expression levels or improper tissue-specificity of particular promoters, or rely on the use of multiple copies of the same promoter, which can result in DNA instability or transgene silencing. We describe here a flexible system of plasmids that allows for expression of 1-7 genes per binary plasmid, and up to 18 genes altogether after multiple rounds of transformation or sexual crosses. This vector system includes six seed-specific promoters and two constitutive promoters. Effective constitutive and seed-specific RNA interference gene-suppression cloning vectors were also constructed for silencing of endogenous genes. Taken together, this molecular toolkit allows combinatorial cloning for multiple transgene expression in seeds, vegetative organs, or both simultaneously, while also providing the means to coordinately overexpress some genes while silencing others.

  16. Impacts of diversification of cytochrome P450 on plant metabolism.

    Science.gov (United States)

    Mizutani, Masaharu

    2012-01-01

    Cytochrome P450 monooxygenases (P450s) catalyze a wide variety of monooxygenation reactions in primary and secondary metabolism in plants. The share of P450 genes in each plant genome is estimated to be up to 1%. This implies that the diversification of P450 has made a significant contribution to the ability to acquire the emergence of new metabolic pathways during land plant evolution. The P450 families conserved universally in land plants contribute to their chemical defense mechanisms. Several P450s are involved in the biosynthesis and catabolism of plant hormones. Species-specific P450 families are essential for the biosynthetic pathways of phytochemicals such as terpenoids and alkaloids. Genome wide analysis of the gene clusters including P450 genes will provide a clue to defining the metabolic roles of orphan P450s. Metabolic engineering with plant P450s is an important technology for large-scale production of valuable phytochemicals such as medicines.

  17. Tools for metabolic engineering in Streptomyces.

    Science.gov (United States)

    Bekker, Valerie; Dodd, Amanda; Brady, Dean; Rumbold, Karl

    2014-01-01

    During the last few decades, Streptomycetes have shown to be a very important and adaptable group of bacteria for the production of various beneficial secondary metabolites. These secondary metabolites have been of great interest in academia and the pharmaceutical industries. To date, a vast variety of techniques and tools for metabolic engineering of relevant structural biosynthetic gene clusters have been developed. The main aim of this review is to summarize and discuss the published literature on tools for metabolic engineering of Streptomyces over the last decade. These strategies involve precursor engineering, structural and regulatory gene engineering, and the up or downregulation of genes, as well as genome shuffling and the use of genome scale metabolic models, which can reconstruct bacterial metabolic pathways to predict phenotypic changes and hence rationalize engineering strategies. These tools are continuously being developed to simplify the engineering strategies for this vital group of bacteria.

  18. Tools for metabolic engineering in Streptomyces

    Science.gov (United States)

    Bekker, Valerie; Dodd, Amanda; Brady, Dean; Rumbold, Karl

    2014-01-01

    During the last few decades, Streptomycetes have shown to be a very important and adaptable group of bacteria for the production of various beneficial secondary metabolites. These secondary metabolites have been of great interest in academia and the pharmaceutical industries. To date, a vast variety of techniques and tools for metabolic engineering of relevant structural biosynthetic gene clusters have been developed. The main aim of this review is to summarize and discuss the published literature on tools for metabolic engineering of Streptomyces over the last decade. These strategies involve precursor engineering, structural and regulatory gene engineering, and the up or downregulation of genes, as well as genome shuffling and the use of genome scale metabolic models, which can reconstruct bacterial metabolic pathways to predict phenotypic changes and hence rationalize engineering strategies. These tools are continuously being developed to simplify the engineering strategies for this vital group of bacteria. PMID:25482230

  19. Genome-scale modeling for metabolic engineering

    Energy Technology Data Exchange (ETDEWEB)

    Simeonidis, E; Price, ND

    2015-01-13

    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.

  20. Genome-scale modeling for metabolic engineering.

    Science.gov (United States)

    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.

  1. Lessons learned from metabolic engineering of cyanogenic glucosides

    DEFF Research Database (Denmark)

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

    2007-01-01

    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....... The interplay of a multitude of biosynthetic pathways and the possibility of metabolic cross-talk combined with an incomplete understanding of the regulation of these pathways, explain why metabolic engineering of plant secondary metabolism is still in its infancy and subject to much trial and error. Cyanogenic...... glucosides are ancient defense compounds that release toxic HCN upon tissue disruption caused e.g. by chewing insects. The committed steps of the cyanogenic glucoside biosynthetic pathway are encoded by three genes. This unique genetic simplicity and the availability of the corresponding cDNAs have given...

  2. 植物类胡萝卜素代谢工程与应用%Metabolic engineering and applications of plant carotenoid biosynthesis

    Institute of Scientific and Technical Information of China (English)

    陈中媛; 卢山; 黄继荣

    2011-01-01

    Carotenoids are essential nutrients for human beings. In addition to their antioxidant properties and their functions in preventing cancer and cardiovascular diseases, some carotenoids are also precursors of vitamin A,which our bodies cannot de novo synthesize. There are globally about 2.8~3.3 million preschool children with clinical vitamin A deficiency (VAD), with another ~200 million in subclinical VAD situation. With the deciphering of plant carotenoid metabolic pathway and the characterization of enzymes, genes and regulatory mechanisms functioning in this pathway, it is now feasible to promote the biosynthesis and accumulation of carotenoids in specific tissues of staple crops by means of metabolic engineering and gene transformation. Theoretically, it is ready to relief, at least in part, VAD with transgenic crops. Major progresses in this field are briefly summarized here.%类胡萝卜素是人类所需要的重要营养成分之一,不仅具有抗氧化、预防肿瘤和心血管等疾病的作用,而且还是人体合成维生素A的前体.全球大约有280万~330万学龄前儿童出现维生素缺乏(vitamin A deficiency,VAD)的临床症状;近2亿儿童处于半缺乏状态.通过对植物类胡萝卜素生物合成途径的解析,以及对参与这一代谢过程的酶及其调控机制的深入了解,目前已经可以通过基因工程在主要农作物中组织特异性地促进类胡萝卜素的合成与积累.从理论上已经可以利用转基因植物来减少VAD的出现.该文简要回顾近年来这一领域的研究进展.

  3. Expanding the docosahexaenoic acid food web for sustainable production: engineering lower plant pathways into higher plants.

    Science.gov (United States)

    Petrie, James R; Singh, Surinder P

    2011-01-01

    Algae are becoming an increasingly important component of land plant metabolic engineering projects. Land plants and algae have similar enough genetics to allow relatively straightforward gene transfer and they also share enough metabolic similarities that algal enzymes often function in a plant cell environment. Understanding metabolic systems in algae can provide insights into homologous systems in land plants. As examples, algal models are currently being used by several groups to better understand starch and lipid metabolism and catabolism, fields which have relevance in land plants. Importantly, land plants and algae also have enough metabolic divergence that algal genes can often provide new metabolic traits to plants. Furthermore, many algal genomes have now been sequenced, with many more in progress, and this easy access to genome-wide information has revealed that algal genomes are often relatively simple when compared with plants. One example of the importance of algal, and in particular microalgal, resources to land plant research is the metabolic engineering of long-chain polyunsaturated fatty acids into oilseed crops which typically uses microalgal genes to extend existing natural plant biosynthetic pathways. This review describes both recent progress and remaining challenges in this field.

  4. Engineering of sugar metabolism in Lactococcus lactis

    NARCIS (Netherlands)

    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 int

  5. Harnessing yeast organelles for metabolic engineering.

    Science.gov (United States)

    Hammer, Sarah K; Avalos, José L

    2017-08-01

    Each subcellular compartment in yeast offers a unique physiochemical environment and metabolite, enzyme, and cofactor composition. While yeast metabolic engineering has focused on assembling pathways in the cell cytosol, there is growing interest in embracing subcellular compartmentalization. Beyond harnessing distinct organelle properties, physical separation of organelles from the cytosol has the potential to eliminate metabolic crosstalk and enhance compartmentalized pathway efficiency. In this Perspective we review the state of the art in yeast subcellular engineering, highlighting the benefits of targeting biosynthetic pathways to subcellular compartments, including mitochondria, peroxisomes, the ER and/or Golgi, vacuoles, and the cell wall, in different yeast species. We compare the performances of strains developed with subcellular engineering to those of native producers or yeast strains previously engineered with cytosolic pathways. We also identify important challenges that lie ahead, which need to be addressed for organelle engineering to become as mainstream as cytosolic engineering in academia and industry.

  6. Engineering plants for spaceflight environments

    Science.gov (United States)

    Bugbee, B.

    1999-01-01

    The conversion efficiency of radiation into biomass and yield has steadily increased for centuries because of continued improvement in both plant genetics and environmental control. Considerable effort has gone into improving the environment for plant growth in space, but work has only begun to engineer plants for spaceflight. Genetic manipulation offers tremendous potential to improve our ability to study gravitational effects. Genetic manipulation will also be necessary to build an efficient regenerative life support system. We cannot fully characterize plant response to the spaceflight environment without understanding and manipulating their genetic composition. Identification and selection of the existing germplasm is the first step. There are thousands of cultivars of each of our major crop plants, each specifically adapted to a unique environment on our planet. Thousands of additional lines are held in national germplasm collections to maintain genetic diversity. Spaceflight imposes the need to tap this diversity. Existing lines need to be evaluated in the environment that is characteristic of closed-system spaceflight conditions. Many of the plant growth challenges we confront in space can be better solved through genetic change than by hardware engineering. Ten thousand years of plant breeding has demonstrated the value of matching genetics with the environment. For example, providing continuous light can increase plant growth in space, but this often induces calcium deficiencies because Ca is not supplied by guttation during a dark period. This deficiency cannot be eliminated through increased root-zone and foliar Ca applications. It can be solved, in wheat, through genetic selection of lines that do not have the deficiency. Subsequent comparison of lines with and without the Ca deficiency has also helped us understand the nature of the problem.

  7. Transporters in plant sulfur metabolism

    Directory of Open Access Journals (Sweden)

    Tamara eGigolashvili

    2014-09-01

    Full Text Available Sulfur is an essential nutrient, necessary for synthesis of many metabolites. The uptake of sulfate, primary and secondary assimilation, the biosynthesis, storage and final utilisation of sulfur (S containing compounds requires a lot of movement between organs, cells, and organelles. Efficient transport systems of S-containing compounds across the internal barriers or the plasma membrane and organellar membranes are therefore required. Here, we review a current state of knowledge of the transport of a range of S-containing metabolites within and between the cells as well as of their long distance transport. An improved understanding of mechanisms and regulation of transport will facilitate successful engineering of the respective pathways, to improve the plant yield, biotic interaction and nutritional properties of crops.

  8. An engineered plant peroxisome and its application in biotechnology.

    Science.gov (United States)

    Kessel-Vigelius, Sarah K; Wiese, Jan; Schroers, Martin G; Wrobel, Thomas J; Hahn, Florian; Linka, Nicole

    2013-09-01

    Plant metabolic engineering is a promising tool for biotechnological applications. Major goals include enhancing plant fitness for an increased product yield and improving or introducing novel pathways to synthesize industrially relevant products. Plant peroxisomes are favorable targets for metabolic engineering, because they are involved in diverse functions, including primary and secondary metabolism, development, abiotic stress response, and pathogen defense. This review discusses targets for manipulating endogenous peroxisomal pathways, such as fatty acid β-oxidation, or introducing novel pathways, such as the synthesis of biodegradable polymers. Furthermore, strategies to bypass peroxisomal pathways for improved energy efficiency and detoxification of environmental pollutants are discussed. In sum, we highlight the biotechnological potential of plant peroxisomes and indicate future perspectives to exploit peroxisomes as biofactories. Copyright © 2013 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  9. Systematic Applications of Metabolomics in Metabolic Engineering

    Directory of Open Access Journals (Sweden)

    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.

  10. Systematic Applications of Metabolomics in Metabolic Engineering

    Science.gov (United States)

    Dromms, Robert A.; Styczynski, Mark P.

    2012-01-01

    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. PMID:24957776

  11. Cytochrome P450-mediated metabolic engineering

    DEFF Research Database (Denmark)

    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...

  12. Engineered Gold Nanoparticles and Plant Adaptation Potential

    Science.gov (United States)

    Siddiqi, Khwaja Salahuddin; Husen, Azamal

    2016-09-01

    Use of metal nanoparticles in biological system has recently been recognised although little is known about their possible effects on plant growth and development. Nanoparticles accumulation, translocation, growth response and stress modulation in plant system is not well understood. Plants exposed to gold and gold nanoparticles have been demonstrated to exhibit both positive and negative effects. Their growth and yield vary from species to species. Cytoxicity of engineered gold nanoparticles depends on the concentration, particle size and shape. They exhibit increase in vegetative growth and yield of fruit/seed at lower concentration and decrease them at higher concentration. Studies have shown that the gold nanoparticles exposure has improved free radical scavenging potential and antioxidant enzymatic activities and alter micro RNAs expression that regulate different morphological, physiological and metabolic processes in plants. These modulations lead to improved plant growth and yields. Prior to the use of gold nanoparticles, it has been suggested that its cost may be calculated to see if it is economically feasible.

  13. Metabolic engineering for improved microbial pentose fermentation.

    Science.gov (United States)

    Fernandes, Sara; Murray, Patrick

    2010-01-01

    Global concern over the depletion of fossil fuel reserves, and the detrimental impact that combustion of these materials has on the environment, is focusing attention on initiatives to create sustainable approaches for the production and use of biofuels from various biomass substrates. The development of a low-cost, safe and eco-friendly process for the utilization of renewable resources to generate value-added products with biotechnological potential as well as robust microorganisms capable of efficient fermentation of all types of sugars are essential to underpin the economic production of biofuels from biomass feedstocks. Saccharomyces cerevisiae, the most established fermentation yeast used in large scale bioconversion strategies, does not however metabolise the pentose sugars, xylose and arabinose and bioengineering is required for introduction of efficient pentose metabolic pathways and pentose sugar transport proteins for bioconversion of these substrates. Our approach provided a basis for future experiments that may ultimately lead to the development of industrial S. cerevisiae strains engineered to express pentose metabolising proteins from thermophilic fungi living on decaying plant material and here we expand our original article and discuss the strategies implemented to improve pentose fermentation.

  14. Posttranslational Protein Modifications in Plant Metabolism.

    Science.gov (United States)

    Friso, Giulia; van Wijk, Klaas J

    2015-11-01

    Posttranslational modifications (PTMs) of proteins greatly expand proteome diversity, increase functionality, and allow for rapid responses, all at relatively low costs for the cell. PTMs play key roles in plants through their impact on signaling, gene expression, protein stability and interactions, and enzyme kinetics. Following a brief discussion of the experimental and bioinformatics challenges of PTM identification, localization, and quantification (occupancy), a concise overview is provided of the major PTMs and their (potential) functional consequences in plants, with emphasis on plant metabolism. Classic examples that illustrate the regulation of plant metabolic enzymes and pathways by PTMs and their cross talk are summarized. Recent large-scale proteomics studies mapped many PTMs to a wide range of metabolic functions. Unraveling of the PTM code, i.e. a predictive understanding of the (combinatorial) consequences of PTMs, is needed to convert this growing wealth of data into an understanding of plant metabolic regulation.

  15. Systems metabolic engineering in an industrial setting.

    Science.gov (United States)

    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.

  16. Engineering triterpene metabolism in tobacco

    OpenAIRE

    2012-01-01

    Terpenes comprise a distinct class of natural products that serve a diverse range of physiological functions, provide for interactions between plants and their environment and represent a resource for many kinds of practical applications. To better appreciate the importance of terpenes to overall growth and development, and to create a production capacity for specific terpenes of industrial interest, we have pioneered the development of strategies for diverting carbon flow from the native ter...

  17. Plant Specialized Metabolism: the Easy and the Hard

    Institute of Scientific and Technical Information of China (English)

    Shan Lu

    2010-01-01

    @@ Although normally termed" plant secondary metabolism", the phrase" plant specialized metabolism" has become instruments, such as gas chromatography, high performance liquid chromatography, mass spectrometry and nuclear magnetic resonance.

  18. Engineered resistance against fungal plant pathogens

    NARCIS (Netherlands)

    Honée, G.

    1999-01-01

    Development of genetic engineering technology and molecular characterization of plant defense responses have provided strategies for controlling plant diseases additional to those based on chemical control or classical breeding programs. Most of these alternative strategies are based on the overprod

  19. Managing sulfur metabolism in plants

    NARCIS (Netherlands)

    Hawkesford, M.J.; De Kok, LJ

    2006-01-01

    Resolution and analysis of genes encoding components of the pathways of primary sulphur assimilation have provided the potential to elucidate how sulphur is managed by plants. Individual roles for members of gene families and regulatory mechanisms operating at gene, cellular and whole plant levels h

  20. Modulation of sulfur metabolism enables efficient glucosinolate engineering

    Directory of Open Access Journals (Sweden)

    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

  1. Enabling plant synthetic biology through genome engineering.

    Science.gov (United States)

    Baltes, Nicholas J; Voytas, Daniel F

    2015-02-01

    Synthetic biology seeks to create new biological systems, including user-designed plants and plant cells. These systems can be employed for a variety of purposes, ranging from producing compounds of industrial or therapeutic value, to reducing crop losses by altering cellular responses to pathogens or climate change. To realize the full potential of plant synthetic biology, techniques are required that provide control over the genetic code - enabling targeted modifications to DNA sequences within living plant cells. Such control is now within reach owing to recent advances in the use of sequence-specific nucleases to precisely engineer genomes. We discuss here the enormous potential provided by genome engineering for plant synthetic biology.

  2. Convergent evolution in plant specialized metabolism.

    Science.gov (United States)

    Pichersky, Eran; Lewinsohn, Efraim

    2011-01-01

    Plants synthesize a multitude of compounds that contribute to adaptation to their ecological niches. Such compounds serve as attractants of other living organisms beneficial to the plants or as defense against other biotic as well as abiotic agents. Selection for increased fitness, a never-ending process, has resulted in each plant lineage synthesizing a distinct set of specialized metabolites appropriate for its environment. The total number of specialized metabolites found in the plant kingdom far exceeds the capacity of any one plant genome to encode the necessary enzymes, and just as a plant lineage acquires the ability to make new specialized compounds during evolution, it also loses the ability to make others. Although the ability of plants to make novel, specialized metabolites continues to evolve, there are also many examples in which different plants have independently evolved the ability to make compounds already present in other plant lineages or to make different compounds that fulfill the same role-both are examples of convergent evolution. Here, we discuss many examples of convergent evolution in specialized metabolism. There are many genetic and biochemical mechanisms that can give rise to convergent evolution, and we conclude that, overall, convergent evolution in plant specialized metabolism is surprisingly common.

  3. Metabolic Engineering of Microalgal Based Biofuel Production: Prospects and Challenges.

    Science.gov (United States)

    Banerjee, Chiranjib; Dubey, Kashyap K; Shukla, Pratyoosh

    2016-01-01

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  5. Understanding Plant Nitrogen Metabolism through Metabolomics and Computational Approaches

    Directory of Open Access Journals (Sweden)

    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.

  6. Modularization of genetic elements promotes synthetic metabolic engineering.

    Science.gov (United States)

    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.

  7. Expanding the metabolic engineering toolbox with directed evolution.

    Science.gov (United States)

    Abatemarco, Joseph; Hill, Andrew; Alper, Hal S

    2013-12-01

    Cellular systems can be engineered into factories that produce high-value chemicals from renewable feedstock. Such an approach requires an expanded toolbox for metabolic engineering. Recently, protein engineering and directed evolution strategies have started to play a growing and critical role within metabolic engineering. This review focuses on the various ways in which directed evolution can be applied in conjunction with metabolic engineering to improve product yields. Specifically, we discuss the application of directed evolution on both catalytic and non-catalytic traits of enzymes, on regulatory elements, and on whole genomes in a metabolic engineering context. We demonstrate how the goals of metabolic pathway engineering can be achieved in part through evolving cellular parts as opposed to traditional approaches that rely on gene overexpression and deletion. Finally, we discuss the current limitations in screening technology that hinder the full implementation of a metabolic pathway-directed evolution approach.

  8. Metabolic Engineering for Substrate Co-utilization

    Science.gov (United States)

    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

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

    Science.gov (United States)

    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.

  10. Metabolic engineering of Arabidopsis for butanetriol production using bacterial genes.

    Science.gov (United States)

    Abdel-Ghany, Salah E; Day, Irene; Heuberger, Adam L; Broeckling, Corey D; Reddy, Anireddy S N

    2013-11-01

    1,2,4-butanetriol (butanetriol) is a useful precursor for the synthesis of the energetic material butanetriol trinitrate and several pharmaceutical compounds. Bacterial synthesis of butanetriol from xylose or arabinose takes place in a pathway that requires four enzymes. To produce butanetriol in plants by expressing bacterial enzymes, we cloned native bacterial or codon optimized synthetic genes under different promoters into a binary vector and stably transformed Arabidopsis plants. Transgenic lines expressing introduced genes were analyzed for the production of butanetriol using gas chromatography coupled to mass spectrometry (GC-MS). Soil-grown transgenic plants expressing these genes produced up to 20 µg/g of butanetriol. To test if an exogenous supply of pentose sugar precursors would enhance the butanetriol level, transgenic plants were grown in a medium supplemented with either xylose or arabinose and the amount of butanetriol was quantified. Plants expressing synthetic genes in the arabinose pathway showed up to a forty-fold increase in butanetriol levels after arabinose was added to the medium. Transgenic plants expressing either bacterial or synthetic xylose pathways, or the arabinose pathway showed toxicity symptoms when xylose or arabinose was added to the medium, suggesting that a by-product in the pathway or butanetriol affected plant growth. Furthermore, the metabolite profile of plants expressing arabinose and xylose pathways was altered. Our results demonstrate that bacterial pathways that produce butanetriol can be engineered into plants to produce this chemical. This proof-of-concept study for phytoproduction of butanetriol paves the way to further manipulate metabolic pathways in plants to enhance the level of butanetriol production.

  11. Advancing metabolic engineering through systems biology of industrial microorganisms

    DEFF Research Database (Denmark)

    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...... 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. Improving peppermint essential oil yield and composition by metabolic engineering.

    Science.gov (United States)

    Lange, Bernd Markus; Mahmoud, Soheil Seyed; Wildung, Mark R; Turner, Glenn W; Davis, Edward M; Lange, Iris; Baker, Raymond C; Boydston, Rick A; Croteau, Rodney B

    2011-10-11

    Peppermint (Mentha × piperita L.) was transformed with various gene constructs to evaluate the utility of metabolic engineering for improving essential oil yield and composition. Oil yield increases were achieved by overexpressing genes involved in the supply of precursors through the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. Two-gene combinations to enhance both oil yield and composition in a single transgenic line were assessed as well. The most promising results were obtained by transforming plants expressing an antisense version of (+)-menthofuran synthase, which is critical for adjusting the levels of specific undesirable oil constituents, with a construct for the overexpression of the MEP pathway gene 1-deoxy-D-xylulose 5-phosphate reductoisomerase (up to 61% oil yield increase over wild-type controls with low levels of the undesirable side-product (+)-menthofuran and its intermediate (+)-pulegone). Elite transgenic lines were advanced to multiyear field trials, which demonstrated consistent oil yield increases of up to 78% over wild-type controls and desirable effects on oil composition under commercial growth conditions. The transgenic expression of a gene encoding (+)-limonene synthase was used to accumulate elevated levels of (+)-limonene, which allows oil derived from transgenic plants to be recognized during the processing of commercial formulations containing peppermint oil. Our study illustrates the utility of metabolic engineering for the sustainable agricultural production of high quality essential oils at a competitive cost.

  13. Engineering Molecular Immunity Against Plant Viruses

    KAUST Repository

    Zaidi, Syed Shan-e-Ali

    2017-04-26

    Genomic engineering has been used to precisely alter eukaryotic genomes at the single-base level for targeted gene editing, replacement, fusion, and mutagenesis, and plant viruses such as Tobacco rattle virus have been developed into efficient vectors for delivering genome-engineering reagents. In addition to altering the host genome, these methods can target pathogens to engineer molecular immunity. Indeed, recent studies have shown that clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) systems that target the genomes of DNA viruses can interfere with viral activity and limit viral symptoms in planta, demonstrating the utility of this system for engineering molecular immunity in plants. CRISPR/Cas9 can efficiently target single and multiple viral infections and confer plant immunity. Here, we discuss the use of site-specific nucleases to engineer molecular immunity against DNA and RNA viruses in plants. We also explore how to address the potential challenges encountered when producing plants with engineered resistance to single and mixed viral infections.

  14. Chromosome engineering: power tools for plant genetics.

    Science.gov (United States)

    Chan, Simon W L

    2010-12-01

    The term "chromosome engineering" describes technologies in which chromosomes are manipulated to change their mode of genetic inheritance. This review examines recent innovations in chromosome engineering that promise to greatly increase the efficiency of plant breeding. Haploid Arabidopsis thaliana have been produced by altering the kinetochore protein CENH3, yielding instant homozygous lines. Haploid production will facilitate reverse breeding, a method that downregulates recombination to ensure progeny contain intact parental chromosomes. Another chromosome engineering success is the conversion of meiosis into mitosis, which produces diploid gametes that are clones of the parent plant. This is a key step in apomixis (asexual reproduction through seeds) and could help to preserve hybrid vigor in the future. New homologous recombination methods in plants will potentiate many chromosome engineering applications. Copyright © 2010 Elsevier Ltd. All rights reserved.

  15. PLEVIS: plant engineering visual and interactive simulator

    Energy Technology Data Exchange (ETDEWEB)

    Tanaka, Kazuma; Yoshikawa, Eiji; Ohtsuka, Shiroh; Kawakami, Seishiroh (Toshiba Corp., Kawasaki, Kanagawa (Japan))

    1990-07-01

    TOSHIBA has devoted much effort since 1974 to the development of nuclear power plant simulators for operator training, such as replica simulators and compact simulators. These simulators have special purpose consoles or control room equipment for operation training, and run on one or more process computers. Hence they require a large amount of space, e.g. a special room for installation. PLEVIS is a real time plant engineering simulator and operates on a general purpose desk-side engineering workstation (TOSHIBA AS-series) with a high resolution bit-mapped display. The major objectives of PLEVIS are to further improve training performance and efficiency for operators and engineers and to provide a powerful and cost effective plant engineering tool. (author).

  16. Engineering crassulacean acid metabolism to improve water-use efficiency.

    Science.gov (United States)

    Borland, Anne M; Hartwell, James; Weston, David J; Schlauch, Karen A; Tschaplinski, Timothy J; Tuskan, Gerald A; Yang, Xiaohan; Cushman, John C

    2014-05-01

    Climatic extremes threaten agricultural sustainability worldwide. One approach to increase plant water-use efficiency (WUE) is to introduce crassulacean acid metabolism (CAM) into C3 crops. Such a task requires comprehensive systems-level understanding of the enzymatic and regulatory pathways underpinning this temporal CO2 pump. Here we review the progress that has been made in achieving this goal. Given that CAM arose through multiple independent evolutionary origins, comparative transcriptomics and genomics of taxonomically diverse CAM species are being used to define the genetic 'parts list' required to operate the core CAM functional modules of nocturnal carboxylation, diurnal decarboxylation, and inverse stomatal regulation. Engineered CAM offers the potential to sustain plant productivity for food, feed, fiber, and biofuel production in hotter and drier climates.

  17. Flux analysis and metabolomics for systematic metabolic engineering of microorganisms.

    Science.gov (United States)

    Toya, Yoshihiro; Shimizu, Hiroshi

    2013-11-01

    Rational engineering of metabolism is important for bio-production using microorganisms. Metabolic design based on in silico simulations and experimental validation of the metabolic state in the engineered strain helps in accomplishing systematic metabolic engineering. Flux balance analysis (FBA) is a method for the prediction of metabolic phenotype, and many applications have been developed using FBA to design metabolic networks. Elementary mode analysis (EMA) and ensemble modeling techniques are also useful tools for in silico strain design. The metabolome and flux distribution of the metabolic pathways enable us to evaluate the metabolic state and provide useful clues to improve target productivity. Here, we reviewed several computational applications for metabolic engineering by using genome-scale metabolic models of microorganisms. We also discussed the recent progress made in the field of metabolomics and (13)C-metabolic flux analysis techniques, and reviewed these applications pertaining to bio-production development. Because these in silico or experimental approaches have their respective advantages and disadvantages, the combined usage of these methods is complementary and effective for metabolic engineering.

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

    NARCIS (Netherlands)

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

    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' ar

  19. Modular Engineering of Production Plants

    DEFF Research Database (Denmark)

    Miller, Thomas Dedenroth

    1998-01-01

    Based on a case-study on design of pharmaceutical production plants, this paper suggests that modularity may support business efficiency for companies with one-of-a-kind production and without in-house manufacturing. Modularity may support efficient management of design knowledge and may facilitate...

  20. Modular Engineering of Production Plants

    DEFF Research Database (Denmark)

    Miller, Thomas Dedenroth

    1998-01-01

    Based on a case-study on design of pharmaceutical production plants, this paper suggests that modularity may support business efficiency for companies with one-of-a-kind production and without in-house manufacturing. Modularity may support efficient management of design knowledge and may facilitate...

  1. The sweet branch of metabolic engineering: cherry-picking the low-hanging sugary fruits.

    Science.gov (United States)

    Chen, Rachel

    2015-12-09

    In the first science review on the then nascent Metabolic Engineering field in 1991, Dr. James E. Bailey described how improving erythropoietin (EPO) glycosylation can be achieved via metabolic engineering of Chinese hamster ovary (CHO) cells. In the intervening decades, metabolic engineering has brought sweet successes in glycoprotein engineering, including antibodies, vaccines, and other human therapeutics. Today, not only eukaryotes (CHO, plant, insect, yeast) are being used for manufacturing protein therapeutics with human-like glycosylation, newly elucidated bacterial glycosylation systems are enthusiastically embraced as potential breakthrough to revolutionize the biopharmaceutical industry. Notwithstanding these excitement in glycoprotein, the sweet metabolic engineering reaches far beyond glycoproteins. Many different types of oligo- and poly-saccharides are synthesized with metabolically engineered cells. For example, several recombinant hyaluronan bioprocesses are now in commercial production, and the titer of 2'-fucosyllactose, the most abundant fucosylated trisaccharide in human milk, reaches over 20 g/L with engineered E. coli cells. These successes represent only the first low hanging fruits, which have been appreciated scientifically, medically and fortunately, commercially as well. As one of the four building blocks of life, sugar molecules permeate almost all aspects of life. They are also unique in being intimately associated with all major types of biopolymers (including DNA/RNA, proteins, lipids) meanwhile they stand alone as bioactive polysaccharides, or free soluble oligosaccharides. As such, all sugar moieties in biological components, small or big and free or bound, are important targets for metabolic engineering. Opportunities abound at the interface of glycosciences and metabolic engineering. Continued investment and successes in this branch of metabolic engineering will make vastly diverse sugar-containing molecules (a

  2. Hairy Root and Its Application in Plant Genetic Engineering

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Agrobacterium rhizogenes Conn. causes hairy root disease in plants. Hairy root-infected A. rhizogenes is characterized by a high growth rate and genetic stability. Hairy root cultures have been proven to be an efficient means of producing secondary metabolites that are normally biosynthesized in roots of differentiated plants.Furthermore, a transgenic root system offers tremendous potential for introducing additional genes along with the Ri plasmid, especially with modified genes, into medicinal plant cells with A. rhizogenes vector systems.The cultures have turned out to be a valuable tool with which to study the biochemical properties and the gene expression profile of metabolic pathways. Moreover, the cultures can be used to elucidate the intermediates and key enzymes involved in the biosynthesis of secondary metabolites. The present article discusses various applications of hairy root cultures in plant genetic engineering and potential problems associated with them.

  3. Applied evolutionary theories for engineering of secondary metabolic pathways.

    Science.gov (United States)

    Bachmann, Brian O

    2016-12-01

    An expanded definition of 'secondary metabolism' is emerging. Once the exclusive provenance of naturally occurring organisms, evolved over geological time scales, secondary metabolism increasingly encompasses molecules generated via human engineered biocatalysts and biosynthetic pathways. Many of the tools and strategies for enzyme and pathway engineering can find origins in evolutionary theories. This perspective presents an overview of selected proposed evolutionary strategies in the context of engineering secondary metabolism. In addition to the wealth of biocatalysts provided via secondary metabolic pathways, improving the understanding of biosynthetic pathway evolution will provide rich resources for methods to adapt to applied laboratory evolution. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Impact of 'ome' analyses on inverse metabolic engineering

    DEFF Research Database (Denmark)

    Bro, Christoffer; Nielsen, Jens

    2004-01-01

    Genome-wide or large-scale methodologies employed in functional genomics such as DNA sequencing, transcription profiling, proteomics, and metabolite profiling have become important tools in many metabolic engineering strategies. These techniques allow the identification of genetic differences...... genomics in particular have the potential to promote and expand inverse metabolic engineering. Here, we review the use of functional genomics methods in inverse metabolic engineering, examples are presented, and we discuss the identification of targets for metabolic engineering with low fold changes using...... and insight into their cellular effects. In the field of inverse metabolic engineering mapping of differences between strains with different degree of a certain desired phenotype and subsequent identification of factors conferring that phenotype are an essential part. Therefore, the tools of functional...

  5. Engineered plant biomass feedstock particles

    Science.gov (United States)

    Dooley, James H [Federal Way, WA; Lanning, David N [Federal Way, WA; Broderick, Thomas F [Lake Forest Park, WA

    2012-04-17

    A new class of plant biomass feedstock particles characterized by consistent piece size and shape uniformity, high skeletal surface area, and good flow properties. The particles of plant biomass material having fibers aligned in a grain are characterized by a length dimension (L) aligned substantially parallel to the grain and defining a substantially uniform distance along the grain, a width dimension (W) normal to L and aligned cross grain, and a height dimension (H) normal to W and L. In particular, the L.times.H dimensions define a pair of substantially parallel side surfaces characterized by substantially intact longitudinally arrayed fibers, the W.times.H dimensions define a pair of substantially parallel end surfaces characterized by crosscut fibers and end checking between fibers, and the L.times.W dimensions define a pair of substantially parallel top and bottom surfaces. The L.times.W surfaces of particles with L/H dimension ratios of 4:1 or less are further elaborated by surface checking between longitudinally arrayed fibers. The length dimension L is preferably aligned within 30.degree. parallel to the grain, and more preferably within 10.degree. parallel to the grain. The plant biomass material is preferably selected from among wood, agricultural crop residues, plantation grasses, hemp, bagasse, and bamboo.

  6. Metstoich--Teaching Quantitative Metabolism and Energetics in Biochemical Engineering

    Science.gov (United States)

    Wong, Kelvin W. W.; Barford, John P.

    2010-01-01

    Metstoich, a metabolic calculator developed for teaching, can provide a novel way to teach quantitative metabolism to biochemical engineering students. It can also introduce biochemistry/life science students to the quantitative aspects of life science subjects they have studied. Metstoich links traditional biochemistry-based metabolic approaches…

  7. Metstoich--Teaching Quantitative Metabolism and Energetics in Biochemical Engineering

    Science.gov (United States)

    Wong, Kelvin W. W.; Barford, John P.

    2010-01-01

    Metstoich, a metabolic calculator developed for teaching, can provide a novel way to teach quantitative metabolism to biochemical engineering students. It can also introduce biochemistry/life science students to the quantitative aspects of life science subjects they have studied. Metstoich links traditional biochemistry-based metabolic approaches…

  8. Regulation of primary plant metabolism during plant-pathogen interactions and its contribution to plant defense

    Directory of Open Access Journals (Sweden)

    Clemencia M Rojas

    2014-02-01

    Full Text Available Plants are constantly exposed to microorganisms in the environment and, as a result, have evolved intricate mechanisms to recognize and defend themselves against potential pathogens. One of these responses is the downregulation of photosynthesis and other processes associated with primary metabolism that are essential for plant growth. It has been suggested that the energy saved by downregulation of primary metabolism is diverted and used for defense responses. However, several studies have shown that upregulation of primary metabolism also occurs during plant-pathogen interactions. We propose that upregulation of primary metabolism modulates signal transduction cascades that lead to plant defense responses. In support of this thought, we here compile evidence from the literature to show that upon exposure to pathogens or elicitors, plants induce several genes associated with primary metabolic pathways, such as those involved in the synthesis or degradation of carbohydrates, amino acids and lipids. In addition, genetic studies have confirmed the involvement of these metabolic pathways in plant defense responses. This review provides a new perspective highlighting the relevance of primary metabolism in regulating plant defense against pathogens with the hope to stimulate further research in this area.

  9. Engineering photosynthesis in plants and synthetic microorganisms.

    Science.gov (United States)

    Maurino, Veronica G; Weber, Andreas P M

    2013-01-01

    Photosynthetic organisms, such as cyanobacteria, algae, and plants, sustain life on earth by converting light energy, water, and CO(2) into chemical energy. However, due to global change and a growing human population, arable land is becoming scarce and resources, including water and fertilizers, are becoming exhausted. It will therefore be crucial to design innovative strategies for sustainable plant production to maintain the food and energy bases of human civilization. Several different strategies for engineering improved photosynthesis in crop plants and introducing novel photosynthetic capacity into microorganisms have been reviewed.

  10. Kinematic amplification strategies in plants and engineering

    Science.gov (United States)

    Charpentier, Victor; Hannequart, Philippe; Adriaenssens, Sigrid; Baverel, Olivier; Viglino, Emmanuel; Eisenman, Sasha

    2017-06-01

    While plants are primarily sessile at the organismal level, they do exhibit a vast array of movements at the organ or sub-organ level. These movements can occur for reasons as diverse as seed dispersal, nutrition, protection or pollination. Their advanced mechanisms generate a myriad of movement typologies, many of which are not fully understood. In recent years, there has been a renewal of interest in understanding the mechanical behavior of plants from an engineering perspective, with an interest in developing novel applications by up-sizing these mechanisms from the micro- to the macro-scale. This literature review identifies the main strategies used by plants to create and amplify movements and anatomize the most recent mechanical understanding of compliant engineering mechanics. The paper ultimately demonstrates that plant movements, rooted in compliance and multi-functionality, can effectively inspire better kinematic/adaptive structures and materials. In plants, the actuators and the deployment structures are fused into a single system. The understanding of those natural movements therefore starts with an exploration of mechanisms at the origins of movements. Plant movements, whether slow or fast, active or passive, reversible or irreversible, are presented and detailed for their mechanical significance. With a focus on displacement amplification, the most recent promising strategies for actuation and adaptive systems are examined with respect to the mechanical principles of shape morphing plant tissues.

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

    DEFF Research Database (Denmark)

    Nielsen, Jens Bredal

    1998-01-01

    enzymes. Despite the prospect of obtaining major improvement through metabolic engineering, this approach is, however, not expected to completely replace the classical approach to strain improvement-random mutagenesis followed by screening. Identification of the optimal genetic changes for improvement......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...

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

    Science.gov (United States)

    Angermayr, S Andreas; Gorchs Rovira, Aleix; Hellingwerf, Klaas J

    2015-06-01

    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 CO(2), light, and water directly. Such cyanobacterial 'cell factories' are constructed to produce biofuels, bioplastics, and commodity chemicals. Efforts of metabolic engineers and synthetic biologists allow the modification of the intermediary metabolism at various branching points, expanding the product range. The new biosynthesis routes 'tap' the metabolism ever more efficiently, particularly through the engineering of driving forces and utilization of cofactors generated during the light reactions of photosynthesis, resulting in higher product titers. High rates of carbon rechanneling ultimately allow an almost-complete allocation of fixed carbon to product above biomass.

  13. Metabolic engineering of microorganisms: general strategies and drug production.

    Science.gov (United States)

    Lee, Sang Yup; Kim, Hyun Uk; Park, Jin Hwan; Park, Jong Myung; Kim, Tae Yong

    2009-01-01

    Many drugs and drug precursors found in natural organisms are rather difficult to synthesize chemically and to extract in large amounts. Metabolic engineering is playing an increasingly important role in the production of these drugs and drug precursors. This is typically achieved by establishing new metabolic pathways leading to the product formation, and enforcing or removing the existing metabolic pathways toward enhanced product formation. Recent advances in system biology and synthetic biology are allowing us to perform metabolic engineering at the whole cell level, thus enabling optimal design of a microorganism for the efficient production of drugs and drug precursors. In this review, we describe the general strategies for the metabolic engineering of microorganisms for the production of drugs and drug precursors. As successful examples of metabolic engineering, the approaches taken toward strain development for the production of artemisinin, an antimalarial drug, and benzylisoquinoline alkaloids, a family of antibacterial and anticancer drugs, are described in detail. Also, systems metabolic engineering of Escherichia coli for the production of L-valine, an important drug precursor, is showcased as an important strategy of future metabolic engineering effort.

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

    Science.gov (United States)

    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.

  15. Transcriptional Regulation of Plant Secondary Metabolism

    Institute of Scientific and Technical Information of China (English)

    Chang-Qing Yang; Xin Fang; Xiu-Ming Wu; Ying-Bo Mao; Ling-Jian Wang; Xiao-Ya Chen

    2012-01-01

    Plant secondary metabolites play critical roles in plant-environment interactions.They are synthesized in different organs or tissues at particular developmental stages,and in response to various environmental stimuli,both biotic and abiotic.Accordingly,corresponding genes are regulated at the transcriptional level by multiple transcription factors.Several families of transcription factors have been identified to participate in controlling the biosynthesis and accumulation of secondary metabolites.These regulators integrate internal (often developmental) and external signals,bind to corresponding cis-elements — which are often in the promoter regions — to activate or repress the expression of enzyme-coding genes,and some of them interact with other transcription factors to form a complex.In this review,we summarize recent research in these areas,with an emphasis on newly-identified transcription factors and their functions in metabolism regulation.

  16. Engineered plant biomass feedstock particles

    Science.gov (United States)

    Dooley, James H.; Lanning, David N.; Broderick, Thomas F.

    2011-10-11

    A novel class of flowable biomass feedstock particles with unusually large surface areas that can be manufactured in remarkably uniform sizes using low-energy comminution techniques. The feedstock particles are roughly parallelepiped in shape and characterized by a length dimension (L) aligned substantially with the grain direction and defining a substantially uniform distance along the grain, a width dimension (W) normal to L and aligned cross grain, and a height dimension (H) normal to W and L. The particles exhibit a disrupted grain structure with prominent end and surface checks that greatly enhances their skeletal surface area as compared to their envelope surface area. The L.times.H dimensions define a pair of substantially parallel side surfaces characterized by substantially intact longitudinally arrayed fibers. The W.times.H dimensions define a pair of substantially parallel end surfaces characterized by crosscut fibers and end checking between fibers. The L.times.W dimensions define a pair of substantially parallel top surfaces characterized by some surface checking between longitudinally arrayed fibers. The feedstock particles are manufactured from a variety of plant biomass materials including wood, crop residues, plantation grasses, hemp, bagasse, and bamboo.

  17. Engineered plant biomass feedstock particles

    Science.gov (United States)

    Dooley, James H [Federal Way, WA; Lanning, David N [Federal Way, WA; Broderick, Thomas F [Lake Forest Park, WA

    2011-10-18

    A novel class of flowable biomass feedstock particles with unusually large surface areas that can be manufactured in remarkably uniform sizes using low-energy comminution techniques. The feedstock particles are roughly parallelepiped in shape and characterized by a length dimension (L) aligned substantially with the grain direction and defining a substantially uniform distance along the grain, a width dimension (W) normal to L and aligned cross grain, and a height dimension (H) normal to W and L. The particles exhibit a disrupted grain structure with prominent end and surface checks that greatly enhances their skeletal surface area as compared to their envelope surface area. The L.times.H dimensions define a pair of substantially parallel side surfaces characterized by substantially intact longitudinally arrayed fibers. The W.times.H dimensions define a pair of substantially parallel end surfaces characterized by crosscut fibers and end checking between fibers. The L.times.W dimensions define a pair of substantially parallel top surfaces characterized by some surface checking between longitudinally arrayed fibers. At least 80% of the particles pass through a 1/4 inch screen having a 6.3 mm nominal sieve opening but are retained by a No. 10 screen having a 2 mm nominal sieve opening. The feedstock particles are manufactured from a variety of plant biomass materials including wood, crop residues, plantation grasses, hemp, bagasse, and bamboo.

  18. PLEVIS (Plant Engineering Visual and Interactive Simulator), a plant engineering visual and interactive simulator

    Energy Technology Data Exchange (ETDEWEB)

    Mizukami, Masumi; Yoshikawa, Eiji; Tanaka, Kazuma; Otsuka, Shiro

    1989-01-01

    Toshiba and Nippon Atomic Industry Group Company (NAIG) have devoted much effort to the development of nuclear power simulators for operator training, such as replica simulators and compact simulators since 1973. These simulators have special-purpose consoles or control room equipment for operation training and run on one or more process computers; therefore, they require a large amount of space, e.g., a special room for installation. The Plant Engineering Visual and Interactive Simulator (PLEVIS) is a real-time plant engineering simulator and operates on a general-purpose desk-side engineering workstation (Sun4 series) with a high-resolution bit-mapped display. The major objectives of PLEVIS are to further improve training performance and efficiency, not only for operators, but also for engineers, and to provide a powerful and cost-effective plant engineering tool. The PELVIS development objectives and background technologies are shown. System configuration and major features are discussed.

  19. Animal and plant stem cells concepts, propagation and engineering

    CERN Document Server

    Pavlović, Mirjana

    2017-01-01

    This book provides a multifaceted look into the world of stem cells and explains the similarities and differences between plant and human stem cells. It explores the intersection between animals and plants and explains their cooperative role in bioengineering studies. The book treats both theoretical and practical aspects of stem cell research. It covers the advantages and limitations of many common applications related to stem cells: their sources, categories, engineering of these cells, reprogramming of their functions, and their role as novel cellular therapeutic approach. Written by experts in the field, the book focuses on aspects of stem cells ranging from expansion-propagation to metabolic reprogramming. It introduces the emergence of cancer stem cells and different modalities in targeted cancer stem cell therapies. It is a valuable source of fresh information for academics and researchers, examining molecular mechanisms of animal and plant stem cell regulation and their usage for therapeutic applicati...

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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

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

    Science.gov (United States)

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

    2013-05-01

    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. We review advances in 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.

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

    Science.gov (United States)

    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.

  4. Metabolic engineering as a tool for enhanced lactic acid production.

    Science.gov (United States)

    Upadhyaya, Bikram P; DeVeaux, Linda C; Christopher, Lew P

    2014-12-01

    Metabolic engineering is a powerful biotechnological tool that finds, among others, increased use in constructing microbial strains for higher lactic acid productivity, lower costs and reduced pollution. Engineering the metabolic pathways has concentrated on improving the lactic acid fermentation parameters, enhancing the acid tolerance of production organisms and their abilities to utilize a broad range of substrates, including fermentable biomass-derived sugars. Recent efforts have focused on metabolic engineering of lactic acid bacteria as they produce high yields and have a small genome size that facilitates their genetic manipulation. We summarize here the current trends in metabolic engineering techniques and strategies for manipulating lactic acid producing organisms developed to address and overcome major challenges in the lactic acid production process.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

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

    DEFF Research Database (Denmark)

    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....

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

    KAUST Repository

    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.

  9. PanDaTox: A tool for accelerated metabolic engineering

    Energy Technology Data Exchange (ETDEWEB)

    Amitai, Gil; Sorek, Rotem

    2012-07-18

    Metabolic engineering is often facilitated by cloning of genes encoding enzymes from various heterologous organisms into E. coli. Such engineering efforts are frequently hampered by foreign genes that are toxic to the E. coli host. We have developed PanDaTox (www.weizmann.ac.il/pandatox), a web-based resource that provides experimental toxicity information for more than 1.5 million genes from hundreds of different microbial genomes. The toxicity predictions, which were extensively experimentally verified, are based on serial cloning of genes into E. coli as part of the Sanger whole genome shotgun sequencing process. PanDaTox can accelerate metabolic engineering projects by allowing researchers to exclude toxic genes from the engineering plan and verify the clonability of selected genes before the actual metabolic engineering experiments are conducted.

  10. Metabolic engineering of micronutrients in crop plants

    OpenAIRE

    Blancquaert, Dieter; De Steur, Hans; Gellynck, Xavier; Straeten, Dominique Van Der

    2017-01-01

    Micronutrient deficiency is a widespread phenomenon, most prevalent in developing countries. Being causally linked to the occurrence of a range of diseases, it affects billions of people worldwide. Enhancing the content of micronutrients in crop products through biotechnology is a promising technique to fight micronutrient malnutrition worldwide. Micronutrient fortification of food products has been implemented in a number of Western countries, but remains inaccessible for poor rural populati...

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

    DEFF Research Database (Denmark)

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

    2004-01-01

    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.......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 metabolic...

  12. Arsenic uptake and metabolism in plants.

    Science.gov (United States)

    Zhao, F J; Ma, J F; Meharg, A A; McGrath, S P

    2009-03-01

    Arsenic (As) is an element that is nonessential for and toxic to plants. Arsenic contamination in the environment occurs in many regions, and, depending on environmental factors, its accumulation in food crops may pose a health risk to humans.Recent progress in understanding the mechanisms of As uptake and metabolism in plants is reviewed here. Arsenate is taken up by phosphate transporters. A number of the aquaporin nodulin26-like intrinsic proteins (NIPs) are able to transport arsenite,the predominant form of As in reducing environments. In rice (Oryza sativa), arsenite uptake shares the highly efficient silicon (Si) pathway of entry to root cells and efflux towards the xylem. In root cells arsenate is rapidly reduced to arsenite, which is effluxed to the external medium, complexed by thiol peptides or translocated to shoots. One type of arsenate reductase has been identified, but its in planta functions remain to be investigated. Some fern species in the Pteridaceae family are able to hyperaccumulate As in above-ground tissues. Hyperaccumulation appears to involve enhanced arsenate uptake, decreased arsenite-thiol complexation and arsenite efflux to the external medium, greatly enhanced xylem translocation of arsenite, and vacuolar sequestration of arsenite in fronds. Current knowledge gaps and future research directions are also identified.

  13. Metabolic engineering of Yarrowia lipolytica for industrial applications.

    Science.gov (United States)

    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.

  14. Microbial metabolic engineering for L-threonine production.

    Science.gov (United States)

    Dong, Xunyan; Quinn, Peter J; Wang, Xiaoyuan

    2012-01-01

    L-threonine, one of the three major amino acids produced throughout the world, has a wide application in industry, as an additive or as a precursor for the biosynthesis of other chemicals. It is predominantly produced through microbial fermentation the efficiency of which largely depends on the quality of strains. Metabolic engineering based on a cogent understanding of the metabolic pathways of L-threonine biosynthesis and regulation provides an effective alternative to the traditional breeding for strain development. Continuing efforts have been made in revealing the mechanisms and regulation of L-threonine producing strains, as well as in metabolic engineering of suitable organisms whereby genetically-defined, industrially competitive L-threonine producing strains have been successfully constructed. This review focuses on the global metabolic and regulatory networks responsible for L-threonine biosynthesis, the molecular mechanisms of regulation, and the strategies employed in strain engineering.

  15. Metabolic engineering for amino-, oligo-, and polysugar production in microbes.

    Science.gov (United States)

    Hossain, Gazi Sakir; Shin, Hyun-Dong; Li, Jianghua; Wang, Miao; Du, Guocheng; Chen, Jian; Liu, Long

    2016-03-01

    Amino-, oligo-, and polysugars are important for both medicinal and industrial applications. Microbial processes used in production of such sugars are not only carbon-intensive and energy-demanding processes but also have other distinct disadvantages such as low productivity, low yields, and by-product contamination. Therefore, metabolic engineering has emerged as an effective tool for developing engineered strains to deliver production strategies for many valuable sugars, which were previously difficult to manufacture by other means, in necessary amounts to support their applications. In this review, the recent strategies used for metabolic engineering are summarized and future prospects of this technique are discussed. We hope that this review will contribute to the development of functional and high-value sugar production by metabolic engineering strategies.

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

    Directory of Open Access Journals (Sweden)

    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.

  17. Engineering of complex protein sialylation in plants

    Science.gov (United States)

    Kallolimath, Somanath; Castilho, Alexandra; Strasser, Richard; Grünwald-Gruber, Clemens; Altmann, Friedrich; Strubl, Sebastian; Galuska, Christina Elisabeth; Zlatina, Kristina; Galuska, Sebastian Peter; Werner, Stefan; Thiesler, Hauke; Werneburg, Sebastian; Hildebrandt, Herbert; Gerardy-Schahn, Rita; Steinkellner, Herta

    2016-01-01

    Sialic acids (Sias) are abundant terminal modifications of protein-linked glycans. A unique feature of Sia, compared with other monosaccharides, is the formation of linear homo-polymers, with its most complex form polysialic acid (polySia). Sia and polySia mediate diverse biological functions and have great potential for therapeutic use. However, technological hurdles in producing defined protein sialylation due to the enormous structural diversity render their precise investigation a challenge. Here, we describe a plant-based expression platform that enables the controlled in vivo synthesis of sialylated structures with different interlinkages and degree of polymerization (DP). The approach relies on a combination of stably transformed plants with transient expression modules. By the introduction of multigene vectors carrying the human sialylation pathway into glycosylation-destructed mutants, transgenic plants that sialylate glycoproteins in α2,6- or α2,3-linkage were generated. Moreover, by the transient coexpression of human α2,8-polysialyltransferases, polySia structures with a DP >40 were synthesized in these plants. Importantly, plant-derived polySia are functionally active, as demonstrated by a cell-based cytotoxicity assay and inhibition of microglia activation. This pathway engineering approach enables experimental investigations of defined sialylation and facilitates a rational design of glycan structures with optimized biotechnological functions. PMID:27444013

  18. Genetic engineering of novel flower colors in floricultural plants: recent advances via transgenic approaches.

    Science.gov (United States)

    Nishihara, Masahiro; Nakatsuka, Takashi

    2010-01-01

    Since the first successful genetic engineering of flower color in petunia, several new techniques have been developed and applied to modify flower color not only in model plants but also in floricultural plants. A typical example is the commercial violet-flowered carnation "Moondust series" developed by Suntry Ltd. and Florigene Ltd. More recently, blue-flowered roses have been successfully produced and are expected to be commercially available in the near future. In recent years, successful modification of flower color by sophisticated regulation of flower-pigment metabolic pathways has become possible. In this chapter, we review recent advances in flower color modification by genetic engineering, especially focusing on the methodology. We have included our own recent results on successful production of flower-color-modified transgenic plants in a model plant, tobacco and an ornamental plant, gentian. Based on these results, genetic engineering of flower color for improvement of floricultural plants is discussed.

  19. Thiamine in plants: aspects of its metabolism and functions.

    Science.gov (United States)

    Goyer, Aymeric

    2010-10-01

    Thiamine diphosphate (vitamin B(1)) plays a fundamental role as an enzymatic cofactor in universal metabolic pathways including glycolysis, the pentose phosphate pathway, and the tricarboxylic acid cycle. In addition, thiamine diphosphate has recently been shown to have functions other than as a cofactor in response to abiotic and biotic stress in plants. Recently, several steps of the plant thiamine biosynthetic pathway have been characterized, and a mechanism of feedback regulation of thiamine biosynthesis via riboswitch has been unraveled. This review focuses on these most recent advances made in our understanding of thiamine metabolism and functions in plants. Phenotypes of plant mutants affected in thiamine biosynthesis are described, and genomics, proteomics, and metabolomics data that have increased further our knowledge of plant thiamine metabolic pathways and functions are summarized. Aspects of thiamine metabolism such as catabolism, salvage, and transport in plants are discussed.

  20. Genetic-Metabolic Coupling for Targeted Metabolic Engineering

    DEFF Research Database (Denmark)

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

    2017-01-01

    Production of chemicals in microbes often employs potent biosynthetic enzymes, which can interact with the microbial native metabolism to affect cell fitness and product yield. However, production optimization largely relies on data collected from wild-type strains in the absence of metabolic...... 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...

  1. LAILAPS: the plant science search engine.

    Science.gov (United States)

    Esch, Maria; Chen, Jinbo; Colmsee, Christian; Klapperstück, Matthias; Grafahrend-Belau, Eva; Scholz, Uwe; Lange, Matthias

    2015-01-01

    With the number of sequenced plant genomes growing, the number of predicted genes and functional annotations is also increasing. The association between genes and phenotypic traits is currently of great interest. Unfortunately, the information available today is widely scattered over a number of different databases. Information retrieval (IR) has become an all-encompassing bioinformatics methodology for extracting knowledge from complex, heterogeneous and distributed databases, and therefore can be a useful tool for obtaining a comprehensive view of plant genomics, from genes to traits. Here we describe LAILAPS (http://lailaps.ipk-gatersleben.de), an IR system designed to link plant genomic data in the context of phenotypic attributes for a detailed forward genetic research. LAILAPS comprises around 65 million indexed documents, encompassing >13 major life science databases with around 80 million links to plant genomic resources. The LAILAPS search engine allows fuzzy querying for candidate genes linked to specific traits over a loosely integrated system of indexed and interlinked genome databases. Query assistance and an evidence-based annotation system enable time-efficient and comprehensive information retrieval. An artificial neural network incorporating user feedback and behavior tracking allows relevance sorting of results. We fully describe LAILAPS's functionality and capabilities by comparing this system's performance with other widely used systems and by reporting both a validation in maize and a knowledge discovery use-case focusing on candidate genes in barley.

  2. Synergizing (13)C Metabolic Flux Analysis and Metabolic Engineering for Biochemical Production.

    Science.gov (United States)

    Guo, Weihua; Sheng, Jiayuan; Feng, Xueyang

    2017-04-20

    Metabolic engineering of industrial microorganisms to produce chemicals, fuels, and drugs has attracted increasing interest as it provides an environment-friendly and renewable route that does not depend on depleting petroleum sources. However, the microbial metabolism is so complex that metabolic engineering efforts often have difficulty in achieving a satisfactory yield, titer, or productivity of the target chemical. To overcome this challenge, (13)C Metabolic Flux Analysis ((13)C-MFA) has been developed to investigate rigorously the cell metabolism and quantify the carbon flux distribution in central metabolic pathways. In the past decade, (13)C-MFA has been widely used in academic labs and the biotechnology industry to pinpoint the key issues related to microbial-based chemical production and to guide the development of the appropriate metabolic engineering strategies for improving the biochemical production. In this chapter we introduce the basics of (13)C-MFA and illustrate how (13)C-MFA has been applied to synergize with metabolic engineering to identify and tackle the rate-limiting steps in biochemical production.

  3. Sulfur containing air pollutants and their effects on plant metabolism

    NARCIS (Netherlands)

    De Kok, L.J.; Maas, F.M.; Stulen, I.; Kuiper, P.J.C.; Mathy, P.

    1988-01-01

    H2S and SO2 affected plant yield at relative low levels without causing visible injury. In general H2S was more toxic than SO2. Short-term exposure (24 to 48 h) of plants to the pollutants affected sulfur, nitrogen and energy metabolism. Both exposure of plants to H2S and SO2 resulted in an accumula

  4. Sulfur containing air pollutants and their effects on plant metabolism

    NARCIS (Netherlands)

    De Kok, L.J.; Maas, F.M.; Stulen, I.; Kuiper, P.J.C.; Mathy, P.

    1988-01-01

    H2S and SO2 affected plant yield at relative low levels without causing visible injury. In general H2S was more toxic than SO2. Short-term exposure (24 to 48 h) of plants to the pollutants affected sulfur, nitrogen and energy metabolism. Both exposure of plants to H2S and SO2 resulted in an

  5. Corynebacterium glutamicum Metabolic Engineering with CRISPR Interference (CRISPRi).

    Science.gov (United States)

    Cleto, Sara; Jensen, Jaide Vk; Wendisch, Volker F; Lu, Timothy K

    2016-05-20

    Corynebacterium glutamicum is an important organism for the industrial production of amino acids. Metabolic pathways in this organism are usually engineered by conventional methods such as homologous recombination, which depends on rare double-crossover events. To facilitate the mapping of gene expression levels to metabolic outputs, we applied CRISPR interference (CRISPRi) technology using deactivated Cas9 (dCas9) to repress genes in C. glutamicum. We then determined the effects of target repression on amino acid titers. Single-guide RNAs directing dCas9 to specific targets reduced expression of pgi and pck up to 98%, and of pyk up to 97%, resulting in titer enhancement ratios of l-lysine and l-glutamate production comparable to levels achieved by gene deletion. This approach for C. glutamicum metabolic engineering, which only requires 3 days, indicates that CRISPRi can be used for quick and efficient metabolic pathway remodeling without the need for gene deletions or mutations and subsequent selection.

  6. Plant Hormones: Metabolism, Signaling and Crosstalk

    Institute of Scientific and Technical Information of China (English)

    Li-Jia Qu; Yunde Zhao

    2011-01-01

    @@ Plants synthesize various hormones in response to environmental cues and developmental signals to ensure their proper growth and development.Elucidation of the molecular mechanisms by which plant hormones control growth and development contributes to our understanding of fundamental plant biology and provides tools to improve crops.Because of their critical roles in plant growth and development, plant hormones have been studied extensively since the early days of plant biology.

  7. Corynebacterium glutamicum Metabolic Engineering with CRISPR Interference (CRISPRi)

    OpenAIRE

    Cleto, Sara; Jensen, Jaide VK; Wendisch, Volker F; Lu, Timothy K.

    2016-01-01

    Corynebacterium glutamicum is an important organism for the industrial production of amino acids. Metabolic pathways in this organism are usually engineered by conventional methods such as homologous recombination, which depends on rare double-crossover events. To facilitate the mapping of gene expression levels to metabolic outputs, we applied CRISPR interference (CRISPRi) technology using deactivated Cas9 (dCas9) to repress genes in C. glutamicum. We then determined the effects of target re...

  8. Metabolic engineering of bacteria for ethanol production

    Energy Technology Data Exchange (ETDEWEB)

    Ingram, L.O.; Gomez, P.F.; Lai, X.; Moniruzzaman, M.; Wood, B.E.; Yomano, L.P.; York, S.W. [Univ. of Florida, Gainesville, FL (United States). Dept. of Microbiology and Cell Science

    1998-04-20

    Technologies are available which will allow the conversion of lignocellulose into fuel ethanol using genetically engineered bacteria. Assembling these into a cost-effective process remains a challenge. The authors` work has focused primarily on the genetic engineering of enteric bacteria using a portable ethanol production pathway. Genes encoding Zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase have been integrated into the chromosome of Escherichia coli B to produce strain KO11 for the fermentation of hemicellulose-derived syrups. This organism can efficiently ferment all hexose and pentose sugars present in the polymers of hemicellulose. Klebsiella oxytoca M5A1 has been genetically engineered in a similar manner to produce strain P2 for ethanol production from cellulose. This organism has the native ability to ferment cellobiose and cellotriose, eliminating the need for one class of cellulase enzymes. The optimal pH for cellulose fermentation with this organism is near that of fungal cellulases. The general approach for the genetic engineering of new biocatalysts has been most successful with enteric bacteria thus far. However, this approach may also prove useful with gram-positive bacteria which have other important traits for lignocellulose conversion. Many opportunities remain for further improvements in the biomass to ethanol processes.

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

    DEFF Research Database (Denmark)

    Jensen, Michael Krogh; Keasling, Jay

    2015-01-01

    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...... 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...... 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....

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

    Science.gov (United States)

    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.

  11. Enzymes and Metabolites in Carbohydrate Metabolism of Desiccation Tolerant Plants

    Directory of Open Access Journals (Sweden)

    Qingwei Zhang

    2016-12-01

    Full Text Available Resurrection plants can tolerate extreme water loss. Substantial sugar accumulation is a phenomenon in resurrection plants during dehydration. Sugars have been identified as one important factor contributing to desiccation tolerance. Phylogenetic diversity of resurrection plants reflects the diversity of sugar metabolism in response to dehydration. Sugars, which accumulate during dehydration, have been shown to protect macromolecules and membranes and to scavenge reactive oxygen species. This review focuses on the performance of enzymes participating in sugar metabolism during dehydration stress. The relation between sugar metabolism and other biochemical activities is discussed and open questions as well as potential experimental approaches are proposed.

  12. Genome editing with engineered nucleases in plants.

    Science.gov (United States)

    Osakabe, Yuriko; Osakabe, Keishi

    2015-03-01

    Numerous examples of successful 'genome editing' now exist. Genome editing uses engineered nucleases as powerful tools to target specific DNA sequences to edit genes precisely in the genomes of both model and crop plants, as well as a variety of other organisms. The DNA-binding domains of zinc finger (ZF) proteins were the first to be used as genome editing tools, in the form of designed ZF nucleases (ZFNs). More recently, transcription activator-like effector nucleases (TALENs), as well as the clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9) system, which utilizes RNA-DNA interactions, have proved useful. A key step in genome editing is the generation of a double-stranded DNA break that is specific to the target gene. This is achieved by custom-designed endonucleases, which enable site-directed mutagenesis via a non-homologous end-joining (NHEJ) repair pathway and/or gene targeting via homologous recombination (HR) to occur efficiently at specific sites in the genome. This review provides an overview of recent advances in genome editing technologies in plants, and discusses how these can provide insights into current plant molecular biology research and molecular breeding technology.

  13. High specificity in plant leaf metabolic responses to arbuscular mycorrhiza.

    Science.gov (United States)

    Schweiger, Rabea; Baier, Markus C; Persicke, Marcus; Müller, Caroline

    2014-05-22

    The chemical composition of plants (phytometabolome) is dynamic and modified by environmental factors. Understanding its modulation allows to improve crop quality and decode mechanisms underlying plant-pest interactions. Many studies that investigate metabolic responses to the environment focus on single model species and/or few target metabolites. However, comparative studies using environmental metabolomics are needed to evaluate commonalities of chemical responses to certain challenges. We assessed the specificity of foliar metabolic responses of five plant species to the widespread, ancient symbiosis with a generalist arbuscular mycorrhizal fungus. Here we show that plant species share a large 'core metabolome' but nevertheless the phytometabolomes are modulated highly species/taxon-specifically. Such a low conservation of responses across species highlights the importance to consider plant metabolic prerequisites and the long time of specific plant-fungus coevolution. Thus, the transferability of findings regarding phytometabolome modulation by an identical AM symbiont is severely limited even between closely related species.

  14. Metabolic engineering of the shikimate pathway

    Science.gov (United States)

    Juminaga, Darmawi; Keasling, Jay D.

    2017-01-10

    The present disclosure relates to engineered microorganisms that produce amino acids and amino acid intermediates. In particular, the disclosure relates to recombinant nucleic acids encoding operons that increase production of aromatic amino acids and the aromatic amino acid intermediate shikimate; microorganisms with increased production of aromatic amino acids and the aromatic amino acid intermediate shikimate; and methods related to the production of aromatic amino acids, the aromatic amino acid intermediate shikimate, and commodity chemicals derived therefrom.

  15. Metabolic engineering of the shikimate pathway

    Energy Technology Data Exchange (ETDEWEB)

    Juminaga, Darmawi; Keasling, Jay D.

    2017-01-10

    The present disclosure relates to engineered microorganisms that produce amino acids and amino acid intermediates. In particular, the disclosure relates to recombinant nucleic acids encoding operons that increase production of aromatic amino acids and the aromatic amino acid intermediate shikimate; microorganisms with increased production of aromatic amino acids and the aromatic amino acid intermediate shikimate; and methods related to the production of aromatic amino acids, the aromatic amino acid intermediate shikimate, and commodity chemicals derived therefrom.

  16. Metabolic engineering of Corynebacterium glutamicum for methanol metabolism.

    Science.gov (United States)

    Witthoff, Sabrina; Schmitz, Katja; Niedenführ, Sebastian; Nöh, Katharina; Noack, Stephan; Bott, Michael; Marienhagen, Jan

    2015-03-01

    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, [(13)C]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.

  17. [Research progress of genetic engineering on medicinal plants].

    Science.gov (United States)

    Teng, Zhong-qiu; Shen, Ye

    2015-02-01

    The application of genetic engineering technology in modern agriculture shows its outstanding role in dealing with food shortage. Traditional medicinal plant cultivation and collection have also faced with challenges, such as lack of resources, deterioration of environment, germplasm of recession and a series of problems. Genetic engineering can be used to improve the disease resistance, insect resistance, herbicides resistant ability of medicinal plant, also can improve the medicinal plant yield and increase the content of active substances in medicinal plants. Thus, the potent biotechnology can play an important role in protection and large area planting of medicinal plants. In the development of medicinal plant genetic engineering, the safety of transgenic medicinal plants should also be paid attention to. A set of scientific safety evaluation and judgment standard which is suitable for transgenic medicinal plants should be established based on the recognition of the particularity of medicinal plants.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    DEFF Research Database (Denmark)

    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...... 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...... production. The involvement of directed metabolic engineering through the integration of tools from genetic engineering, systems biology and mathematical modeling, is also discussed....

  20. Computer Modeling of Carbon Metabolism Enables Biofuel Engineering (Fact Sheet)

    Energy Technology Data Exchange (ETDEWEB)

    2011-09-01

    In an effort to reduce the cost of biofuels, the National Renewable Energy Laboratory (NREL) has merged biochemistry with modern computing and mathematics. The result is a model of carbon metabolism that will help researchers understand and engineer the process of photosynthesis for optimal biofuel production.

  1. Metabolically engineered cells for the production of pinosylvin

    DEFF Research Database (Denmark)

    2008-01-01

    A genetically engineered micro-organism having an operative metabolic pathway producing cinnamoyl-CoA and producing pinosylvin therefrom by the action of a stilbene synthase is used for pinosylvin production. Said cinnamic acid may be formed from L-phenylalanine by a L-phenylalanine ammonia lyase...

  2. Engineering yeast metabolism for production of fuels and chemicals

    DEFF Research Database (Denmark)

    Nielsen, Jens

    2016-01-01

    of metabolic engineering designs, in particular for development of platform strains that can be used for production of a fatty acid derived products, e.g. fatty alcohols and alkanes. It will be argued that with advancement in genome-editing technologies and novel methods for rapid phenotypic screening...

  3. Metabolic engineering for the production of hydrocarbon fuels.

    Science.gov (United States)

    Lee, Sang Yup; Kim, Hye Mi; Cheon, Seungwoo

    2015-06-01

    Biofuels have been attracting increasing attention to provide a solution to the problems of climate change and our dependence on limited fossil oil. During the last decade, metabolic engineering has been performed to develop superior microorganisms for the production of so called advanced biofuels. Among the advanced biofuels, hydrocarbons possess high-energy content and superior fuel properties to other biofuels, and thus have recently been attracting much research interest. Here we review the recent advances in the microbial production of hydrocarbon fuels together with the metabolic engineering strategies employed to develop their production strains. Strategies employed for the production of long-chain and short-chain hydrocarbons derived from fatty acid metabolism along with the isoprenoid-derived hydrocarbons are reviewed. Also, the current limitations and future prospects in hydrocarbon-based biofuel production are discussed.

  4. Applications of computational modeling in metabolic engineering of yeast

    DEFF Research Database (Denmark)

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

    2015-01-01

    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...

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

    DEFF Research Database (Denmark)

    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...... optimal genetic modifications that improve the rate and yield of chemical production. A new generation of COBRA models and methods is now being developed. -. encompassing many biological processes and simulation strategies. -. and next-generation models enable new types of predictions. Here, three key...... 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....

  6. Terpenoid Indole Alkaloids Biosynthesis and Metabolic Engineering in Catharanthus roseus

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Catharanthus roseus L. (Madagascar periwinkle) biosynthesizes a diverse array of secondary metabolites including anticancer dimeric alkaloids (vinblastine and vincristine) and antihypertensive alkaloids (ajmalicine and serpentine). The multi-step terpenoid indole alkaloids (TIAs) biosynthetic pathway in C. roseus is complex and is under strict molecular regulation. Many enzymes and genes involved in the TIAs biosynthesis have been studied in recent decades. Moreover,some regulatory proteins were found recently to control the production of TIAs in C. roseus. Based on mastering the rough scheme of the pathway and cloning the related genes, metabolic engineering of TIAs biosynthesis has been studied in C.roseus aiming at increasing the desired secondary metabolites in the past few years. The present article summarizes recent advances in isolation and characterization of TIAs biosynthesis genes and transcriptional regulators involved in the second metabolic control in C. roseus. Metabolic engineering applications in TIAs pathway via overexpression of these genes and regulators in C. roseus are also discussed.

  7. Cyanobacterial metabolic engineering for biofuel and chemical production.

    Science.gov (United States)

    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 CO2 are contributing to the global greenhouse effect. Large scale use of atmospheric CO2 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 CO2 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.

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

    Science.gov (United States)

    King, Zachary A; Lloyd, Colton J; Feist, Adam M; Palsson, Bernhard O

    2015-12-01

    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 optimal genetic modifications that improve the rate and yield of chemical production. A new generation of COBRA models and methods is now being developed--encompassing many biological processes and simulation strategies-and next-generation models enable new types of predictions. Here, three key 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.

  9. Application of new metabolic engineering tools for Clostridium acetobutylicum.

    Science.gov (United States)

    Lütke-Eversloh, Tina

    2014-07-01

    The renewed interests in clostridial acetone-butanol-ethanol (ABE) fermentation as a next-generation biofuel source led to significantly intensified research in the past few years. This mini-review focuses on the current status of metabolic engineering techniques available for the model organism of ABE fermentation, Clostridium acetobutylicum. A comprehensive survey of various application examples covers two general issues related to both basic and applied research questions: (i) how to improve biofuel production and (ii) what information can be deduced from respective genotype/phenotype manipulations. Recently developed strategies to engineer C. acetobutylicum are summarized including the current portfolio of altered gene expression methodologies, as well as systematic (rational) and explorative (combinatorial) metabolic engineering approaches.

  10. Iodine biofortification of crops: agronomic biofortification, metabolic engineering and iodine bioavailability.

    Science.gov (United States)

    Gonzali, Silvia; Kiferle, Claudia; Perata, Pierdomenico

    2016-10-27

    Iodine deficiency is a widespread micronutrient malnutrition problem, and the addition of iodine to table salt represents the most common prophylaxis tool. The biofortification of crops with iodine is a recent strategy to further enrich the human diet with a potentially cost-effective, well accepted and bioavailable iodine source. Understanding how iodine functions in higher plants is key to establishing suitable biofortification approaches. This review describes the current knowledge regarding iodine physiology in higher plants, and provides updates on recent agronomic and metabolic engineering strategies of biofortification. Whereas the direct administration of iodine is effective to increase the iodine content in many plant species, a more sophisticated genetic engineering approach seems to be necessary for the iodine biofortification of some important staple crops.

  11. Engineering the spatial organization of metabolic pathways

    DEFF Research Database (Denmark)

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

    does however often depend on both heterologous and host enzymes. In this case, no spatial coordination of the biosynthetic enzymes can be expected to be in place. Presumably this contributes to the low productivity regularly observed for heterologous pathways. In one test case, we investigated whether...... 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......, as enzyme fusion combined with down-regulation of a competing pathway and up-regulation of a selected pathway enzyme resulted in a five-fold higher sesquiterpene production. This simple test case demonstrates that engineering of the spatial organization of pathways has great potential for diverting flux...

  12. Metabolic engineering of Saccharomyces cerevisiae to improve succinic acid production based on metabolic profiling.

    Science.gov (United States)

    Ito, Yuma; Hirasawa, Takashi; Shimizu, Hiroshi

    2014-01-01

    We performed metabolic engineering on the budding yeast Saccharomyces cerevisiae for enhanced production of succinic acid. Aerobic succinic acid production in S. cerevisiae was achieved by disrupting the SDH1 and SDH2 genes, which encode the catalytic subunits of succinic acid dehydrogenase. Increased succinic acid production was achieved by eliminating the ethanol biosynthesis pathways. Metabolic profiling analysis revealed that succinic acid accumulated intracellularly following disruption of the SDH1 and SDH2 genes, which suggests that enhancing the export of intracellular succinic acid outside of cells increases succinic acid production in S. cerevisiae. The mae1 gene encoding the Schizosaccharomyces pombe malic acid transporter was introduced into S. cerevisiae, and as a result, succinic acid production was successfully improved. Metabolic profiling analysis is useful in producing chemicals for metabolic engineering of microorganisms.

  13. [Review: plant polyphenols modulate lipid metabolism and related molecular mechanism].

    Science.gov (United States)

    Dai, Yan-li; Zou, Yu-xiao; Liu, Fan; Li, Hong-zhi

    2015-11-01

    Lipid metabolism disorder is an important risk factor to obesity, hyperlipidemia and type 2 diabetes as well as other chronic metabolic disease. It is also a key target in preventing metabolic syndrome, chronic disease prevention. Plant polyphenol plays an important role in maintaining or improving lipid profile in a variety of ways. including regulating cholesterol absorption, inhibiting synthesis and secretion of triglyceride, and lowering plasma low density lipoprotein oxidation, etc. The purpose of this article is to review the lipid regulation effects of plant polyphenols and its related mechanisms.

  14. Compartmentalized metabolic engineering for biochemical and biofuel production.

    Science.gov (United States)

    Huttanus, Herbert M; Feng, Xueyang

    2017-06-01

    Sub-cellular compartments create specialized reaction chambers in eukaryotes. These compartments provide favorable micro-environments for many metabolic processes. Recently, metabolic engineers have explored the concept of pathway compartmentalization to enhance the performance of metabolic pathways. This strategy offers many unique advantages, including (i) increased local concentrations of enzymes and substrates, (ii) accessing alternate substrate pools, (iii) separation from competing reactions, and (iv) isolation of harmful intermediates or conditions needed for the pathway. In this review, the method of localizing metabolic pathways into specific organelles as well as the benefits of pathway compartmentalization in terms of enhancing the production of value-added chemicals is discussed. Copyright © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Toward metabolic engineering in the context of system biology and synthetic biology: advances and prospects.

    Science.gov (United States)

    Liu, Yanfeng; Shin, Hyun-dong; Li, Jianghua; Liu, Long

    2015-02-01

    Metabolic engineering facilitates the rational development of recombinant bacterial strains for metabolite overproduction. Building on enormous advances in system biology and synthetic biology, novel strategies have been established for multivariate optimization of metabolic networks in ensemble, spatial, and dynamic manners such as modular pathway engineering, compartmentalization metabolic engineering, and metabolic engineering guided by genome-scale metabolic models, in vitro reconstitution, and systems and synthetic biology. Herein, we summarize recent advances in novel metabolic engineering strategies. Combined with advancing kinetic models and synthetic biology tools, more efficient new strategies for improving cellular properties can be established and applied for industrially important biochemical production.

  16. Debottlenecking the 1,3-propanediol pathway by metabolic engineering.

    Science.gov (United States)

    Celińska, E

    2010-01-01

    The history of 1,3-propanediol (1,3-PD) conversion from being a specialty chemical to being a bulk chemical illustrates that the concerted effort of different metabolic engineering approaches brings the most successful results. In order to metabolically tailor the 1,3-PD production pathway multiple strategies have been pursued. Knocking-out genes responsible for by-products formation, intergeneric transfer and overexpression of the genes directly involved in the pathway, manipulation with internal redox balance, introduction of a synthetic flux control point, and modification of the substrate mechanism of transport are some of the strategies applied. The metabolic engineering of the microbial 1,3-PD production exploits both native producers and microorganisms with acquired ability to produce the diol via genetic manipulations. Combination of the appropriate genes from homologous and heterologous hosts is expected to bring a desired objective of production of 1,3-PD cheaply, efficiently and independently from non-renewable resources. The state-of-the-art of the 1,3-PD pathway metabolic engineering is reviewed in this paper.

  17. Auxin metabolism rates and implications for plant development

    Directory of Open Access Journals (Sweden)

    Eric M Kramer

    2015-03-01

    Full Text Available Studies of auxin metabolism rarely express their results as a metabolic rate, although the data obtained would often permit such a calculation to be made. We analyze data from 31 previously published papers to quantify the rates of auxin biosynthesis, conjugation, conjugate hydrolysis, and catabolism in seed plants. Most metabolic pathways have rates in the range 10 nM/h to 1 μM/h, with the exception of auxin conjugation, which has rates as high as ~100 μM/h. The highest rates of auxin conjugation suggests that auxin metabolic sinks may be very small, perhaps as small as a single cell. By contrast, the relatively low rate of auxin biosynthesis requires plants to conserve and recycle auxin during long-distance transport. The consequences for plant development are discussed.

  18. Effects of Engineered Nanomaterials on Plants Growth: An Overview

    OpenAIRE

    Farzad Aslani; Samira Bagheri; Nurhidayatullaili Muhd Julkapli; Abdul Shukor Juraimi; Farahnaz Sadat Golestan Hashemi; Ali Baghdadi

    2014-01-01

    Rapid development and wide applications of nanotechnology brought about a significant increment on the number of engineered nanomaterials (ENs) inevitably entering our living system. Plants comprise of a very important living component of the terrestrial ecosystem. Studies on the influence of engineered nanomaterials (carbon and metal/metal oxides based) on plant growth indicated that in the excess content, engineered nanomaterials influences seed germination. It assessed the shoot-to-root ra...

  19. Effects of engineered nanomaterials on plants growth: an overview.

    Science.gov (United States)

    Aslani, Farzad; Bagheri, Samira; Muhd Julkapli, Nurhidayatullaili; Juraimi, Abdul Shukor; Hashemi, Farahnaz Sadat Golestan; Baghdadi, Ali

    2014-01-01

    Rapid development and wide applications of nanotechnology brought about a significant increment on the number of engineered nanomaterials (ENs) inevitably entering our living system. Plants comprise of a very important living component of the terrestrial ecosystem. Studies on the influence of engineered nanomaterials (carbon and metal/metal oxides based) on plant growth indicated that in the excess content, engineered nanomaterials influences seed germination. It assessed the shoot-to-root ratio and the growth of the seedlings. From the toxicological studies to date, certain types of engineered nanomaterials can be toxic once they are not bound to a substrate or if they are freely circulating in living systems. It is assumed that the different types of engineered nanomaterials affect the different routes, behavior, and the capability of the plants. Furthermore, different, or even opposing conclusions, have been drawn from most studies on the interactions between engineered nanomaterials with plants. Therefore, this paper comprehensively reviews the studies on the different types of engineered nanomaterials and their interactions with different plant species, including the phytotoxicity, uptakes, and translocation of engineered nanomaterials by the plant at the whole plant and cellular level.

  20. Metabolic engineering of biosynthesis and sequestration of artemisinin

    NARCIS (Netherlands)

    Wang, B.

    2016-01-01

    The sesquiterpenoid artemisinin (AN) is the most important medicine for the treatment of malaria in humans. The industrial production of AN still mainly depends on extraction from the plant Artemisia annua. However, the concentration of AN in A. annua is low. Although different engineering

  1. Metabolic engineering of biosynthesis and sequestration of artemisinin

    NARCIS (Netherlands)

    Wang, B.

    2016-01-01

    The sesquiterpenoid artemisinin (AN) is the most important medicine for the treatment of malaria in humans. The industrial production of AN still mainly depends on extraction from the plant Artemisia annua. However, the concentration of AN in A. annua is low. Although different engineering strategie

  2. Plant-derived therapeutics for the treatment of metabolic syndrome.

    Science.gov (United States)

    Graf, Brittany L; Raskin, Ilya; Cefalu, William T; Ribnicky, David M

    2010-10-01

    Metabolic syndrome is defined as a set of coexisting metabolic disorders that increase an individual's likelihood of developing type 2 diabetes, cardiovascular disease and stroke. Medicinal plants, some of which have been used for thousands of years, serve as an excellent source of bioactive compounds for the treatment of metabolic syndrome because they contain a wide range of phytochemicals with diverse metabolic effects. In order for botanicals to be effectively used against metabolic syndrome, however, botanical preparations must be characterized and standardized through the identification of their active compounds and respective modes of action, followed by validation in controlled clinical trials with clearly defined endpoints. This review assesses examples of commonly known and partially characterized botanicals to describe specific considerations for the phytochemical, preclinical and clinical characterization of botanicals associated with metabolic syndrome.

  3. Flux analysis in plant metabolic networks: increasing throughput and coverage.

    Science.gov (United States)

    Junker, Björn H

    2014-04-01

    Quantitative information about metabolic networks has been mainly obtained at the level of metabolite contents, transcript abundance, and enzyme activities. However, the active process of metabolism is represented by the flow of matter through the pathways. These metabolic fluxes can be predicted by Flux Balance Analysis or determined experimentally by (13)C-Metabolic Flux Analysis. These relatively complicated and time-consuming methods have recently seen significant improvements at the level of coverage and throughput. Metabolic models have developed from single cell models into whole-organism dynamic models. Advances in lab automation and data handling have significantly increased the throughput of flux measurements. This review summarizes advances to increase coverage and throughput of metabolic flux analysis in plants.

  4. Genetic engineering of syringyl-enriched lignin in plants

    Science.gov (United States)

    Chiang, Vincent Lee; Li, Laigeng

    2004-11-02

    The present invention relates to a novel DNA sequence, which encodes a previously unidentified lignin biosynthetic pathway enzyme, sinapyl alcohol dehydrogenase (SAD) that regulates the biosynthesis of syringyl lignin in plants. Also provided are methods for incorporating this novel SAD gene sequence or substantially similar sequences into a plant genome for genetic engineering of syringyl-enriched lignin in plants.

  5. Metabolic engineering of Saccharomyces cerevisiae for lactose/whey fermentation.

    Science.gov (United States)

    Domingues, Lucília; Guimarães, Pedro M R; Oliveira, Carla

    2010-01-01

    Lactose is an interesting carbon source for the production of several bio-products by fermentation, primarily because it is the major component of cheese whey, the main by-product of dairy activities. However, the microorganism more widely used in industrial fermentation processes, the yeast Saccharomyces cerevisiae, does not have a lactose metabolization system. Therefore, several metabolic engineering approaches have been used to construct lactose-consuming S. cerevisiae strains, particularly involving the expression of the lactose genes of the phylogenetically related yeast Kluyveromyces lactis, but also the lactose genes from Escherichia coli and Aspergillus niger, as reviewed here. Due to the existing large amounts of whey, the production of bio-ethanol from lactose by engineered S. cerevisiae has been considered as a possible route for whey surplus. Emphasis is given in the present review on strain improvement for lactose-to-ethanol bioprocesses, namely flocculent yeast strains for continuous high-cell-density systems with enhanced ethanol productivity.

  6. Improving Metabolic Pathway Efficiency by Statistical Model-Based Multivariate Regulatory Metabolic Engineering.

    Science.gov (United States)

    Xu, Peng; Rizzoni, Elizabeth Anne; Sul, Se-Yeong; Stephanopoulos, Gregory

    2017-01-20

    Metabolic engineering entails target modification of cell metabolism to maximize the production of a specific compound. For empowering combinatorial optimization in strain engineering, tools and algorithms are needed to efficiently sample the multidimensional gene expression space and locate the desirable overproduction phenotype. We addressed this challenge by employing design of experiment (DoE) models to quantitatively correlate gene expression with strain performance. By fractionally sampling the gene expression landscape, we statistically screened the dominant enzyme targets that determine metabolic pathway efficiency. An empirical quadratic regression model was subsequently used to identify the optimal gene expression patterns of the investigated pathway. As a proof of concept, our approach yielded the natural product violacein at 525.4 mg/L in shake flasks, a 3.2-fold increase from the baseline strain. Violacein production was further increased to 1.31 g/L in a controlled benchtop bioreactor. We found that formulating discretized gene expression levels into logarithmic variables (Linlog transformation) was essential for implementing this DoE-based optimization procedure. The reported methodology can aid multivariate combinatorial pathway engineering and may be generalized as a standard procedure for accelerating strain engineering and improving metabolic pathway efficiency.

  7. Cajal bodies and plant RNA metabolism

    Science.gov (United States)

    Intracellular location of components associated with RNA metabolism is a key feature of gene expression regulation, although it is often neglected in favor of more direct factors influencing expression. A number of nuclear and subnuclear compartments exist, each carrying out specific functions. Th...

  8. Metabolic engineering of microbes for oligosaccharide and polysaccharide synthesis

    Directory of Open Access Journals (Sweden)

    Chen Rachel

    2006-07-01

    Full Text Available Abstract Metabolic engineering has recently been embraced as an effective tool for developing whole-cell biocatalysts for oligosaccharide and polysaccharide synthesis. Microbial catalysts now provide a practical means to derive many valuable oligosaccharides, previously inaccessible through other methods, in sufficient quantities to support research and clinical applications. The synthesis process based upon these microbes is scalable as it avoids expensive starting materials. Most impressive is the high product concentrations (up to 188 g/L achieved through microbe-catalyzed synthesis. The overall cost for selected molecules has been brought to a reasonable range (estimated $ 30–50/g. Microbial synthesis of oligosaccharides and polysaccharides is a carbon-intensive and energy-intensive process, presenting some unique challenges in metabolic engineering. Unlike nicotinamide cofactors, the required sugar nucleotides are products of multiple interacting pathways, adding significant complexity to the metabolic engineering effort. Besides the challenge of providing the necessary mammalian-originated glycosyltransferases in active form, an adequate uptake of sugar acceptors can be an issue when another sugar is necessary as a carbon and energy source. These challenges are analyzed, and various strategies used to overcome these difficulties are reviewed in this article. Despite the impressive success of the microbial coupling strategy, there is a need to develop a single strain that can achieve at least the same efficiency. Host selection and the manner with which the synthesis interacts with the central metabolism are two important factors in the design of microbial catalysts. Additionally, unlike in vitro enzymatic synthesis, product degradation and byproduct formation are challenges of whole-cell systems that require additional engineering. A systematic approach that accounts for various and often conflicting requirements of the synthesis holds

  9. Culinary plants and their potential impact on metabolic overload.

    Science.gov (United States)

    Kim, Ji Yeon; Kwon, Oran

    2011-07-01

    Contemporary human behavior has led a large proportion of the population to metabolic overload and obesity. Postprandial hyperlipidemia and hyperglycemia evoke redox imbalance in the short term and lead to complex chronic disease in the long term with repeated occurrence. Complex diseases are best prevented with complex components of plants; thus, current nutrition research has begun to focus on the development of plant-based functional foods and dietary supplements for health and well-being. Furthermore, given the wide range of species, parts, and secondary metabolites, culinary plants can contribute significant variety and complexity to the human diet. Although understanding the health benefits of culinary plants has been one of the great challenges in nutritional science due to their inherent complexity, it is an advantageous pursuit. This review will address the challenges and opportunities relating to studies of the health benefits of culinary plants, with an emphasis on obesity attributed to metabolic overload. © 2011 New York Academy of Sciences.

  10. Metabolic stimulation of plant phenolics for food preservation and health.

    Science.gov (United States)

    Sarkar, Dipayan; Shetty, Kalidas

    2014-01-01

    Plant phenolics as secondary metabolites are key to a plant's defense response against biotic and abiotic stresses. These phytochemicals are also increasingly relevant to food preservation and human health in terms of chronic disease management. Phenolic compounds from different food crops with different chemical structures and biological functions have the potential to act as natural antioxidants. Plant-based human foods are rich with these phenolic phytochemicals and can be used effectively for food preservation and bioactive enrichments through metabolic stimulation of key pathways. Phenolic metabolites protect against microbial degradation of plant-based foods during postharvest storage. Phenolics not only provide biotic protection but also help to counter biochemical and physical food deteriorations and to enhance shelf life and nutritional quality. This review summarizes the role of metabolically stimulated plant phenolics in food preservation and their impact on the prevention of oxidative stress-induced human diseases.

  11. Improving peppermint essential oil yield and composition by metabolic engineering

    OpenAIRE

    Lange, Bernd Markus; Mahmoud, Soheil Seyed; Wildung, Mark R.; Turner, Glenn W.; Davis, Edward M.; Lange, Iris; Baker, Raymond C.; Boydston, Rick A.; Croteau, Rodney B.

    2011-01-01

    Peppermint (Mentha × piperita L.) was transformed with various gene constructs to evaluate the utility of metabolic engineering for improving essential oil yield and composition. Oil yield increases were achieved by overexpressing genes involved in the supply of precursors through the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. Two-gene combinations to enhance both oil yield and composition in a single transgenic line were assessed as well. The most promising results were obtained by tr...

  12. Metabolic engineering of lactate dehydrogenase rescues mice from acidosis.

    Science.gov (United States)

    Acharya, Abhinav P; Rafi, Mohammad; Woods, Elliot C; Gardner, Austin B; Murthy, Niren

    2014-06-05

    Acidosis causes millions of deaths each year and strategies for normalizing the blood pH in acidosis patients are greatly needed. The lactate dehydrogenase (LDH) pathway has great potential for treating acidosis due to its ability to convert protons and pyruvate into lactate and thereby raise blood pH, but has been challenging to develop into a therapy because there are no pharmaceutical-based approaches for engineering metabolic pathways in vivo. In this report we demonstrate that the metabolic flux of the LDH pathway can be engineered with the compound 5-amino-2-hydroxymethylphenyl boronic acid (ABA), which binds lactate and accelerates the consumption of protons by converting pyruvate to lactate and increasing the NAD(+)/NADH ratio. We demonstrate here that ABA can rescue mice from metformin induced acidosis, by binding lactate, and increasing the blood pH from 6.7 to 7.2 and the blood NAD(+)/NADH ratio by 5 fold. ABA is the first class of molecule that can metabolically engineer the LDH pathway and has the potential to have a significant impact on medicine, given the large number of patients that suffer from acidosis.

  13. Synthetic metabolism: engineering biology at the protein and pathway scales.

    Science.gov (United States)

    Martin, Collin H; Nielsen, David R; Solomon, Kevin V; Prather, Kristala L Jones

    2009-03-27

    Biocatalysis has become a powerful tool for the synthesis of high-value compounds, particularly so in the case of highly functionalized and/or stereoactive products. Nature has supplied thousands of enzymes and assembled them into numerous metabolic pathways. Although these native pathways can be use to produce natural bioproducts, there are many valuable and useful compounds that have no known natural biochemical route. Consequently, there is a need for both unnatural metabolic pathways and novel enzymatic activities upon which these pathways can be built. Here, we review the theoretical and experimental strategies for engineering synthetic metabolic pathways at the protein and pathway scales, and highlight the challenges that this subfield of synthetic biology currently faces.

  14. Metabolic engineering for improved fermentation of pentoses by yeasts.

    Science.gov (United States)

    Jeffries, T W; Jin, Y-S

    2004-02-01

    The fermentation of xylose is essential for the bioconversion of lignocellulose to fuels and chemicals, but wild-type strains of Saccharomyces cerevisiae do not metabolize xylose, so researchers have engineered xylose metabolism in this yeast. Glucose transporters mediate xylose uptake, but no transporter specific for xylose has yet been identified. Over-expressing genes for aldose (xylose) reductase, xylitol dehydrogenase and moderate levels of xylulokinase enable xylose assimilation and fermentation, but a balanced supply of NAD(P) and NAD(P)H must be maintained to avoid xylitol production. Reducing production of NADPH by blocking the oxidative pentose phosphate cycle can reduce xylitol formation, but this occurs at the expense of xylose assimilation. Respiration is critical for growth on xylose by both native xylose-fermenting yeasts and recombinant S, cerevisiae. Anaerobic growth by recombinant mutants has been reported. Reducing the respiration capacity of xylose-metabolizing yeasts increases ethanol production. Recently, two routes for arabinose metabolism have been engineered in S. cerevisiae and adapted strains of Pichia stipitis have been shown to ferment hydrolysates with ethanol yields of 0.45 g g(-1) sugar consumed, so commercialization seems feasible for some applications.

  15. Metabolic engineering of Torulopsis glabrata for malate production.

    Science.gov (United States)

    Chen, Xiulai; Xu, Guoqiang; Xu, Nan; Zou, Wei; Zhu, Pan; Liu, Liming; Chen, Jian

    2013-09-01

    The yeast Torulopsis glabrata CCTCC M202019, which is used for industrial pyruvate production, was chosen to explore the suitability of engineering this multi-vitamin auxotrophic yeast for increased malate production. Various metabolic engineering strategies were used to manipulate carbon flux from pyruvate to malate: (i) overexpression of pyruvate carboxylase and malate dehydrogenase; (ii) identification of the bottleneck in malate production by model iNX804; (iii) simultaneous overexpression of genes RoPYC, RoMDH and SpMAE1. Using these strategies, 8.5gL(-1) malate was accumulated in the engineered strain T.G-PMS, which was about 10-fold greater than that of the control strain T.G-26. The results presented here suggest that T. glabrata CCTCC M202019 is a promising candidate for industrial malate production.

  16. Arsenomics: Omics of Arsenic Metabolism in Plants

    Directory of Open Access Journals (Sweden)

    Rudra Deo eTripathi

    2012-07-01

    Full Text Available AbstractArsenic (As contamination of drinking water and groundwater used for irrigation can lead to contamination of the food chain and poses serious health risk to people worldwide. To reduce As intake through the consumption of contaminated food, identification of the mechanisms for As accumulation and detoxification in plant is a prerequisite to develop efficient phytoremediation methods and safer crops with reduced As levels. Transcriptome, proteome and metabolome analysis of any organism reflects the total biological activities at any given time which are responsible for the adaptation of the organism to the surrounding environmental conditions. As these approaches are very important in analyzing plant As transport and accumulation, we termed Arsenomics as approach which deals transcriptome, proteome and metabolome alterations during As exposure. Although, various studies have been performed to understand modulation in transcriptome in response to As, many important questions need to be addressed regarding the translated proteins of plants at proteomic and metabolomic level, resulting in various ecophysiological responses. In this review, the comprehensive knowledge generated in this area has been compiled and analyzed. There is a need to strengthen Arsenomics which will lead to develop of tools to develop As-free plants for safe consumption.

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

    Science.gov (United States)

    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.

  18. Metabolic engineering of Escherichia coli to improve recombinant protein production.

    Science.gov (United States)

    Liu, Min; Feng, Xinjun; Ding, Yamei; Zhao, Guang; Liu, Huizhou; Xian, Mo

    2015-12-01

    Escherichia coli is one of the most widely used strains for recombinant protein production. However, obstacles also exist in both academic researches and industrial applications, such as the metabolic burden, the carbon source waste, and the cells' physiological deterioration. This article reviews recent approaches for improving recombinant protein production in metabolic engineering, including workhorse selection, stress factor application, and carbon flux regulation. Selecting a suitable host is the first key point for recombinant protein production. In general, it all depends on characteristics of the strains and the target proteins. It will be triggered cells physiological deterioration when the medium is significantly different from the cell's natural environment. Coexpression of stress factors can help proteins to fold into their native conformation. Carbon flux regulation is a direct approach for redirecting more carbon flux toward the desirable pathways and products. However, some undesirable consequences are usually found in metabolic engineering, such as glucose transport inhibition, cell growth retardation, and useless metabolite accumulation. More efficient regulators and platform cell factories should be explored to meet a variety of production demands.

  19. Analysis and Engineering of Metabolic Pathway Fluxes in Corynebacterium glutamicum

    Science.gov (United States)

    Wittmann, Christoph

    The Gram-positive soil bacterium Corynebacterium glutamicum was discovered as a natural overproducer of glutamate about 50 years ago. Linked to the steadily increasing economical importance of this microorganism for production of glutamate and other amino acids, the quest for efficient production strains has been an intense area of research during the past few decades. Efficient production strains were created by applying classical mutagenesis and selection and especially metabolic engineering strategies with the advent of recombinant DNA technology. Hereby experimental and computational approaches have provided fascinating insights into the metabolism of this microorganism and directed strain engineering. Today, C. glutamicum is applied to the industrial production of more than 2 million tons of amino acids per year. The huge achievements in recent years, including the sequencing of the complete genome and efficient post genomic approaches, now provide the basis for a new, fascinating era of research - analysis of metabolic and regulatory properties of C. glutamicum on a global scale towards novel and superior bioprocesses.

  20. Label-free selection of RNA aptamers for metabolic engineering.

    Science.gov (United States)

    Hwang, Chuhern; Carothers, James M

    2016-08-15

    RNA aptamers can be assembled into genetic regulatory devices that sense and respond to levels of specific cellular metabolites and thus serve an integral part of designing dynamic control into engineered metabolic pathways. Here, we describe a practical method for generating specific and high affinity aptamers to enable the wider use of in vitro selection and a broader application of aptamers for metabolic engineering. Conventional selection methods involving either radioactive labeling of RNA or the use of label-free methods such as SPR to track aptamer enrichment require resources that are not widely accessible to research groups. We present a label-free selection method that uses small volume spectrophotometers to track RNA enrichment paired with previously characterized affinity chromatography methods. Borrowing techniques used in solid phase peptide synthesis, we present an approach for immobilizing a wide range of metabolites to an amino PEGA matrix. As an illustration, we detail laboratory techniques employed to generate aptamers that bind p-aminophenylalanine, a metabolic precursor for bio-based production of plastics and the pristinamycin family of antibiotics. We focused on the development of methods for ligand immobilization, selection via affinity chromatography, and nucleic acid quantification that can be performed with common laboratory equipment.

  1. Engineering nonphosphorylative metabolism to generate lignocellulose-derived products.

    Science.gov (United States)

    Tai, Yi-Shu; Xiong, Mingyong; Jambunathan, Pooja; Wang, Jingyu; Wang, Jilong; Stapleton, Cole; Zhang, Kechun

    2016-04-01

    Conversion of lignocellulosic biomass into value-added products provides important environmental and economic benefits. Here we report the engineering of an unconventional metabolism for the production of tricarboxylic acid (TCA)-cycle derivatives from D-xylose, L-arabinose and D-galacturonate. We designed a growth-based selection platform to identify several gene clusters functional in Escherichia coli that can perform this nonphosphorylative assimilation of sugars into the TCA cycle in less than six steps. To demonstrate the application of this new metabolic platform, we built artificial biosynthetic pathways to 1,4-butanediol (BDO) with a theoretical molar yield of 100%. By screening and engineering downstream pathway enzymes, 2-ketoacid decarboxylases and alcohol dehydrogenases, we constructed E. coli strains capable of producing BDO from D-xylose, L-arabinose and D-galacturonate. The titers, rates and yields were higher than those previously reported using conventional pathways. This work demonstrates the potential of nonphosphorylative metabolism for biomanufacturing with improved biosynthetic efficiencies.

  2. EWS based visual and interactive simulator for plant engineering

    Energy Technology Data Exchange (ETDEWEB)

    Ohtsuka, Shiroh [Toshiba Corp. (Japan). Isogo Nuclear Engineering Center; Tanaka, Kazuma; Yoshikawa, Eiji [Toshiba Corp. (Japan). Nuclear Engineering Lab.

    1994-12-31

    The `Plant Engineering Visual and Interactive Simulator (PLEVIS)` is a realtime plant engineering simulator and runs on a general-purpose desk-top engineering workstation with a high-resolution bit-mapped display. PLEVIS is unique in that simulation models are integrated with a control/interlock model editor. PLEVIS can be used in a wide variety of applications, some of which are: (1) Design and modification studies of a control and interlock system, (2) Plant response evaluation for plant start-up testing and troubleshooting, (3) Transient recognition and mitigation studies, and (4) Familiarization with the plant process and control/ interlock system concept. The basic features of PLEVIS in order to realize the above applications are described in the presentation. (1 ref., 6 figs.).

  3. Metabolic engineering of resveratrol and other longevity boosting compounds.

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Y; Chen, H; Yu, O

    2010-09-16

    Resveratrol, a compound commonly found in red wine, has attracted many attentions recently. It is a diphenolic natural product accumulated in grapes and a few other species under stress conditions. It possesses a special ability to increase the life span of eukaryotic organisms, ranging from yeast, to fruit fly, to obese mouse. The demand for resveratrol as a food and nutrition supplement has increased significantly in recent years. Extensive work has been carried out to increase the production of resveratrol in plants and microbes. In this review, we will discuss the biosynthetic pathway of resveratrol and engineering methods to heterologously express the pathway in various organisms. We will outline the shortcuts and limitations of common engineering efforts. We will also discuss briefly the features and engineering challenges of other longevity boosting compounds.

  4. Cardio-Metabolic Benefits of Plant-Based Diets

    Directory of Open Access Journals (Sweden)

    Hana Kahleova

    2017-08-01

    Full Text Available Cardio-metabolic disease, namely ischemic heart disease, stroke, obesity, and type 2 diabetes, represent substantial health and economic burdens. Almost one half of cardio-metabolic deaths in the U.S. might be prevented through proper nutrition. Plant-based (vegetarian and vegan diets are an effective strategy for improving nutrient intake. At the same time, they are associated with decreased all-cause mortality and decreased risk of obesity, type 2 diabetes, and coronary heart disease. Evidence suggests that plant-based diets may reduce the risk of coronary heart disease events by an estimated 40% and the risk of cerebral vascular disease events by 29%. These diets also reduce the risk of developing metabolic syndrome and type 2 diabetes by about one half. Properly planned vegetarian diets are healthful, effective for weight and glycemic control, and provide metabolic and cardiovascular benefits, including reversing atherosclerosis and decreasing blood lipids and blood pressure. The use of plant-based diets as a means of prevention and treatment of cardio-metabolic disease should be promoted through dietary guidelines and recommendations.

  5. Dynamic metabolic flux analysis of plant cell wall synthesis.

    Science.gov (United States)

    Chen, Xuewen; Alonso, Ana P; Shachar-Hill, Yair

    2013-07-01

    The regulation of plant cell wall synthesis pathways remains poorly understood. This has become a bottleneck in designing bioenergy crops. The goal of this study was to analyze the regulation of plant cell wall precursor metabolism using metabolic flux analysis based on dynamic labeling experiments. Arabidopsis T87 cells were cultured heterotrophically with (13)C labeled sucrose. The time course of ¹³C labeling patterns in cell wall precursors and related sugar phosphates was monitored using liquid chromatography tandem mass spectrometry until steady state labeling was reached. A kinetic model based on mass action reaction mechanisms was developed to simulate the carbon flow in the cell wall synthesis network. The kinetic parameters of the model were determined by fitting the model to the labeling time course data, cell wall composition, and synthesis rates. A metabolic control analysis was performed to predict metabolic regulations that may improve plant biomass composition for biofuel production. Our results describe the routes and rates of carbon flow from sucrose to cell wall precursors. We found that sucrose invertase is responsible for the entry of sucrose into metabolism and UDP-glucose-4-epimerase plays a dominant role in UDP-Gal synthesis in heterotrophic Aradidopsis cells under aerobic conditions. We also predicted reactions that exert strong regulatory influence over carbon flow to cell wall synthesis and its composition.

  6. Connecting proline metabolism and signaling pathways in plant senescence

    Directory of Open Access Journals (Sweden)

    Lu eZhang

    2015-07-01

    Full Text Available The amino acid proline has a unique biological role in stress adaptation. Proline metabolism is manipulated under stress by multiple and complex regulatory pathways and can profoundly influence cell death and survival in microorganisms, plants, and animals. Though the effects of proline are mediated by diverse signaling pathways, a common theme appears to be the generation of reactive oxygen species (ROS due to proline oxidation being coupled to the respiratory electron transport chain. Considerable research has been devoted to understand how plants exploit proline metabolism in response to abiotic and biotic stress. Here, we review potential mechanisms by which proline metabolism influences plant senescence, namely in the petal and leaf. Recent studies of petal senescence suggest proline content is manipulated to meet energy demands of senescing cells. In the flower and leaf, proline metabolism may influence ROS signaling pathways that delay senescence progression. Future studies focusing on the mechanisms by which proline metabolic shifts occur during senescence may lead to novel methods to rescue crops under stress and to preserve post-harvest agricultural products.

  7. Quantifying plant phenotypes with isotopic labeling and metabolic flux analysis

    Science.gov (United States)

    Analyses of metabolic flux using stable isotopes in plants have traditionally been restricted to tissues with presumed homogeneous cell populations such as developing seeds, cell suspensions, or cultured roots and root tips. It is now possible to describe these and other more complex tissues such a...

  8. Biochemistry and occurrence of O-demethylation in plant metabolism

    Directory of Open Access Journals (Sweden)

    Jillian Hagel

    2010-07-01

    Full Text Available Demethylases play a pivitol role in numerous biological processes from covalent histone modification and DNA repair to specialized metabolism in plants and microorganisms. Enzymes that catalyze O- and N-demethylation include 2-oxoglutarate (2OG/Fe(II-dependent dioxygenases, cytochromes P450, Rieske-domain proteins and flavin adenine dinucleotide (FAD-dependent oxidases. Proposed mechanisms for demethylation by 2OG/Fe(II-dependent enzymes involve hydroxylation at the O- or N-linked methyl group followed by formaldehyde elimination. Members of this enzyme family catalyze a wide variety of reactions in diverse plant metabolic pathways. Recently, we showed that 2OG/Fe(II-dependent dioxygenases catalyze the unique O-demethylation steps of morphine biosynthesis in opium poppy, which provides a rational basis for the widespread occurrence of demethylases in benzylisoquinoline alkaloid metabolism.

  9. Whole genome sequencing of Saccharomyces cerevisiae: from genotype to phenotype for improved metabolic engineering applications

    DEFF Research Database (Denmark)

    Otero, José Manuel; Vongsangnak, Wanwipa; Asadollahi, Mohammadali;

    2010-01-01

    BACKGROUND: The need for rapid and efficient microbial cell factory design and construction are possible through the enabling technology, metabolic engineering, which is now being facilitated by systems biology approaches. Metabolic engineering is often complimented by directed evolution, where s...... that provides clear and high-probability of success metabolic engineering targets. The genome sequence, annotation, and a SNP viewer of CEN.PK113-7D are deposited at http://www.sysbio.se/cenpk.......BACKGROUND: The need for rapid and efficient microbial cell factory design and construction are possible through the enabling technology, metabolic engineering, which is now being facilitated by systems biology approaches. Metabolic engineering is often complimented by directed evolution, where...... selective pressure is applied to a partially genetically engineered strain to confer a desirable phenotype. The exact genetic modification or resulting genotype that leads to the improved phenotype is often not identified or understood to enable further metabolic engineering. RESULTS: In this work we...

  10. Development of biosensors and their application in metabolic engineering.

    Science.gov (United States)

    Zhang, Jie; Jensen, Michael K; Keasling, Jay D

    2015-10-01

    In a sustainable bioeconomy, many commodities and high value chemicals, including pharmaceuticals, will be manufactured using microbial cell factories from renewable feedstocks. These cell factories can be efficiently generated by constructing libraries of diversified genomes followed by screening 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 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 on the metabolism for improved performance of cell factories. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Crossing kingdoms: Using decellularized plants as perfusable tissue engineering scaffolds.

    Science.gov (United States)

    Gershlak, Joshua R; Hernandez, Sarah; Fontana, Gianluca; Perreault, Luke R; Hansen, Katrina J; Larson, Sara A; Binder, Bernard Y K; Dolivo, David M; Yang, Tianhong; Dominko, Tanja; Rolle, Marsha W; Weathers, Pamela J; Medina-Bolivar, Fabricio; Cramer, Carole L; Murphy, William L; Gaudette, Glenn R

    2017-05-01

    Despite significant advances in the fabrication of bioengineered scaffolds for tissue engineering, delivery of nutrients in complex engineered human tissues remains a challenge. By taking advantage of the similarities in the vascular structure of plant and animal tissues, we developed decellularized plant tissue as a prevascularized scaffold for tissue engineering applications. Perfusion-based decellularization was modified for different plant species, providing different geometries of scaffolding. After decellularization, plant scaffolds remained patent and able to transport microparticles. Plant scaffolds were recellularized with human endothelial cells that colonized the inner surfaces of plant vasculature. Human mesenchymal stem cells and human pluripotent stem cell derived cardiomyocytes adhered to the outer surfaces of plant scaffolds. Cardiomyocytes demonstrated contractile function and calcium handling capabilities over the course of 21 days. These data demonstrate the potential of decellularized plants as scaffolds for tissue engineering, which could ultimately provide a cost-efficient, "green" technology for regenerating large volume vascularized tissue mass. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

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

    DEFF Research Database (Denmark)

    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...

  13. Development of biosensors and their application in metabolic engineering

    DEFF Research Database (Denmark)

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

    2015-01-01

    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...... 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...

  14. Metabolic engineering for isoprenoid-based biofuel production.

    Science.gov (United States)

    Gupta, P; Phulara, S C

    2015-09-01

    Sustainable economic and industrial growth is the need of the hour and it requires renewable energy resources having better performance and compatibility with existing fuel infrastructure from biological routes. Isoprenoids (C ≥ 5) can be a potential alternative due to their diverse nature and physiochemical properties similar to that of petroleum based fuels. In the past decade, extensive research has been done to utilize metabolic engineering strategies in micro-organisms primarily, (i) to overcome the limitations associated with their natural and non-natural production and (ii) to develop commercially competent microbial strain for isoprenoid-based biofuel production. This review briefly describes the engineered isoprenoid biosynthetic pathways in well-characterized microbial systems for the production of several isoprenoid-based biofuels and fuel precursors.

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

    DEFF Research Database (Denmark)

    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...... enzymes involved in biosynthesis of the precursor for PUFAs, or codon optimization of the heterologous genes, or expression of heterologous enzymes involved in the biosynthesis of the precursor for PUFAs....

  16. Development of nuclear power plant real-time engineering simulator

    Institute of Scientific and Technical Information of China (English)

    LIN Meng; YANG Yan-Hua; ZHANG Rong-Hua; HU Rui

    2005-01-01

    A nuclear power plant real-time engineering simulator was developed based on general-purpose thermal-hydraulic system simulation code RELAP5. It main1y consists of three parts: improved thermal-hydraulic system simulation code RELAP5, control and protection system and human-machine interface. A normal transient of CHASHMA nuclear power plant turbine step load change from 100% to 90% of full power, was simulated by the engineering simulator as an application example. This paper presents structure and main features of the engineering simulator, and application results are shown and discussed.

  17. Metabolic engineering of Arabidopsis for remediation of different polycyclic aromatic hydrocarbons using a hybrid bacterial dioxygenase complex.

    Science.gov (United States)

    Peng, Rihe; Fu, Xiaoyan; Tian, Yongsheng; Zhao, Wei; Zhu, Bo; Xu, Jing; Wang, Bo; Wang, Lijuan; Yao, Quanhong

    2014-11-01

    The widespread presence of polycyclic aromatic hydrocarbons (PAHs) and their potential harm to various organisms has generated interest in efficiently eliminating these compounds from the environment. Phytoremediation is an efficient technology for cleaning up pollutants. However, unlike microorganisms, plants lack the catabolic pathway for complete degradation of these dangerous groups of compounds. One way to enhance the potential of plants for remediation of these compounds is by transferring genes involved in xenobiotic degradation from microbes to plants. In this paper, four genes, namely nidA and nidB (encoding the large and small subunits of naphthalene dioxygenase of Mycobacterium vanbaalenii PYR-1) as well as NahAa and NahAb (encoding flavoprotein reductase and ferredoxin of the electron-transport chain of the Pseudomonas putida G7 naphthalene dioxygenase system), were transferred and ectopically expressed in Arabidopsis thaliana. Transgenic Arabidopsis plants overexpressing the heterozygous naphthalene dioxygenase system exhibited enhanced tolerance toward 2-4 rings PAHs. Transgenic plants assimilated PAHs from the culture media faster and accumulated less in vivo than wild-type plants. Furthermore, examination of metabolic intermediates by gas chromatography-mass spectrometry revealed that the naphthalene metabolic pathway in transgenic plants mainly involves the dioxygenase pathway. Taken together, our findings suggest that grafting the naphthalene dioxygenase complex into plants is a possible strategy to breed PAH-tolerant plants to efficiently degrade PAHs in the environment. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  18. Shedding light on ethylene metabolism in higher plants

    Directory of Open Access Journals (Sweden)

    Maria Aurineide Rodrigues

    2014-12-01

    Full Text Available Ethylene metabolism in higher plants is regulated by a wide array of endogenous and environmental factors. During most physiological processes, ethylene levels are mainly determined by a strict control of the rate-limiting biosynthetic steps responsible for the production of 1-aminocyclopropane-1-carboxylic acid (ACC and its subsequent conversion to ethylene. Responsible for these reactions, the key enzymes ACC synthase and ACC oxidase are encoded by multigene families formed by members that can be differentially regulated at the transcription and post-translational levels by specific developmental and environmental signals. Among the wide variety of environmental cues controlling plant ethylene production, light quality, duration and intensity have consistently been demonstrated to influence the metabolism of this plant hormone in diverse plant tissues, organs and species. Although still not completely elucidated, the mechanisms underlying the interaction between light signal transduction and ethylene evolution appears to involve a complex network that includes central transcription factors connecting multiple signaling pathways, which can be reciprocally modulated by ethylene itself, other phytohormones, and specific light wavelengths. Accumulating evidence has indicated particular photoreceptors as essential mediators in light-induced signaling cascades affecting ethylene levels. Therefore, this review specifically focuses on discussing the current knowledge of the potential molecular mechanisms implicated in the light-induced responses affecting ethylene metabolism during the regulation of developmental and metabolic plant responses. Besides presenting the state of the art in this research field, some overlooked mechanisms and future directions to elucidate the exact nature of the light-ethylene interplay in higher plants will also be compiled and discussed.

  19. Transfer of engineered genes from crop to wild plants

    DEFF Research Database (Denmark)

    Bagger Jørgensen, Rikke; Hauser, T.P.; Mikkelsen, T.R.;

    1996-01-01

    The escape of engineered genes - genes inserted using recombinant DNA techniques - from cultivated plants to wild or weedy relatives has raised concern about possible risks to the environment or to health. The media have added considerably to public concern by suggesting that such gene escape...... is a new and rather unexpected phenomenon. However, transfer of engineered genes between plants is not at-all surprising, because it is mediated by exactly the same mechanisms as those responsible for transferring endogenous plant genes: it takes place by sexual crosses, with pollen as the carrier...

  20. Changing trends in biotechnology of secondary metabolism in medicinal and aromatic plants.

    Science.gov (United States)

    Gandhi, Sumit G; Mahajan, Vidushi; Bedi, Yashbir S

    2015-02-01

    Medicinal and aromatic plants are known to produce secondary metabolites that find uses as flavoring agents, fragrances, insecticides, dyes and drugs. Biotechnology offers several choices through which secondary metabolism in medicinal plants can be altered in innovative ways, to overproduce phytochemicals of interest, to reduce the content of toxic compounds or even to produce novel chemicals. Detailed investigation of chromatin organization and microRNAs affecting biosynthesis of secondary metabolites as well as exploring cryptic biosynthetic clusters and synthetic biology options, may provide additional ways to harness this resource. Plant secondary metabolites are a fascinating class of phytochemicals exhibiting immense chemical diversity. Considerable enigma regarding their natural biological functions and the vast array of pharmacological activities, amongst other uses, make secondary metabolites interesting and important candidates for research. Here, we present an update on changing trends in the biotechnological approaches that are used to understand and exploit the secondary metabolism in medicinal and aromatic plants. Bioprocessing in the form of suspension culture, organ culture or transformed hairy roots has been successful in scaling up secondary metabolite production in many cases. Pathway elucidation and metabolic engineering have been useful to get enhanced yield of the metabolite of interest; or, for producing novel metabolites. Heterologous expression of putative plant secondary metabolite biosynthesis genes in a microbe is useful to validate their functions, and in some cases, also, to produce plant metabolites in microbes. Endophytes, the microbes that normally colonize plant tissues, may also produce the phytochemicals produced by the host plant. The review also provides perspectives on future research in the field.

  1. Human factor engineering applied to nuclear power plant design

    Energy Technology Data Exchange (ETDEWEB)

    Manrique, A. [TECNATOM SA, BWR General Electric Business Manager, Madrid (Spain); Valdivia, J.C. [TECNATOM SA, Operation Engineering Project Manager, Madrid (Spain); Jimenez, A. [TECNATOM SA, Operation Engineering Div. Manager, Madrid (Spain)

    2001-07-01

    For the design and construction of new nuclear power plants as well as for maintenance and operation of the existing ones new man-machine interface designs and modifications are been produced. For these new designs Human Factor Engineering must be applied the same as for any other traditional engineering discipline. Advantages of implementing adequate Human Factor Engineering techniques in the design of nuclear reactors have become not only a fact recognized by the majority of engineers and operators but also an explicit requirement regulated and mandatory for the new designs of the so called advanced reactors. Additionally, the big saving achieved by a nuclear power plant having an operating methodology which significantly decreases the risk of operating errors makes it necessary and almost vital its implementation. The first step for this is preparing a plan to incorporate all the Human Factor Engineering principles and developing an integral design of the Instrumentation and Control and Man-machine interface systems. (author)

  2. Metabolic engineering of the diterpenoid sclareol in the moss Physcomitrella patens

    DEFF Research Database (Denmark)

    Pan, Xiwu

    Plant terpenoids play indispensable roles in primary metabolism as the vital constituents in photosynthesis (chlorophylls, carotenoids and plastoquinones), respiration (ubiquinone) and development regulation (gibberellins, abscisic acid, cytokinin and brassinosteroids). They are also the membrane...... sclarea). Metabolic engineering is an attractive solution to achieve cost-effective and sustainable production of these valuable terpenoids. In this study, we introduced the sclareol biosynthetic genes into the moss Physcomitrella patens and obtained the novel compound successfully. As a proof......-of-concept, the best yield of sclareol was 0.96 μg/mg dry weights (DW) in liquid culture, whereas a larger proportion was released into media that leading to a total production of 2.83 μg/mg DW. We propose that the sclareol precursor (GGPP) pool is capable of flexibly responding to either PpCPS/KS disruption...

  3. Apparatus and plant engineering. Apparate- und Anlagentechnik

    Energy Technology Data Exchange (ETDEWEB)

    Klapp, E. (Erlangen-Nuernberg Univ., Erlangen (Germany, F.R.). Lehrstuhl fuer Apparatetechnik und Anlagenbau)

    1980-01-01

    The book has been written to help in planning economically components and systems for materials and energy conversion, in calculating and designing and, finally, commissioning those plants, especially with a view to practical design aspects, - on the basis of sufficient background information about process and materials characteristics, the behavior of materials and plants, fault conditions, and safety evaluation and - with the help of references appropriate to the problems to be dealt with.

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

    DEFF Research Database (Denmark)

    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 prod......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...

  5. Impeded Carbohydrate Metabolism in Rice Plants under Submergence Stress

    Institute of Scientific and Technical Information of China (English)

    Malay Kumar ADAK; Nirmalya GHOSH; Dilip Kumar DASGUPTA; Sudha GUPTA

    2011-01-01

    The detrimental effects of submergence on physiological performances of some rice varieties with special references to carbohydrate metabolisms and their allied enzymes during post-flowering stages have been documented and clarified in the present investigation.It was found that photosynthetic rate and concomitant translocation of sugars into the panicles were both related to the yield.The detrimental effects of the complete submergence were recorded in generation of sucrose,starch,sucrose phosphate synthase and phosphorylase activity in the developing panicles of the plants as compared to those under normal or control (i.e.non-submerged) condition.The accumulation of starch was significantly lower in plants under submergence and that was correlated with ADP-glucose pyrophosphorylase activity.Photosynthetic rate was most affected under submergence in varying days of post-flowering and was also related to the down regulation of Ribulose bisphosphate carboxylase activity.However,under normal or control condition,there recorded a steady maintenance of photosynthetic rate at the post-flowering stages and significantly higher values of Ribulose bisphosphate carboxylase activity.Still,photosynthetic rate of the plants under both control and submerged conditions had hardly any significant correlation with sugar accumulation and other enzymes of carbohydrate metabolism like invertase with grain yield.Finally,plants under submergence suffered significant loss of yield by poor grain filling which was related to impeded carbohydrate metabolism in the tissues.It is evident that loss of yield under submergence is attributed both by lower sink size or sink capacity (number of panicles,in this case) as well as subdued carbohydrate metabolism in plants and its subsequent partitioning into the grains.

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

    Science.gov (United States)

    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. Process engineering for bioflavour production with metabolically active yeasts - a mini-review.

    Science.gov (United States)

    Carlquist, Magnus; Gibson, Brian; Karagul Yuceer, Yonca; Paraskevopoulou, Adamantini; Sandell, Mari; Angelov, Angel I; Gotcheva, Velitchka; Angelov, Angel D; Etschmann, Marlene; de Billerbeck, Gustavo M; Lidén, Gunnar

    2015-01-01

    Flavours are biologically active molecules of large commercial interest in the food, cosmetics, detergent and pharmaceutical industries. The production of flavours can take place by either extraction from plant materials, chemical synthesis, biological conversion of precursor molecules or de novo biosynthesis. The latter alternatives are gaining importance through the rapidly growing fields of systems biology and metabolic engineering, giving efficient production hosts for the so-called 'bioflavours', which are natural flavour and/or fragrance compounds obtained with cell factories or enzymatic systems. Yeasts are potential production hosts for bioflavours. In this mini-review, we give an overview of bioflavour production in yeasts from the process-engineering perspective. Two specific examples, production of 2-phenylethanol and vanillin, are used to illustrate the process challenges and strategies used.

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

    Energy Technology Data Exchange (ETDEWEB)

    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: the ultimate paradigm for continuous pharmaceutical manufacturing.

    Science.gov (United States)

    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.

  10. Rhizobia species: A Boon for "Plant Genetic Engineering".

    Science.gov (United States)

    Patel, Urmi; Sinha, Sarika

    2011-10-01

    Since past three decades new discoveries in plant genetic engineering have shown remarkable potentials for crop improvement. Agrobacterium Ti plasmid based DNA transfer is no longer the only efficient way of introducing agronomically important genes into plants. Recent studies have explored a novel plant genetic engineering tool, Rhizobia sp., as an alternative to Agrobacterium, thereby expanding the choice of bacterial species in agricultural plant biotechnology. Rhizobia sp. serve as an open license source with no major restrictions in plant biotechnology and help broaden the spectrum for plant biotechnologists with respect to the use of gene transfer vehicles in plants. New efficient transgenic plants can be produced by transferring genes of interest using binary vector carrying Rhizobia sp. Studies focusing on the interactions of Rhizobia sp. with their hosts, for stable and transient transformation and expression of genes, could help in the development of an adequate gene transfer vehicle. Along with being biologically beneficial, it may also bring a new means for fast economic development of transgenic plants, thus giving rise to a new era in plant biotechnology, viz. "Rhizobia mediated transformation technology."

  11. Metabolic Engineering of Eukaryotic Microalgae: Potential and Challenges come with great diversity

    Directory of Open Access Journals (Sweden)

    Javier A Gimpel

    2015-12-01

    Full Text Available The great phylogenetic diversity of microalgae is corresponded by a wide arrange of interesting and useful metabolites. Nonetheless metabolic engineering in microalgae has been limited, since specific transformation tools must be developed for each species for either the nuclear or chloroplast genomes. Microalgae as production platforms for metabolites offer several advantages over plants and other microorganisms, like the ability of GMO containment and reduced costs in culture media, respectively. Currently, microalgae have proved particularly well suited for the commercial production of omega-3 fatty acids and carotenoids. Therefore most metabolic engineering strategies have been developed for these metabolites. Microalgal biofuels have also drawn great attention recently, resulting in efforts for improving the production of hydrogen and photosynthates, particularly triacylglycerides. Metabolic pathways of microalgae have also been manipulated in order to improve photosynthetic growth under specific conditions and for achieving trophic conversion. Although these pathways are not strictly related to secondary metabolites, the synthetic biology approaches could potentially be translated to this field and will also be discussed.

  12. In Metabolic Engineering of Eukaryotic Microalgae: Potential and Challenges Come with Great Diversity

    Science.gov (United States)

    Gimpel, Javier A.; Henríquez, Vitalia; Mayfield, Stephen P.

    2015-01-01

    The great phylogenetic diversity of microalgae is corresponded by a wide arrange of interesting and useful metabolites. Nonetheless metabolic engineering in microalgae has been limited, since specific transformation tools must be developed for each species for either the nuclear or chloroplast genomes. Microalgae as production platforms for metabolites offer several advantages over plants and other microorganisms, like the ability of GMO containment and reduced costs in culture media, respectively. Currently, microalgae have proved particularly well suited for the commercial production of omega-3 fatty acids and carotenoids. Therefore most metabolic engineering strategies have been developed for these metabolites. Microalgal biofuels have also drawn great attention recently, resulting in efforts for improving the production of hydrogen and photosynthates, particularly triacylglycerides. Metabolic pathways of microalgae have also been manipulated in order to improve photosynthetic growth under specific conditions and for achieving trophic conversion. Although these pathways are not strictly related to secondary metabolites, the synthetic biology approaches could potentially be translated to this field and will also be discussed. PMID:26696985

  13. Metabolically Engineered Fungal Cells With Increased Content Of Polyunsaturated Fatty Acids

    DEFF Research Database (Denmark)

    2008-01-01

    This invention relates to the production of fatty acids and particularly to the production of the polyunsaturated fatty acids (PUFAs) arachidonic acid (ARA) and eicosapentaenoic acid (EPA) in genetically engineered fungal cells, in particular, to metabolically engineered Saccharomyces cerevisiae...

  14. Metabolic engineering of yeast for fermentative production of flavonoids

    DEFF Research Database (Denmark)

    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...... biosynthetic pathways, comprising up to eight heterologous genes from plants. The obtained titers of kaempferol 26.57±2.66mgL-1 and quercetin 20.38±2.57mgL-1 exceed the previously reported titers in yeast. This is also the first report of de novo biosynthesis of resokaempferol and fisetin in yeast. The work...

  15. An integrated network visualization framework towards metabolic engineering applications.

    Science.gov (United States)

    Noronha, Alberto; Vilaça, Paulo; Rocha, Miguel

    2014-12-30

    Over the last years, several methods for the phenotype simulation of microorganisms, under specified genetic and environmental conditions have been proposed, in the context of Metabolic Engineering (ME). These methods provided insight on the functioning of microbial metabolism and played a key role in the design of genetic modifications that can lead to strains of industrial interest. On the other hand, in the context of Systems Biology research, biological network visualization has reinforced its role as a core tool in understanding biological processes. However, it has been scarcely used to foster ME related methods, in spite of the acknowledged potential. In this work, an open-source software that aims to fill the gap between ME and metabolic network visualization is proposed, in the form of a plugin to the OptFlux ME platform. The framework is based on an abstract layer, where the network is represented as a bipartite graph containing minimal information about the underlying entities and their desired relative placement. The framework provides input/output support for networks specified in standard formats, such as XGMML, SBGN or SBML, providing a connection to genome-scale metabolic models. An user-interface makes it possible to edit, manipulate and query nodes in the network, providing tools to visualize diverse effects, including visual filters and aspect changing (e.g. colors, shapes and sizes). These tools are particularly interesting for ME, since they allow overlaying phenotype simulation results or elementary flux modes over the networks. The framework and its source code are freely available, together with documentation and other resources, being illustrated with well documented case studies.

  16. Systems metabolic engineering of microorganisms to achieve large-scale production of flavonoid scaffolds.

    Science.gov (United States)

    Wu, Junjun; Du, Guocheng; Zhou, Jingwen; Chen, Jian

    2014-10-20

    Flavonoids possess pharmaceutical potential due to their health-promoting activities. The complex structures of these products make extraction from plants difficult, and chemical synthesis is limited because of the use of many toxic solvents. Microbial production offers an alternate way to produce these compounds on an industrial scale in a more economical and environment-friendly manner. However, at present microbial production has been achieved only on a laboratory scale and improvements and scale-up of these processes remain challenging. Naringenin and pinocembrin, which are flavonoid scaffolds and precursors for most of the flavonoids, are the model molecules that are key to solving the current issues restricting industrial production of these chemicals. The emergence of systems metabolic engineering, which combines systems biology with synthetic biology and evolutionary engineering at the systems level, offers new perspectives on strain and process optimization. In this review, current challenges in large-scale fermentation processes involving flavonoid scaffolds and the strategies and tools of systems metabolic engineering used to overcome these challenges are summarized. This will offer insights into overcoming the limitations and challenges of large-scale microbial production of these important pharmaceutical compounds.

  17. Engineering phenolics metabolism in the grasses using transcription factors

    Energy Technology Data Exchange (ETDEWEB)

    Grotewold, Erich [The Ohio State University

    2013-07-26

    The economical competitiveness of agriculture-derived biofuels can be significantly enhanced by increasing biomass/acre yields and by furnishing the desired carbon balance for facilitating liquid fuel production (e.g., ethanol) or for high-energy solid waste availability to be used as biopower (e.g., for electricity production). Biomass production and carbon balance are tightly linked to the biosynthesis of phenolic compounds, which are found in crops and in agricultural residues either as lignins, as part of the cell wall, or as soluble phenolics which play a variety of functions in the biology of plants. The grasses, in particular maize, provide the single major source of agricultural biomass, offering significant opportunities for increasing renewable fuel production. Our laboratory has pioneered the use of transcription factors for manipulating plant metabolic pathways, an approach that will be applied here towards altering the composition of phenolic compounds in maize. Previously, we identified a small group of ten maize R2R3-MYB transcription factors with all the characteristics of regulators of different aspects of phenolic biosynthesis. Here, we propose to investigate the participation of these R2R3-MYB factors in the regulation of soluble and insoluble maize phenolics, using a combination of over-expression and down-regulation of these transcription factors in transgenic maize cultured cells and in maize plants. Maize cells and plants altered in the activity of these regulatory proteins will be analyzed for phenolic composition by targeted metabolic profiling. Specifically, we will I) Investigate the effect of gain- and loss-of-function of a select group of R2R3-MYB transcription factors on the phenolic composition of maize plants and II) Identify the biosynthetic genes regulated by each of the selected R2R3-MYB factors. While a likely outcome of these studies are transgenic maize plants with altered phenolic composition, this research will significantly

  18. Structural mechanisms of plant glucan phosphatases in starch metabolism.

    Science.gov (United States)

    Meekins, David A; Vander Kooi, Craig W; Gentry, Matthew S

    2016-07-01

    Glucan phosphatases are a recently discovered class of enzymes that dephosphorylate starch and glycogen, thereby regulating energy metabolism. Plant genomes encode two glucan phosphatases, called Starch EXcess4 (SEX4) and Like Sex Four2 (LSF2), that regulate starch metabolism by selectively dephosphorylating glucose moieties within starch glucan chains. Recently, the structures of both SEX4 and LSF2 were determined, with and without phosphoglucan products bound, revealing the mechanism for their unique activities. This review explores the structural and enzymatic features of the plant glucan phosphatases, and outlines how they are uniquely adapted to perform their cellular functions. We outline the physical mechanisms used by SEX4 and LSF2 to interact with starch glucans: SEX4 binds glucan chains via a continuous glucan-binding platform comprising its dual-specificity phosphatase domain and carbohydrate-binding module, while LSF2 utilizes surface binding sites. SEX4 and LSF2 both contain a unique network of aromatic residues in their catalytic dual-specificity phosphatase domains that serve as glucan engagement platforms and are unique to the glucan phosphatases. We also discuss the phosphoglucan substrate specificities inherent to SEX4 and LSF2, and outline structural features within the active site that govern glucan orientation. This review defines the structural mechanism of the plant glucan phosphatases with respect to phosphatases, starch metabolism and protein-glucan interaction, thereby providing a framework for their application in both agricultural and industrial settings.

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

    Energy Technology Data Exchange (ETDEWEB)

    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)

  20. Systematic Comparison of C3 and C4 Plants Based on Metabolic Network Analysis

    Directory of Open Access Journals (Sweden)

    Wang Chuanli

    2012-12-01

    Full Text Available Abstract Background The C4 photosynthetic cycle supercharges photosynthesis by concentrating CO2 around ribulose-1,5-bisphosphate carboxylase and significantly reduces the oxygenation reaction. Therefore engineering C4 feature into C3 plants has been suggested as a feasible way to increase photosynthesis and yield of C3 plants, such as rice, wheat, and potato. To identify the possible transition from C3 to C4 plants, the systematic comparison of C3 and C4 metabolism is necessary. Results We compared C3 and C4 metabolic networks using the improved constraint-based models for Arabidopsis and maize. By graph theory, we found the C3 network exhibit more dense topology structure than C4. The simulation of enzyme knockouts demonstrated that both C3 and C4 networks are very robust, especially when optimizing CO2 fixation. Moreover, C4 plant has better robustness no matter the objective function is biomass synthesis or CO2 fixation. In addition, all the essential reactions in C3 network are also essential for C4, while there are some other reactions specifically essential for C4, which validated that the basic metabolism of C4 plant is similar to C3, but C4 is more complex. We also identified more correlated reaction sets in C4, and demonstrated C4 plants have better modularity with complex mechanism coordinates the reactions and pathways than that of C3 plants. We also found the increase of both biomass production and CO2 fixation with light intensity and CO2 concentration in C4 is faster than that in C3, which reflected more efficient use of light and CO2 in C4 plant. Finally, we explored the contribution of different C4 subtypes to biomass production by setting specific constraints. Conclusions All results are consistent with the actual situation, which indicate that Flux Balance Analysis is a powerful method to study plant metabolism at systems level. We demonstrated that in contrast to C3, C4 plants have more dense topology, higher robustness

  1. The use of metabolic engineering to produce fatty acid-derived biofuel and chemicals in Saccharomyces cerevisiae: a review

    Directory of Open Access Journals (Sweden)

    Liwei Chen

    2016-11-01

    Full Text Available Production of fatty acid-derived biofuels and chemicals have garnered attention in recent years owing to their potential to replace petroleum and plant oil-derived products. Through the metabolic engineering of the fatty acid metabolism pathway, advanced fuels and chemicals such as free fatty acid, triacylglycerol, biodiesel, fatty alcohols, alkanes/alkene, R-3-hydroxybutyric acid, polyhydroxyalkanoates and flavonoids have been produced. The robustness, high tolerance to organic solvent, good reputation in industrial fermentations and excellent availability of genetic tools make the yeast Saccharomyces cerevisiae a suitable cell factory for fatty acid-derived biofuels and chemicals production. This review will describe the successful metabolic engineering strategies employed to produce the fatty acid-derived bio-products in S. cerevisiae, including the enhancement of precursors and co-factors supply, promotion of the enzyme expression and activity, elimination of competing pathways, and the improvement of strain tolerance.

  2. LAILAPS: The Plant Science Search Engine

    OpenAIRE

    2014-01-01

    With the number of sequenced plant genomes growing, the number of predicted genes and functional annotations is also increasing. The association between genes and phenotypic traits is currently of great interest. Unfortunately, the information available today is widely scattered over a number of different databases. Information retrieval (IR) has become an all-encompassing bioinformatics methodology for extracting knowledge from complex, heterogeneous and distributed databases, and therefore ...

  3. Biobased organic acids production by metabolically engineered microorganisms

    DEFF Research Database (Denmark)

    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 ex...... performance microbes for production of succinic acid and 3-hydroxypropionic acid. Also, the key limitations and challenges in microbial organic acids production are discussed......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...

  4. Metabolic Engineering for Expanding the Substrate Range of Yarrowia lipolytica.

    Science.gov (United States)

    Ledesma-Amaro, Rodrigo; Nicaud, Jean-Marc

    2016-10-01

    Economically viable biotechnology processes must be characterized by a favorable ratio between the production costs and the product market price. In the bioproduction of bulk chemicals, costs must be minimized so that the process is competitive relative to petroleum-based production. The substrate costs must thus be reduced by employing inexpensive carbon sources, such as industrial wastes. Unfortunately, the most convenient microorganisms for a bioconversion are typically unable to degrade such substrates. Fortunately, the discovery of new enzymes together with advances in synthetic biology has moved metabolic engineering forward, expanding substrate ranges. Here we review the latest advances made using the industrial yeast Yarrowia lipolytica, which can exploit various carbon sources to produce biofuels and chemicals.

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

    DEFF Research Database (Denmark)

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

    2005-01-01

    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......, 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...

  6. Metabolic engineering to improve ethanol production in Thermoanaerobacter mathranii

    DEFF Research Database (Denmark)

    Yao, Shuo; Mikkelsen, Marie Just

    2010-01-01

    Thermoanaerobacter mathranii can produce ethanol from lignocellulosic biomass at high temperatures, but its biotechnological exploitation will require metabolic engineering to increase its ethanol yield. With a cofactor-dependent ethanol production pathway in T. mathranii, 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 yield beyond that obtained with glucose and xylose. The ldh gene coding for lactate dehydrogenase was previously deleted from T. mathranii to eliminate...... an NADH oxidation pathway. To further facilitate NADH regeneration used for ethanol formation, a heterologous gene gldA encoding an NAD+-dependent glycerol dehydrogenase was expressed in T. mathranii. One of the resulting recombinant strains, T. mathranii BG1G1 (Δldh, P xyl GldA), showed increased ethanol...

  7. Metabolic engineering of Saccharomyces cerevisiae for production of butanol isomers.

    Science.gov (United States)

    Generoso, Wesley Cardoso; Schadeweg, Virginia; Oreb, Mislav; Boles, Eckhard

    2015-06-01

    Saccharomyces cerevisiae has decisive advantages in industrial processes due to its tolerance to alcohols and fermentation conditions. Butanol isomers are considered as suitable fuel substitutes and valuable biomass-derived chemical building blocks. Whereas high production was achieved with bacterial systems, metabolic engineering of yeast for butanol production is in the beginning. For isobutanol synthesis, combination of valine biosynthesis and degradation, and complete pathway re-localisation into cytosol or mitochondria gave promising results. However, competing pathways, co-factor imbalances and FeS cluster assembly are still major issues. 1-Butanol production via the Clostridium pathway seems to be limited by cytosolic acetyl-CoA, its central precursor. Endogenous 1-butanol pathways have been discovered via threonine or glycine catabolism. 2-Butanol production was established but was limited by B12-dependence.

  8. Metabolic engineering strategies for improving xylitol production from hemicellulosic sugars.

    Science.gov (United States)

    Su, Buli; Wu, Mianbin; Lin, Jianping; Yang, Lirong

    2013-11-01

    Xylitol is a five-carbon sugar alcohol with potential for use as a sweetener. Industrially, xylitol is currently produced by chemical hydrogenation of D-xylose using Raney nickel catalysts and this requires expensive separation and purification steps as well as high pressure and temperature that lead to environmental pollution. Highly efficient biotechnological production of xylitol using microorganisms is gaining more attention and has been proposed as an alternative process. Although the biotechnological method has not yet surpassed the advantages of chemical reduction in terms of yield and cost, various strategies offer promise for the biotechnological production of xylitol. In this review, the focus is on the most recent developments of the main metabolic engineering strategies for improving the production of xylitol.

  9. Engineering microbes for plant polyketide biosynthesis.

    Science.gov (United States)

    Lussier, François-Xavier; Colatriano, David; Wiltshire, Zach; Page, Jonathan E; Martin, Vincent J J

    2012-01-01

    Polyketides are an important group of secondary metabolites, many of which have important industrial applications in the food and pharmaceutical industries. Polyketides are synthesized from one of three classes of enzymes differentiated by their biochemical features and product structure: type I, type II or type III polyketide synthases (PKSs). Plant type III PKS enzymes, which will be the main focus of this review, are relatively small homodimeric proteins that catalyze iterative decarboxylative condensations of malonyl units with a CoA-linked starter molecule. This review will describe the plant type III polyketide synthetic pathway, including the synthesis of chalcones, stilbenes and curcuminoids, as well as recent work on the synthesis of these polyketides in heterologous organisms. The limitations and bottlenecks of heterologous expression as well as attempts at creating diversity through the synthesis of novel "unnatural" polyketides using type III PKSs will also be discussed. Although synthetic production of plant polyketides is still in its infancy, their potential as useful bioactive compounds makes them an extremely interesting area of study.

  10. ENGINEERING MICROBES FOR PLANT POLYKETIDE BIOSYNTHESIS

    Directory of Open Access Journals (Sweden)

    François-Xavier Lussier

    2012-10-01

    Full Text Available Polyketides are an important group of secondary metabolites, many of which have important industrial applications in the food and pharmaceutical industries. Polyketides are synthesized from one of three classes of enzymes differentiated by their biochemical features and product structure: type I, type II or type III polyketide synthases (PKSs. Plant type III PKS enzymes, which will be the main focus of this review, are relatively small homodimeric proteins that catalyze iterative decarboxylative condensations of malonyl units with a CoA-linked starter molecule. This review will describe the plant type III polyketide synthetic pathway, including the synthesis of chalcones, stilbenes and curcuminoids, as well as recent work on the synthesis of these polyketides in heterologous organisms. The limitations and bottlenecks of heterologous expression as well as attempts at creating diversity through the synthesis of novel “unnatural” polyketides using type III PKSs will also be discussed. Although synthetic production of plant polyketides is still in its infancy, their potential as useful bioactive compounds makes them an extremely interesting area of study.

  11. Engineering Microbes for Plant Polyketide Biosynthesis

    Directory of Open Access Journals (Sweden)

    Vincent J. J. Martin

    2012-10-01

    Full Text Available Polyketides are an important group of secondary metabolites, many of which have important industrial applications, including in the food and pharmaceutical industries. Polyketides are synthesized from one of three classes of enzymes differentiated by their biochemical features and product structure: type I, type II or type III polyketide synthases (PKSs. Plant type III PKS enzymes, which will be the main focus of this review, are relatively small homodimeric proteins that catalyze iterative decarboxylative condensations of malonyl units with a CoA-linked starter molecule. This review will describe the plant type III polyketide synthetic pathway, including the synthesis of chalcones, stilbenes and curcuminoids, as well as recent work on the synthesis of these polyketides in heterologous organisms. The limitations and bottlenecks of heterologous expression as well as attempts at creating diversity through the synthesis of novel “unnatural” polyketides using type III PKSs will also be discussed. Although synthetic production of plant polyketides is still in its infancy, their potential as useful bioactive compounds makes them an extremely interesting area of study.

  12. Genetic elements of plant viruses as tools for genetic engineering.

    OpenAIRE

    Mushegian, A R; Shepherd, R J

    1995-01-01

    Viruses have developed successful strategies for propagation at the expense of their host cells. Efficient gene expression, genome multiplication, and invasion of the host are enabled by virus-encoded genetic elements, many of which are well characterized. Sequences derived from plant DNA and RNA viruses can be used to control expression of other genes in vivo. The main groups of plant virus genetic elements useful in genetic engineering are reviewed, including the signals for DNA-dependent a...

  13. Plant interactions alter the predictions of metabolic scaling theory.

    Directory of Open Access Journals (Sweden)

    Yue Lin

    Full Text Available Metabolic scaling theory (MST is an attempt to link physiological processes of individual organisms with macroecology. It predicts a power law relationship with an exponent of -4/3 between mean individual biomass and density during density-dependent mortality (self-thinning. Empirical tests have produced variable results, and the validity of MST is intensely debated. MST focuses on organisms' internal physiological mechanisms but we hypothesize that ecological interactions can be more important in determining plant mass-density relationships induced by density. We employ an individual-based model of plant stand development that includes three elements: a model of individual plant growth based on MST, different modes of local competition (size-symmetric vs. -asymmetric, and different resource levels. Our model is consistent with the observed variation in the slopes of self-thinning trajectories. Slopes were significantly shallower than -4/3 if competition was size-symmetric. We conclude that when the size of survivors is influenced by strong ecological interactions, these can override predictions of MST, whereas when surviving plants are less affected by interactions, individual-level metabolic processes can scale up to the population level. MST, like thermodynamics or biomechanics, sets limits within which organisms can live and function, but there may be stronger limits determined by ecological interactions. In such cases MST will not be predictive.

  14. Transcription factors as tools to engineer enhanced drought stress tolerance in plants.

    Science.gov (United States)

    Hussain, Syed Sarfraz; Kayani, Mahmood Akhtar; Amjad, Muhammad

    2011-01-01

    Plant growth and productivity are greatly affected by abiotic stresses such as drought, salinity, and temperature. Drought stress is one of the major limitations to crop productivity worldwide due to its multigene nature, making the production of transgenic crops a challenging prospect. To develop crop plant with enhanced tolerance of drought stress, a basic understanding of physiological, biochemical, and gene regulatory networks is essential. In the signal transduction network that leads from the perception of stress signals to the expression of stress-responsive genes, transcription factors (TFs) play an essential role. Because TFs, as opposed to most structural genes, tend to control multiple pathways steps, they have emerged as powerful tools for the manipulation of complex metabolic pathways in plants. One such class of TFs is DREB/CBF that binds to drought responsive cis-acting elements. Transgenic plants have been developed with enhanced stress tolerance by manipulating the expression of DREB/CBF. Recently the functions of an increasing number of plant TFs are being elucidated and increased understanding of these factors in controlling drought stress response has lead to practical approaches for engineering stress tolerance in plants. The utility of the various TFs in plant stress research we review is illustrated by several published examples. The manipulation of native plant regularity networks therefore represents a new era for genetically modified crops. This review focuses on the recent understanding, latest advancements related to TFs and present status of their deployment in developing stress tolerant transgenic plants.

  15. Plant interactions alter the predictions of metabolic scaling theory

    DEFF Research Database (Denmark)

    Lin, Yue; Berger, Uta; Grimm, Volker;

    2013-01-01

    Metabolic scaling theory (MST) is an attempt to link physiological processes of individual organisms with macroecology. It predicts a power law relationship with an exponent of 24/3 between mean individual biomass and density during densitydependent mortality (self-thinning). Empirical tests have...... produced variable results, and the validity of MST is intensely debated. MST focuses on organisms’ internal physiological mechanisms but we hypothesize that ecological interactions can be more important in determining plant mass-density relationships induced by density. We employ an individual-based model...... of plant stand development that includes three elements: a model of individual plant growth based on MST, different modes of local competition (size-symmetric vs. -asymmetric), and different resource levels. Our model is consistent with the observed variation in the slopes of self-thinning trajectories...

  16. Plant-microbe interactions to probe regulation of plant carbon metabolism.

    Science.gov (United States)

    Biemelt, Sophia; Sonnewald, Uwe

    2006-02-01

    Plant growth and development is dependent on coordinated assimilate production, distribution and allocation. Application of biochemical and molecular techniques substantially contributed to a better understanding of these processes, although the underlying regulatory mechanisms are still not fully elucidated and attempts to improve crop yield by modulating carbon partitioning were only partially successful. Plant pathogens also interfere with source-sink interaction. To this end they have evolved a wide range of sophisticated strategies to allow their systemic spread, suppression of plant defence and induction of sink function to support nutrient acquisition for their growth. Studying compatible interactions of plants and pathogens like viruses, bacteria and fungi can be exploited to investigate different levels of source-sink regulation. The identification of microbial factors and their host targets involved in regulation of plant primary metabolism may allow developing novel strategies to increase crop yield. Here we will discuss recent studies on plant-microbe interactions aimed at elucidating mechanisms of compatibility.

  17. Metabolic engineering for improved production of ethanol by Corynebacterium glutamicum.

    Science.gov (United States)

    Jojima, Toru; Noburyu, Ryoji; Sasaki, Miho; Tajima, Takahisa; Suda, Masako; Yukawa, Hideaki; Inui, Masayuki

    2015-02-01

    Recombinant Corynebacterium glutamicum harboring genes for pyruvate decarboxylase (pdc) and alcohol dehydrogenase (adhB) can produce ethanol under oxygen deprivation. We investigated the effects of elevating the expression levels of glycolytic genes, as well as pdc and adhB, on ethanol production. Overexpression of four glycolytic genes (pgi, pfkA, gapA, and pyk) in C. glutamicum significantly increased the rate of ethanol production. Overexpression of tpi, encoding triosephosphate isomerase, further enhanced productivity. Elevated expression of pdc and adhB increased ethanol yield, but not the rate of production. Fed-batch fermentation using an optimized strain resulted in ethanol production of 119 g/L from 245 g/L glucose with a yield of 95% of the theoretical maximum. Further metabolic engineering, including integration of the genes for xylose and arabinose metabolism, enabled consumption of glucose, xylose, and arabinose, and ethanol production (83 g/L) at a yield of 90 %. This study demonstrated that C. glutamicum has significant potential for the production of cellulosic ethanol.

  18. Metabolization of the bacteriostatic agent triclosan in edible plants and its consequences for plant uptake assessment.

    Science.gov (United States)

    Macherius, André; Eggen, Trine; Lorenz, Wilhelm; Moeder, Monika; Ondruschka, Jelka; Reemtsma, Thorsten

    2012-10-02

    Persistent environmental contaminants may enter agricultural fields via the application of sewage sludge, by irrigation with treated municipal wastewater or by manuring. It has been shown that such contaminants can be incorporated into crop plants. The metabolism of the bacteriostatic agents triclocarban, triclosan, and its transformation product methyl triclosan was investigated after their uptake into carrot cell cultures. A fast metabolization of triclosan was observed and eight so far unknown phase II metabolites, conjugates with saccharides, disaccharides, malonic acid, and sulfate, were identified by liquid chromatography-mass spectrometry. Triclocarban and methyl triclosan lack a phenolic group and remained unaltered in the cell cultures. Phase I metabolization was not observed for any of the compounds. All eight triclosan conjugates identified in the cell cultures were also detected in extracts of intact carrot plants cultivated on triclosan contaminated soils. Their total amount in the plants was assessed to exceed the amount of the triclosan itself by a factor of 5. This study shows that a disregard of conjugates in studies on plant uptake of environmental contaminants may severely underestimates the extent of uptake into plants and, eventually, the potential human exposure to contaminants via food of plant origin.

  19. Metabolic engineering of tomato fruit organic acid content guided by biochemical analysis of an introgression line.

    Science.gov (United States)

    Morgan, Megan J; Osorio, Sonia; Gehl, Bernadette; Baxter, Charles J; Kruger, Nicholas J; Ratcliffe, R George; Fernie, Alisdair R; Sweetlove, Lee J

    2013-01-01

    Organic acid content is regarded as one of the most important quality traits of fresh tomato (Solanum lycopersicum). However, the complexity of carboxylic acid metabolism and storage means that it is difficult to predict the best way to engineer altered carboxylic acid levels. Here, we used a biochemical analysis of a tomato introgression line with increased levels of fruit citrate and malate at breaker stage to identify a metabolic engineering target that was subsequently tested in transgenic plants. Increased carboxylic acid levels in introgression line 2-5 were not accompanied by changes in the pattern of carbohydrate oxidation by pericarp discs or the catalytic capacity of tricarboxylic acid cycle enzymes measured in isolated mitochondria. However, there was a significant decrease in the maximum catalytic activity of aconitase in total tissue extracts, suggesting that a cytosolic isoform of aconitase was affected. To test the role of cytosolic aconitase in controlling fruit citrate levels, we analyzed fruit of transgenic lines expressing an antisense construct against SlAco3b, one of the two tomato genes encoding aconitase. A green fluorescent protein fusion of SlAco3b was dual targeted to cytosol and mitochondria, while the other aconitase, SlAco3a, was exclusively mitochondrial when transiently expressed in tobacco (Nicotiana tabacum) leaves. Both aconitase transcripts were decreased in fruit from transgenic lines, and aconitase activity was reduced by about 30% in the transgenic lines. Other measured enzymes of carboxylic acid metabolism were not significantly altered. Both citrate and malate levels were increased in ripe fruit of the transgenic plants, and as a consequence, total carboxylic acid content was increased by 50% at maturity.

  20. Membrane engineering of S. cerevisiae targeting sphingolipid metabolism.

    Science.gov (United States)

    Lindahl, Lina; Santos, Aline X S; Olsson, Helén; Olsson, Lisbeth; Bettiga, Maurizio

    2017-02-01

    The sustainable production of fuels and chemicals using microbial cell factories is now well established. However, many microbial production processes are still limited in scale due to inhibition from compounds that are present in the feedstock or are produced during fermentation. Some of these inhibitors interfere with cellular membranes and change the physicochemical properties of the membranes. Another group of molecules is dependent on their permeation rate through the membrane for their inhibition. We have investigated the use of membrane engineering to counteract the negative effects of inhibitors on the microorganism with focus on modulating the abundance of complex sphingolipids in the cell membrane of Saccharomyces cerevisiae. Overexpression of ELO3, involved in fatty acid elongation, and AUR1, which catalyses the formation of complex sphingolipids, had no effect on the membrane lipid profile or on cellular physiology. Deletion of the genes ORM1 and ORM2, encoding negative regulators of sphingolipid biosynthesis, decreased cell viability and considerably reduced phosphatidylinositol and complex sphingolipids. Additionally, combining ELO3 and AUR1 overexpression with orm1/2Δ improved cell viability and increased fatty acyl chain length compared with only orm1/2Δ. These findings can be used to further study the sphingolipid metabolism, as well as giving guidance in membrane engineering.

  1. Polyphenols: planting the seeds of treatment for the metabolic syndrome.

    Science.gov (United States)

    Cherniack, E Paul

    2011-06-01

    Greater understanding about the pathogenesis of metabolic syndrome and potential causes suggests that plant polyphenols might be useful as a treatment. Dietary excess energy can be stored in adipocytes, leading to the release of proinflammatory cytokines and adipose-related hormones that cause vascular injury. Plant polyphenols, organic compounds found in numerous plant species and their fruits, are being actively studied as potential treatments for components of the metabolic syndrome. Individual polyphenols that have been examined include resveratrol, quercetin, epigallocathechin-3-gallate, and curcumin. Resveratrol lowers weight, blood pressure, glucose, and insulin resistance in rodents, and a human trial is currently underway. Quercetin decreases lipid and glucose levels in obese rats, and in a human investigation of subjects with the metabolic syndrome has lowered blood pressure without significant alteration of lipids. Epigallocathechin-3-gallate-induced weight loss has attenuated glucose levels and insulin resistance in rodents and improved hemoglobin A(1c) and lipid in human studies. Plant extracts also can be used. Grape seed and chokeberry extracts have decreased blood pressure and lipid levels in small human trials. Other human investigations have shown the beneficial effects of cocoa, coffee, carob, and Momordica charantia. Thus far, most studies have involved a small number of subjects and have been of short duration. Future studies should be designed to account for a disease process in which the pathogenic factors may take place for years before disease manifestations take place, the possibly limited bioavailability of polyphenols, and the potential need to provide combinations or modifications of polyphenols. Published by Elsevier Inc.

  2. Higher plant metabolism and energetics in hypogravity: Amino acid metabolism in higher plants

    Science.gov (United States)

    Mazelis, M.

    1976-01-01

    Laboratory's investigation into the amino acid metabolism of dwarf marigolds exposed to an environment of simulated hypogravity is summarized. Using both in vivo, and/or in vitro studies, the following effects of hypogravitational stress have been shown: (1) increased proline incorporation into cell wall protein, (2) inhibition of amino acid decarboxylation, (3) decrease in glutamic acid decarboxylase activity; and (4) decrease in the relative amount of a number of soluble amino acids present in deproteinized extracts of marigold leaves. It is concluded from these data there are several rapid, major alterations in amino acid metabolism associated with hypogravitational stress in marigolds. The mechanism(s) and generality of these effects with regard to other species is still unknown.

  3. Overcoming the plasticity of plant specialized metabolism for selective diterpene production in yeast.

    Science.gov (United States)

    Ignea, Codruta; Athanasakoglou, Anastasia; Andreadelli, Aggeliki; Apostolaki, Maria; Iakovides, Minas; Stephanou, Euripides G; Makris, Antonios M; Kampranis, Sotirios C

    2017-08-18

    Plants synthesize numerous specialized metabolites (also termed natural products) to mediate dynamic interactions with their surroundings. The complexity of plant specialized metabolism is the result of an inherent biosynthetic plasticity rooted in the substrate and product promiscuity of the enzymes involved. The pathway of carnosic acid-related diterpenes in rosemary and sage involves promiscuous cytochrome P450s whose combined activity results in a multitude of structurally related compounds. Some of these minor products, such as pisiferic acid and salviol, have established bioactivity, but their limited availability prevents further evaluation. Reconstructing carnosic acid biosynthesis in yeast achieved significant titers of the main compound but could not specifically yield the minor products. Specific production of pisiferic acid and salviol was achieved by restricting the promiscuity of a key enzyme, CYP76AH24, through a single-residue substitution (F112L). Coupled with additional metabolic engineering interventions, overall improvements of 24 and 14-fold for pisiferic acid and salviol, respectively, were obtained. These results provide an example of how synthetic biology can help navigating the complex landscape of plant natural product biosynthesis to achieve heterologous production of useful minor metabolites. In the context of plant adaptation, these findings also suggest a molecular basis for the rapid evolution of terpene biosynthetic pathways.

  4. Saab claims world's most modern engine-assembly plant

    Science.gov (United States)

    Bergmark, U.

    1984-01-01

    The modernization of an engine assembly plant using computer aided manufacturing techniques and robotics is described. The development of associated tools and production procedures used to assemble the Light Component Project (LCP) modular automobile are discussd. Structural design criteria used to reduce the weight and increase the fuel efficiency of the vehicle are explained.

  5. Pyridine metabolism in tea plants: salvage, conjugate formation and catabolism.

    Science.gov (United States)

    Ashihara, Hiroshi; Deng, Wei-Wei

    2012-11-01

    Pyridine compounds, including nicotinic acid and nicotinamide, are key metabolites of both the salvage pathway for NAD and the biosynthesis of related secondary compounds. We examined the in situ metabolic fate of [carbonyl-(14)C]nicotinamide, [2-(14)C]nicotinic acid and [carboxyl-(14)C]nicotinic acid riboside in tissue segments of tea (Camellia sinensis) plants, and determined the activity of enzymes involved in pyridine metabolism in protein extracts from young tea leaves. Exogenously supplied (14)C-labelled nicotinamide was readily converted to nicotinic acid, and some nicotinic acid was salvaged to nicotinic acid mononucleotide and then utilized for the synthesis of NAD and NADP. The nicotinic acid riboside salvage pathway discovered recently in mungbean cotyledons is also operative in tea leaves. Nicotinic acid was converted to nicotinic acid N-glucoside, but not to trigonelline (N-methylnicotinic acid), in any part of tea seedlings. Active catabolism of nicotinic acid was observed in tea leaves. The fate of [2-(14)C]nicotinic acid indicates that glutaric acid is a major catabolite of nicotinic acid; it was further metabolised, and carbon atoms were finally released as CO(2). The catabolic pathway observed in tea leaves appears to start with the nicotinic acid N-glucoside formation; this pathway differs from catabolic pathways observed in microorganisms. Profiles of pyridine metabolism in tea plants are discussed.

  6. Does iodine biofortification affect oxidative metabolism in lettuce plants?

    Science.gov (United States)

    Blasco, Begoña; Ríos, Juan Jose; Leyva, Rocío; Cervilla, Luis Miguel; Sánchez-Rodríguez, Eva; Rubio-Wilhelmi, María Mar; Rosales, Miguel Angel; Ruiz, Juan Manuel; Romero, Luis

    2011-09-01

    Plants produce low levels of reactive oxygen species (ROS), which form part of basic cell chemical communication; however, different types of stress can lead to an overexpression of ROS that can damage macromolecules essential for plant growth and development. Iodine is vital to human health, and iodine biofortification programs help improve the human intake through plant consumption. This biofortification process has been shown to influence the antioxidant capacity of lettuce plants, suggesting that the oxidative metabolism of the plant may be affected. The results of this study demonstrate that the response to oxidative stress is variable and depends on the form of iodine applied. Application of iodide (I(-)) to lettuce plants produces a reduction in superoxide dismutase (SOD) activity and an increase in catalase (CAT) and L-galactono dehydrogenase enzyme activities and in the activity of antioxidant compounds such as ascorbate (AA) and glutathione. This did not prove a very effective approach since a dose of 80 μM produced a reduction in the biomass of the plants. For its part, application of iodate (IO (3) (-) ) produced an increase in the activities of SOD, ascorbate peroxidase, and CAT, the main enzymes involved in ROS detoxification; it also increased the concentration of AA and the regenerative activities of the Halliwell-Asada cycle. These data confirm the non-phytotoxicity of IO (3) (-) since there is no lipid peroxidation or biomass reduction. According to our results, the ability of IO (3) (-) to induce the antioxidant system indicates that application of this form of iodine may be an effective strategy to improve the response of plants to different types of stress.

  7. Systems metabolic engineering of microorganisms for natural and non-natural chemicals.

    Science.gov (United States)

    Lee, Jeong Wook; Na, Dokyun; Park, Jong Myoung; Lee, Joungmin; Choi, Sol; Lee, Sang Yup

    2012-05-17

    Growing concerns over limited fossil resources and associated environmental problems are motivating the development of sustainable processes for the production of chemicals, fuels and materials from renewable resources. Metabolic engineering is a key enabling technology for transforming microorganisms into efficient cell factories for these compounds. Systems metabolic engineering, which incorporates the concepts and techniques of systems biology, synthetic biology and evolutionary engineering at the systems level, offers a conceptual and technological framework to speed the creation of new metabolic enzymes and pathways or the modification of existing pathways for the optimal production of desired products. Here we discuss the general strategies of systems metabolic engineering and examples of its application and offer insights as to when and how each of the different strategies should be used. Finally, we highlight the limitations and challenges to be overcome for the systems metabolic engineering of microorganisms at more advanced levels.

  8. TALENs: Customizable Molecular DNA Scissors for Genome Engineering of Plants

    Institute of Scientific and Technical Information of China (English)

    Kunling Chen; Caixia Gao

    2013-01-01

    Precise genome modification with engineered nucleases is a powerful tool for studying basic biology and applied biotechnology.Transcription activator-like effector nucleases (TALENs),consisting of an engineered specific (TALE) DNA binding domain and a Fok I cleavage domain,are newly developed versatile reagents for genome engineering in different organisms.Because of the simplicity of the DNA recognition code and their modular assembly,TALENs can act as customizable molecular DNA scissors inducing double-strand breaks (DSBs) at given genomic location.Thus,they provide a valuable approach to targeted genome modifications such as mutations,insertions,replacements or chromosome rearrangements.In this article,we review the development of TALENs,and summarize the principles and tools for TALEN-mediated gene targeting in plant cells,as well as current and potential strategies for use in plant research and crop improvement.

  9. Modulation of Phytoalexin Biosynthesis in Engineered Plants for Disease Resistance

    Directory of Open Access Journals (Sweden)

    Sylvain Cordelier

    2013-07-01

    Full Text Available Phytoalexins are antimicrobial substances of low molecular weight produced by plants in response to infection or stress, which form part of their active defense mechanisms. Starting in the 1950’s, research on phytoalexins has begun with biochemistry and bio-organic chemistry, resulting in the determination of their structure, their biological activity as well as mechanisms of their synthesis and their catabolism by microorganisms. Elucidation of the biosynthesis of numerous phytoalexins has permitted the use of molecular biology tools for the exploration of the genes encoding enzymes of their synthesis pathways and their regulators. Genetic manipulation of phytoalexins has been investigated to increase the disease resistance of plants. The first example of a disease resistance resulting from foreign phytoalexin expression in a novel plant has concerned a phytoalexin from grapevine which was transferred to tobacco. Transformations were then operated to investigate the potential of other phytoalexin biosynthetic genes to confer resistance to pathogens. Unexpectedly, engineering phytoalexins for disease resistance in plants seem to have been limited to exploiting only a few phytoalexin biosynthetic genes, especially those encoding stilbenes and some isoflavonoids. Research has rather focused on indirect approaches which allow modulation of the accumulation of phytoalexin employing transcriptional regulators or components of upstream regulatory pathways. Genetic approaches using gain- or less-of functions in phytoalexin engineering together with modulation of phytoalexin accumulation through molecular engineering of plant hormones and defense-related marker and elicitor genes have been reviewed.

  10. Altered sucrose metabolism impacts plant biomass production and flower development.

    Science.gov (United States)

    Coleman, Heather D; Beamish, Leigh; Reid, Anya; Park, Ji-Young; Mansfield, Shawn D

    2010-04-01

    Nicotiana tabacum (tobacco) was transformed with three genes involved in sucrose metabolism, UDP-glucose pyrophosphorylase (UGPase, EC 2.7.7.9), sucrose synthase (SuSy, EC 2.4.1.13) and sucrose phosphate synthase (SPS, EC 2.4.1.14). Plants harbouring the single transgenes were subsequently crossed to produce double and triple transgenic lines, including: 2 x 35S::UGPase x SPS, 4CL::UGPase x SPS, 2 x 35S::SuSy x SPS, 4CL::SuSy x SPS, 2 x 35S::UGPase x SuSy x SPS, and 4CL::UGPase x SuSy x SPS. The ultimate aim of the study was to examine whether it is possible to alter cellulose production through the manipulation of sucrose metabolism genes. While altering sucrose metabolism using UGPase, SuSy and SPS does not have an end effect on cellulose production, their simultaneous overexpression resulted in enhanced primary growth as seen in an increase in height growth, in some cases over 50%. Furthermore, the pyramiding strategy of simultaneously altering the expression of multiple genes in combination resulted in increased time to reproductive bud formation as well as altered flower morphology and foliar stipule formation in 4CL lines. Upregulation of these sucrose metabolism genes appears to directly impact primary growth and therefore biomass production in tobacco.

  11. Advances in Research on Genetically Engineered Plants for Metal Resistance

    Institute of Scientific and Technical Information of China (English)

    Ri-Qing Zhang; Chun-Fang Tang; Shi-Zhi Wen; Yun-Guo Liu; Ke-Lin Li

    2006-01-01

    The engineering application of natural hyperaccumulators in removing or inactivating metal pollutants from soil and surface water in field trials mostly presents the insurmountable shortcoming of low efficiency owing to their little biomass and slow growth. Based on further understanding of the molecular mechanism of metal uptake, translocation, and also the separation, identification, and cloning of some related functional genes, this article highlights and summarizes in detail the advances in research on transgenic techniques, such as Agrobacterium tumefaciens-mediated transformation and particle bombardment, in breeding of plants for metal resistance and accumulation, and points out that deepening the development of transgenic plants is one of the efficient approaches to improving phytoremediation efficiency of metal-contaminated environments. From the viewpoint of sustainable development, governments should strengthen support to the development of genetic engineering for metal resistance and accumulation in plants.

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

    NARCIS (Netherlands)

    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

  13. Metabolic engineering of tomato for high-yield production of astaxanthin.

    Science.gov (United States)

    Huang, Jun-Chao; Zhong, Yu-Juan; Liu, Jin; Sandmann, Gerhard; Chen, Feng

    2013-05-01

    Dietary carotenoids have been shown to be beneficial to health by decreasing the risk of many diseases. Attempts to enhance carotenoids in food crops have been successful although higher plants appear to resist big changes of carotenoid biosynthesis by metabolic engineering. Here we report the generation of a more nutritious tomato by modifying the intrinsic carotenes to astaxanthin, a high-value ketocarotenoid rarely found in plants. This was achieved by co-expression of the algal β-carotene ketolase from Chlamydomonas reinhardtii and β-carotene hydroxylase from Haematococcus pluvialis, a unique pair of enzymes identified to co-operate perfectly in converting β-carotene to astaxanthin by functional complementation in Escherichia coli. Expression of the two enzymes in tomato up-regulated most intrinsic carotenogenic genes, and efficiently directed carbon flux into carotenoids, leading to massive accumulations of mostly free astaxanthin in leaves (3.12mg/g) but esterified astaxanthin in fruits (16.1mg/g) and a 16-fold increase of total carotenoid capacity therein without affecting the plant normal growth and development. This study opened up the possibility of employing crop plants as green factories for economical production of astaxanthin.

  14. Metabolic engineering of Pichia pastoris to produce ricinoleic acid, a hydroxy fatty acid of industrial importance.

    Science.gov (United States)

    Meesapyodsuk, Dauenpen; Chen, Yan; Ng, Siew Hon; Chen, Jianan; Qiu, Xiao

    2015-11-01

    Ricinoleic acid (12-hydroxyoctadec-cis-9-enoic acid) has many specialized uses in bioproduct industries, while castor bean is currently the only commercial source for the fatty acid. This report describes metabolic engineering of a microbial system (Pichia pastoris) to produce ricinoleic acid using a "push" (synthesis) and "pull" (assembly) strategy. CpFAH, a fatty acid hydroxylase from Claviceps purpurea, was used for synthesis of ricinoleic acid, and CpDGAT1, a diacylglycerol acyl transferase for the triacylglycerol synthesis from the same species, was used for assembly of the fatty acid. Coexpression of CpFAH and CpDGAT1 produced higher lipid contents and ricinoleic acid levels than expression of CpFAH alone. Coexpression in a mutant haploid strain defective in the Δ12 desaturase activity resulted in a higher level of ricinoleic acid than that in the diploid strain. Intriguingly, the ricinoleic acid produced was mainly distributed in the neutral lipid fractions, particularly the free fatty acid form, but with little in the polar lipids. This work demonstrates the effectiveness of the metabolic engineering strategy and excellent capacity of the microbial system for production of ricinoleic acid as an alternative to plant sources for industrial uses. Copyright © 2015 by the American Society for Biochemistry and Molecular Biology, Inc.

  15. Weedy lignocellulosic feedstock and microbial metabolic engineering: advancing the generation of 'Biofuel'.

    Science.gov (United States)

    Chandel, Anuj K; Singh, Om V

    2011-03-01

    Lignocellulosic materials are the most abundant renewable organic resources (~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.

  16. Computational Modeling of Auxin: A Foundation for Plant Engineering

    Directory of Open Access Journals (Sweden)

    Alejandro Morales-Tapia

    2016-12-01

    Full Text Available Since the development of agriculture, humans have relied on the cultivation of plants to satisfy our increasing demand for food, natural products, and other raw materials. As we understand more about plant development, we can better manipulate plants to fulfill our particular needs.Auxins are a class of simple metabolites that coordinate many developmental activities like growth and the appearance of functional structures in plants. Computational modeling of auxin has proven to be an excellent tool in elucidating many mechanisms that underlie these developmental events. Due to the complexity of these mechanisms, current modelling efforts are concerned only with single phenomena focused on narrow spatial and developmental contexts; but a general model of plant development could be assembled by integrating the insights from all of them.In this perspective, we summarize the current collection of auxin-driven computational models, focusing on how they could come together into a single model for plant development. A model of this nature would allow researchers to test hypotheses in silico and yield accurate predictions about the behavior of a plant under a given set of physical and biochemical constraints. It would also provide a solid foundation towards the establishment of plant engineering, a proposed discipline intended to enable the design and production of plants that exhibit an arbitrarily defined set of features.

  17. Different Narrow-Band Light Ranges Alter Plant Secondary Metabolism and Plant Defense Response to Aphids.

    Science.gov (United States)

    Rechner, Ole; Neugart, Susanne; Schreiner, Monika; Wu, Sasa; Poehling, Hans-Michael

    2016-10-01

    Light of different wavelengths affects various physiological processes in plants. Short-wavelength radiation (like UV) can activate defense pathways in plants and enhance the biosynthesis of secondary metabolites (such as flavonoids and glucosinolates) responsible for resistance against certain herbivorous insects. The intensity of light-induced, metabolite-based resistance is plant- and insect species-specific and depends on herbivore feeding guild and specialization. In this study, broccoli (Brassica oleracea var. italica) plants were grown for 4 weeks in a climate chamber under conventional fluorescent tubes and were additionally treated with UV-B (310 nm), UV-A (365 or 385 nm), or violet (420 nm) light generated with UV-B tubes or light-emitting diodes (LEDs). The objective was to determine the influence of narrow bandwidths of light (from UV-B to violet) on plant secondary metabolism and on the performance of the cabbage aphid Brevicoryne brassicae (a specialist) and the green peach aphid Myzus persicae (a generalist). Among flavonol glycosides, specific quercetin and kaempferol glycosides increased markedly under UV-B, while among glucosinolates only 4-methoxy-3-indolylmethyl showed a 2-fold increase in plants exposed to UV-B and UV-A. The concentration of 3-indolylmethyl glucosinolate in broccoli plants increased with UV-B treatment. Brevicoryne brassicae adult weights and fecundity were lower on UV-B treated plants compared to UV-A or violet light-treated plants. Adult weights and fecundity of M. persicae were increased under UV-B and UV-A treatments. When specific light wavelengths are used to induce metabolic changes in plants, the specificity of the induced effects on herbivores should be considered.

  18. Roles for glutathione transferases in plant secondary metabolism.

    Science.gov (United States)

    Dixon, David P; Skipsey, Mark; Edwards, Robert

    2010-03-01

    Plant glutathione transferases (GSTs) are classified as enzymes of secondary metabolism, but while their roles in catalysing the conjugation and detoxification of herbicides are well known, their endogenous functions are largely obscure. Thus, while the presence of GST-derived S-glutathionylated xenobiotics have been described in many plants, there is little direct evidence for the accumulation of similarly conjugated natural products, despite the presence of a complex and dichotomous metabolic pathway which processes these reaction products. The conservation in glutathione conjugating and processing pathways, the co-regulation of GSTs with inducible plant secondary metabolism and biochemical studies showing the potential of these enzymes to conjugate reactive natural products are all suggestive of important endogenous functions. As a framework for addressing these enigmatic functions we postulate that either: (a) the natural reaction products of GSTs are unstable and undergo reversible S-glutathionylation; (b) the conjugation products of GSTs are very rapidly processed to derived metabolites; (c) GSTs do not catalyse conventional conjugation reactions but instead use glutathione as a cofactor rather than co-substrate; or (d) GSTs are non-catalytic and function as transporter proteins for secondary metabolites and their unstable intermediates. In this review, we describe how enzyme biochemistry and informatics are providing clues as to GST function allowing for the critical evaluation of each of these hypotheses. We also present evidence for the involvement of GSTs in the synthesis of sulfur-containing secondary metabolites such as volatiles and glucosinolates, and the conjugation, transport and storage of reactive oxylipins, phenolics and flavonoids.

  19. Understanding regulatory networks and engineering for enhanced drought tolerance in plants.

    Science.gov (United States)

    Valliyodan, Babu; Nguyen, Henry T

    2006-04-01

    Drought stress is one of the major limitations to crop productivity. To develop crop plants with enhanced tolerance of drought stress, a basic understanding of physiological, biochemical and gene regulatory networks is essential. Various functional genomics tools have helped to advance our understanding of stress signal perception and transduction, and of the associated molecular regulatory network. These tools have revealed several stress-inducible genes and various transcription factors that regulate the drought-stress-inducible systems. Translational genomics of these candidate genes using model plants provided encouraging results, but the field testing of transgenic crop plants for better performance and yield is still minimal. Better understanding of the specific roles of various metabolites in crop stress tolerance will give rise to a strategy for the metabolic engineering of crop tolerance of drought.

  20. Butinge Wind Power Plant - Pre-engineering

    Energy Technology Data Exchange (ETDEWEB)

    Mollestad, Knut A.

    1997-03-01

    A preliminary study has been done for a wind farm at Butinge in Lithuania consisting of six 600 kW wind turbines. The location of the wind farm is planned near the border with Latvia and near the coast of the Baltic Sea. The preliminary study has through a relatively detailed technical examination of the project, primarily aimed at gathering basic data for budget estimation and cost calculations. Other relevant aspects of the project are also considered. These include an examination of the basis for the project, an analysis of wind conditions, consideration of the various environmental conditions, discussion about the form of organisation and division of responsibility, as well as an examination of the operational and maintenance framework. Based on wind measurements taken at IFE (Institute for Energy Technology), the annual mean wind is estimated to be 7.3 m/s at 45 m height (equivalent to the hub height for the proposed wind turbines). A production calculation has also been done, which shows that the total energy production of the entire wind farm is expected to 9.47 GWh per year. This is equivalent to a use time of 2630 hours respective to a load factor of 30%. The total cost is estimated at 30.2 mill NOK, and the energy price (including operational costs) delivered from the ''power plant wall'', based on this figure and the production estimate, is calculated at 0,32 NOK/kWh. This is based on a lifetime of 20 years for the facility and a 6.5% required rate of return. In addition, there is outlined a possible financial solution based on various price-scenarios and support arrangements. With an equity capital of 8% and 28% external support, the project meets the current energy price in Lithuania (3,4 US cents/kWh). Without support the sales price of the produced energy is increased by approx. 50%. (author)

  1. Analysis of requirements for teaching materials based on the course bioinformatics for plant metabolism

    Science.gov (United States)

    Balqis, Widodo, Lukiati, Betty; Amin, Mohamad

    2017-05-01

    A way to improve the quality of learning in the course of Plant Metabolism in the Department of Biology, State University of Malang, is to develop teaching materials. This research evaluates the needs of bioinformatics-based teaching material in the course Plant Metabolism by the Analyze, Design, Develop, Implement, and Evaluate (ADDIE) development model. Data were collected through questionnaires distributed to the students in the Plant Metabolism course of the Department of Biology, University of Malang, and analysis of the plan of lectures semester (RPS). Learning gains of this course show that it is not yet integrated into the field of bioinformatics. All respondents stated that plant metabolism books do not include bioinformatics and fail to explain the metabolism of a chemical compound of a local plant in Indonesia. Respondents thought that bioinformatics can explain examples and metabolism of a secondary metabolite analysis techniques and discuss potential medicinal compounds from local plants. As many as 65% of the respondents said that the existing metabolism book could not be used to understand secondary metabolism in lectures of plant metabolism. Therefore, the development of teaching materials including plant metabolism-based bioinformatics is important to improve the understanding of the lecture material in plant metabolism.

  2. Fusion and metabolism of plant cells as affected by microgravity.

    Science.gov (United States)

    Hampp, R; Hoffmann, E; Schönherr, K; Johann, P; De Filippis, L

    1997-01-01

    Plant cell protoplasts derived from leaf tissue of two different tobacco species (Nicotiana tabacum., N. rustica L.) were exposed to short-term (sounding rocket experiments) and long-term (spacelab) microgravity environments in order to study both (electro) cell fusion and cell metabolism during early and later stages of tissue regeneration. The period of exposure to microgravity varied from 10 min (sounding rocket) to 10 d (space shuttle). The process of electro fusion of protoplasts was improved under conditions of microgravity: the time needed to establish close membrane contact between protoplasts (alignment time) was reduced (5 as compared to 15 s under 1 g) and numbers of fusion products between protoplasts of different specific density were increased by a factor of about 10. In addition, viability of fusion products, as shown by the ability to form callus, increased from about 60% to more than 90%. Regenerated fusion products obtained from both sounding-rocket and spacelab experiments showed a wide range of intermediate properties between the two parental plants. This was verified by isozyme analysis and random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). In order to address potential metabolic responses, more general markers such as the overall energy state (ATP/ADP ratio), the redox charge of the diphosphopyridine nucleotide system (NADH/NAD ratio), and the pool size of fructose-2,6-bisphosphate (Fru 2,6 bisp), a regulator of the balance between glycolysis and gluconeogenesis, were determined. Responses of these parameters were different with regard to short-term and long-term exposure. Shortly after transition to reduced gravitation (sounding rocket) ratios of ATP/ADP exhibited strong fluctuation while the pool size of NAD decreased (indicating an increased NADH/NAD ratio) and that of Fru 2,6 bisp increased. As similar changes can be observed under stress conditions, this response is probably indicative of a metabolic stress

  3. Metabolic engineering of the phenylpropanoid pathway enhances the antioxidant capacity of Saussurea involucrata.

    Directory of Open Access Journals (Sweden)

    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.

  4. Methylcyclopentadienyl manganese tricarbonyl (MMT), plant uptake and effects on metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Jones, A.R.; Lytle, C.M.; Stone, R.L.; Smith, B.N [Department of Botany and Range Science, Brigham Young University, Provo (United States); Hansen, L.D. [Department of Chemistry and Biochemistry, Brigham Young University, Provo (United States)

    2000-04-01

    In the USA and Canada, methylcyclopentadienyl manganese (MMT) is currently added to gasoline to replace tetraethyl lead as an antiknock fuel additive. Manganese concentrations in roadside soil and plants are increasing and correlated with distance from the roadway, traffic volume, plant type, and microhabitat. Radish (Raphanus sativus L.) seedlings were treated for either five or thirty-five days with different levels of manganous chloride (0-1000ppm). Metabolic heat rates (q) and respiration rates (R{sub CO{sub 2}}), measured calorimetrically, indicated severe stress at Mn concentrations between 10 and 100ppm and at temperatures above 20C. Predicted growth rates (R{sub SG}) also decreased in these circumstances.

  5. Plant diterpene synthases: exploring modularity and metabolic diversity for bioengineering.

    Science.gov (United States)

    Zerbe, Philipp; Bohlmann, Jörg

    2015-07-01

    Plants produce thousands of diterpenoid natural products; some of which are of significant industrial value as biobased pharmaceuticals (taxol), fragrances (sclareol), food additives (steviosides), and commodity chemicals (diterpene resin acids). In nature, diterpene synthase (diTPS) enzymes are essential for generating diverse diterpene hydrocarbon scaffolds. While some diTPSs also form oxygenated compounds, more commonly, oxygenation is achieved by cytochrome P450-dependent mono-oxygenases. Recent genome-, transcriptome-, and metabolome-guided gene discovery and enzyme characterization identified novel diTPS functions that form the core of complex modular pathway systems. Insights into diterpene metabolism may translate into the development of new bioengineered microbial and plant-based production systems.

  6. Metabolism of fluoranthene in different plant cell cultures and intact plants

    Energy Technology Data Exchange (ETDEWEB)

    Kolb, M.; Harms, H.

    2000-05-01

    The metabolism of fluoranthene was investigated in 11 cell cultures of different plant species using a [{sup 14}C]-labeled standard. Most species metabolized less than 5% of fluoranthene to soluble metabolites and formed less than 5% nonextractable residues during the standardized 48-h test procedure. Higher metabolic rates were observed in lettuce (Lactuca sativa, 6%), wheat (Tricitum aestivum, 9%), and tomato (Lycopersicon esculentum, 15%). A special high metabolic rate of nearly 50% was determined for the rose species Paul's Scarlet. Chromatographic analysis of metabolites extracted from aseptically grown tomato plants proved that the metabolites detected in the cell cultures were also formed in the intact plants. Metabolites produced in tomato and rose cells from [{sup 14}C]-fluoranthene were conjugated with glucose, glucuronic acid, and other cell components. After acid hydrolyses, the main metabolite of both species was 1-hydroxyfluoranthene as identified by gas chromatography-mass spectrometry and high-performance liquid chromatography with diode array detection. The second metabolite formed by both species was 8-hydroxyfluoranthene. A third metabolite in tomatoes was 3-hydroxyfluoranthene.

  7. Plant-Derived Human Collagen Scaffolds for Skin Tissue Engineering

    Science.gov (United States)

    Willard, James J.; Drexler, Jason W.; Das, Amitava; Roy, Sashwati; Shilo, Shani; Shoseyov, Oded

    2013-01-01

    Tissue engineering scaffolds are commonly formed using proteins extracted from animal tissues, such as bovine hide. Risks associated with the use of these materials include hypersensitivity and pathogenic contamination. Human-derived proteins lower the risk of hypersensitivity, but possess the risk of disease transmission. Methods engineering recombinant human proteins using plant material provide an alternate source of these materials without the risk of disease transmission or concerns regarding variability. To investigate the utility of plant-derived human collagen (PDHC) in the development of engineered skin (ES), PDHC and bovine hide collagen were formed into tissue engineering scaffolds using electrospinning or freeze-drying. Both raw materials were easily formed into two common scaffold types, electrospun nonwoven scaffolds and lyophilized sponges, with similar architectures. The processing time, however, was significantly lower with PDHC. PDHC scaffolds supported primary human cell attachment and proliferation at an equivalent or higher level than the bovine material. Interleukin-1 beta production was significantly lower when activated THP-1 macrophages where exposed to PDHC electrospun scaffolds compared to bovine collagen. Both materials promoted proper maturation and differentiation of ES. These data suggest that PDHC may provide a novel source of raw material for tissue engineering with low risk of allergic response or disease transmission. PMID:23298216

  8. Photoreceptor effects on plant biomass, resource allocation, and metabolic state.

    Science.gov (United States)

    Yang, Deyue; Seaton, Daniel D; Krahmer, Johanna; Halliday, Karen J

    2016-07-05

    Plants sense the light environment through an ensemble of photoreceptors. Members of the phytochrome class of light receptors are known to play a critical role in seedling establishment, and are among the best-characterized plant signaling components. Phytochromes also regulate adult plant growth; however, our knowledge of this process is rather fragmented. This study demonstrates that phytochrome controls carbon allocation and biomass production in the developing plant. Phytochrome mutants have a reduced CO2 uptake, yet overaccumulate daytime sucrose and starch. This finding suggests that even though carbon fixation is impeded, the available carbon resources are not fully used for growth during the day. Supporting this notion, phytochrome depletion alters the proportion of day:night growth. In addition, phytochrome loss leads to sizeable reductions in overall growth, dry weight, total protein levels, and the expression of CELLULOSE SYNTHASE-LIKE genes. Because cellulose and protein are major constituents of plant biomass, our data point to an important role for phytochrome in regulating these fundamental components of plant productivity. We show that phytochrome loss impacts core metabolism, leading to elevated levels of tricarboxylic acid cycle intermediates, amino acids, sugar derivatives, and notably the stress metabolites proline and raffinose. Furthermore, the already growth-retarded phytochrome mutants are less responsive to growth-inhibiting abiotic stresses and have elevated expression of stress marker genes. This coordinated response appears to divert resources from energetically costly biomass production to improve resilience. In nature, this strategy may be activated in phytochrome-disabling, vegetation-dense habitats to enhance survival in potentially resource-limiting conditions.

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

    Science.gov (United States)

    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.

  10. Metabolic Engineering of Oleaginous Yeasts for Fatty Alcohol Production

    Energy Technology Data Exchange (ETDEWEB)

    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.

  11. Metabolic engineering of Bacillus subtilis for terpenoid production.

    Science.gov (United States)

    Guan, Zheng; Xue, Dan; Abdallah, Ingy I; Dijkshoorn, Linda; Setroikromo, Rita; Lv, Guiyuan; Quax, Wim J

    2015-11-01

    Terpenoids are the largest group of small-molecule natural products, with more than 60,000 compounds made from isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP). As the most diverse group of small-molecule natural products, terpenoids play an important role in the pharmaceutical, food, and cosmetic industries. For decades, Escherichia coli (E. coli) and Saccharomyces cerevisiae (S. cerevisiae) were extensively studied to biosynthesize terpenoids, because they are both fully amenable to genetic modifications and have vast molecular resources. On the other hand, our literature survey (20 years) revealed that terpenoids are naturally more widespread in Bacillales. In the mid-1990s, an inherent methylerythritol phosphate (MEP) pathway was discovered in Bacillus subtilis (B. subtilis). Since B. subtilis is a generally recognized as safe (GRAS) organism and has long been used for the industrial production of proteins, attempts to biosynthesize terpenoids in this bacterium have aroused much interest in the scientific community. This review discusses metabolic engineering of B. subtilis for terpenoid production, and encountered challenges will be discussed. We will summarize some major advances and outline future directions for exploiting the potential of B. subtilis as a desired "cell factory" to produce terpenoids.

  12. Metabolic engineering of Yarrowia lipolytica for itaconic acid production.

    Science.gov (United States)

    Blazeck, John; Hill, Andrew; Jamoussi, Mariam; Pan, Anny; Miller, Jarrett; Alper, Hal S

    2015-11-01

    Itaconic acid is a naturally produced organic acid with diverse applications as a replacement for petroleum derived products. However, its industrial viability as a bio-replacement has been restricted due to limitations with native producers. In this light, Yarrowia lipolytica is an excellent potential candidate for itaconic acid production due to its innate capacity to accumulate citric acid cycle intermediates and tolerance to lower pH. Here, we demonstrate the capacity to produce itaconic acid in Y. lipolytica through heterologous expression of the itaconic acid synthesis enzyme, resulting in an initial titer of 33 mg/L. Further optimizations of this strain via metabolic pathway engineering, enzyme localization, and media optimization strategies enabled 4.6g/L of itaconic acid to be produced in bioreactors, representing a 140-fold improvement over initial titer. Moreover, these fermentation conditions did not require additional nutrient supplementation and utilized a low pH condition that enabled the acid form of itaconic acid to be produced. Overall yields (0.058 g/g yield from glucose) and maximum productivity of 0.045 g/L/h still provide areas for future strain improvement. Nevertheless, this work demonstrates that Y. lipolytica has the potential to serve as an industrially relevant platform for itaconic acid production.

  13. Advancing oleaginous microorganisms to produce lipid via metabolic engineering technology.

    Science.gov (United States)

    Liang, Ming-Hua; Jiang, Jian-Guo

    2013-10-01

    With the depletion of global petroleum and its increasing price, biodiesel has been becoming one of the most promising biofuels for global fuels market. Researchers exploit oleaginous microorganisms for biodiesel production due to their short life cycle, less labor required, less affection by venue, and easier to scale up. Many oleaginous microorganisms can accumulate lipids, especially triacylglycerols (TAGs), which are the main materials for biodiesel production. This review is covering the related researches on different oleaginous microorganisms, such as yeast, mold, bacteria and microalgae, which might become the potential oil feedstocks for biodiesel production in the future, showing that biodiesel from oleaginous microorganisms has a great prospect in the development of biomass energy. Microbial oils biosynthesis process includes fatty acid synthesis approach and TAG synthesis approach. In addition, the strategies to increase lipids accumulation via metabolic engineering technology, involving the enhancement of fatty acid synthesis approach, the enhancement of TAG synthesis approach, the regulation of related TAG biosynthesis bypass approaches, the blocking of competing pathways and the multi-gene approach, are discussed in detail. It is suggested that DGAT and ME are the most promising targets for gene transformation, and reducing PEPC activity is observed to be beneficial for lipid production.

  14. Engineering a catabolic pathway in plants for the degradation of 1,2-dichloroethane.

    Science.gov (United States)

    Mena-Benitez, Gilda L; Gandia-Herrero, Fernando; Graham, Stuart; Larson, Tony R; McQueen-Mason, Simon J; French, Christopher E; Rylott, Elizabeth L; Bruce, Neil C

    2008-07-01

    Plants are increasingly being employed to clean up environmental pollutants such as heavy metals; however, a major limitation of phytoremediation is the inability of plants to mineralize most organic pollutants. A key component of organic pollutants is halogenated aliphatic compounds that include 1,2-dichloroethane (1,2-DCA). Although plants lack the enzymatic activity required to metabolize this compound, two bacterial enzymes, haloalkane dehalogenase (DhlA) and haloacid dehalogenase (DhlB) from the bacterium Xanthobacter autotrophicus GJ10, have the ability to dehalogenate a range of halogenated aliphatics, including 1,2-DCA. We have engineered the dhlA and dhlB genes into tobacco (Nicotiana tabacum 'Xanthi') plants and used 1,2-DCA as a model substrate to demonstrate the ability of the transgenic tobacco to remediate a range of halogenated, aliphatic hydrocarbons. DhlA converts 1,2-DCA to 2-chloroethanol, which is then metabolized to the phytotoxic 2-chloroacetaldehyde, then chloroacetic acid, by endogenous plant alcohol dehydrogenase and aldehyde dehydrogenase activities, respectively. Chloroacetic acid is dehalogenated by DhlB to produce the glyoxylate cycle intermediate glycolate. Plants expressing only DhlA produced phytotoxic levels of chlorinated intermediates and died, while plants expressing DhlA together with DhlB thrived at levels of 1,2-DCA that were toxic to DhlA-expressing plants. This represents a significant advance in the development of a low-cost phytoremediation approach toward the clean-up of halogenated organic pollutants from contaminated soil and groundwater.

  15. Nucleobase and nucleoside transport and integration into plant metabolism

    Directory of Open Access Journals (Sweden)

    Christopher eGirke

    2014-09-01

    Full Text Available Nucleotide metabolism is an essential process in all living organisms. Besides newly synthesized nucleotides, the recycling (salvage of partially degraded nucleotides i.e. nucleosides and nucleobases serves to keep the homeostasis of the nucleotide pool. Both types of metabolites are substrates of at least six families of transport proteins in Arabidopsis thaliana (Arabidopsis with a total of 49 members. In the last years several members of such transport proteins have been analyzed allowing to present a more detailed picture of nucleoside and nucleobase transport and the physiological function of these processes. Besides functioning in nucleotide metabolism it turned out that individual members of the before named transporters exhibit the capacity to transport a wide range of different substrates including vitamins and phytohormones. The aim of this review is to summarize the current knowledge on nucleobase and nucleoside transport processes in plants and integrate this into nucleotide metabolism in general. Thereby, we will focus on those proteins which have been characterized at the biochemical level.

  16. Nucleobase and nucleoside transport and integration into plant metabolism.

    Science.gov (United States)

    Girke, Christopher; Daumann, Manuel; Niopek-Witz, Sandra; Möhlmann, Torsten

    2014-01-01

    Nucleotide metabolism is an essential process in all living organisms. Besides newly synthesized nucleotides, the recycling (salvage) of partially degraded nucleotides, i.e., nucleosides and nucleobases serves to keep the homeostasis of the nucleotide pool. Both types of metabolites are substrates of at least six families of transport proteins in Arabidopsis thaliana (Arabidopsis) with a total of 49 members. In the last years several members of such transport proteins have been analyzed allowing to present a more detailed picture of nucleoside and nucleobase transport and the physiological function of these processes. Besides functioning in nucleotide metabolism it turned out that individual members of the before named transporters exhibit the capacity to transport a wide range of different substrates including vitamins and phytohormones. The aim of this review is to summarize the current knowledge on nucleobase and nucleoside transport processes in plants and integrate this into nucleotide metabolism in general. Thereby, we will focus on those proteins which have been characterized at the biochemical level.

  17. Metabolic engineering of lactic acid bacteria, the combined approach: kinetic modelling, metabolic control and experimental analysis.

    Science.gov (United States)

    Hoefnagel, Marcel H N; Starrenburg, Marjo J C; Martens, Dirk E; Hugenholtz, Jeroen; Kleerebezem, Michiel; Van Swam, Iris I; Bongers, Roger; Westerhoff, Hans V; Snoep, Jacky L

    2002-04-01

    Everyone who has ever tried to radically change metabolic fluxes knows that it is often harder to determine which enzymes have to be modified than it is to actually implement these changes. In the more traditional genetic engineering approaches 'bottle-necks' are pinpointed using qualitative, intuitive approaches, but the alleviation of suspected 'rate-limiting' steps has not often been successful. Here the authors demonstrate that a model of pyruvate distribution in Lactococcus lactis based on enzyme kinetics in combination with metabolic control analysis clearly indicates the key control points in the flux to acetoin and diacetyl, important flavour compounds. The model presented here (available at http://jjj.biochem.sun.ac.za/wcfs.html) showed that the enzymes with the greatest effect on this flux resided outside the acetolactate synthase branch itself. Experiments confirmed the predictions of the model, i.e. knocking out lactate dehydrogenase and overexpressing NADH oxidase increased the flux through the acetolactate synthase branch from 0 to 75% of measured product formation rates.

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

    Science.gov (United States)

    Ye, Xiaoting; Honda, Kohsuke; Sakai, Takaaki; Okano, Kenji; Omasa, Takeshi; Hirota, Ryuichi; Kuroda, Akio; Ohtake, Hisao

    2012-09-06

    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. 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. 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 potentially applicable to the biocatalytic manufacture of any chemicals

  19. Genetic engineering of plant food with reduced allergenicity.

    Science.gov (United States)

    Scheurer, Stephan; Sonnewald, Sophia

    2009-01-01

    Food allergies are a major health concern in industrialized countries. Since a specific immunotherapy for food allergies is not available in clinical routine praxis till now, reduction of allergens in foods, either by food processing or genetic engineering are strategies to minimize the risk of adverse reactions for food allergic patients. This review summarizes biotechnological approaches, especially the RNA interference (RNAi) technology, for the reduction of selected allergens in plant foods. So far, only a limited number of reports showing proof-of-concept of this methodology are available. Using RNAi an impressive reduction of allergen accumulation was obtained which was stable in the next generations of plants. Since threshold doses for most food allergens are not known, the beneficial effect has to be evaluated by oral challenge tests in the future. The article critically addresses the potential and limitations of genetic engineering, as well as of alternative strategies to generate "low allergic" foods.

  20. Computational approaches to metabolic engineering utilizing systems biology and synthetic biology.

    Science.gov (United States)

    Fong, Stephen S

    2014-08-01

    Metabolic engineering modifies cellular function to address various biochemical applications. Underlying metabolic engineering efforts are a host of tools and knowledge that are integrated to enable successful outcomes. Concurrent development of computational and experimental tools has enabled different approaches to metabolic engineering. One approach is to leverage knowledge and computational tools to prospectively predict designs to achieve the desired outcome. An alternative approach is to utilize combinatorial experimental tools to empirically explore the range of cellular function and to screen for desired traits. This mini-review focuses on computational systems biology and synthetic biology tools that can be used in combination for prospective in silico strain design.

  1. Shikimic acid production in Escherichia coli: From classical metabolic engineering strategies to omics applied to improve its production

    Directory of Open Access Journals (Sweden)

    Juan Andrés Martínez

    2015-09-01

    Full Text Available Shikimic acid (SA is an intermediate of the SA pathway that is present in bacteria and plants. SA has gained great interest because it is a precursor in the synthesis of the drug oseltamivir phosphate (OSF, an efficient inhibitor of the neuraminidase enzyme of diverse seasonal influenza viruses, the avian influenza virus H5N1, and the human influenza virus H1N1. For the purposes of OSF production, SA is extracted from the pods of Chinese star anise plants (Illicium spp., yielding up to 17% of SA (dry basis content. The high demand for OSF necessary to manage a major influenza outbreak is not adequately met by industrial production using SA from plants sources. As the SA pathway is present in the model bacteria Escherichia coli, several intuitive metabolically engineered strains have been applied for its successful overproduction by biotechnological processes, resulting in strains producing up to 71 g/L of SA, with high conversion yields of up to 0.42 (mol SA/mol Glc, in both batch and fed-batch cultures using complex fermentation broths, including glucose as a carbon source and yeast extract. Global transcriptomic analyses have been performed in SA producing strains, resulting in the identification of possible key target genes for the design of a rational strain improvement strategy. Because possible target genes are involved in the transport, catabolism and interconversion of different carbon sources and metabolic intermediates outside the central carbon metabolism and SA pathways, as genes involved in diverse cellular stress responses, the development of rational cellular strain improvement strategies based on omics data constitutes a challenging task to improve SA production in currently overproducing engineered strains. In this review, we discuss the main metabolic engineering strategies that have been applied for the development of efficient SA producing strains, as the perspective of omics analysis has focused on further strain improvement

  2. Metabolic adaptation in transplastomic plants massively accumulating recombinant proteins.

    Directory of Open Access Journals (Sweden)

    Julia Bally

    Full Text Available BACKGROUND: Recombinant chloroplasts are endowed with an astonishing capacity to accumulate foreign proteins. However, knowledge about the impact on resident proteins of such high levels of recombinant protein accumulation is lacking. METHODOLOGY/PRINCIPAL FINDINGS: Here we used proteomics to characterize tobacco (Nicotiana tabacum plastid transformants massively accumulating a p-hydroxyphenyl pyruvate dioxygenase (HPPD or a green fluorescent protein (GFP. While under the conditions used no obvious modifications in plant phenotype could be observed, these proteins accumulated to even higher levels than ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco, the most abundant protein on the planet. This accumulation occurred at the expense of a limited number of leaf proteins including Rubisco. In particular, enzymes involved in CO(2 metabolism such as nuclear-encoded plastidial Calvin cycle enzymes and mitochondrial glycine decarboxylase were found to adjust their accumulation level to these novel physiological conditions. CONCLUSIONS/SIGNIFICANCE: The results document how protein synthetic capacity is limited in plant cells. They may provide new avenues to evaluate possible bottlenecks in recombinant protein technology and to maintain plant fitness in future studies aiming at producing recombinant proteins of interest through chloroplast transformation.

  3. Nuclear magnetic resonance spectroscopy of dense cell populations for metabolic studies and bioreactor engineering: a synergistic partnership.

    Science.gov (United States)

    Dale, B E; Gillies, R J

    1991-01-01

    Commercial exploitation of the fruits of recombinant DNA and cell fusion technologies is significantly limited by the lack of fundamental metabolic information on the cell lines of interest, whether these are plant, animal, insect, or microbial cells. NMR can help to provide this information and thereby improve bioreactor design and operation. However, in the case of on-line NMR of dense cell culture devices for metabolic studies, these devices are inherently heterogeneous bioreactors. To ensure that the metabolic information generated is reliable, a number of precautions should be taken. These are the same precautions that should be taken to ensure that commercial bioreactors operate in a reaction-controlled regime. Therefore, reactor engineering methodologies, particularly diffusion and reaction analyses and reaction monitoring by whole-cell NMR must go hand in hand, each extending, complementing, and validating the other.

  4. In vivo synthesis of nanomaterials in plants: location of silver nanoparticles and plant metabolism

    Science.gov (United States)

    Marchiol, Luca; Mattiello, Alessandro; Pošćić, Filip; Giordano, Cristiana; Musetti, Rita

    2014-03-01

    Metallic nanoparticles (MeNPs) can be formed in living plants by reduction of the metal ions absorbed as soluble salts. It is very likely that plant metabolism has an important role in MeNP biosynthesis. The in vivo formation of silver nanoparticles (AgNPs) was observed in Brassica juncea, Festuca rubra and Medicago sativa. Plants were grown in Hoagland's solution for 30 days and then exposed for 24 h to a solution of 1,000 ppm AgNO3. In the leaf extracts of control plants, the concentrations of glucose, fructose, ascorbic acid, citric acid and total polyphenols were determined. Total Ag content in plant fractions was determined by inductively coupled plasma atomic emission spectroscopy. Despite the short exposure time, the Ag uptake and translocation to plant leaves was very high, reaching 6,156 and 2,459 mg kg-1 in B. juncea and F. rubra, respectively. Ultrastructural analysis was performed by transmission electron microscopy (TEM), and AgNPs were detected by TEM X-ray microanalysis. TEM images of plant fractions showed the in vivo formation of AgNPs in the roots, stems and leaves of the plants. In the roots, AgNPs were present in the cortical parenchymal cells, on the cell wall of the xylem vessels and in regions corresponding to the pits. In leaf tissues, AgNPs of different sizes and shapes were located close to the cell wall, as well as in the cytoplasm and within chloroplasts. AgNPs were not observed in the phloem of the three plant species. This is the first report of AgNP synthesis in living plants of F. rubra. The contents of reducing sugars and antioxidant compounds, proposed as being involved in the biosynthesis of AgNPs, were quite different between the species, thus suggesting that it is unlikely that a single substance is responsible for this process.

  5. Metabolic engineering of Saccharomyces cerevisiae for production of ginsenosides.

    Science.gov (United States)

    Dai, Zhubo; Liu, Yi; Zhang, Xianan; Shi, Mingyu; Wang, Beibei; Wang, Dong; Huang, Luqi; Zhang, Xueli

    2013-11-01

    Ginsenosides are the primary bioactive components of ginseng, which is a popular medicinal herb and exhibits diverse pharmacological activities. Protopanaxadiol is the aglycon of several dammarane-type ginsenosides, which also has anticancer activity. For microbial production of protopanaxadiol, dammarenediol-II synthase and protopanaxadiol synthase genes of Panax ginseng, together with a NADPH-cytochrome P450 reductase gene of Arabidopsis thaliana, were introduced into Saccharomyces cerevisiae, resulting in production of 0.05 mg/g DCW protopanaxadiol. Increasing squalene and 2,3-oxidosqualene supplies through overexpressing truncated 3-hydroxyl-3-methylglutaryl-CoA reductase, farnesyl diphosphate synthase, squalene synthase and 2,3-oxidosqualene synthase genes, together with increasing protopanaxadiol synthase activity through codon optimization, led to 262-fold increase of protopanaxadiol production. Finally, using two-phase extractive fermentation resulted in production of 8.40 mg/g DCW protopanaxadiol (1189 mg/L), together with 10.94 mg/g DCW dammarenediol-II (1548 mg/L). The yeast strains engineered in this work can serve as the basis for creating an alternative way for production of ginsenosides in place of extraction from plant sources.

  6. Cytochrome P450-Mediated Phytoremediation using Transgenic Plants: A Need for Engineered Cytochrome P450 Enzymes

    Science.gov (United States)

    Kumar, Santosh; Jin, Mengyao; Weemhoff, James L

    2013-01-01

    There is an increasing demand for versatile and ubiquitous Cytochrome P450 (CYP) biocatalysts for biotechnology, medicine, and bioremediation. In the last decade there has been an increase in realization of the power of CYP biocatalysts for detoxification of soil and water contaminants using transgenic plants. However, the major limitations of mammalian CYP enzymes are that they require CYP reductase (CPR) for their activity, and they show relatively low activity, stability, and expression. On the other hand, bacterial CYP enzymes show limited substrate diversity and usually do not metabolize herbicides and industrial contaminants. Therefore, there has been a considerable interest for biotechnological industries and the scientific community to design CYP enzymes to improve their catalytic efficiency, stability, expression, substrate diversity, and the suitability of P450-CPR fusion enzymes. Engineered CYP enzymes have potential for transgenic plants-mediated phytoremediation of herbicides and environmental contaminants. In this review we discuss: 1) the role of CYP enzymes in phytoremediation using transgenic plants, 2) problems associated with wild-type CYP enzymes in phytoremediation, and 3) examples of engineered CYP enzymes and their potential role in transgenic plant-mediated phytoremediation. PMID:25298920

  7. Metabolic engineering of Clostridium tyrobutyricum for n-butanol production.

    Science.gov (United States)

    Yu, Mingrui; Zhang, Yali; Tang, I-Ching; Yang, Shang-Tian

    2011-07-01

    Clostridium tyrobutyricum ATCC 25755, a butyric acid producing bacterium, has been engineered to overexpress aldehyde/alcohol dehydrogenase 2 (adhE2, Genebank no. AF321779) from Clostridium acetobutylicum ATCC 824, which converts butyryl-CoA to butanol, under the control of native thiolase (thl) promoter. Butanol titer of 1.1g/L was obtained in C. tyrobutyricum overexpressing adhE2. The effects of inactivating acetate kinase (ack) and phosphotransbutyrylase (ptb) genes in the host on butanol production were then studied. A high C4/C2 product ratio of 10.6 (mol/mol) was obtained in ack knockout mutant, whereas a low C4/C2 product ratio of 1.4 (mol/mol) was obtained in ptb knockout mutant, confirming that ack and ptb genes play important roles in controlling metabolic flux distribution in C. tyrobutyricum. The highest butanol titer of 10.0g/L and butanol yield of 27.0% (w/w, 66% of theoretical yield) were achieved from glucose in the ack knockout mutant overexpressing adhE2. When a more reduced substrate mannitol was used, the butanol titer reached 16.0 g/L with 30.6% (w/w) yield (75% theoretical yield). Moreover, C. tyrobutyricum showed good butanol tolerance, with >80% and ∼60% relative growth rate at 1.0% and 1.5% (v/v) butanol. These results suggest that C. tyrobutyricum is a promising heterologous host for n-butanol production from renewable biomass.

  8. Human factors engineering plan for reviewing nuclear plant modernization programs

    Energy Technology Data Exchange (ETDEWEB)

    O' Hara, John; Higgins, James [Brookhaven National Laboratory, Upton, NY (United States)

    2004-12-01

    The Swedish Nuclear Power Inspectorate reviews the human factors engineering (HFE) aspects of nuclear power plants (NPPs) involved in the modernization of the plant systems and control rooms. The purpose of a HFE review is to help ensure personnel and public safety by verifying that accepted HFE practices and guidelines are incorporated into the program and nuclear power plant design. Such a review helps to ensure the HFE aspects of an NPP are developed, designed, and evaluated on the basis of a structured top-down system analysis using accepted HFE principles. The review addresses eleven HFE elements: HFE Program Management, Operating Experience Review, Functional Requirements Analysis and Allocation, Task Analysis, Staffing, Human Reliability Analysis, Human-System Interface Design, Procedure Development, Training Program Development, Human Factors Verification and Validation, and Design Implementation.

  9. Engineering plant-microbe symbiosis for rhizoremediation of heavy metals.

    Science.gov (United States)

    Wu, Cindy H; Wood, Thomas K; Mulchandani, Ashok; Chen, Wilfred

    2006-02-01

    The use of plants for rehabilitation of heavy-metal-contaminated environments is an emerging area of interest because it provides an ecologically sound and safe method for restoration and remediation. Although a number of plant species are capable of hyperaccumulation of heavy metals, the technology is not applicable for remediating sites with multiple contaminants. A clever solution is to combine the advantages of microbe-plant symbiosis within the plant rhizosphere into an effective cleanup technology. We demonstrated that expression of a metal-binding peptide (EC20) in a rhizobacterium, Pseudomonas putida 06909, not only improved cadmium binding but also alleviated the cellular toxicity of cadmium. More importantly, inoculation of sunflower roots with the engineered rhizobacterium resulted in a marked decrease in cadmium phytotoxicity and a 40% increase in cadmium accumulation in the plant root. Owing to the significantly improved growth characteristics of both the rhizobacterium and plant, the use of EC20-expressing P. putida endowed with organic-degrading capabilities may be a promising strategy to remediate mixed organic-metal-contaminated sites.

  10. Lessons learned: Are engineered nanomaterials toxic to terrestrial plants?

    Science.gov (United States)

    Reddy, P Venkata Laxma; Hernandez-Viezcas, J A; Peralta-Videa, J R; Gardea-Torresdey, J L

    2016-10-15

    The expansion of nanotechnology and its ubiquitous applications has fostered unavoidable interaction between engineered nanomaterials (ENMs) and plants. Recent research has shown ambiguous results with regard to the impact of ENMs in plants. On one hand, there are reports that show hazardous effects, while on the other hand, some reports highlight positive effects. This uncertainty whether the ENMs are primarily hazardous or whether they have a potential for propitious impact on plants, has raised questions in the scientific community. In this review, we tried to demystify this ambiguity by citing various exposure studies of different ENMs (nano-Ag, nano-Au, nano-Si, nano-CeO2, nano-TiO2, nano-CuO, nano-ZnO, and CNTs, among others) and their effects on various groups of plant families. After scrutinizing the most recent literature, it seems that the divergence in the research results may be possibly attributed to multiple factors such as ENM properties, plant species, soil dynamics, and soil microbial community. The analysis of the literature also suggests that there is a knowledge gap on the effects of ENMs towards changes in color, texture, shape, and nutritional aspects on ENM exposed plants. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Altered Levels of Aroma and Volatiles by Metabolic Engineering of Shikimate Pathway Genes in Tomato Fruits

    Directory of Open Access Journals (Sweden)

    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

  12. How to Do It. Plant Eco-Physiology: Experiments on Crassulacean Acid Metabolism, Using Minimal Equipment.

    Science.gov (United States)

    Friend, Douglas J. C.

    1990-01-01

    Features of Crassulacean Acid Metabolism plants are presented. Investigations of a complex eco-physiological plant adaptation to the problems of growth in an arid environment are discussed. Materials and procedures for these investigations are described. (CW)

  13. Overcoming the plasticity of plant specialized metabolism for selective diterpene production in yeast

    DEFF Research Database (Denmark)

    Ignea, Codruta; Athanasakoglou, Anastasia; Andreadelli, Aggeliki

    2017-01-01

    Plants synthesize numerous specialized metabolites (also termed natural products) to mediate dynamic interactions with their surroundings. The complexity of plant specialized metabolism is the result of an inherent biosynthetic plasticity rooted in the substrate and product promiscuity...

  14. Metabolic engineering of ammonium assimilation in xylose-fermenting Saccharomyes cerevisiae improves ethanol production

    DEFF Research Database (Denmark)

    Roca, Christophe Francois Aime; Nielsen, Jens; Olsson, Lisbeth

    2003-01-01

    Cofactor imbalance impedes xylose assimilation in Saccharomyces cerevisiae that has been metabolically engineered for xylose utilization. To improve cofactor use, we modified ammonia assimilation in recombinant S. cerevisiae by deleting GDH1, which encodes an NADPH-dependent glutamate dehydrogenase...

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

    Directory of Open Access Journals (Sweden)

    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.

  16. Gene expression in plant lipid metabolism in Arabidopsis seedlings.

    Directory of Open Access Journals (Sweden)

    An-Shan Hsiao

    Full Text Available Events in plant lipid metabolism are important during seedling establishment. As it has not been experimentally verified whether lipid metabolism in 2- and 5-day-old Arabidopsis thaliana seedlings is diurnally-controlled, quantitative real-time PCR analysis was used to investigate the expression of target genes in acyl-lipid transfer, β-oxidation and triacylglycerol (TAG synthesis and hydrolysis in wild-type Arabidopsis WS and Col-0. In both WS and Col-0, ACYL-COA-BINDING PROTEIN3 (ACBP3, DIACYLGLYCEROL ACYLTRANSFERASE1 (DGAT1 and DGAT3 showed diurnal control in 2- and 5-day-old seedlings. Also, COMATOSE (CTS was diurnally regulated in 2-day-old seedlings and LONG-CHAIN ACYL-COA SYNTHETASE6 (LACS6 in 5-day-old seedlings in both WS and Col-0. Subsequently, the effect of CIRCADIAN CLOCK ASSOCIATED1 (CCA1 and LATE ELONGATED HYPOCOTYL (LHY from the core clock system was examined using the cca1lhy mutant and CCA1-overexpressing (CCA1-OX lines versus wild-type WS and Col-0, respectively. Results revealed differential gene expression in lipid metabolism between 2- and 5-day-old mutant and wild-type WS seedlings, as well as between CCA1-OX and wild-type Col-0. Of the ACBPs, ACBP3 displayed the most significant changes between cca1lhy and WS and between CCA1-OX and Col-0, consistent with previous reports that ACBP3 is greatly affected by light/dark cycling. Evidence of oil body retention in 4- and 5-day-old seedlings of the cca1lhy mutant in comparison to WS indicated the effect of cca1lhy on storage lipid reserve mobilization. Lipid profiling revealed differences in primary lipid metabolism, namely in TAG, fatty acid methyl ester and acyl-CoA contents amongst cca1lhy, CCA1-OX, and wild-type seedlings. Taken together, this study demonstrates that lipid metabolism is subject to diurnal regulation in the early stages of seedling development in Arabidopsis.

  17. Metabolic engineering for microbial production of aromatic amino acids and derived compounds.

    Science.gov (United States)

    Bongaerts, J; Krämer, M; Müller, U; Raeven, L; Wubbolts, M

    2001-10-01

    Metabolic engineering to design and construct microorganisms suitable for the production of aromatic amino acids and derivatives thereof requires control of a complicated network of metabolic reactions that partly act in parallel and frequently are in rapid equilibrium. Engineering the regulatory circuits, the uptake of carbon, the glycolytic pathway, the pentose phosphate pathway, and the common aromatic amino acid pathway as well as amino acid importers and exporters that have all been targeted to effect higher productivities of these compounds are discussed.

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

    DEFF Research Database (Denmark)

    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....

  19. Survey on the technological development issues for large-scale methanol engine power generation plant; Ogata methanol engine hatsuden plant ni kansuru gijutsu kaihatsu kadai chosa

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1994-03-01

    Based on the result of `Survey on the feasibility of large-scale methanol engine power generation plant` in fiscal 1992, concrete technological development issues were studied for its practical use, and the technological R & D scheme was prepared for large-scale methanol engine power plant featured by low NOx and high efficiency. Technological development issues of this plant were as follows: improvement of thermal efficiency, reduction of NOx emission, improvement of the reliability and durability of ignition and fuel injection systems, and reduction of vibration. As the economical effect of the technological development, the profitability of NOx control measures was compared between this methanol engine and conventional heavy oil diesel engines or gas engines. As a result, this engine was more economical than conventional engines. It was suggested that development of the equipment will be completed in nearly 4 years through every component study, single-cylinder model experiment and real engine test. 21 refs., 43 figs., 19 tabs.

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

    Science.gov (United States)

    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.

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

    NARCIS (Netherlands)

    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

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

    NARCIS (Netherlands)

    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 compou

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

    NARCIS (Netherlands)

    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

  4. Perspectives for a better understanding of the metabolic integration of photorespiration within a complex plant primary metabolism network.

    Science.gov (United States)

    Hodges, Michael; Dellero, Younès; Keech, Olivier; Betti, Marco; Raghavendra, Agepati S; Sage, Rowan; Zhu, Xin-Guang; Allen, Doug K; Weber, Andreas P M

    2016-05-01

    Photorespiration is an essential high flux metabolic pathway that is found in all oxygen-producing photosynthetic organisms. It is often viewed as a closed metabolic repair pathway that serves to detoxify 2-phosphoglycolic acid and to recycle carbon to fuel the Calvin-Benson cycle. However, this view is too simplistic since the photorespiratory cycle is known to interact with several primary metabolic pathways, including photosynthesis, nitrate assimilation, amino acid metabolism, C1 metabolism and the Krebs (TCA) cycle. Here we will review recent advances in photorespiration research and discuss future priorities to better understand (i) the metabolic integration of the photorespiratory cycle within the complex network of plant primary metabolism and (ii) the importance of photorespiration in response to abiotic and biotic stresses.

  5. Metabolic engineering of Escherichia coli for efficient free fatty acid production from glycerol.

    Science.gov (United States)

    Wu, Hui; Karanjikar, Mukund; San, Ka-Yiu

    2014-09-01

    Crude glycerol, generated as waste by-product in biodiesel production process, has been considered as an important carbon source for converting to value-added bioproducts recently. Free fatty acids (FFAs) can be used as precursors for the production of biofuels or biochemicals. Microbial biosynthesis of FFAs can be achieved by introducing an acyl-acyl carrier protein thioesterase into Escherichia coli. In this study, the effect of metabolic manipulation of FFAs synthesis cycle, host genetic background and cofactor engineering on FFAs production using glycerol as feed stocks was investigated. The highest concentration of FFAs produced by the engineered stain reached 4.82g/L with the yield of 29.55% (g FFAs/g glycerol), about 83% of the maximum theoretical pathway value by the type II fatty acid synthesis pathway. In addition, crude glycerol from biodiesel plant was also used as feedstock in this study. The FFA production was 3.53g/L with a yield of 24.13%. The yield dropped slightly when crude glycerol was used as a carbon source instead of pure glycerol, while it still can reach about 68% of the maximum theoretical pathway yield.

  6. In-depth metabolic phenotyping of genetically engineered mouse models in obesity and diabetes.

    Science.gov (United States)

    Lee, Hui-Young; Jeong, Kyeong-Hoon; Choi, Cheol Soo

    2014-10-01

    The world-wide prevalence of obesity and diabetes has increased sharply during the last two decades. Accordingly, the metabolic phenotyping of genetically engineered mouse models is critical for evaluating the functional roles of target genes in obesity and diabetes, and for developing new therapeutic targets. In this review, we discuss the practical meaning of metabolic phenotyping, the strategy of choosing appropriate tests, and considerations when designing and performing metabolic phenotyping in mice.

  7. Metabolic engineering of ethanol production in Thermoanaerobacter mathranii

    Energy Technology Data Exchange (ETDEWEB)

    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

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

    Directory of Open Access Journals (Sweden)

    McAnulty Michael J

    2012-05-01

    Full Text Available Abstract Background 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. Results 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. Conclusions 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.

  9. Examining strategies to facilitate vitamin B1 biofortification of plants by genetic engineering

    Directory of Open Access Journals (Sweden)

    Lucille ePourcel

    2013-05-01

    Full Text Available Thiamin (vitamin B1 is made by plants and microorganisms but is an essential micronutrient in the human diet. All organisms require it as a cofactor in its form as thiamin pyrophosphate (TPP for the activity of key enzymes of central metabolism. In humans, deficiency is widespread particularly in populations where polished rice is a major component of the diet. Considerable progress has been made on the elucidation of the biosynthesis pathway within the last few years enabling concrete strategies for biofortification purposes to be devised, with a particular focus here on genetic engineering. Furthermore, the vitamin has been shown to play a role in both abiotic and biotic stress responses. The precursors for de novo biosynthesis of thiamin differ between microorganisms and plants. Bacteria use intermediates derived from purine and isoprenoid biosynthesis, whereas the pathway in yeast involves the use of compounds from the vitamin B3 and B6 groups. Plants on the other hand use a combination of the bacterial and yeast pathways and there is subcellular partitioning of the biosynthesis steps. Specifically, thiamin biosynthesis occurs in the chloroplast of plants through the separate formation of the pyrimidine and thiazole moieties, which are then coupled to form thiamin monophosphate (TMP. Phosphorylation of thiamin to form TPP occurs in the cytosol. Therefore, thiamin (or TMP must be exported from the chloroplast to the cytosol for the latter step to be executed. The regulation of biosynthesis is mediated through riboswitches, where binding of the product TPP to the pre-mRNA of a biosynthetic gene modulates expression. Here we examine and hypothesize on genetic engineering approaches attempting to increase the thiamin content employing knowledge gained with the model plant Arabidopsis thaliana. We will discuss the regulatory steps that need to be taken into consideration and can be used a prerequisite for devising such strategies in crop plants.

  10. Plant Glandular Trichomes as Targets for Breeding or Engineering of Resistance to Herbivores

    Directory of Open Access Journals (Sweden)

    Merijn R. Kant

    2012-12-01

    Full Text Available Glandular trichomes are specialized hairs found on the surface of about 30% of all vascular plants and are responsible for a significant portion of a plant’s secondary chemistry. Glandular trichomes are an important source of essential oils, i.e., natural fragrances or products that can be used by the pharmaceutical industry, although many of these substances have evolved to provide the plant with protection against herbivores and pathogens. The storage compartment of glandular trichomes usually is located on the tip of the hair and is part of the glandular cell, or cells, which are metabolically active. Trichomes and their exudates can be harvested relatively easily, and this has permitted a detailed study of their metabolites, as well as the genes and proteins responsible for them. This knowledge now assists classical breeding programs, as well as targeted genetic engineering, aimed to optimize trichome density and physiology to facilitate customization of essential oil production or to tune biocide activity to enhance crop protection. We will provide an overview of the metabolic diversity found within plant glandular trichomes, with the emphasis on those of the Solanaceae, and of the tools available to manipulate their activities for enhancing the plant’s resistance to pests.

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

    Energy Technology Data Exchange (ETDEWEB)

    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

  12. Analysis of systemic sulfur metabolism in plants using integrated '-omics' strategies.

    Science.gov (United States)

    Hirai, Masami Yokota; Saito, Kazuki

    2008-10-01

    Sulfur (S) assimilation by plants plays a key role in the S cycle in nature, and metabolism of the assimilated S provides various compounds that are useful for animals, including humans. It is important to understand the mechanisms involved in systemic S metabolism in order to improve crop agronomy and the production of dietary and nutraceutical plants. Recent advances in '-omic' studies have provided comprehensive insights into S metabolism. These studies may be regarded as important case studies that provide information on the complicated regulatory mechanisms involved in plant metabolism.

  13. Rice folate enhancement through metabolic engineering has an impact on rice seed metabolism, but does not affect the expression of the endogenous folate biosynthesis genes.

    Science.gov (United States)

    Blancquaert, Dieter; Van Daele, Jeroen; Storozhenko, Sergei; Stove, Christophe; Lambert, Willy; Van Der Straeten, Dominique

    2013-11-01

    Folates are key-players in one-carbon metabolism in all organisms. However, only micro-organisms and plants are able to synthesize folates de novo and humans rely entirely on their diet as a sole folate source. As a consequence, folate deficiency is a global problem. Although different strategies are currently implemented to fight folate deficiency, up until now, all of them have their own drawbacks. As an alternative and complementary means to those classical strategies, folate biofortification of rice by metabolic engineering was successfully achieved a couple of years ago. To gain more insight into folate biosynthesis regulation and the effect of folate enhancement on general rice seed metabolism, a transcriptomic study was conducted in developing transgenic rice seeds, overexpressing 2 genes of the folate biosynthetic pathway. Upon folate enhancement, the expression of 235 genes was significantly altered. Here, we show that rice folate biofortification has an important effect on folate dependent, seed developmental and plant stress response/defense processes, but does not affect the expression of the endogenous folate biosynthesis genes.

  14. From plant surface to plant metabolism: the uncertain fate of foliar-applied nutrients.

    Science.gov (United States)

    Fernández, Victoria; Brown, Patrick H

    2013-01-01

    The application of agrochemical sprays to the aerial parts of crop plants is an important agricultural practice world-wide. While variable effectiveness is often seen in response to foliar treatments, there is abundant evidence showing the beneficial effect of foliar fertilizers in terms of improving the metabolism, quality, and yields of crops. This mini-review is focused on the major bottlenecks associated with the uptake and translocation of foliar-applied nutrient solutions. A better understanding of the complex scenario surrounding the ultimate delivery of foliar-applied nutrients to sink cells and organs is essential for improving the effectiveness and performance of foliar fertilizers.

  15. Fermentative production of branched chain amino acids: a focus on metabolic engineering.

    Science.gov (United States)

    Park, Jin Hwan; Lee, Sang Yup

    2010-01-01

    The branched chain amino acids (BCAAs), L-valine, L-leucine, and L-isoleucine, have recently been attracting much attention as their potential to be applied in various fields, including animal feed additive, cosmetics, and pharmaceuticals, increased. Strategies for developing microbial strains efficiently producing BCAAs are now in transition toward systems metabolic engineering from random mutagenesis. The metabolism and regulatory circuits of BCAA biosynthesis need to be thoroughly understood for designing system-wide metabolic engineering strategies. Here we review the current knowledge on BCAAs including their biosynthetic pathways, regulations, and export and transport systems. Recent advances in the development of BCAA production strains are also reviewed with a particular focus on L-valine production strain. At the end, the general strategies for developing BCAA overproducers by systems metabolic engineering are suggested.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  17. Rapid Optimization of Engineered Metabolic Pathways with Serine Integrase Recombinational Assembly (SIRA).

    Science.gov (United States)

    Merrick, C A; Wardrope, C; Paget, J E; Colloms, S D; Rosser, S J

    2016-01-01

    Metabolic pathway engineering in microbial hosts for heterologous biosynthesis of commodity compounds and fine chemicals offers a cheaper, greener, and more reliable method of production than does chemical synthesis. However, engineering metabolic pathways within a microbe is a complicated process: levels of gene expression, protein stability, enzyme activity, and metabolic flux must be balanced for high productivity without compromising host cell viability. A major rate-limiting step in engineering microbes for optimum biosynthesis of a target compound is DNA assembly, as current methods can be cumbersome and costly. Serine integrase recombinational assembly (SIRA) is a rapid DNA assembly method that utilizes serine integrases, and is particularly applicable to rapid optimization of engineered metabolic pathways. Using six pairs of orthogonal attP and attB sites with different central dinucleotide sequences that follow SIRA design principles, we have demonstrated that ΦC31 integrase can be used to (1) insert a single piece of DNA into a substrate plasmid; (2) assemble three, four, and five DNA parts encoding the enzymes for functional metabolic pathways in a one-pot reaction; (3) generate combinatorial libraries of metabolic pathway constructs with varied ribosome binding site strengths or gene orders in a one-pot reaction; and (4) replace and add DNA parts within a construct through targeted postassembly modification. We explain the mechanism of SIRA and the principles behind designing a SIRA reaction. We also provide protocols for making SIRA reaction components and practical methods for applying SIRA to rapid optimization of metabolic pathways.

  18. Cloning and selection of carotenoid ketolase genes for the engineering of high-yield astaxanthin in plants.

    Science.gov (United States)

    Huang, Junchao; Zhong, Yujuan; Sandmann, Gerhard; Liu, Jin; Chen, Feng

    2012-08-01

    β-Carotene ketolase (BKT) catalyzes the rate-limiting steps for the biosynthesis of astaxanthin. Several bkt genes have been isolated and explored to modify plant carotenoids to astaxanthin with limited success. In this study, five algal BKT cDNAs were isolated and characterized for the engineering of high-yield astaxanthin in plants. The products of the cDNAs showed high similarity in sequence and enzymatic activity of converting β-carotene into canthaxanthin. However, the enzymes exhibited extremely different activities in converting zeaxanthin into astaxanthin. Chlamydomonas reinhardtii BKT showed the highest conversion rate (ca 85%), whereas, Neochloris wimmeri BKT exhibited very poor activity of ketolating zeaxanthin. Expression of C. reinhardtii BKT in tobacco led to a twofold increase of total carotenoids in the leaves with astaxanthin being the predominant. The bkt genes described here provide a valuable resource for metabolic engineering of plants as cell factories for astaxanthin production.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. Enhancing Carbon Fixation by Metabolic Engineering: A Model System of Complex Network Modulation

    Energy Technology Data Exchange (ETDEWEB)

    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

  1. Metabolic engineering of volatile isoprenoids in plants and microbes

    NARCIS (Netherlands)

    Vickers, C.; Bongers, M.; Liu, Q.; Delatte, T.L.; Bouwmeester, H.J.

    2014-01-01

    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

  2. Something Old, Something New: Conserved Enzymes and the Evolution of Novelty in Plant Specialized Metabolism.

    Science.gov (United States)

    Moghe, Gaurav D; Last, Robert L

    2015-11-01

    Plants produce hundreds of thousands of small molecules known as specialized metabolites, many of which are of economic and ecological importance. This remarkable variety is a consequence of the diversity and rapid evolution of specialized metabolic pathways. These novel biosynthetic pathways originate via gene duplication or by functional divergence of existing genes, and they subsequently evolve through selection and/or drift. Studies over the past two decades revealed that diverse specialized metabolic pathways have resulted from the incorporation of primary metabolic enzymes. We discuss examples of enzyme recruitment from primary metabolism and the variety of paths taken by duplicated primary metabolic enzymes toward integration into specialized metabolism. These examples provide insight into processes by which plant specialized metabolic pathways evolve and suggest approaches to discover enzymes of previously uncharacterized metabolic networks.

  3. Something Old, Something New: Conserved Enzymes and the Evolution of Novelty in Plant Specialized Metabolism1

    Science.gov (United States)

    Moghe, Gaurav D.; Last, Robert L.

    2015-01-01

    Plants produce hundreds of thousands of small molecules known as specialized metabolites, many of which are of economic and ecological importance. This remarkable variety is a consequence of the diversity and rapid evolution of specialized metabolic pathways. These novel biosynthetic pathways originate via gene duplication or by functional divergence of existing genes, and they subsequently evolve through selection and/or drift. Studies over the past two decades revealed that diverse specialized metabolic pathways have resulted from the incorporation of primary metabolic enzymes. We discuss examples of enzyme recruitment from primary metabolism and the variety of paths taken by duplicated primary metabolic enzymes toward integration into specialized metabolism. These examples provide insight into processes by which plant specialized metabolic pathways evolve and suggest approaches to discover enzymes of previously uncharacterized metabolic networks. PMID:26276843

  4. Metabolic engineering from a cybernetic perspective. 1. Theoretical preliminaries

    Science.gov (United States)

    Varner; Ramkrishna

    1999-05-01

    The theoretical basis of a cybernetic metabolic network design and analysis framework, which has been subsequently successfully applied to predict system response to genetic alteration, is presented. This conceptual methodology consists of three main branches, namely, a model realization framework, a representation of genetic alteration, and lastly, a metabolic design component. These concepts are introduced as a series of postulates that describe the basic tenets of the approach. Each branch is discussed in turn, starting with the cybernetic representation of arbitrarily complex metabolic networks. A set of postulates is put forth that affords the modular construction of cybernetic models of metabolic networks using as a base a library of elementary pathways. This is followed by a discussion of the representation of genetic alterations within the cybernetic framework. It is postulated that the objective of the base network and the altered system are identical (at least on the time scale required for the organism to "learn" new objectives). This implies, with respect to resource allocation, that the base network and its genetically altered counterpart may still be treated as optimal systems; however, the set of competing physiological choices open to the altered network expands or contracts depending upon the nature of the genetic perturbation. Lastly, to add a predictive design aspect to the methodology, we present a set of postulates that outline the application of metabolic control analysis to cybernetic model systems. We postulate that sensitivity coefficients computed from a cybernetic model, although still local in scope, have the added benefit of a systematic representation of regulatory function as described by the cybernetic variables. Thus, information gained from sensitivity measurements stemming from a cybernetic model include the explicit input of metabolic regulation, a component that is lacking in a purely kinetic representation of metabolic function

  5. 10 CFR Appendix S to Part 50 - Earthquake Engineering Criteria for Nuclear Power Plants

    Science.gov (United States)

    2010-01-01

    ... 10 Energy 1 2010-01-01 2010-01-01 false Earthquake Engineering Criteria for Nuclear Power Plants S... FACILITIES Pt. 50, App. S Appendix S to Part 50—Earthquake Engineering Criteria for Nuclear Power Plants..., as specified in § 50.54(ff), nuclear power plants that have implemented the earthquake...

  6. Metabolic engineering of the iodine content in Arabidopsis

    OpenAIRE

    Martina Landini; Silvia Gonzali; Claudia Kiferle; Massimo Tonacchera; Patrizia Agretti; Antonio Dimida; Paolo Vitti; Amedeo Alpi; Aldo Pinchera; Pierdomenico Perata

    2012-01-01

    Plants are a poor source of iodine, an essential micronutrient for human health. Several attempts of iodine biofortification of crops have been carried out, but the scarce knowledge on the physiology of iodine in plants makes results often contradictory and not generalizable. In this work, we used a molecular approach to investigate how the ability of a plant to accumulate iodine can be influenced by different mechanisms. In particular, we demonstrated that the iodine content in Arabidopsis t...

  7. Phenolic metabolism of Matricaria chamomilla plants exposed to nickel.

    Science.gov (United States)

    Kovácik, Jozef; Klejdus, Borivoj; Backor, Martin

    2009-09-01

    We examined accumulation of phenolic acids, total soluble phenolics and flavonoids, and activities of phenolic metabolism-related enzymes (shikimate dehydrogenase (SKDH), phenylalanine ammonia-lyase (PAL), cinnamyl alcohol dehydrogenase (CAD), polyphenol oxidase (PPO)) in Matricaria chamomilla plants exposed to 3, 60 and 120 microM of nickel (Ni) for 10 days. Ni showed low toxicity as indicated by unaltered content of total soluble phenolics in the leaf rosettes. In the roots, the effects of Ni were more visible, including increased total phenolics and PAL activity, but a decrease in PPO activity was observed. CAD activity was not affected by any of the Ni concentrations. Cinnamic acid derivatives were affected more than benzoic acid derivatives. Accumulation of chlorogenic acid, an important antioxidant compound, was enhanced by Ni treatment (ca. 4-fold in 120 microM Ni). Accumulation of protocatechuic acid, a phenol with high chelating strength, even decreased in the leaf rosettes. These observations are discussed in connection to antioxidative properties of phenolic metabolites and previously tested metals (cadmium and copper).

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

    Science.gov (United States)

    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.

  9. [Advances in studies on growth metabolism and response mechanisms of medicinal plants under drought stress].

    Science.gov (United States)

    Si, Can; Zhang, Jun-Yi; Xu, Hu-Chao

    2014-07-01

    Drought stress exerts a considerable effect on growth, physiology and secondary metabolisms of the medicinal plants. It could inhabit the growth of the medicinal plants but promote secretion of secondary metabolites. Other researches indicated that the medicinal plants could depend on the ABA signaling pathway and secreting osmotic substances to resist the drought stress and reduce the damage by it. The article concludes the changes in growth, physiology, secondary metabolisms and response mechanisms of medicinal plants to drought stress that provides a theoretical basis for exploring the relationship between medicinal plants and drought stress.

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

    DEFF Research Database (Denmark)

    Chen, Xiao

    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......-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...

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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

  14. Systems biology as an integrated platform for bioinformatics, systems synthetic biology, and systems metabolic engineering.

    Science.gov (United States)

    Chen, Bor-Sen; Wu, Chia-Chou

    2013-10-11

    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.

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

    Science.gov (United States)

    Guo, Weihua; Sheng, Jiayuan; Feng, Xueyang

    2017-01-01

    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.

  16. Tracer methods for investigating biosynthetic pathways and the metabolism of bioactive substances in plants. [Herbicides; Plant growth regulators

    Energy Technology Data Exchange (ETDEWEB)

    Schuette, H.R. (Akademie der Wissenschaften der DDR, Halle/Saale. Inst. fuer Biochemie der Pflanzen)

    1984-03-01

    Proceeding from the general terms of investigating the courses of reactions in plants by means of tracer methods, problems and possibilities of the methods are discussed on the basis of examples referring in particular to double labelling techniques and to the determination of the distribution of radioactivity in the resulting products. Examples of herbicides and plant growth regulators are used for describing metabolism studies.

  17. Metabolic engineering of essential oil yield and composition in mint by altering expression of deoxyxylulose phosphate reductoisomerase and menthofuran synthase.

    Science.gov (United States)

    Mahmoud, S S; Croteau, R B

    2001-07-17

    Peppermint (Mentha x piperita L.) was independently transformed with a homologous sense version of the 1-deoxy-d-xylulose-5-phosphate reductoisomerase cDNA and with a homologous antisense version of the menthofuran synthase cDNA, both driven by the CaMV 35S promoter. Two groups of transgenic plants were regenerated in the reductoisomerase experiments, one of which remained normal in appearance and development; another was deficient in chlorophyll production and grew slowly. Transgenic plants of normal appearance and growth habit expressed the reductoisomerase transgene strongly and constitutively, as determined by RNA blot analysis and direct enzyme assay, and these plants accumulated substantially more essential oil (about 50% yield increase) without change in monoterpene composition compared with wild-type. Chlorophyll-deficient plants did not afford detectable reductoisomerase mRNA or enzyme activity and yielded less essential oil than did wild-type plants, indicating cosuppression of the reductoisomerase gene. Plants transformed with the antisense version of the menthofuran synthase cDNA were normal in appearance but produced less than half of this undesirable monoterpene oil component than did wild-type mint grown under unstressed or stressed conditions. These experiments demonstrate that essential oil quantity and quality can be regulated by metabolic engineering. Thus, alteration of the committed step of the mevalonate-independent pathway for supply of terpenoid precursors improves flux through the pathway that leads to increased monoterpene production, and antisense manipulation of a selected downstream monoterpene biosynthetic step leads to improved oil composition.

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

    Directory of Open Access Journals (Sweden)

    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.

  19. Modulation of plant growth and metabolism in cadmium-enriched environments.

    Science.gov (United States)

    Qadir, Shaista; Jamshieed, Sumiya; Rasool, Saiema; Ashraf, Muhammad; Akram, Nudrat Aisha; Ahmad, Parvaiz

    2014-01-01

    Cadmium (Cd) is a water soluble metal pollutant that is not essential to plant growth.It has attracted attention from soil scientists and plant nutritionists in recent years because of its toxicity and mobility in the soil-plant continuum. Even low levels of Cd (0.1-1 J.!M) cause adverse effects on plant growth and metabolism. Cadmium is known to trigger the synthesis of reactive oxygen species, hinder utilization, uptake and transport of essential nutrients and water, and modify photosynthetic machinery,thereby resulting in plant tissue death. Although the effects of Cd are dose- as well as plant species-dependent, some plants show Cd tolerance through a wide range of cellular responses. Such tolerance results from synthesis of osmolytes,generation of enzymatic and non-enzymatic antioxidants and metal-detoxifying peptides, changes in gene expression, and metal ion homeostasis and compartmentalization of ligand-metal complexes. Cd toxicity in plants produces effects on chlorophyllbio synthesis, reduces photosynthesis, and upsets plant water relations and hormonal and/or nutritional balances. All of these effects on plants and on plant metabolism ultimately reduce growth and productivity.In this review, we describe the extent to which Cd affects underlying metabolic processes in plants and how such altered processes affect plant growth. We review the sources of Cd contamination, its uptake, transportation and bioavailability and accumulation in plants, and its antagonistic and synergistic effects with other metals and compounds. We further address the effects of Cd on plant genetics and metabolism,and how plants respond to mitigate the adverse effects of Cd exposure, as well as strategies(e.g., plant breeding) that can reduce the impact of Cd contamination on plants.

  20. Small molecules that interact with RNA: riboswitch-based gene control and its involvement in metabolic regulation in plants and algae.

    Science.gov (United States)

    Bocobza, Samuel E; Aharoni, Asaph

    2014-08-01

    Riboswitches are RNA elements that bind small molecules and in turn regulate gene expression. This mechanism allows the cell to sense the intracellular concentration of these small molecules. A particular riboswitch typically regulates its adjacent gene by altering the transcription, the translation or the splicing of this gene. Recently, a riboswitch that binds thiamin pyrophosphate (TPP) was characterized and found to regulate thiamin biosynthesis in plants and algae. Furthermore, it appears that this element is an essential regulator of primary metabolism in plants. Manipulation of endogenous riboswitch activity resulted in metabolic phenotypes that underlined the role of these elements and their ligands in preserving metabolic homeostasis. This situation supports the hypothesis that riboswitches could be remnants of the most ancient metabolic regulators. Here, we review the mode of action of the plant and algal TPP riboswitch and its relevance to the metabolic network. We also discuss the potential engineering of riboswitches as metabolite sensors in plants and platforms for gene control. Whether additional such RNA-based mechanisms exist in plants and in algae is still an open question, yet, the importance of these elements to metabolic regulation is beyond doubt.

  1. Combinatorial metabolic engineering of Saccharomyces cerevisiae for terminal alkene production.

    Science.gov (United States)

    Chen, Binbin; Lee, Dong-Yup; Chang, Matthew Wook

    2015-09-01

    Biological production of terminal alkenes has garnered a significant interest due to their industrial applications such as lubricants, detergents and fuels. Here, we engineered the yeast Saccharomyces cerevisiae to produce terminal alkenes via a one-step fatty acid decarboxylation pathway and improved the alkene production using combinatorial engineering strategies. In brief, we first characterized eight fatty acid decarboxylases to enable and enhance alkene production. We then increased the production titer 7-fold by improving the availability of the precursor fatty acids. We additionally increased the titer about 5-fold through genetic cofactor engineering and gene expression tuning in rich medium. Lastly, we further improved the titer 1.8-fold to 3.7 mg/L by optimizing the culturing conditions in bioreactors. This study represents the first report of terminal alkene biosynthesis in S. cerevisiae, and the abovementioned combinatorial engineering approaches collectively increased the titer 67.4-fold. We envision that these approaches could provide insights into devising engineering strategies to improve the production of fatty acid-derived biochemicals in S. cerevisiae.

  2. Metabolic engineering of the iodine content in Arabidopsis.

    Science.gov (United States)

    Landini, Martina; Gonzali, Silvia; Kiferle, Claudia; Tonacchera, Massimo; Agretti, Patrizia; Dimida, Antonio; Vitti, Paolo; Alpi, Amedeo; Pinchera, Aldo; Perata, Pierdomenico

    2012-01-01

    Plants are a poor source of iodine, an essential micronutrient for human health. Several attempts of iodine biofortification of crops have been carried out, but the scarce knowledge on the physiology of iodine in plants makes results often contradictory and not generalizable. In this work, we used a molecular approach to investigate how the ability of a plant to accumulate iodine can be influenced by different mechanisms. In particular, we demonstrated that the iodine content in Arabidopsis thaliana can be increased either by facilitating its uptake with the overexpression of the human sodium-iodide symporter (NIS) or through the reduction of its volatilization by knocking-out HOL-1, a halide methyltransferase. Our experiments show that the iodine content in plants results from a balance between intake and retention. A correct manipulation of this mechanism could improve iodine biofortification of crops and prevent the release of the ozone layer-threatening methyl iodide into the atmosphere.

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

    NARCIS (Netherlands)

    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 con

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

    NARCIS (Netherlands)

    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

  5. Engineering metabolic highways in Lactococci and other lactic acid bacteria

    NARCIS (Netherlands)

    Vos, de W.M.; Hugenholtz, J.

    2004-01-01

    Lactic acid bacteria (LAB) are widely used in industrial food fermentations and are receiving increased attention for use as cell factories for the production of food and pharmaceutical products. Glycolytic conversion of sugars into lactic acid is the main metabolic highway in these Gram-positive

  6. Engineering metabolic highways in Lactococci and other lactic acid bacteria

    NARCIS (Netherlands)

    Vos, de W.M.; Hugenholtz, J.

    2004-01-01

    Lactic acid bacteria (LAB) are widely used in industrial food fermentations and are receiving increased attention for use as cell factories for the production of food and pharmaceutical products. Glycolytic conversion of sugars into lactic acid is the main metabolic highway in these Gram-positive ba

  7. Production of isopropanol by metabolically engineered Escherichia coli.

    Science.gov (United States)

    Jojima, Toru; Inui, Masayuki; Yukawa, Hideaki

    2008-01-01

    A genetically engineered strain of Escherichia coli JM109 harboring the isopropanol-producing pathway consisting of five genes encoding four enzymes, thiolase, coenzyme A (CoA) transferase, acetoacetate decarboxylase from Clostridium acetobutylicum ATCC 824, and primary-secondary alcohol dehydrogenase from C. beijerinckii NRRL B593, produced up to 227 mM of isopropanol from glucose under aerobic fed-batch culture conditions. Acetate production by the engineered strain was approximately one sixth that produced by a control E. coli strain bearing an expression vector without the clostridial genes. These results demonstrate a functional isopropanol-producing pathway in E. coli and consequently carbon flux from acetyl-CoA directed to isopropanol instead of acetate. This is the first report on isopropanol production by genetically engineered microorganism under aerobic culture conditions.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  9. Microbial isoprenoid production: an example of green chemistry through metabolic engineering.

    Science.gov (United States)

    Maury, Jérôme; Asadollahi, Mohammad A; Møller, Kasper; Clark, Anthony; Nielsen, Jens

    2005-01-01

    Saving energy, cost efficiency, producing less waste, improving the biodegradability of products, potential for producing novel and complex molecules with improved properties, and reducing the dependency on fossil fuels as raw materials are the main advantages of using biotechnological processes to produce chemicals. Such processes are often referred to as green chemistry or white biotechnology. Metabolic engineering, which permits the rational design of cell factories using directed genetic modifications, is an indispensable strategy for expanding green chemistry. In this chapter, the benefits of using metabolic engineering approaches for the development of green chemistry are illustrated by the recent advances in microbial production of isoprenoids, a diverse and important group of natural compounds with numerous existing and potential commercial applications. Accumulated knowledge on the metabolic pathways leading to the synthesis of the principal precursors of isoprenoids is reviewed, and recent investigations into isoprenoid production using engineered cell factories are described.

  10. Optimization of a yeast RNA interference system for controlling gene expression and enabling rapid metabolic engineering.

    Science.gov (United States)

    Crook, Nathan C; Schmitz, Alexander C; Alper, Hal S

    2014-05-16

    Reduction of endogenous gene expression is a fundamental operation of metabolic engineering, yet current methods for gene knockdown (i.e., genome editing) remain laborious and slow, especially in yeast. In contrast, RNA interference allows facile and tunable gene knockdown via a simple plasmid transformation step, enabling metabolic engineers to rapidly prototype knockdown strategies in multiple strains before expending significant cost to undertake genome editing. Although RNAi is naturally present in a myriad of eukaryotes, it has only been recently implemented in Saccharomyces cerevisiae as a heterologous pathway and so has not yet been optimized as a metabolic engineering tool. In this study, we elucidate a set of design principles for the construction of hairpin RNA expression cassettes in yeast and implement RNA interference to quickly identify routes for improvement of itaconic acid production in this organism. The approach developed here enables rapid prototyping of knockdown strategies and thus accelerates and reduces the cost of the design-build-test cycle in yeast.

  11. 2009 Plant Lipids: Structure, Metabolism & Function Gordon Research Conference - February 1- 6 ,2009

    Energy Technology Data Exchange (ETDEWEB)

    Kent D. Chapman

    2009-02-06

    The Gordon Research Conference on 'Plant Lipids: Structure, Metabolism and Function' has been instituted to accelerate research productivity in the field of plant lipids. This conference will facilitate wide dissemination of research breakthroughs, support recruitment of young scientists to the field of plant lipid metabolism and encourage broad participation of the plant lipid community in guiding future directions for research in plant lipids. This conference will build upon the strengths of the successful, previous biannual meetings of the National Plant Lipid Cooperative (www.plantlipids.org) that began in 1993, but will reflect a broader scope of topics to include the biochemistry, cell biology, metabolic regulation, and signaling functions of plant acyl lipids. Most importantly, this conference also will serve as a physical focal point for the interaction of the plant lipid research community. Applications to attend this conference will be open to all researchers interested in plant lipids and will provide a venue for the presentation of the latest research results, networking opportunities for young scientists, and a forum for the development and exchange of useful lipid resources and new ideas. By bringing together senior- and junior-level scientists involved in plant lipid metabolism, a broad range of insights will be shared and the community of plant lipid researchers will function more as a network of vested partners. This is important for the vitality of the research community and for the perceived value that will encourage conference attendance into the future.

  12. Variation of glucosinolates and quinone reductase activity among different varieties of Chinese kale and improvement of glucoraphanin by metabolic engineering.

    Science.gov (United States)

    Qian, Hongmei; Sun, Bo; Miao, Huiying; Cai, Congxi; Xu, Chaojiong; Wang, Qiaomei

    2015-02-01

    The variation of glucosinolates and quinone reductase (QR) activity in fourteen varieties of Chinese kale (Brassica oleracea var. alboglabra Bailey) was investigated in the present study. Results showed that gluconapin (GNA), instead of glucoraphanin (GRA), was the most predominant glucosinolate in all varieties, and QR activity was remarkably positively correlated with the glucoraphanin level. AOP2, a tandem 2-oxoglutarate-dependent dioxygenase, catalyzes the conversion of glucoraphanin to gluconapin in glucosinolate biosynthesis. Here, antisense AOP2 was transformed into Gailan-04, the variety with the highest gluconapin content and ratio of GNA/GRA. The glucoraphanin content and corresponding QR activity were notably increased in transgenic plants, while no significant difference at the level of other main nutritional compounds (total phenolics, vitamin C, carotenoids and chlorophyll) was observed between the transgenic lines and the wide-type plants. Taken together, metabolic engineering is a good practice for improvement of glucoraphanin in Chinese kale.

  13. Metabolic engineering of microbial pathways for advanced biofuels production.

    Science.gov (United States)

    Zhang, Fuzhong; Rodriguez, Sarah; Keasling, Jay D

    2011-12-01

    Production of biofuels from renewable resources such as cellulosic biomass provides a source of liquid transportation fuel to replace petroleum-based fuels. This endeavor requires the conversion of cellulosic biomass into simple sugars, and the conversion of simple sugars into biofuels. Recently, microorganisms have been engineered to convert simple sugars into several types of biofuels, such as alcohols, fatty acid alkyl esters, alkanes, and terpenes, with high titers and yields. Here, we review recently engineered biosynthetic pathways from the well-characterized microorganisms Escherichia coli and Saccharomyces cerevisiae for the production of several advanced biofuels. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. (Im)Perfect robustness and adaptation of metabolic networks subject to metabolic and gene-expression regulation: marrying control engineering with metabolic control analysis.

    Science.gov (United States)

    He, Fei; Fromion, Vincent; Westerhoff, Hans V

    2013-11-21

    Metabolic control analysis (MCA) and supply-demand theory have led to appreciable understanding of the systems properties of metabolic networks that are subject exclusively to metabolic regulation. Supply-demand theory has not yet considered gene-expression regulation explicitly whilst a variant of MCA, i.e. Hierarchical Control Analysis (HCA), has done so. Existing analyses based on control engineering approaches have not been very explicit about whether metabolic or gene-expression regulation would be involved, but designed different ways in which regulation could be organized, with the potential of causing adaptation to be perfect. This study integrates control engineering and classical MCA augmented with supply-demand theory and HCA. Because gene-expression regulation involves time integration, it is identified as a natural instantiation of the 'integral control' (or near integral control) known in control engineering. This study then focuses on robustness against and adaptation to perturbations of process activities in the network, which could result from environmental perturbations, mutations or slow noise. It is shown however that this type of 'integral control' should rarely be expected to lead to the 'perfect adaptation': although the gene-expression regulation increases the robustness of important metabolite concentrations, it rarely makes them infinitely robust. For perfect adaptation to occur, the protein degradation reactions should be zero order in the concentration of the protein, which may be rare biologically for cells growing steadily. A proposed new framework integrating the methodologies of control engineering and metabolic and hierarchical control analysis, improves the understanding of biological systems that are regulated both metabolically and by gene expression. In particular, the new approach enables one to address the issue whether the intracellular biochemical networks that have been and are being identified by genomics and systems

  15. A survey of mortality at two automotive engine manufacturing plants.

    Science.gov (United States)

    Park, R M; Mirer, F E

    1996-12-01

    Mortality at two engine plants was analyzed using proportional mortality and logistic regression models of mortality odds ratios to expand previous observations of increased cancers of the stomach, pancreas, and bladder, and cirrhosis of the liver among workers exposed to machining fluids. Causes of death and work histories were available for 1,870 decendents. There was a significant excess of deaths coded as diabetes for white men in both plants (PMR = 25/16.7 = 1.5, 95% CI = 1.02, 2.20), and a deficit of respiratory diseases. Black men had fewer than expected diabetes deaths and more emphysema deaths. Elevated PMRs for cancers of the stomach, pancreas, prostate, bladder, and kidney were not statistically significant in plantwide populations. However, stomach cancer mortality increased with duration in camshaft and crankshaft production at Plant 1 (OR = 5.1, 95% CI = 1.6, 17; at mean duration of exposed cases), and among tool room workers (OR = 6.3, 95% CI = 1.3, 31), but these results were based on five cases. Nitrosamines were probably present in camshaft and crankshaft grinding at Plant 1. Pancreas cancer risk increased among workers at both plants ever employed in inspection (OR = 2.5, 16), in machining with straight oil (OR = 3.6, 95% CI = 1.04, 12), or in skilled trades (OR = 2.9, 95% CI = 1.1, 7.5). Lung cancer increased in cylinder head machining (OR = 3.9, 95% CI = 1.4, 11), millwright work (OR = 3.8, 95% CI = 1.6, 9.0), and in Plant 2 generally (OR = 1.45, 95% CI = 0.97, 2.2). Potential lung carcinogens included heat treatment emissions, chlorinated oils, and coal tar fumes (millwrights). Bladder cancer increased with duration among workers grinding in straight oil MF (OR = 3.0, 95% CI = 1.15, 7.8) and in machining/heat-treat operations (OR = 2.9, 95% CI = 1.14, 7.2).

  16. Perspectives on the interactions between metabolism, redox, and epigenetics in plants.

    Science.gov (United States)

    Shen, Yuan; Issakidis-Bourguet, Emmanuelle; Zhou, Dao-Xiu

    2016-10-01

    Epigenetic modifications of chromatin usually involve consumption of key metabolites and redox-active molecules. Primary metabolic flux and cellular redox states control the activity of enzymes involved in chromatin modifications, such as DNA methylation, histone acetylation, and histone methylation, which in turn regulate gene expression and/or enzymatic activity of specific metabolic and redox pathways. Thus, coordination of metabolism and epigenetic regulation of gene expression is critical to control growth and development in response to the cellular environment. Much has been learned from animal and yeast cells with regard to the interplay between metabolism and epigenetic regulation, and now the metabolic control of epigenetic pathways in plants is an increasing area of study. Epigenetic mechanisms are largely similar between plant and mammalian cells, but plants display very important differences in both metabolism and metabolic/redox signaling pathways. In this review, we summarize recent developments in the field and discuss perspectives of studying interactions between plant epigenetic and metabolism/redox systems, which are essential for plant adaptation to environmental conditions.

  17. Plant oxidosqualene metabolism: cycloartenol synthase-dependent sterol biosynthesis in Nicotiana benthamiana.

    Science.gov (United States)

    Gas-Pascual, Elisabet; Berna, Anne; Bach, Thomas J; Schaller, Hubert

    2014-01-01

    The plant sterol pathway exhibits a major biosynthetic difference as compared with that of metazoans. The committed sterol precursor is the pentacyclic cycloartenol (9β,19-cyclolanost-24-en-3β-ol) and not lanosterol (lanosta-8,24-dien-3β-ol), as it was shown in the late sixties. However, plant genome mining over the last years revealed the general presence of lanosterol synthases encoding sequences (LAS1) in the oxidosqualene cyclase repertoire, in addition to cycloartenol synthases (CAS1) and to non-steroidal triterpene synthases that contribute to the metabolic diversity of C30H50O compounds on earth. Furthermore, plant LAS1 proteins have been unambiguously identified by peptidic signatures and by their capacity to complement the yeast lanosterol synthase deficiency. A dual pathway for the synthesis of sterols through lanosterol and cycloartenol was reported in the model Arabidopsis thaliana, though the contribution of a lanosterol pathway to the production of 24-alkyl-Δ(5)-sterols was quite marginal (Ohyama et al. (2009) PNAS 106, 725). To investigate further the physiological relevance of CAS1 and LAS1 genes in plants, we have silenced their expression in Nicotiana benthamiana. We used virus induced gene silencing (VIGS) based on gene specific sequences from a Nicotiana tabacum CAS1 or derived from the solgenomics initiative (http://solgenomics.net/) to challenge the respective roles of CAS1 and LAS1. In this report, we show a CAS1-specific functional sterol pathway in engineered yeast, and a strict dependence on CAS1 of tobacco sterol biosynthesis.

  18. Plant Oxidosqualene Metabolism: Cycloartenol Synthase–Dependent Sterol Biosynthesis in Nicotiana benthamiana

    Science.gov (United States)

    Gas-Pascual, Elisabet; Berna, Anne; Bach, Thomas J.; Schaller, Hubert

    2014-01-01

    The plant sterol pathway exhibits a major biosynthetic difference as compared with that of metazoans. The committed sterol precursor is the pentacyclic cycloartenol (9β,19-cyclolanost-24-en-3β-ol) and not lanosterol (lanosta-8,24-dien-3β-ol), as it was shown in the late sixties. However, plant genome mining over the last years revealed the general presence of lanosterol synthases encoding sequences (LAS1) in the oxidosqualene cyclase repertoire, in addition to cycloartenol synthases (CAS1) and to non-steroidal triterpene synthases that contribute to the metabolic diversity of C30H50O compounds on earth. Furthermore, plant LAS1 proteins have been unambiguously identified by peptidic signatures and by their capacity to complement the yeast lanosterol synthase deficiency. A dual pathway for the synthesis of sterols through lanosterol and cycloartenol was reported in the model Arabidopsis thaliana, though the contribution of a lanosterol pathway to the production of 24-alkyl-Δ5-sterols was quite marginal (Ohyama et al. (2009) PNAS 106, 725). To investigate further the physiological relevance of CAS1 and LAS1 genes in plants, we have silenced their expression in Nicotiana benthamiana. We used virus induced gene silencing (VIGS) based on gene specific sequences from a Nicotiana tabacum CAS1 or derived from the solgenomics initiative (http://solgenomics.net/) to challenge the respective roles of CAS1 and LAS1. In this report, we show a CAS1-specific functional sterol pathway in engineered yeast, and a strict dependence on CAS1 of tobacco sterol biosynthesis. PMID:25343375

  19. Plutonium Finishing Plant. Interim plutonium stabilization engineering study

    Energy Technology Data Exchange (ETDEWEB)

    Sevigny, G.J.; Gallucci, R.H.; Garrett, S.M.K.; Geeting, J.G.H.; Goheen, R.S.; Molton, P.M.; Templeton, K.J.; Villegas, A.J. [Pacific Northwest Lab., Richland, WA (United States); Nass, R. [Nuclear Fuel Services, Inc. (United States)

    1995-08-01

    This report provides the results of an engineering study that evaluated the available technologies for stabilizing the plutonium stored at the Plutonium Finishing Plant located at the hanford Site in southeastern Washington. Further processing of the plutonium may be required to prepare the plutonium for interim (<50 years) storage. Specifically this document provides the current plutonium inventory and characterization, the initial screening process, and the process descriptions and flowsheets of the technologies that passed the initial screening. The conclusions and recommendations also are provided. The information contained in this report will be used to assist in the preparation of the environmental impact statement and to help decision makers determine which is the preferred technology to process the plutonium for interim storage.

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

    Science.gov (United States)

    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.

  1. Power plant technology 2014. Strategies, systems engineering and operation; Kraftwerkstechnik 2014. Strategien, Anlagentechnik und Betrieb

    Energy Technology Data Exchange (ETDEWEB)

    Beckmann, Michael; Hurtado, Antonio

    2014-07-01

    The book on power plant technology 2014 (strategies, systems engineering and operation) covers the following issues: Climate, politics and economy; wind power; fossil-fuel power plants, flexible power plants - plant operation, flexible power plants- materials, materials for energy technology, fuel feed and incineration, modeling of the water-vapor-circuit, corrosion, deposits and cleaning, vapor turbines, GUD power plants, fluidized bed combustion, energetic biomass use, combined heat and power generation and decentralized units, storage facilities, emissions - mitigation and measuring techniques.

  2. Camelina sativa: An ideal platform for the metabolic engineering and field production of industrial lipids.

    Science.gov (United States)

    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.

  3. Toward glycerol biorefinery: metabolic engineering for the production of biofuels and chemicals from glycerol.

    Science.gov (United States)

    Chen, Zhen; Liu, Dehua

    2016-01-01

    As an inevitable by-product of the biofuel industry, glycerol is becoming an attractive feedstock for biorefinery due to its abundance, low price and high degree of reduction. Converting crude glycerol into value-added products is important to increase the economic viability of the biofuel industry. Metabolic engineering of industrial strains to improve its performance and to enlarge the product spectrum of glycerol biotransformation process is highly important toward glycerol biorefinery. This review focuses on recent metabolic engineering efforts as well as challenges involved in the utilization of glycerol as feedstock for the production of fuels and chemicals, especially for the production of diols, organic acids and biofuels.

  4. Accessing Nature’s diversity through metabolic engineering and synthetic biology [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Jason R. King

    2016-03-01

    Full Text Available In this perspective, we highlight recent examples and trends in metabolic engineering and synthetic biology that demonstrate the synthetic potential of enzyme and pathway engineering for natural product discovery. In doing so, we introduce natural paradigms of secondary metabolism whereby simple carbon substrates are combined into complex molecules through “scaffold diversification”, and subsequent “derivatization” of these scaffolds is used to synthesize distinct complex natural products. We provide examples in which modern pathway engineering efforts including combinatorial biosynthesis and biological retrosynthesis can be coupled to directed enzyme evolution and rational enzyme engineering to allow access to the “privileged” chemical space of natural products in industry-proven microbes. Finally, we forecast the potential to produce natural product-like discovery platforms in biological systems that are amenable to single-step discovery, validation, and synthesis for streamlined discovery and production of biologically active agents.

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

    Science.gov (United States)

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  7. Towards Increasing State Support Efficiency of Mechanical Engineering Plants

    Directory of Open Access Journals (Sweden)

    L. I. Kushnarev

    2016-01-01

    Full Text Available The article discusses a hot topic concerning the organization and implementation of a proprietary method of the technical service aimed at solving the problems of technological modernization of manufacturing facilities, machines and equipment and more efficient use of financial means allocated by the government of the Russian Federation for these purposes through increasing a level of competitiveness thanks to growing quality and reliability of equipment. Based on the analysis the paper gives an accent on the factors that are capable to provide a rate of return sufficient for additional, along with the state support, investments of innovative development of the enterprise because of technological modernization of production. Such an approach to technological modernization of mechanical engineering plants allows 3–5 times enhancement of their development rates provided that a careful scheduling of activities is based on the organizational and technological design. The researches have allowed us to formulate the proposals for the project implementation on innovative development of mechanical engineering.

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

    OpenAIRE

    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...

  9. Photorespiration: current status and approaches for metabolic engineering.

    Science.gov (United States)

    Maurino, Veronica G; Peterhansel, Christoph

    2010-06-01

    Photorespiration results from the oxygenase reaction catalysed by ribulose-1,5-bisphosphate carboxylase/oxygenase and serves as a carbon recovery system. It comprises enzymatic reactions distributed in chloroplasts, peroxisomes and mitochondria. The recent discovery of a cytosolic bypass and the requirement of complex formation between some photorespiratory proteins added additional levels of complexity to the known pathway. Photorespiration may have evolved in both, C(3) and C(4) plants, to prevent an accumulation of toxic levels of glycolate. Moreover, it is suggested that photorespiration evolved in cyanobacteria before the origin of chloroplasts. Synthetic detours, reminiscent of secondary photorespiratory pathways naturally occurring in cyanobacteria, were installed in Arabidopsis thaliana to bypass photorespiration. An enrichment of CO(2) in the chloroplast and positive effects on plant growth raised the question why these pathways have been lost from higher plants.

  10. Dynamic control of gene expression in Saccharomyces cerevisiae engineered for the production of plant sesquitepene α-santalene in a fed-batch mode

    DEFF Research Database (Denmark)

    Scalcinati, Gionata; Knuf, Christoph; Partow, Siavash;

    2012-01-01

    Microbial cells engineered for efficient production of plant sesquiterpenes may allow for sustainable and scalable production of these compounds that can be used as e.g. perfumes and pharmaceuticals. Here, for the first time a Saccharomyces cerevisiae strain capable of producing high levels of α......-santalene, the precursor of a commercially interesting compound, was constructed through a rationally designed metabolic engineering approach. Optimal sesquiterpene production was obtained by modulating the expression of one of the key metabolic steps of the mevalonate (MVA) pathway, squalene synthase (Erg9). To couple...

  11. The molecular components of phospho- and glycolipid metabolism in plant cell membranes under the phosphorus deficiency

    Directory of Open Access Journals (Sweden)

    Svietlova N. B.

    2012-01-01

    Full Text Available One of the aspects of molecular regulation of phosphorus metabolism in plants, the lipid components of membrane structures, has been reviewed. The refocusing of phosphoand glycolipid metabolism is an indicator of phosphorus accessibility in plants. The compensatory mechanisms of substitution of phospholipids with non-phosphorus containing glycolipids in membranes, allow plants to adapt to the phosphate (Pi starvation. Phospholipids are the reserve pool of cellular phosphorus at reutilization of ions in the donor-acceptor system of plants. The mechanisms of transcriptional regulation of genes involved in the synthesis of phospholipids and glycolipids under Pi deficit have been analyzed.

  12. Plant Resistance to Virus Diseases through Genetic Engineering: Can a Similar Approach Control Plant-parasitic Nematodes?

    OpenAIRE

    Reimann-Philipp, Ulrich; Beachy, Roger N.

    1993-01-01

    Genetically engineered resistance against plant virus diseases has been achieved by transforming plants with gene constructs that encode viral sequences. Several successful field trials of virus-resistant transgenic plants have been carried out. Specific features of virus infection make it possible to interfere with different steps of the infection and disease cycle by accumulating products of chimeric genes introduced into transgenic plants. In this paper we describe the most common methods ...

  13. An outbreak of extrinsic alveolitis at a car engine plant.

    Science.gov (United States)

    Dawkins, Paul; Robertson, Alastair; Robertson, Wendy; Moore, Vicky; Reynolds, John; Langman, Gerald; Robinson, Edward; Harris-Roberts, Joanne; Crook, Brian; Burge, Sherwood

    2006-12-01

    Twelve workers from a car engine-manufacturing plant presented with extrinsic allergic alveolitis (EAA), with heterogeneous clinical, radiological and pathological findings. They were exposed to metalworking fluids (MWF) that cooled, lubricated and cleaned the machines. They were characterized by history, examination, lung function testing, radiology, bronchoscopic lavage, lung biopsy and serology. Sera were tested for precipitins to a crude extract of used MWF and to reference cultures of bacteria suspected to be implicated. All were males and none were current smokers. All had dyspnoea, many had weight loss and cough, but only half had influenza-like symptoms. Only half had auscultatory crackles. Five had peak flow variability, four with an occupational component. There was overall restrictive spirometry, decreased lung volumes and reduced gas transfers. Ten had radiological evidence of interstitial lung disease. Seven (of eight) had lymphocytosis on bronchial lavage, including the two with inconclusive radiology. Seven (of 11) had lung biopsies showing inflammatory infiltrates, two with fibrosis and one with granulomas. Three (of 11) had strong positive precipitins to an extract of the used MWF from the plant. Molecular biological analysis of the MWF revealed Acinetobacter and Ochrobactrum. Precipitins to Acinetobacter were detected in seven of 11 workers tested (and four of 11 control workers). Precipitins to Ochrobactrum were detected in three of 11 workers tested (and three of 11 control workers). This is the largest series reported in Europe of EAA due to an aerosol of microbiologically contaminated MWF in heavy manufacturing industry.

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

    Science.gov (United States)

    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.

  15. Fatty Acid Biosynthesis Revisited: Structure Elucidation and Metabolic Engineering

    Science.gov (United States)

    Beld, Joris; Lee, D. John

    2014-01-01

    Fatty acids are primary metabolites synthesized by complex, elegant, and essential biosynthetic machinery. Fatty acid synthases resemble an iterative assembly line, with an acyl carrier protein conveying the growing fatty acid to necessary enzymatic domains for modification. Each catalytic domain is a unique enzyme spanning a wide range of folds and structures. Although they harbor the same enzymatic activities, two different types of fatty acid synthase architectures are observed in nature. During recent years, strained petroleum supplies have driven interest in engineering organisms to either produce more fatty acids or specific high value products. Such efforts require a fundamental understanding of the enzymatic activities and regulation of fatty acid synthases. Despite more than one hundred years of research, we continue to learn new lessons about fatty acid synthases’ many intricate structural and regulatory elements. In this review, we summarize each enzymatic domain and discuss efforts to engineer fatty acid synthases, providing some clues to important challenges and opportunities in the field. PMID:25360565

  16. LTR-retrotransposons in plants: Engines of evolution.

    Science.gov (United States)

    Galindo-González, Leonardo; Mhiri, Corinne; Deyholos, Michael K; Grandbastien, Marie-Angèle

    2017-08-30

    LTR retrotransposons are the most abundant group of transposable elements (TEs) in plants. These elements can fall inside or close to genes, and therefore influence their expression and evolution. This review aims to examine how LTR retrotransposons, especially Ty1-copia elements, mediate gene regulation and evolution. Various stimuli, including polyploidization and biotic and abiotic elicitors, result in the transcription and movement of these retrotransposons, and can facilitate adaptation. The presence of cis-regulatory motifs in the LTRs are central to their stress-mediated responses and are shared with host stress-responsive genes, showing a complex evolutionary history in which TEs provide new regulatory units to genes. The presence of retrotransposon remnants in genes that are necessary for normal gene function, demonstrates the importance of exaptation and co-option, and is also a consequence of the abundance of these elements in plant genomes. Furthermore, insertions of LTR retrotransposons in and around genes provide potential for alternative splicing, epigenetic control, transduction, duplication and recombination. These characteristics can become an active part of the evolution of gene families as in the case of resistance genes (R-genes). The character of TEs as exclusively selfish is now being re-evaluated. Since genome-wide reprogramming via TEs is a long evolutionary process, the changes we can examine are case-specific and their fitness advantage may not be evident until TE-derived motifs and domains have been completely co-opted and fixed. Nevertheless, the presence of LTR retrotransposons inside genes and as part of gene promoter regions is consistent with their roles as engines of plant genome evolution. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Metabolic cartography: experimental quantification of metabolic fluxes from isotopic labelling studies.

    Science.gov (United States)

    O'Grady, John; Schwender, Jörg; Shachar-Hill, Yair; Morgan, John A

    2012-03-01

    For the past decade, flux maps have provided researchers with an in-depth perspective on plant metabolism. As a rapidly developing field, significant headway has been made recently in computation, experimentation, and overall understanding of metabolic flux analysis. These advances are particularly applicable to the study of plant metabolism. New dynamic computational methods such as non-stationary metabolic flux analysis are finding their place in the toolbox of metabolic engineering, allowing more organisms to be studied and decreasing the time necessary for experimentation, thereby opening new avenues by which to explore the vast diversity of plant metabolism. Also, improved methods of metabolite detection and measurement have been developed, enabling increasingly greater resolution of flux measurements and the analysis of a greater number of the multitude of plant metabolic pathways. Methods to deconvolute organelle-specific metabolism are employed with increasing effectiveness, elucidating the compartmental specificity inherent in plant metabolism. Advances in metabolite measurements have also enabled new types of experiments, such as the calculation of metabolic fluxes based on (13)CO(2) dynamic labelling data, and will continue to direct plant metabolic engineering. Newly calculated metabolic flux maps reveal surprising and useful information about plant metabolism, guiding future genetic engineering of crops to higher yields. Due to the significant level of complexity in plants, these methods in combination with other systems biology measurements are necessary to guide plant metabolic engineering in the future.

  18. Metabolic cartography: experimental quantification of metabolic fluxes from isotopic labelling studies

    Energy Technology Data Exchange (ETDEWEB)

    O' Grady, J; Schwender, J; Shachar-Hill, Y; Morgan, JA

    2012-03-26

    For the past decade, flux maps have provided researchers with an in-depth perspective on plant metabolism. As a rapidly developing field, significant headway has been made recently in computation, experimentation, and overall understanding of metabolic flux analysis. These advances are particularly applicable to the study of plant metabolism. New dynamic computational methods such as non-stationary metabolic flux analysis are finding their place in the toolbox of metabolic engineering, allowing more organisms to be studied and decreasing the time necessary for experimentation, thereby opening new avenues by which to explore the vast diversity of plant metabolism. Also, improved methods of metabolite detection and measurement have been developed, enabling increasingly greater resolution of flux measurements and the analysis of a greater number of the multitude of plant metabolic pathways. Methods to deconvolute organelle-specific metabolism are employed with increasing effectiveness, elucidating the compartmental specificity inherent in plant metabolism. Advances in metabolite measurements have also enabled new types of experiments, such as the calculation of metabolic fluxes based on (CO2)-C-13 dynamic labelling data, and will continue to direct plant metabolic engineering. Newly calculated metabolic flux maps reveal surprising and useful information about plant metabolism, guiding future genetic engineering of crops to higher yields. Due to the significant level of complexity in plants, these methods in combination with other systems biology measurements are necessary to guide plant metabolic engineering in the future.

  19. Metabolic cartography: experimental quantification of metabolic fluxes from isotopic labelling studies

    Energy Technology Data Exchange (ETDEWEB)

    O' Grady J.; Schwender J.; Shachar-Hill, Y.; Morgan, J. A.

    2012-03-01

    For the past decade, flux maps have provided researchers with an in-depth perspective on plant metabolism. As a rapidly developing field, significant headway has been made recently in computation, experimentation, and overall understanding of metabolic flux analysis. These advances are particularly applicable to the study of plant metabolism. New dynamic computational methods such as non-stationary metabolic flux analysis are finding their place in the toolbox of metabolic engineering, allowing more organisms to be studied and decreasing the time necessary for experimentation, thereby opening new avenues by which to explore the vast diversity of plant metabolism. Also, improved methods of metabolite detection and measurement have been developed, enabling increasingly greater resolution of flux measurements and the analysis of a greater number of the multitude of plant metabolic pathways. Methods to deconvolute organelle-specific metabolism are employed with increasing effectiveness, elucidating the compartmental specificity inherent in plant metabolism. Advances in metabolite measurements have also enabled new types of experiments, such as the calculation of metabolic fluxes based on {sup 13}CO{sub 2} dynamic labelling data, and will continue to direct plant metabolic engineering. Newly calculated metabolic flux maps reveal surprising and useful information about plant metabolism, guiding future genetic engineering of crops to higher yields. Due to the significant level of complexity in plants, these methods in combination with other systems biology measurements are necessary to guide plant metabolic engineering in the future.

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

    Directory of Open Access Journals (Sweden)

    Yujin Cao

    Full Text Available Xylonate is a valuable chemical for versatile applications. Although the chemical synthesis route and microbial conversion pathway were established decades ago, no commercial production of xylonate has been obtained so far. In this study, the industrially important microorganism Escherichia coli was engineered to produce xylonate from xylose. Through the coexpression of a xylose dehydrogenase (xdh and a xylonolactonase (xylC from Caulobacter crescentus, the recombinant strain could convert 1 g/L xylose to 0.84 g/L xylonate and 0.10 g/L xylonolactone after being induced for 12 h. Furthermore, the competitive pathway for xylose catabolism in E. coli was blocked by disrupting two genes (xylA and xylB encoding xylose isomerase and xylulose kinase. Under fed-batch conditions, the finally engineered strain produced up to 27.3 g/L xylonate and 1.7 g/L xylonolactone from 30 g/L xylose, about 88% of the theoretical yield. These results suggest that the engineered E. coli strain has a promising perspective for large-scale production of xylonate.

  1. Metabolic engineering of Candida utilis for isopropanol production.

    Science.gov (United States)

    Tamakawa, Hideyuki; Mita, Tokiko; Yokoyama, Aki; Ikushima, Shigehito; Yoshida, Satoshi

    2013-07-01

    A genetically-engineered strain of the yeast Candida utilis harboring genes encoding (1) an acetoacetyl-CoA transferase from Clostridium acetobutylicum ATCC 824, (2) an acetoacetate decarboxylase, and (3) a primary-secondary alcohol dehydrogenase derived from Clostridium beijerinckii NRRL B593 produced up to 0.21 g/L of isopropanol. Because the engineered strain accumulated acetate, isopropanol titer was improved to 1.2 g/L under neutralized fermentation conditions. Optimization of isopropanol production was attempted by the overexpression and disruption of several endogenous genes. Simultaneous overexpression of two genes encoding acetyl-CoA synthetase and acetyl-CoA acetyltransferase increased isopropanol titer to 9.5 g/L. Moreover, in fed-batch cultivation, the resultant recombinant strain produced 27.2 g/L of isopropanol from glucose with a yield of 41.5 % (mol/mol). This is the first demonstration of the production of isopropanol by genetically engineered yeast.

  2. Harnessing biodiesel-producing microbes: from genetic engineering of lipase to metabolic engineering of fatty acid biosynthetic pathway.

    Science.gov (United States)

    Yan, Jinyong; Yan, Yunjun; Madzak, Catherine; Han, Bingnan

    2017-02-01

    Microbial production routes, notably whole-cell lipase-mediated biotransformation and fatty-acids-derived biosynthesis, offer new opportunities for synthesizing biodiesel. They compare favorably to immobilized lipase and chemically catalyzed processes. Genetically modified whole-cell lipase-mediated in vitro route, together with in vivo and ex vivo microbial biosynthesis routes, constitutes emerging and rapidly developing research areas for effective production of biodiesel. This review presents recent advances in customizing microorganisms for producing biodiesel, via genetic engineering of lipases and metabolic engineering (including system regulation) of fatty-acids-derived pathways. Microbial hosts used include Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris and Aspergillus oryzae. These microbial cells can be genetically modified to produce lipases under different forms: intracellularly expressed, secreted or surface-displayed. They can be metabolically redesigned and systematically regulated to obtain balanced biodiesel-producing cells, as highlighted in this study. Such genetically or metabolically modified microbial cells can support not only in vitro biotransformation of various common oil feedstocks to biodiesel, but also de novo biosynthesis of biodiesel from glucose, glycerol or even cellulosic biomass. We believe that the genetically tractable oleaginous yeast Yarrowia lipolytica could be developed to an effective biodiesel-producing microbial cell factory. For this purpose, we propose several engineered pathways, based on lipase and wax ester synthase, in this promising oleaginous host.

  3. Whole genome sequencing of Saccharomyces cerevisiae: from genotype to phenotype for improved metabolic engineering applications

    Directory of Open Access Journals (Sweden)

    Asadollahi Mohammad A

    2010-12-01

    Full Text Available Abstract Background The need for rapid and efficient microbial cell factory design and construction are possible through the enabling technology, metabolic engineering, which is now being facilitated by systems biology approaches. Metabolic engineering is often complimented by directed evolution, where selective pressure is applied to a partially genetically engineered strain to confer a desirable phenotype. The exact genetic modification or resulting genotype that leads to the improved phenotype is often not identified or understood to enable further metabolic engineering. Results In this work we performed whole genome high-throughput sequencing and annotation can be used to identify single nucleotide polymorphisms (SNPs between Saccharomyces cerevisiae strains S288c and CEN.PK113-7D. The yeast strain S288c was the first eukaryote sequenced, serving as the reference genome for the Saccharomyces Genome Database, while CEN.PK113-7D is a preferred laboratory strain for industrial biotechnology research. A total of 13,787 high-quality SNPs were detected between both strains (reference strain: S288c. Considering only metabolic genes (782 of 5,596 annotated genes, a total of 219 metabolism specific SNPs are distributed across 158 metabolic genes, with 85 of the SNPs being nonsynonymous (e.g., encoding amino acid modifications. Amongst metabolic SNPs detected, there was pathway enrichment in the galactose uptake pathway (GAL1, GAL10 and ergosterol biosynthetic pathway (ERG8, ERG9. Physiological characterization confirmed a strong deficiency in galactose uptake and metabolism in S288c compared to CEN.PK113-7D, and similarly, ergosterol content in CEN.PK113-7D was significantly higher in both glucose and galactose supplemented cultivations compared to S288c. Furthermore, DNA microarray profiling of S288c and CEN.PK113-7D in both glucose and galactose batch cultures did not provide a clear hypothesis for major phenotypes observed, suggesting that

  4. Biofortification of plants with altered antioxidant content and composition: genetic engineering strategies.

    Science.gov (United States)

    Zhu, Changfu; Sanahuja, Georgina; Yuan, Dawei; Farré, Gemma; Arjó, Gemma; Berman, Judit; Zorrilla-López, Uxue; Banakar, Raviraj; Bai, Chao; Pérez-Massot, Eduard; Bassie, Ludovic; Capell, Teresa; Christou, Paul

    2013-02-01

    Antioxidants are protective molecules that neutralize reactive oxygen species and prevent oxidative damage to cellular components such as membranes, proteins and nucleic acids, therefore reducing the rate of cell death and hence the effects of ageing and ageing-related diseases. The fortification of food with antioxidants represents an overlap between two diverse environments, namely fortification of staple foods with essential nutrients that happen to have antioxidant properties (e.g. vitamins C and E) and the fortification of luxury foods with health-promoting but non-essential antioxidants such as flavonoids as part of the nutraceuticals/functional foods industry. Although processed foods can be artificially fortified with vitamins, minerals and nutraceuticals, a more sustainable approach is to introduce the traits for such health-promoting compounds at source, an approach known as biofortification. Regardless of the target compound, the same challenges arise when considering the biofortification of plants with antioxidants, that is the need to modulate endogenous metabolic pathways to increase the production of specific antioxidants without affecting plant growth and development and without collateral effects on other metabolic pathways. These challenges become even more intricate as we move from the engineering of individual pathways to several pathways simultaneously. In this review, we consider the state of the art in antioxidant biofortification and discuss the challenges that remain to be overcome in the development of nutritionally complete and health-promoting functional foods. © 2012 The Authors Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  5. Engineering Plants for Geminivirus Resistance with CRISPR/Cas9 System.

    Science.gov (United States)

    Zaidi, Syed Shan-E-Ali; Mansoor, Shahid; Ali, Zahir; Tashkandi, Manal; Mahfouz, Magdy M

    2016-04-01

    The CRISPR/Cas9 system is an efficient genome-editing platform for diverse eukaryotic species, including plants. Recent work harnessed CRISPR/Cas9 technology to engineer resistance to geminiviruses. Here, we discuss opportunities, emerging developments, and potential pitfalls for using this technology to engineer resistance against single and multiple geminivirus infections in plants.

  6. Engineering Plants for Geminivirus Resistance with CRISPR/Cas9 System

    KAUST Repository

    Zaidi, Syed Shan-e-Ali

    2016-02-14

    The CRISPR/Cas9 system is an efficient genome-editing platform for diverse eukaryotic species, including plants. Recent work harnessed CRISPR/Cas9 technology to engineer resistance to geminiviruses. Here, we discuss opportunities, emerging developments, and potential pitfalls for using this technology to engineer resistance against single and multiple geminivirus infections in plants.

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

    Science.gov (United States)

    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.

  8. Phase I: the pipeline-gas demonstration plant. Demonstration plant engineering and design. Volume 17. Plant section 2500 - Plant and Instrument Air

    Energy Technology Data Exchange (ETDEWEB)

    None

    1981-05-01

    Contract No. EF-77-C-01-2542 between Conoco Inc. and the US Department of Energy provides for the design, construction, and operation of a demonstration plant capable of processing bituminous caking coals into clean pipeline quality gas. The project is currently in the design phase (Phase I). This phase is scheduled to be completed in June 1981. One of the major efforts of Phase I is the process and project engineering design of the Demonstration Plant. The design has been completed and is being reported in 24 volumes. This is Volume 17 which reports the design of Plant Section 2500 - Plant and Instrument Air. The plant and instrument air system is designed to provide dry, compressed air for a multitude of uses in plant operations and maintenance. A single centrifugal air compressor provides the total plant and instrument air requirements. An air drying system reduces the dew point of the plant and instrument air. Plant Section 2500 is designed to provide air at 100/sup 0/F and 100 psig. Both plant and instrument air are dried to a -40/sup 0/F dew point. Normal plant and instrument air requirements total 1430 standard cubic feet per minute.

  9. All-regime combined-cycle plant: Engineering solutions

    Science.gov (United States)

    Berezinets, P. A.; Tumanovskii, G. G.; Tereshina, G. E.; Krylova, I. N.; Markina, V. N.; Migun, E. N.

    2016-12-01

    The development of distributed power generation systems as a supplement to the centralized unified power grid increases the operational stability and efficiency of the entire power generation industry and improves the power supply to consumers. An all-regime cogeneration combined-cycle plant with a power of 20-25 mW (PGU-20/25T) and an electrical efficiency above 50% has been developed at the All-Russia Thermal Engineering Institute (ATEI) as a distributed power generation object. The PGU-20/25T two-circuit cogeneration plant provides a wide electrical and thermal power adjustment range and the absence of the mutual effect of electrical and thermal power output regimes at controlled frequency and power in a unified or isolated grid. The PGU-20/25T combined-cycle plant incorporates a gas-turbine unit (GTU) with a power of 16 MW, a heat recovery boiler (HRB) with two burners (before the boiler and the last heating stage), and a cogeneration steam turbine with a power of 6/9 MW. The PGU-20/25T plant has a maximum electrical power of 22 MW and an efficiency of 50.8% in the heat recovery regime and a maximum thermal power output of 16.3 MW (14 Gcal/h) in the cogeneration regime. The use of burners can increase the electrical power to 25 MW in the steam condensation regime at an efficiency of 49% and the maximum thermal power output to 29.5 MW (25.4 Gcal/h). When the steam turbine is shut down, the thermal power output can grow to 32.6 MW (28 Gcal/h). The innovative equipment, which was specially developed for PGU-20/25T, improves the reliability of this plant and simplifies its operation. Among this equipment are microflame burners in the heat recovery boiler, a vacuum system based on liquid-ring pumps, and a vacuum deaerator. To enable the application of PGU-20/25T in water-stressed regions, an air condenser preventing the heat-transfer tubes from the risk of covering with ice during operation in frost air has been developed. The vacuum system eliminates the need for

  10. Draft Genome Sequence of the Oleaginous Yeast Yarrowia lipolytica PO1f, a Commonly Used Metabolic Engineering Host

    OpenAIRE

    Liu, Leqian; Alper, Hal S.

    2014-01-01

    The draft genome sequence of the oleaginous yeast Yarrowia lipolytica stain PO1f, a commonly used metabolic engineering host, is presented here. The approximately 20.3-Mb genome sequence of PO1f will greatly facilitate research efforts in metabolic engineering of Yarrowia lipolytica for value-added chemical production.

  11. Metabolic flux phenotype of tobacco hairy roots engineered for increased geraniol production

    NARCIS (Netherlands)

    Masakapalli, S.K.; Ritala, A.; Dong, L.M.; Krol, van der A.R.; Oksman-Caldentey, K.M.; Ratcliffe, R.G.; Sweetlove, L.J.

    2014-01-01

    The goal of this study was to characterise the metabolic flux phenotype of transgenic tobacco (Nicotiana tabacum) hairy roots engineered for increased biosynthesis of geraniol, an intermediate of the terpenoid indole alkaloid pathway. Steady state, stable isotope labelling was used to determine flux

  12. An expanded role for microbial physiology in metabolic engineering and functional genomics: moving towards systems biology

    DEFF Research Database (Denmark)

    Nielsen, Jens; Olsson, Lisbeth

    2002-01-01

    . With the progress in molecular biology it has become possible to optimize industrial fermentations through introduction of directed genetic modification - an approach referred to as metabolic engineering. Furthermore, as a consequence of large sequencing programs the complete genomic sequence has become available...

  13. Getting a grip on glycans : A current overview of the metabolic oligosaccharide engineering toolbox

    NARCIS (Netherlands)

    Sminia, Tjerk J.; Zuilhof, Han; Wennekes, Tom

    2016-01-01

    This review discusses the advances in metabolic oligosaccharide engineering (MOE) from 2010 to 2016 with a focus on the structure, preparation, and reactivity of its chemical probes. A brief historical overview of MOE is followed by a comprehensive overview of the chemical probes currently available

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

    NARCIS (Netherlands)

    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 be

  15. Engineering metabolic pathways in Escherichia coli for constructing a "microbial chassis" for biochemical production.

    Science.gov (United States)

    Matsumoto, Takuya; Tanaka, Tsutomu; Kondo, Akihiko

    2017-05-04

    The present work reviews literature describing the re-design of the metabolic pathways of a microbial host using sophisticated genetic tools, yielding strains for producing value-added chemicals including fuels, building-block chemicals, pharmaceuticals, and derivatives. This work employed Escherichia coli, a well-studied microorganism that has been successfully engineered to produce various chemicals. E. coli has several advantages compared with other microorganisms, including robustness, and handling. To achieve efficient productivities of target compounds, an engineered E. coli should accumulate metabolic precursors of target compounds. Multiple researchers have reported the use of pathway engineering to generate strains capable of accumulating various metabolic precursors, including pyruvate, acetyl-CoA, malonyl-CoA, mevalonate and shikimate. The aim of this review provides a promising guideline for designing E. coli strains capable of producing a variety of useful chemicals. Herein, the present work reviews their common and unique strategies, treating metabolically engineered E. coli as a "microbial chassis". Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Metabolic Engineering of the Moss Physcomitrella patens as a Green Cell Factory to Produce Terpenoids

    DEFF Research Database (Denmark)

    Zhan, Xin

    also achieved with the yields of 1.3 and 0.035 mg/g dry weight respectively, after several metabolic engineering strategies were tried, including HMGR overexpression, CPS/KS gene disruption and plastidic localization of the terpene synthases. In order to synthesize more valuable perfumery ingredient (Z...

  17. Quantitative trait analysis of yeast biodiversity yields novel gene tools for metabolic engineering

    NARCIS (Netherlands)

    Hubmann, Georg; Foulquié-Moreno, Maria R.; Nevoigt, Elke; Duitama, Jorge; Meurens, Nicolas; Pais, Thiago M.; Mathé, Lotte; Saerens, Sofie; Nguyen, Huyen Thi Thanh; Swinnen, Steve; Verstrepen, Kevin J.; Concilio, Luigi; de Troostembergh, Jean-Claude; Thevelein, Johan M.

    2013-01-01

    Engineering of metabolic pathways by genetic modification has been restricted largely to enzyme-encoding structural genes. The product yield of such pathways is a quantitative genetic trait. Out of 52 Saccharomyces cerevisiae strains phenotyped in small-scale fermentations, we identified strain CBS6

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

    NARCIS (Netherlands)

    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 β-lactam antibiotics: - Regulation of secondary me

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

    NARCIS (Netherlands)

    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

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

    Science.gov (United States)

    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.

  1. (13)C Metabolic Flux Analysis for Systematic Metabolic Engineering of S. cerevisiae for Overproduction of Fatty Acids.

    Science.gov (United States)

    Ghosh, Amit; Ando, David; Gin, Jennifer; Runguphan, Weerawat; Denby, Charles; Wang, George; Baidoo, Edward E K; Shymansky, Chris; Keasling, Jay D; García Martín, Héctor

    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 (13)C 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 Yarrowia lipolytica as a robust source of cytoplasmic acetyl-CoA and malate synthase as a desirable target for downregulation 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 downregulation of malate synthase, the engineered strain produced 26% more free fatty acids. Further increases in free fatty acid production of 33% 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%.

  2. 13C Metabolic Flux Analysis for systematic metabolic engineering of S. cerevisiae for overproduction of fatty acids.

    Directory of Open Access Journals (Sweden)

    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.

  3. Coordinations between gene modules control the operation of plant amino acid metabolic networks

    Directory of Open Access Journals (Sweden)

    Galili Gad

    2009-01-01

    Full Text Available Abstract Background Being sessile organisms, plants should adjust their metabolism to dynamic changes in their environment. Such adjustments need particular coordination in branched metabolic networks in which a given metabolite can be converted into multiple other metabolites via different enzymatic chains. In the present report, we developed a novel "Gene Coordination" bioinformatics approach and use it to elucidate adjustable transcriptional interactions of two branched amino acid metabolic networks in plants in response to environmental stresses, using publicly available microarray results. Results Using our "Gene Coordination" approach, we have identified in Arabidopsis plants two oppositely regulated groups of "highly coordinated" genes within the branched Asp-family network of Arabidopsis plants, which metabolizes the amino acids Lys, Met, Thr, Ile and Gly, as well as a single group of "highly coordinated" genes within the branched aromatic amino acid metabolic network, which metabolizes the amino acids Trp, Phe and Tyr. These genes possess highly coordinated adjustable negative and positive expression responses to various stress cues, which apparently regulate adjustable metabolic shifts between competing branches of these networks. We also provide evidence implying that these highly coordinated genes are central to impose intra- and inter-network interactions between the Asp-family and aromatic amino acid metabolic networks as well as differential system interactions with other growth promoting and stress-associated genome-wide genes. Conclusion Our novel Gene Coordination elucidates that branched amino acid metabolic networks in plants are regulated by specific groups of highly coordinated genes that possess adjustable intra-network, inter-network and genome-wide transcriptional interactions. We also hypothesize that such transcriptional interactions enable regulatory metabolic adjustments needed for adaptation to the stresses.

  4. Engineering microbial cell factories for the production of plant natural products: from design principles to industrial-scale production.

    Science.gov (United States)

    Liu, Xiaonan; Ding, Wentao; Jiang, Huifeng

    2017-07-19

    Plant natural products (PNPs) are widely used as pharmaceuticals, nutraceuticals, seasonings, pigments, etc., with a huge commercial value on the global market. However, most of these PNPs are still being extracted from plants. A resource-conserving and environment-friendly synthesis route for PNPs that utilizes microbial cell factories has attracted increasing attention since the 1940s. However, at the present only a handful of PNPs are being produced by microbial cell factories at an industrial scale, and there are still many challenges in their large-scale application. One of the challenges is that most biosynthetic pathways of PNPs are still unknown, which largely limits the number of candidate PNPs for heterologous microbial production. Another challenge is that the metabolic fluxes toward the target products in microbial hosts are often hindered by poor precursor supply, low catalytic activity of enzymes and obstructed product transport. Consequently, despite intensive studies on the metabolic engineering of microbial hosts, the fermentation costs of most heterologously produced PNPs are still too high for industrial-scale production. In this paper, we review several aspects of PNP production in microbial cell factories, including important design principles and recent progress in pathway mining and metabolic engineering. In addition, implemented cases of industrial-scale production of PNPs in microbial cell factories are also highlighted.

  5. What drives the career choice among engineers? A case in Malaysian manufacturing plant

    Directory of Open Access Journals (Sweden)

    Choo, Ling Suan

    2012-01-01

    Full Text Available This empirical study was conducted to determine factors that influence the career choice among engineers in a Malaysian manufacturing plant. Subjects of this study were taken from engineers in one of manufacturing plant in Malaysia. As hypothesized, the results suggest that perceptions of person-job fit and financial rewards are significant factors that influence the career choice of engineers. Contrast to our hypothesis, influence from family members does not have any impact in determining the career choice among engineers. Interventions in attracting more engineers to remain in their career path and limitations of the study are also being discussed in the paper.

  6. The importance of sourcing enzymes from non-conventional fungi for metabolic engineering and biomass breakdown.

    Science.gov (United States)

    Seppälä, Susanna; Wilken, St Elmo; Knop, Doriv; Solomon, Kevin V; O'Malley, Michelle A

    2017-09-21

    A wealth of fungal enzymes has been identified from nature, which continue to drive strain engineering and bioprocessing for a range of industries. However, while a number of clades have been investigated, the vast majority of the fungal kingdom remains unexplored for industrial applications. Here, we discuss selected classes of fungal enzymes that are currently in biotechnological use, and explore more basal, non-conventional fungi and their underexploited biomass-degrading mechanisms as promising agents in the transition towards a bio-based society. Of special interest are anaerobic fungi like the Neocallimastigomycota, which were recently found to harbor the largest diversity of biomass-degrading enzymes among the fungal kingdom. Enzymes sourced from these basal fungi have been used to metabolically engineer substrate utilization in yeast, and may offer new paths to lignin breakdown and tunneled biocatalysis. We also contrast classic enzymology approaches with emerging 'omics'-based tools to decipher function within novel fungal isolates and identify new promising enzymes. Recent developments in genome editing are expected to accelerate discovery and metabolic engineering within these systems, yet are still limited by a lack of high-resolution genomes, gene regulatory regions, and even appropriate culture conditions. Finally, we present new opportunities to harness the biomass-degrading potential of undercharacterized fungi via heterologous expression and engineered microbial consortia. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

    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.

  8. Metabolic engineering and synthetic biology approaches driving isoprenoid production in Escherichia coli.

    Science.gov (United States)

    Wang, Chonglong; Zada, Bakht; Wei, Gongyuan; Kim, Seon-Won

    2017-10-01

    Isoprenoids comprise the largest family of natural organic compounds with many useful applications in the pharmaceutical, nutraceutical, and industrial fields. Rapid developments in metabolic engineering and synthetic biology have facilitated the engineering of isoprenoid biosynthetic pathways in Escherichia coli to induce high levels of production of many different isoprenoids. In this review, the stem pathways for synthesizing isoprene units as well as the branch pathways deriving diverse isoprenoids from the isoprene units have been summarized. The review also highlights the metabolic engineering efforts made for the biosynthesis of hemiterpenoids, monoterpenoids, sesquiterpenoids, diterpenoids, carotenoids, retinoids, and coenzyme Q10 in E. coli. Perspectives and future directions for the synthesis of novel isoprenoids, decoration of isoprenoids using cytochrome P450 enzymes, and secretion or storage of isoprenoids in E. coli have also been included. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Discovering the role of mitochondria in the iron deficiency-induced metabolic responses of plants.

    Science.gov (United States)

    Vigani, Gianpiero

    2012-01-01

    In plants, iron (Fe) deficiency-induced chlorosis is a major problem, affecting both yield and quality of crops. Plants have evolved multifaceted strategies, such as reductase activity, proton extrusion, and specialised storage proteins, to mobilise Fe from the environment and distribute it within the plant. Because of its fundamental role in plant productivity, several issues concerning Fe homeostasis in plants are currently intensively studied. The activation of Fe uptake reactions requires an overall adaptation of the primary metabolism because these activities need the constant supply of energetic substrates (i.e., NADPH and ATP). Several studies concerning the metabolism of Fe-deficient plants have been conducted, but research focused on mitochondrial implications in adaptive responses to nutritional stress has only begun in recent years. Mitochondria are the energetic centre of the root cell, and they are strongly affected by Fe deficiency. Nevertheless, they display a high level of functional flexibility, which allows them to maintain the viability of the cell. Mitochondria represent a crucial target of studies on plant homeostasis, and it might be of interest to concentrate future research on understanding how mitochondria orchestrate the reprogramming of root cell metabolism under Fe deficiency. In this review, I summarise what it is known about the effect of Fe deficiency on mitochondrial metabolism and morphology. Moreover, I present a detailed view of the possible roles of mitochondria in the development of plant responses to Fe deficiency, integrating old findings with new and discussing new hypotheses for future investigations.

  10. Engineering a plant community to deliver multiple ecosystem services.

    Science.gov (United States)

    Storkey, Jonathan; Döring, Thomas; Baddeley, John; Collins, Rosemary; Roderick, Stephen; Jones, Hannah; Watson, Christine

    2015-06-01

    The sustainable delivery of multiple ecosystem services requires the management of functionally diverse biological communities. In an agricultural context, an emphasis on food production has often led to a loss of biodiversity to the detriment of other ecosystem services such as the maintenance of soil health and pest regulation. In scenarios where multiple species can be grown together, it may be possible to better balance environmental and agronomic services through the targeted selection of companion species. We used the case study of legume-based cover crops to engineer a plant community that delivered the optimal balance of six ecosystem services: early productivity, regrowth following mowing, weed suppression, support of invertebrates, soil fertility building (measured as yield of following crop), and conservation of nutrients in the soil. An experimental species pool of 12 cultivated legume species was screened for a range of functional traits and ecosystem services at five sites across a geographical gradient in the United Kingdom. All possible species combinations were then analyzed, using a process-based model of plant competition, to identify the community that delivered the best balance of services at each site. In our system, low to intermediate levels of species richness (one to four species) that exploited functional contrasts in growth habit and phenology were identified as being optimal. The optimal solution was determined largely by the number of species and functional diversity represented by the starting species pool, emphasizing the importance of the initial selection of species for the screening experiments. The approach of using relationships between functional traits and ecosystem services to design multifunctional biological communities has the potential to inform the design of agricultural systems that better balance agronomic and environmental services and meet the current objective of European agricultural policy to maintain viable food

  11. Plant biostimulants: physiological responses induced by protein hydrolyzed-based products and humic substances in plant metabolism

    Directory of Open Access Journals (Sweden)

    Serenella Nardi

    2016-02-01

    Full Text Available ABSTRACT In recent years, the use of biostimulants in sustainable agriculture has been growing. Biostimulants can be obtained from different organic materials and include humic substances (HS, complex organic materials, beneficial chemical elements, peptides and amino acids, inorganic salts, seaweed extracts, chitin and chitosan derivatives, antitranspirants, amino acids and other N-containing substances. The application of biostimulants to plants leads to higher content of nutrients in their tissue and positive metabolic changes. For these reasons, the development of new biostimulants has become a focus of scientific interest. Among their different functions, biostimulants influence plant growth and nitrogen metabolism, especially because of their content in hormones and other signalling molecules. A significant increase in root hair length and density is often observed in plants treated with biostimulants, suggesting that these substances induce a “nutrient acquisition response” that favors nutrient uptake in plants via an increase in the absorptive surface area. Furthermore, biostimulants positively influence the activity and gene expression of enzymes functioning in the primary and secondary plant metabolism. This article reviews the current literature on two main classes of biostimulants: humic substances and protein-based biostimulants. The characteristic of these biostimulants and their effects on plants are thoroughly described.

  12. NATURAL PLANT TOXICANT – CYANOGENIC GLYCOSIDE AMYGDALIN: CHARACTERISTIC, METABOLISM AND THE EFFECT ON ANIMAL REPRODUCTION

    OpenAIRE

    Eduard Kolesár; Marek Halenár; Adriana Kolesárová; Peter Massányi

    2015-01-01

    The amount of cyanogenic glycosides, as natural plant toxicants, in plants varies with plant species and environmental effects. Cyanogenic glycoside as an amygdalin was detected in apricot kernels, bitter almonds and peach, plum, pear and apple seeds. Amygdalin itself is non-toxic, but its HCN production decomposed by some enzymes is toxic substance. Target of this review was to describe the characteristic, metabolism and possible effects of amygdalin on reproductive processes. Previous studi...

  13. Towards a metabolic engineering strain "commons": an Escherichia coli platform strain for ethanol production.

    Science.gov (United States)

    Woodruff, Lauren B A; May, Brian L; Warner, Joseph R; Gill, Ryan T

    2013-05-01

    In the genome-engineering era, it is increasingly important that researchers have access to a common set of platform strains that can serve as debugged production chassis and the basis for applying new metabolic engineering strategies for modeling and characterizing flux, engineering complex traits, and optimizing overall performance. Here, we describe such a platform strain of E. coli engineered for ethanol production. Starting with a fully characterized host strain (BW25113), we site-specifically integrated the genes required for homoethanol production under the control of a strong inducible promoter into the genome and deleted the genes encoding four enzymes from competing pathways. This strain is capable of producing >30 g/L of ethanol in minimal media with production or tolerance was lost when grown under production conditions. Thus, our findings reinforce the need for a metabolic engineering "commons" that could provide a set of platform strains for use in more sophisticated genome-engineering strategies. Towards this end, we have made this production strain available to the scientific community.

  14. Carbon and nitrogen metabolism in arbuscular mycorrhizal maize plants under low-temperature stress

    DEFF Research Database (Denmark)

    Zhu, Xian-Can; Song, Feng-Bin; Liu, Fulai

    2015-01-01

    Effects of the arbuscular mycorrhizal (AM) fungus Glomus tortuosum on carbon (C) and nitrogen (N) metabolism of Zea mays L. grown under low-temperature stress was investigated. Maize plants inoculated or not inoculated with AM fungus were grown in a growth chamber at 258C for 4 weeks...... phosphate synthase and amylase activities at low temperature. Moreover, low-temperature stress increased theC :Nratio in the leaves of maize plants, and AM colonisation decreased the root C :N ratio. These results suggested a difference in the C and N metabolism of maize plants at ambient and low...... temperature regimes. AM symbiosis modulated C metabolic enzymes, thereby inducing an accumulation of soluble sugars, which may have contributed to an increased tolerance to low temperature, and therefore higher Pn in maize plants....

  15. Plant metabolic clusters – from genetics to genomics

    National Research Council Canada - National Science Library

    Nützmann, Hans‐Wilhelm; Huang, Ancheng; Osbourn, Anne

    2016-01-01

    ..., and for evolutionary biology more widely, lies in understanding how and why different plants make different kinds of chemicals, and how new natural product pathways are formed. The discovery of new plant natural product pathways and chemistries is now being revolutionized by two key developments. First, breakthroughs in sequencing techn...

  16. Metabolic changes in Arabidopsis thaliana plants overexpressing chalcone synthase

    NARCIS (Netherlands)

    Dao, Thi Thanh Hien

    2010-01-01

    The study has shown that it is possible to introduce the heterologous CHS gene in Arabidopsis thaliana and common multicopies of transgenes containing plants were obtained. Analysis of the change in metabolome of CHS transgenic plants, high expression transgenic lines can be identified by markers su

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

    Energy Technology Data Exchange (ETDEWEB)

    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

  18. Metabolically engineered Saccharomyces cerevisiae for branched-chain ester productions.

    Science.gov (United States)

    Yuan, Jifeng; Mishra, Pranjul; Ching, Chi Bun

    2016-12-10

    Medium branched-chain esters can be used not only as a biofuel but are also useful chemicals with various industrial applications. The development of economically feasible and environment friendly bio-based fuels requires efficient cell factories capable of producing desired products in high yield. Herein, we sought to use a number of strategies to engineer Saccharomyces cerevisiae for high-level production of branched-chain esters. Mitochondrion-based expression of ATF1 gene in a base strain with an overexpressed valine biosynthetic pathway together with expression of mitochondrion-relocalized α-ketoacid decarboxylase (encoded by ARO10) and alcohol dehydrogenase (encoded by ADH7) not only produced isobutyl acetate, but also 3-methyl-1-butyl acetate and 2-methyl-1-butyl acetate. Further segmentation of the downstream esterification step into the cytosol to utilize the cytosolic acetyl-CoA pool for acetyltransferase (ATF)-mediated condensation enabled an additional fold improvement of ester productions. The best titre attained in the present study is 260.2mg/L isobutyl acetate, 296.1mg/L 3-methyl-1-butyl acetate and 289.6mg/L 2-methyl-1-butyl acetate.

  19. Physiological and molecular implications of plant polyamine metabolism during biotic interactions

    Directory of Open Access Journals (Sweden)

    Juan Francisco Jiménez Bremont

    2014-03-01

    Full Text Available During ontogeny, plants interact with a wide variety of microorganisms. The association with mutualistic microbes results in benefits for the plant. By contrast, pathogens may cause a remarkable impairment of plant growth and development. Both types of plant-microbe interactions provoke notable changes in the polyamine (PA metabolism of the host and/or the microbe, being each interaction a complex and dynamic process. It has been well documented that the levels of free and conjugated PAs undergo profound changes in plant tissues during the interaction with microorganisms. In general, this is correlated with a precise and coordinated regulation of PA biosynthetic and catabolic enzymes. Interestingly, some evidence suggests that the relative importance of these metabolic pathways may depend on the nature of the microorganism, a concept that stems from the fact that these amines mediate the activation of plant defense mechanisms. This effect is mediated mostly through PA oxidation, even though part of the response is activated by non-oxidized PAs. In the last years, a great deal of effort has been devoted to profile plant gene expression following microorganism recognition. In addition, the phenotypes of transgenic and mutant plants in PA metabolism genes have been assessed. In this review, we integrated the current knowledge on this field and analyze the possible roles of these amines during the interaction of plants with microbes.

  20. Transient silencing of the KASII genes is feasible in Nicotiana benthamiana for metabolic engineering of wax ester composition.

    Science.gov (United States)

    Aslan, Selcuk; Hofvander, Per; Dutta, Paresh; Sitbon, Folke; Sun, Chuanxin

    2015-06-11

    The beta-ketoacyl-ACP synthase II (KASII) is an enzyme in fatty acid biosynthesis, catalyzing the elongation of 16:0-acyl carrier protein (ACP) to 18:0-ACP in plastids. Mutations in KASII genes in higher plants can lead to lethality, which makes it difficult to utilize the gene for lipid metabolic engineering. We demonstrated previously that transient expression of plastid-directed fatty acyl reductases and wax ester synthases could result in different compositions of wax esters. We hypothesized that changing the ratio between C16 (palmitoyl-compounds) and C18 (stearoyl-compounds) in the plastidic acyl-ACP pool by inhibition of KASII expression would change the yield and composition of wax esters via substrate preference of the introduced enzymes. Here, we report that transient inhibition of KASII expression by three different RNAi constructs in leaves of N. benthamiana results in almost complete inhibition of KASII expression. The transient RNAi approach led to a shift of carbon flux from a pool of C18 fatty acids to C16, which significantly increased wax ester production in AtFAR6-containing combinations. The results demonstrate that transient inhibition of KASII in vegetative tissues of higher plants enables metabolic studies towards industrial production of lipids such as wax esters with specific quality and composition.

  1. Observability of plant metabolic networks is reflected in the correlation of metabolic profiles

    DEFF Research Database (Denmark)

    Schwahn, Kevin; Küken, Anika; Kliebenstein, Daniel James

    2016-01-01

    -of-the-art genome-scale metabolic networks. By using metabolic data profiles from a set of seven environmental perturbations as well as from natural variability, we demonstrate that the data profiles of sensor metabolites are more correlated than those of nonsensor metabolites. This pattern was confirmed...

  2. Exploring the Metabolic Stability of Engineered Hairy Roots after 16 Years Maintenance

    Science.gov (United States)

    Häkkinen, Suvi T.; Moyano, Elisabeth; Cusidó, Rosa M.; Oksman-Caldentey, Kirsi-Marja

    2016-01-01

    Plants remain a major source of new drugs, leads and fine chemicals. Cell cultures deriving from plants offer a fascinating tool to study plant metabolic pathways and offer large scale production systems for valuable compounds – commercial examples include compounds such as paclitaxel. The major constraint with undifferentiated cell cultures is that they are generally considered to be genetically unstable and cultured cells tend to produce low yields of secondary metabolites especially over time. Hairy roots, a tumor tissue caused by infection of Agrobacterium rhizogenes is a relevant alternative for plant secondary metabolite production for being fast growing, able to grow without phytohormones, and displaying higher stability than undifferentiated cells. Although genetic and metabolic stability has often been connected to transgenic hairy roots, there are only few reports on how a very long-term subculturing effects on the production capacity of hairy roots. In this study, hairy roots producing high tropane alkaloid levels were subjected to 16-year follow-up in relation to genetic and metabolic stability. Cryopreservation method for hairy roots of Hyoscyamus muticus was developed to replace laborious subculturing, and although the post-thaw recovery rates remained low, the expression of transgene remained unaltered in cryopreserved roots. It was shown that although displaying some fluctuation in the metabolite yields, even an exceedingly long-term subculturing was successfully applied without significant loss of metabolic activity. PMID:27746806

  3. Exploring the metabolic stability of engineered hairy roots after 16 years maintenance

    Directory of Open Access Journals (Sweden)

    Suvi Tuulikki Häkkinen

    2016-09-01

    Full Text Available Plants remain a major source of new drugs, leads and fine chemicals. Cell cultures deriving from plants offer a fascinating tool to study plant metabolic pathways and offer large scale production systems for valuable compounds – commercial examples include compounds such as paclitaxel. The major constraint with undifferentiated cell cultures is that they are generally considered to be genetically unstable and cultured cells tend to produce low yields of secondary metabolites especially over time. Hairy roots, a tumour tissue caused by infection of Agrobacterium rhizogenes is a relevant alternative for plant secondary metabolite production for being fast growing, able to grow without phytohormones, and displaying higher stability than undifferentiated cells. Although genetic and metabolic stability has often been connected to transgenic hairy roots, there are only few reports on how a very long-term subculturing effects on the production capacity of hairy roots. In this study, hairy roots producing high tropane alkaloid levels were subjected to 16 -year follow-up in relation to genetic and metabolic stability. Cryopreservation method for hairy roots of H. muticus was developed to replace laborious subculturing, and although the post-thaw recovery rates remained low, the expression of transgene remained unaltered in cryopreserved roots. It was shown that although displaying some fluctuation in the metabolite yields, even an exceedingly long-term subculturing was successfully applied without significant loss of metabolic activity.

  4. Metabolic engineering of sugarcane to accumulate energy-dense triacylglycerols in vegetative biomass.

    Science.gov (United States)

    Zale, Janice; Jung, Je Hyeong; Kim, Jae Yoon; Pathak, Bhuvan; Karan, Ratna; Liu, Hui; Chen, Xiuhua; Wu, Hao; Candreva, Jason; Zhai, Zhiyang; Shanklin, John; Altpeter, Fredy

    2016-02-01

    Elevating the lipid content in vegetative tissues has emerged as a new strategy for increasing energy density and biofuel yield of crops. Storage lipids in contrast to structural and signaling lipids are mainly composed of glycerol esters of fatty acids, also known as triacylglycerol (TAG). TAGs are one of the most energy-rich and abundant forms of reduced carbon available in nature. Therefore, altering the carbon-partitioning balance in favour of TAG in vegetative tissues of sugarcane, one of the highest yielding biomass crops, is expected to drastically increase energy yields. Here we report metabolic engineering to elevate TAG accumulation in vegetative tissues of sugarcane. Constitutive co-expression of WRINKLED1 (WRI1), diacylglycerol acyltransferase1-2 (DGAT1-2) and oleosin1 (OLE1) and simultaneous cosuppression of ADP-glucose pyrophosphorylase (AGPase) and a subunit of the peroxisomal ABC transporter1 (PXA1) in transgenic sugarcane elevated TAG accumulation in leaves or stems by 95- or 43-fold to 1.9% or 0.9% of dry weight (DW), respectively, while expression or suppression of one to three of the target genes increased TAG levels by 1.5- to 9.5-fold. Accumulation of TAG in vegetative progeny plants was consistent with the results from primary transgenics and contributed to a total fatty acid content of up to 4.7% or 1.7% of DW in mature leaves or stems, respectively. Lipid droplets were visible within mesophyll cells of transgenic leaves by confocal fluorescence microscopy. These results provide the basis for optimizations of TAG accumulation in sugarcane and other high yielding biomass grasses and will open new prospects for biofuel applications. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  5. [Production of coenzyme Q10 by metabolically engineered Escherichia coli].

    Science.gov (United States)

    Dai, Guanping; Miao, Liangtian; Sun, Tao; Li, Qingyan; Xiao, Dongguang; Zhang, Xueli

    2015-02-01

    Coenzyme Q10 (CoQ10) is a lipophilic antioxidant that improves human immunity, delays senility and enhances the vitality of the human body and has wide applications in pharmaceutical and cosmetic industries. Microbial fermentation is a sustainable way to produce CoQ10, and attracts increased interest. In this work, the native CoQ8 synthetic pathway of Escherichia coli was replaced by the CoQ10 synthetic pathway through integrating decaprenyl diphosphate synthase gene (dps) from Rhodobacter sphaeroides into chromosome of E. coli ATCC 8739, followed by deletion of the native octaprenyl diphosphate synthase gene (ispB). The resulting strain GD-14 produced 0.68 mg/L CoQ10 with a yield of 0.54 mg/g DCW. Modulation of dxs and idi genes of the MEP pathway and ubiCA genes in combination led to 2.46-fold increase of CoQ10 production (from 0.54 to 1.87 mg/g DCW). Recruiting glucose facilitator protein of Zymomonas mobilis to replace the native phosphoenolpyruvate: carbohydrate phosphotransferase systems (PTS) further led to a 16% increase of CoQ10 yield. Finally, fed-batch fermentation of the best strain GD-51 was performed, which produced 433 mg/L CoQ10 with a yield of 11.7 mg/g DCW. To the best of our knowledge, this was the highest CoQ10 titer and yield obtained for engineered E. coli.

  6. Ketocarotenoid Production in Soybean Seeds through Metabolic Engineering.

    Directory of Open Access Journals (Sweden)

    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.

  7. Metabolic engineering of Saccharomyces cerevisiae for caffeine and theobromine production.

    Science.gov (United States)

    Jin, Lu; Bhuiya, Mohammad Wadud; Li, Mengmeng; Liu, XiangQi; Han, Jixiang; Deng, WeiWei; Wang, Min; Yu, Oliver; Zhang, Zhengzhu

    2014-01-01

    Caffeine (1, 3, 7-trimethylxanthine) and theobromine (3, 7-dimethylxanthine) are the major purine alkaloids in plants, e.g., tea (Camellia sinensis) and coffee (Coffea arabica). Caffeine is a major component of coffee and is used widely in food and beverage industries. Most of the enzymes involved in the caffeine biosynthetic pathway have been reported previously. Here, we demonstrated the biosynthesis of caffeine (0.38 mg/L) by co-expression of Coffea arabica xanthosine methyltransferase (CaXMT) and Camellia sinensis caffeine synthase (TCS) in Saccharomyces cerevisiae. Furthermore, we endeavored to develop this production platform for making other purine-based alkaloids. To increase the catalytic activity of TCS in an effort to increase theobromine production, we identified four amino acid residues based on structural analyses of 3D-model of TCS. Two TCS1 mutants (Val317Met and Phe217Trp) slightly increased in theobromine accumulation and simultaneously decreased in caffeine production. The application and further optimization of this biosynthetic platform are discussed.

  8. Metabolic engineering of Saccharomyces cerevisiae for caffeine and theobromine production.

    Directory of Open Access Journals (Sweden)

    Lu Jin

    Full Text Available Caffeine (1, 3, 7-trimethylxanthine and theobromine (3, 7-dimethylxanthine are the major purine alkaloids in plants, e.g., tea (Camellia sinensis and coffee (Coffea arabica. Caffeine is a major component of coffee and is used widely in food and beverage industries. Most of the enzymes involved in the caffeine biosynthetic pathway have been reported previously. Here, we demonstrated the biosynthesis of caffeine (0.38 mg/L by co-expression of Coffea arabica xanthosine methyltransferase (CaXMT and Camellia sinensis caffeine synthase (TCS in Saccharomyces cerevisiae. Furthermore, we endeavored to develop this production platform for making other purine-based alkaloids. To increase the catalytic activity of TCS in an effort to increase theobromine production, we identified four amino acid residues based on structural analyses of 3D-model of TCS. Two TCS1 mutants (Val317Met and Phe217Trp slightly increased in theobromine accumulation and simultaneously decreased in caffeine production. The application and further optimization of this biosynthetic platform are discussed.

  9. Production of salidroside in metabolically engineered Escherichia coli

    Science.gov (United States)

    Bai, Yanfen; Bi, Huiping; Zhuang, Yibin; Liu, Chang; Cai, Tao; Liu, Xiaonan; Zhang, Xueli; Liu, Tao; Ma, Yanhe

    2014-01-01

    Salidroside (1) is the most important bioactive component of Rhodiola (also called as “Tibetan Ginseng”), which is a valuable medicinal herb exhibiting several adaptogenic properties. Due to the inefficiency of plant extraction and chemical synthesis, the supply of salidroside (1) is currently limited. Herein, we achieved unprecedented biosynthesis of salidroside (1) from glucose in a microorganism. First, the pyruvate decarboxylase ARO10 and endogenous alcohol dehydrogenases were recruited to convert 4-hydroxyphenylpyruvate (2), an intermediate of L-tyrosine pathway, to tyrosol (3) in Escherichia coli. Subsequently, tyrosol production was improved by overexpressing the pathway genes, and by eliminating competing pathways and feedback inhibition. Finally, by introducing Rhodiola-derived glycosyltransferase UGT73B6 into the above-mentioned recombinant strain, salidroside (1) was produced with a titer of 56.9 mg/L. Interestingly, the Rhodiola-derived glycosyltransferase, UGT73B6, also catalyzed the attachment of glucose to the phenol position of tyrosol (3) to form icariside D2 (4), which was not reported in any previous literatures. PMID:25323006

  10. Metabolic engineering of microorganisms for the production of L-arginine and its derivatives.

    Science.gov (United States)

    Shin, Jae Ho; Lee, Sang Yup

    2014-12-03

    L-arginine (ARG) is an important amino acid for both medicinal and industrial applications. For almost six decades, the research has been going on for its improved industrial level production using different microorganisms. While the initial approaches involved random mutagenesis for increased tolerance to ARG and consequently higher ARG titer, it is laborious and often leads to unwanted phenotypes, such as retarded growth. Discovery of L-glutamate (GLU) overproducing strains and using them as base strains for ARG production led to improved ARG production titer. Continued effort to unveil molecular mechanisms led to the accumulation of detailed knowledge on amino acid metabolism, which has contributed to better understanding of ARG biosynthesis and its regulation. Moreover, systems metabolic engineering now enables scientists and engineers to efficiently construct genetically defined microorganisms for ARG overproduction in a more rational and system-wide manner. Despite such effort, ARG biosynthesis is still not fully understood and many of the genes in the pathway are mislabeled. Here, we review the major metabolic pathways and its regulation involved in ARG biosynthesis in different prokaryotes including recent discoveries. Also, various strategies for metabolic engineering of bacteria for the overproduction of ARG are described. Furthermore, metabolic engineering approaches for producing ARG derivatives such as L-ornithine (ORN), putrescine and cyanophycin are described. ORN is used in medical applications, while putrescine can be used as a bio-based precursor for the synthesis of nylon-4,6 and nylon-4,10. Cyanophycin is also an important compound for the production of polyaspartate, another important bio-based polymer. Strategies outlined here will serve as a general guideline for rationally designing of cell-factories for overproduction of ARG and related compounds that are industrially valuable.

  11. Metabolic engineering of Clostridium acetobutylicum for butyric acid production with high butyric acid selectivity.

    Science.gov (United States)

    Jang, Yu-Sin; Im, Jung Ae; Choi, So Young; Lee, Jung Im; Lee, Sang Yup

    2014-05-01

    A typical characteristic of the butyric acid-producing Clostridium is coproduction of both butyric and acetic acids. Increasing the butyric acid selectivity important for economical butyric acid production has been rather difficult in clostridia due to their complex metabolic pathways. In this work, Clostridium acetobutylicum was metabolically engineered for highly selective butyric acid production. For this purpose, the second butyrate kinase of C. acetobutylicum encoded by the bukII gene instead of butyrate kinase I encoded by the buk gene was employed. Furthermore, metabolic pathways were engineered to further enhance the NADH-driving force. Batch fermentation of the metabolically engineered C. acetobutylicum strain HCBEKW (pta(-), buk(-), ctfB(-) and adhE1(-)) at pH 6.0 resulted in the production of 32.5g/L of butyric acid with a butyric-to-acetic acid ratio (BA/AA ratio) of 31.3g/g from 83.3g/L of glucose. By further knocking out the hydA gene (encoding hydrogenase) in the HCBEKW strain, the butyric acid titer was not further improved in batch fermentation. However, the BA/AA ratio (28.5g/g) obtained with the HYCBEKW strain (pta(-), buk(-), ctfB(-), adhE1(-) and hydA(-)) was 1.6 times higher than that (18.2g/g) obtained with the HCBEKW strain at pH 5.0, while no improvement was observed at pH 6.0. These results suggested that the buk gene knockout was essential to get a high butyric acid selectivity to acetic acid in C. acetobutylicum. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  12. Engineering resistance to plant viruses: Present status and future prospects

    Science.gov (United States)

    Plant viruses cause severe crop losses across the globe. Resistant cultivars together with pesticide application are commonly used to avoid the losses caused by plant viruses. However, very limited success has been achieved at diminishing the impact of plant viruses. Use of virus resistant plant is ...

  13. Metabolic engineering of Caldicellulosiruptor bescii yields increased hydrogen production from lignocellulosic biomass

    Energy Technology Data Exchange (ETDEWEB)

    Cha, Minseok [University of Georgia, Athens, GA; Chung, Daehwan [University of Georgia, Athens, GA; Elkins, James G [ORNL; Guss, Adam M [ORNL; Westpheling, Janet [University of Georgia, Athens, GA

    2013-01-01

    Background: Members of the anaerobic thermophilic bacterial genus Caldicellulosiruptor are emerging candidates for consolidated bioprocessing (CBP) because they are capable of efficiently growing on biomass without conventional pretreatment. C. bescii produces primarily lactate, acetate and hydrogen as fermentation products, and while some Caldicellulosiruptor strains produce small amounts of ethanol C. bescii does not, making it an attractive background to examine the effects of metabolic engineering. The recent development of methods for genetic manipulation has set the stage for rational engineering of this genus for improved biofuel production. Here, we report the first targeted gene deletion, the gene encoding lactate dehydrogenase (ldh), for metabolic engineering of a member of this genus. Results: A deletion of the C. bescii L-lactate dehydrogenase gene (ldh) was constructed on a non-replicating plasmid and introduced into the C. bescii chromosome by marker replacement. The resulting strain failed to produce detectable levels of lactate from cellobiose and maltose, instead increasing production of acetate and H2 by 21-34% relative to the wild type and pyrFA parent strains. The same phenotype was observed on a real-world substrate switchgrass (Panicum virgatum). Furthermore, the ldh deletion strain grew to a higher maximum optical density than the wild type on maltose and cellobiose, consistent with the prediction that the mutant would gain additional ATP with increased acetate production. Conclusions: Deletion of ldh in C. bescii is the first use of recently developed genetic methods for metabolic engineering of these bacteria. This deletion resulted in a redirection of electron flow from production of lactate to acetate and hydrogen. New capabilities in metabolic engineering combined with intrinsic utilization of lignocellulosic materials position these organisms to provide a new paradigm for consolidated bioprocessing of fuels and other products from

  14. 76 FR 27366 - Chrysler Group, LLC, Power Train Division, Mack Avenue Engine Plants #1 And #2, Including On-Site...

    Science.gov (United States)

    2011-05-11

    ... Employment and Training Administration Chrysler Group, LLC, Power Train Division, Mack Avenue Engine Plants 1... Group, LLC, Power Train Division, Mack Avenue Engine Plant 1, including on-site leased workers of... period at the Mack Avenue Engine Plant 2, Detroit, Michigan location of Chrysler Group, LLC, Power...

  15. Metabolic engineering of geranic acid in maize to achieve fungal resistance is compromised by novel glycosylation patterns.

    Science.gov (United States)

    Yang, Ting; Stoopen, Geert; Yalpani, Nasser; Vervoort, Jacques; de Vos, Ric; Voster, Alessandra; Verstappen, Francel W A; Bouwmeester, Harro J; Jongsma, Maarten A

    2011-07-01

    Many terpenoids are known to have antifungal properties and overexpression of these compounds in crops is a potential tool in disease control. In this study, 15 different mono- and sesquiterpenoids were tested in vitro against two major pathogenic fungi of maize (Zea mays), Colletotrichum graminicola and Fusarium graminearum. Among all tested terpenoids, geranic acid showed very strong inhibitory activity against both fungi (MICgraminicola and F. graminearum did not reveal an effect of these changes in secondary metabolite composition on plant resistance to either fungus. The results demonstrate that metabolic engineering of geraniol into geranic acid can rely on the existing default pathway, but branching glycosylation pathways must be controlled to achieve accumulation of the aglycones. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Natural toxins that affect plant amino acid metabolism

    Science.gov (United States)

    A diverse range of natural compounds interfere with the synthesis and other aspects of amino acid metabolism. Some are amino acid analogues, but most are not. This review covers a number of specific natural phytotoxic compounds by molecular target site. Inhibition of glutamine synthetase is of part...

  17. Transcription activator-like effector nucleases mediated metabolic engineering for enhanced fatty acids production in Saccharomyces cerevisiae.

    Science.gov (United States)

    Aouida, Mustapha; Li, Lixin; Mahjoub, Ali; Alshareef, Sahar; Ali, Zahir; Piatek, Agnieszka; Mahfouz, Magdy M

    2015-10-01

    Targeted engineering of microbial genomes holds much promise for diverse biotechnological applications. Transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/Cas9 systems are capable of efficiently editing microbial genomes, including that of Saccharomyces cerevisiae. Here, we demonstrate the use of TALENs to edit the genome of S. cerevisiae with the aim of inducing the overproduction of fatty acids. Heterodimeric TALENs were designed to simultaneously edit the FAA1 and FAA4 genes encoding acyl-CoA synthetases in S. cerevisiae. Functional yeast double knockouts generated using these TALENs over-produce large amounts of free fatty acids into the cell. This study demonstrates the use of TALENs for targeted engineering of yeast and demonstrates that this technology can be used to stimulate the enhanced production of free fatty acids, which are potential substrates for biofuel production. This proof-of-principle study extends the utility of TALENs as excellent genome editing tools and highlights their potential use for metabolic engineering of yeast and other organisms, such as microalgae and plants, for biofuel production.

  18. Transcription activator-like effector nucleases mediated metabolic engineering for enhanced fatty acids production in Saccharomyces cerevisiae

    KAUST Repository

    Aouida, Mustapha

    2015-04-01

    Targeted engineering of microbial genomes holds much promise for diverse biotechnological applications. Transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/Cas9 systems are capable of efficiently editing microbial genomes, including that of Saccharomyces cerevisiae. Here, we demonstrate the use of TALENs to edit the genome of S.cerevisiae with the aim of inducing the overproduction of fatty acids. Heterodimeric TALENs were designed to simultaneously edit the FAA1 and FAA4 genes encoding acyl-CoA synthetases in S.cerevisiae. Functional yeast double knockouts generated using these TALENs over-produce large amounts of free fatty acids into the cell. This study demonstrates the use of TALENs for targeted engineering of yeast and demonstrates that this technology can be used to stimulate the enhanced production of free fatty acids, which are potential substrates for biofuel production. This proof-of-principle study extends the utility of TALENs as excellent genome editing tools and highlights their potential use for metabolic engineering of yeast and other organisms, such as microalgae and plants, for biofuel production. © 2015 The Society for Biotechnology, Japan.

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

    Science.gov (United States)

    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.

  20. Metabolic regulation of the plant hormone indole-3-acetic acid

    Energy Technology Data Exchange (ETDEWEB)

    Jerry D. Cohen

    2009-11-01

    The phytohormone indole-3-acetic acid (IAA, auxin) is important for many aspects of plant growth, development and responses to the environment yet the routes to is biosynthesis and mechanisms for regulation of IAA levels remain important research questions. A critical issue concerning the biosynthesis if IAA in plants is that redundant pathways for IAA biosynthesis exist in plants. We showed that these redundant pathways and their relative contribution to net IAA production are under both developmental and environmental control. We worked on three fundamental problems related to how plants get their IAA: 1) An in vitro biochemical approach was used to define the tryptophan dependent pathway to IAA using maize endosperm, where relatively large amounts of IAA are produced over a short developmental period. Both a stable isotope dilution and a protein MS approach were used to identify intermediates and enzymes in the reactions. 2) We developed an in vitro system for analysis of tryptophan-independent IAA biosynthesis in maize seedlings and we used a metabolite profiling approach to isolate intermediates in this reaction. 3) Arabidopsis contains a small family of genes that encode potential indolepyruvate decarboxylase enzymes. We cloned these genes and studied plants that are mutant in these genes and that over-express each member in the family in terms of the level and route of IAA biosynthesis. Together, these allowed further development of a comprehensive picture of the pathways and regulatory components that are involved in IAA homeostasis in higher plants.