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Sample records for cerevisiae strain engineered

  1. Enhanced xylose fermentation and ethanol production by engineered Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Vilela, Leonardo de Figueiredo; de Araujo, Verônica Parente Gomes; Paredes, Raquel de Sousa; Bon, Elba Pinto da Silva; Torres, Fernando Araripe Gonçalves; Neves, Bianca Cruz; Eleutherio, Elis Cristina Araújo

    2015-01-01

    We have recently demonstrated that heterologous expression of a bacterial xylose isomerase gene (xylA) of Burkholderia cenocepacia enabled a laboratorial Saccharomyces cerevisiae strain to ferment xylose anaerobically, without xylitol accumulation. However, the recombinant yeast fermented xylose slowly. In this study, an evolutionary engineering strategy was applied to improve xylose fermentation by the xylA-expressing yeast strain, which involved sequential batch cultivation on xylose. The resulting yeast strain co-fermented glucose and xylose rapidly and almost simultaneously, exhibiting improved ethanol production and productivity. It was also observed that when cells were grown in a medium containing higher glucose concentrations before being transferred to fermentation medium, higher rates of xylose consumption and ethanol production were obtained, demonstrating that xylose utilization was not regulated by catabolic repression. Results obtained by qPCR demonstrate that the efficiency in xylose fermentation showed by the evolved strain is associated, to the increase in the expression of genes HXT2 and TAL1, which code for a low-affinity hexose transporter and transaldolase, respectively. The ethanol productivity obtained after the introduction of only one genetic modification and the submission to a one-stage process of evolutionary engineering was equivalent to those of strains submitted to extensive metabolic and evolutionary engineering, providing solid basis for future applications of this strategy in industrial strains. PMID:25852993

  2. Comparison of xylose fermentation by two high-performance engineered strains of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Xin Li

    2016-03-01

    Full Text Available Economical biofuel production from plant biomass requires the conversion of both cellulose and hemicellulose in the plant cell wall. The best industrial fermentation organism, the yeast Saccharomyces cerevisiae, has been developed to utilize xylose by heterologously expressing either a xylose reductase/xylitol dehydrogenase (XR/XDH pathway or a xylose isomerase (XI pathway. Although it has been proposed that the optimal means for fermenting xylose into biofuels would use XI instead of the XR/XDH pathway, no clear comparison of the best publicly-available yeast strains engineered to use XR/XDH or XI has been published. We therefore compared two of the best-performing engineered yeast strains in the public domain—one using the XR/XDH pathway and another using XI—in anaerobic xylose fermentations. We find that, regardless of conditions, the strain using XR/XDH has substantially higher productivity compared to the XI strain. By contrast, the XI strain has better yields in nearly all conditions tested.

  3. Key Process Conditions for Production of C4 Dicarboxylic Acids in Bioreactor Batch Cultures of an Engineered Saccharomyces cerevisiae Strain

    NARCIS (Netherlands)

    Zelle, R.M.; De Hulster, E.; Kloezen, W.; Pronk, J.T.; Van Maris, A.J.A.

    2009-01-01

    A recent effort to improve malic acid production by Saccharomyces cerevisiae by means of metabolic engineering resulted in a strain that produced up to 59 g liter−1 of malate at a yield of 0.42 mol (mol glucose)−1 in calcium carbonate-buffered shake flask cultures. With shake flasks, process paramet

  4. Key Process Conditions for Production of C4 Dicarboxylic Acids in Bioreactor Batch Cultures of an Engineered Saccharomyces cerevisiae Strain

    NARCIS (Netherlands)

    Zelle, R.M.; De Hulster, E.; Kloezen, W.; Pronk, J.T.; Van Maris, A.J.A.

    2010-01-01

    A recent effort to improve malic acid production by Saccharomyces cerevisiae by means of metabolic engineering resulted in a strain that produced up to 59 g liter(-1) of malate at a yield of 0.42 mol (mol glucose)(-1) in calcium carbonate-buffered shake flask cultures. With shake flasks, process par

  5. Improving 2-phenylethanol production via Ehrlich pathway using genetic engineered Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Yin, Sheng; Zhou, Hui; Xiao, Xiao; Lang, Tiandan; Liang, Jingru; Wang, Chengtao

    2015-05-01

    2-phenylethanol (2-PE) is an important aromatic compound with a rose-like fragrance widely used in food industry and cosmetic manufacture. In order to obtain "natural" 2-PE, the genetically modified budding yeasts were developed and applied for the 2-PE production. The gene ARO8 encoding transaminase and the gene ARO10 encoding decarboxylase in the Ehrlich pathway were expressed in Saccharomyces cerevisiae S288c. The activities of transaminase and decarboxylase were both enhanced in the corresponding recombinant strains. Consequently, the 2-PE yield in the recombinant strains with ARO8 and ARO10 were increased by 9.3 and 16.3 %, respectively, than that in the wild strain. A co-expression vector harboring ARO8 and ARO10 was then introduced into S. cerevisiae S288c, generating the recombinant strain SPO810. The fed-batch fermentation results indicated that the 2-PE yield in SPO810 reached 2.61 g L(-1) after 60 h of cultivation, which was 36.8 % higher than that in the wild strain. These results demonstrated that the 2-PE production was significantly improved by enhanced expression of the two key enzymes encoded by ARO8 and ARO10 in the Ehrlich pathway, providing new perspectives for enhancing "natural" 2-PE production in S. cerevisiae. PMID:25681107

  6. Engineering industrial Saccharomyces cerevisiae strains for xylose fermentation and comparison for switchgrass conversion

    Science.gov (United States)

    Saccharomyces physiology and fermentation related properties vary broadly among industrial strains. In this study, six industrial strains of varied genetic background were engineered to ferment xylose. Aerobic growth rates on xylose were 0.040 h**-1 to 0.167 h**-1. Fermentation of xylose, glucose/xy...

  7. Engineering and two-stage evolution of a lignocellulosic hydrolysate-tolerant Saccharomyces cerevisiae strain for anaerobic fermentation of xylose from AFEX pretreated corn stover.

    Science.gov (United States)

    Parreiras, Lucas S; Breuer, Rebecca J; Avanasi Narasimhan, Ragothaman; Higbee, Alan J; La Reau, Alex; Tremaine, Mary; Qin, Li; Willis, Laura B; Bice, Benjamin D; Bonfert, Brandi L; Pinhancos, Rebeca C; Balloon, Allison J; Uppugundla, Nirmal; Liu, Tongjun; Li, Chenlin; Tanjore, Deepti; Ong, Irene M; Li, Haibo; Pohlmann, Edward L; Serate, Jose; Withers, Sydnor T; Simmons, Blake A; Hodge, David B; Westphall, Michael S; Coon, Joshua J; Dale, Bruce E; Balan, Venkatesh; Keating, David H; Zhang, Yaoping; Landick, Robert; Gasch, Audrey P; Sato, Trey K

    2014-01-01

    The inability of the yeast Saccharomyces cerevisiae to ferment xylose effectively under anaerobic conditions is a major barrier to economical production of lignocellulosic biofuels. Although genetic approaches have enabled engineering of S. cerevisiae to convert xylose efficiently into ethanol in defined lab medium, few strains are able to ferment xylose from lignocellulosic hydrolysates in the absence of oxygen. This limited xylose conversion is believed to result from small molecules generated during biomass pretreatment and hydrolysis, which induce cellular stress and impair metabolism. Here, we describe the development of a xylose-fermenting S. cerevisiae strain with tolerance to a range of pretreated and hydrolyzed lignocellulose, including Ammonia Fiber Expansion (AFEX)-pretreated corn stover hydrolysate (ACSH). We genetically engineered a hydrolysate-resistant yeast strain with bacterial xylose isomerase and then applied two separate stages of aerobic and anaerobic directed evolution. The emergent S. cerevisiae strain rapidly converted xylose from lab medium and ACSH to ethanol under strict anaerobic conditions. Metabolomic, genetic and biochemical analyses suggested that a missense mutation in GRE3, which was acquired during the anaerobic evolution, contributed toward improved xylose conversion by reducing intracellular production of xylitol, an inhibitor of xylose isomerase. These results validate our combinatorial approach, which utilized phenotypic strain selection, rational engineering and directed evolution for the generation of a robust S. cerevisiae strain with the ability to ferment xylose anaerobically from ACSH. PMID:25222864

  8. Engineering and two-stage evolution of a lignocellulosic hydrolysate-tolerant Saccharomyces cerevisiae strain for anaerobic fermentation of xylose from AFEX pretreated corn stover.

    Directory of Open Access Journals (Sweden)

    Lucas S Parreiras

    Full Text Available The inability of the yeast Saccharomyces cerevisiae to ferment xylose effectively under anaerobic conditions is a major barrier to economical production of lignocellulosic biofuels. Although genetic approaches have enabled engineering of S. cerevisiae to convert xylose efficiently into ethanol in defined lab medium, few strains are able to ferment xylose from lignocellulosic hydrolysates in the absence of oxygen. This limited xylose conversion is believed to result from small molecules generated during biomass pretreatment and hydrolysis, which induce cellular stress and impair metabolism. Here, we describe the development of a xylose-fermenting S. cerevisiae strain with tolerance to a range of pretreated and hydrolyzed lignocellulose, including Ammonia Fiber Expansion (AFEX-pretreated corn stover hydrolysate (ACSH. We genetically engineered a hydrolysate-resistant yeast strain with bacterial xylose isomerase and then applied two separate stages of aerobic and anaerobic directed evolution. The emergent S. cerevisiae strain rapidly converted xylose from lab medium and ACSH to ethanol under strict anaerobic conditions. Metabolomic, genetic and biochemical analyses suggested that a missense mutation in GRE3, which was acquired during the anaerobic evolution, contributed toward improved xylose conversion by reducing intracellular production of xylitol, an inhibitor of xylose isomerase. These results validate our combinatorial approach, which utilized phenotypic strain selection, rational engineering and directed evolution for the generation of a robust S. cerevisiae strain with the ability to ferment xylose anaerobically from ACSH.

  9. Engineering and Two-Stage Evolution of a Lignocellulosic Hydrolysate-Tolerant Saccharomyces cerevisiae Strain for Anaerobic Fermentation of Xylose from AFEX Pretreated Corn Stover

    OpenAIRE

    Parreiras, Lucas S.; Rebecca J Breuer; Avanasi Narasimhan, Ragothaman; Alan J Higbee; La Reau, Alex; Tremaine, Mary; Qin, Li; Willis, Laura B.; Bice, Benjamin D.; Bonfert, Brandi L.; Pinhancos, Rebeca C.; Balloon, Allison J.; Uppugundla, Nirmal; Liu, Tongjun; Li, Chenlin

    2014-01-01

    The inability of the yeast Saccharomyces cerevisiae to ferment xylose effectively under anaerobic conditions is a major barrier to economical production of lignocellulosic biofuels. Although genetic approaches have enabled engineering of S. cerevisiae to convert xylose efficiently into ethanol in defined lab medium, few strains are able to ferment xylose from lignocellulosic hydrolysates in the absence of oxygen. This limited xylose conversion is believed to result from small molecules genera...

  10. Xylose utilizing recombinant Saccharomyces cerevisiae strains

    Energy Technology Data Exchange (ETDEWEB)

    Walfridsson, M.

    1996-04-01

    Through metabolic engineering, S. cerevisiae was provided with the necessary enzymes required for xylose utilisation during ethanolic fermentation of xylose-rich lignocellulose raw materials. For xylitol production, S. cerevisiae was provided with the Pichia stipitis XYL1 gene encoding xylose reductase (XR). The in-vivo reduction and the following excretion of xylitol, requires a co-substrate for maintenance and cofactor regeneration. Xylitol yields close to 100% were obtained with the XYL1 containing S. cerevisiae. Introducing P. stipitis XYL1 and XYL2 genes, encoding XR and xylitol dehydrogenase (XDH), respectively, enabled S. cerevisiae to convert xylose to xylulose, via xylitol. During the screening work of P. stipitis XDH gene, another gene encoding a polyol dehydrogenase was isolated and cloned in S. cerevisiae. The gene was identified as a D-arabinitol dehydrogenase gene. In P. stipitis it may function as a redox sink by reducing D-ribulose to D-arabinitol. The metabolism through the pentose phosphate pathway (PPP) was enhanced by over-expressing the native genes TKL1 and TAL1 encoding transketolase and transaldolase, respectively, resulting in improved xylose utilisation. The XR and XDH activities in recombinant S. cerevisiae were produced at different levels by constructing yeast vectors in which the PGK1 and ADHI promoters controlled XYL1 and XYL2. With higher XDH than XR activities, less by-products, in the form of xylitol and glycerol, were formed by the recombinant S. cerevisiae strains. The Thermus thermophilus xylA gene encoding a thermostable xylose isomerase was cloned and expressed in S. cerevisiae. The recombinant xylose isomerase was actively produced and a new functional metabolic pathway was established in S. cerevisiae resulting in ethanol production from xylose. 150 refs, 3 figs, 4 tabs

  11. Anaplerotic Role for Cytosolic Malic Enzyme in Engineered Saccharomyces cerevisiae Strains

    NARCIS (Netherlands)

    Zelle, R.M.; Harrison, J.C.; Pronk, J.T.; Van Maris, A.J.A.

    2010-01-01

    Malic enzyme catalyzes the reversible oxidative decarboxylation of malate to pyruvate and CO2. The Saccharomyces cerevisiae MAE1 gene encodes a mitochondrial malic enzyme whose proposed physiological roles are related to the oxidative, malate-decarboxylating reaction. Hitherto, the inability of pyru

  12. Comparison of xylose fermentation by two high-performance engineered strains of Saccharomyces cerevisiae

    OpenAIRE

    Xin Li; Annsea Park; Raissa Estrela; Soo-Rin Kim; Yong-Su Jin; Cate, Jamie H. D.

    2016-01-01

    Economical biofuel production from plant biomass requires the conversion of both cellulose and hemicellulose in the plant cell wall. The best industrial fermentation organism, the yeast Saccharomyces cerevisiae, has been developed to utilize xylose by heterologously expressing either a xylose reductase/xylitol dehydrogenase (XR/XDH) pathway or a xylose isomerase (XI) pathway. Although it has been proposed that the optimal means for fermenting xylose into biofuels would use XI instead of the X...

  13. Improved xylose and arabinose utilization by an industrial recombinant Saccharomyces cerevisiae strain using evolutionary engineering

    DEFF Research Database (Denmark)

    Sanchez, R.G.; Karhumaa, Kaisa; Fonseca, C.;

    2010-01-01

    xylose and arabinose under aerobic and anaerobic conditions. Improved anaerobic ethanol production was achieved at the expense of xylitol and glycerol but arabinose was almost stoichiometrically converted to arabitol. Further characterization of the strain indicated that the selection pressure during...

  14. One-pot production of fructooligosaccharides by a Saccharomyces cerevisiae strain expressing an engineered invertase.

    Science.gov (United States)

    Marín-Navarro, Julia; Talens-Perales, David; Polaina, Julio

    2015-03-01

    We describe a simple, efficient process for the production of 6-kestose, a trisaccharide with well-documented prebiotic properties. A key factor is the use of a yeast transformant expressing an engineered version of Saccharomyces invertase with enhanced transfructosylating activity. When the yeast transformant was grown with 30 % sucrose as the carbon source, 6-kestose accumulated up to ca. 100 g/L in the culture medium. The 6-kestose yield was significantly enhanced (up to 200 g/L) using a two-stage process carried out in the same flask. In the first stage, the culture was grown in 30 % sucrose at physiological temperature (30 °C) to allow overexpression of the invertase. In the second stage, sucrose was added to the culture at high concentration (60 %) and the temperature shifted to 50 °C. In both cases, 6-kestose was synthesized with high specificity, representing more than 95 % of total FOS. PMID:25547837

  15. 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...

  16. Engineering of a Nepetalactol-Producing Platform Strain of Saccharomyces cerevisiae for the Production of Plant Seco-Iridoids.

    Science.gov (United States)

    Campbell, Alex; Bauchart, Philippe; Gold, Nicholas D; Zhu, Yun; De Luca, Vincenzo; Martin, Vincent J J

    2016-05-20

    The monoterpene indole alkaloids (MIAs) are a valuable family of chemicals that include the anticancer drugs vinblastine and vincristine. These compounds are of global significance-appearing on the World Health Organization's list of model essential medicines-but remain exorbitantly priced due to low in planta levels. Chemical synthesis and genetic manipulation of MIA producing plants such as Catharanthus roseus have so far failed to find a solution to this problem. Synthetic biology holds a potential answer, by building the pathway into more tractable organisms such as Saccharomyces cerevisiae. Recent work has taken the first steps in this direction by producing small amounts of the intermediate strictosidine in yeast. In order to help improve on these titers, we aimed to optimize the early biosynthetic steps of the MIA pathway to the metabolite nepetalactol. We combined a number of strategies to create a base strain producing 11.4 mg/L of the precursor geraniol. We also show production of the critical intermediate 10-hydroxygeraniol and demonstrate nepetalactol production in vitro. Lastly we demonstrate that activity of the iridoid synthase toward the intermediates geraniol and 10-hydroxygeraniol results in the synthesis of the nonproductive intermediates citronellol and 10-hydroxycitronellol. This discovery has serious implications for the reconstruction of the MIA in heterologous organisms. PMID:26981892

  17. Metabolic pathway engineering based on metabolomics confers acetic and formic acid tolerance to a recombinant xylose-fermenting strain of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Ishii Jun

    2011-01-01

    Full Text Available Abstract Background The development of novel yeast strains with increased tolerance toward inhibitors in lignocellulosic hydrolysates is highly desirable for the production of bio-ethanol. Weak organic acids such as acetic and formic acids are necessarily released during the pretreatment (i.e. solubilization and hydrolysis of lignocelluloses, which negatively affect microbial growth and ethanol production. However, since the mode of toxicity is complicated, genetic engineering strategies addressing yeast tolerance to weak organic acids have been rare. Thus, enhanced basic research is expected to identify target genes for improved weak acid tolerance. Results In this study, the effect of acetic acid on xylose fermentation was analyzed by examining metabolite profiles in a recombinant xylose-fermenting strain of Saccharomyces cerevisiae. Metabolome analysis revealed that metabolites involved in the non-oxidative pentose phosphate pathway (PPP [e.g. sedoheptulose-7-phosphate, ribulose-5-phosphate, ribose-5-phosphate and erythrose-4-phosphate] were significantly accumulated by the addition of acetate, indicating the possibility that acetic acid slows down the flux of the pathway. Accordingly, a gene encoding a PPP-related enzyme, transaldolase or transketolase, was overexpressed in the xylose-fermenting yeast, which successfully conferred increased ethanol productivity in the presence of acetic and formic acid. Conclusions Our metabolomic approach revealed one of the molecular events underlying the response to acetic acid and focuses attention on the non-oxidative PPP as a target for metabolic engineering. An important challenge for metabolic engineering is identification of gene targets that have material importance. This study has demonstrated that metabolomics is a powerful tool to develop rational strategies to confer tolerance to stress through genetic engineering.

  18. A coniferyl aldehyde dehydrogenase gene from Pseudomonas sp. strain HR199 enhances the conversion of coniferyl aldehyde by Saccharomyces cerevisiae.

    Science.gov (United States)

    Adeboye, Peter Temitope; Olsson, Lisbeth; Bettiga, Maurizio

    2016-07-01

    The conversion of coniferyl aldehyde to cinnamic acids by Saccharomyces cerevisiae under aerobic growth conditions was previously observed. Bacteria such as Pseudomonas have been shown to harbor specialized enzymes for converting coniferyl aldehyde but no comparable enzymes have been identified in S. cerevisiae. CALDH from Pseudomonas was expressed in S. cerevisiae. An acetaldehyde dehydrogenase (Ald5) was also hypothesized to be actively involved in the conversion of coniferyl aldehyde under aerobic growth conditions in S. cerevisiae. In a second S. cerevisiae strain, the acetaldehyde dehydrogenase (ALD5) was deleted. A prototrophic control strain was also engineered. The engineered S. cerevisiae strains were cultivated in the presence of 1.1mM coniferyl aldehyde under aerobic condition in bioreactors. The results confirmed that expression of CALDH increased endogenous conversion of coniferyl aldehyde in S. cerevisiae and ALD5 is actively involved with the conversion of coniferyl aldehyde in S. cerevisiae. PMID:27070284

  19. Aerobic physiology of redox-engineered Saccharomyces cerevisiae strains modified in the ammonium assimilation for increased NADPH availability

    DEFF Research Database (Denmark)

    Santos, Maria Margarida M. dos; Thygesen, G.; Kotter, P.;

    2003-01-01

    Recombinant strains altered in the ammonium assimilation pathways were constructed with the purpose of increasing NADPH availability. The NADPH-dependent glutamate dehydrogenase encoded by GDH1, which accounts for a major fraction of the NADPH consumption during growth on ammonium, was deleted, a...... through the Embden-Meyerhof-Parnas pathway downstream of glucose 6-phosphate. No redox effect was observed in a strain containing a deletion of GLR1, encoding glutathione reductase, an enzyme that is NADPH-consuming....

  20. Automated Yeast Transformation Protocol to Engineer S. cerevisiae Strains for Cellulosic Ethanol Production with Open Reading Frames that Express Proteins Binding to Xylose Isomerase Identified using Robotic Two-hybrid Screen

    Science.gov (United States)

    Commercialization of fuel ethanol production from lignocellulosic biomass has focused on engineering the glucose-fermenting industrial yeast Saccharomyces cerevisiae to utilize pentose sugars. Since S. cerevisiae naturally metabolizes xylulose, one approach involves introducing xylose isomerase (XI...

  1. Glycerol Overproduction by Engineered Saccharomyces cerevisiae Wine Yeast Strains Leads to Substantial Changes in By-Product Formation and to a Stimulation of Fermentation Rate in Stationary Phase

    OpenAIRE

    Remize, F.; Roustan, J. L.; Sablayrolles, J. M.; P. Barre; Dequin, S.

    1999-01-01

    Six commercial wine yeast strains and three nonindustrial strains (two laboratory strains and one haploid strain derived from a wine yeast strain) were engineered to produce large amounts of glycerol with a lower ethanol yield. Overexpression of the GPD1 gene, encoding a glycerol-3-phosphate dehydrogenase, resulted in a 1.5- to 2.5-fold increase in glycerol production and a slight decrease in ethanol formation under conditions simulating wine fermentation. All the strains overexpressing GPD1 ...

  2. Linking Genotype and Phenotype of Saccharomyces cerevisiae Strains Reveals Metabolic Engineering Targets and Leads to Triterpene Hyper-Producers

    DEFF Research Database (Denmark)

    Madsen, Karina Marie; Udatha, Gupta D. B. R. K.; Semba, Saori;

    2011-01-01

    S288C strain, we implemented a strategy for the construction of a beta-amyrin production platform. The genes Erg8, Erg9 and HFA1 contained non-silent SNPs that were computationally analyzed to evaluate the changes that cause in the respective protein structures. Subsequently, Erg8, Erg9 and HFA1...

  3. Aerobic physiology of redox-engineered Saccharomyces cerevisiae strains modified in the ammonium assimilation for increased NADPH availability

    DEFF Research Database (Denmark)

    Santos, Maria Margarida M. dos; Thygesen, G.; Kotter, P.;

    2003-01-01

    Recombinant strains altered in the ammonium assimilation pathways were constructed with the purpose of increasing NADPH availability. The NADPH-dependent glutamate dehydrogenase encoded by GDH1, which accounts for a major fraction of the NADPH consumption during growth on ammonium, was deleted, a...

  4. Fermentation of mixed glucose-xylose substrates by engineered strains of Saccharomyces cerevisiae: role of the coenzyme specificity of xylose reductase, and effect of glucose on xylose utilization

    Directory of Open Access Journals (Sweden)

    Klimacek Mario

    2010-03-01

    Full Text Available Abstract Background In spite of the substantial metabolic engineering effort previously devoted to the development of Saccharomyces cerevisiae strains capable of fermenting both the hexose and pentose sugars present in lignocellulose hydrolysates, the productivity of reported strains for conversion of the naturally most abundant pentose, xylose, is still a major issue of process efficiency. Protein engineering for targeted alteration of the nicotinamide cofactor specificity of enzymes catalyzing the first steps in the metabolic pathway for xylose was a successful approach of reducing xylitol by-product formation and improving ethanol yield from xylose. The previously reported yeast strain BP10001, which expresses heterologous xylose reductase from Candida tenuis in mutated (NADH-preferring form, stands for a series of other yeast strains designed with similar rational. Using 20 g/L xylose as sole source of carbon, BP10001 displayed a low specific uptake rate qxylose (g xylose/g dry cell weight/h of 0.08. The study presented herein was performed with the aim of analysing (external factors that limit qxylose of BP10001 under xylose-only and mixed glucose-xylose substrate conditions. We also carried out a comprehensive investigation on the currently unclear role of coenzyme utilization, NADPH compared to NADH, for xylose reduction during co-fermentation of glucose and xylose. Results BP10001 and BP000, expressing C. tenuis xylose reductase in NADPH-preferring wild-type form, were used. Glucose and xylose (each at 10 g/L were converted sequentially, the corresponding qsubstrate values being similar for each strain (glucose: 3.0; xylose: 0.05. The distribution of fermentation products from glucose was identical for both strains whereas when using xylose, BP10001 showed enhanced ethanol yield (BP10001 0.30 g/g; BP000 0.23 g/g and decreased yields of xylitol (BP10001 0.26 g/g; BP000 0.36 g/g and glycerol (BP10001 0.023 g/g; BP000 0.072 g/g as compared

  5. Heterologous expression of cellulase genes in natural Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Davison, Steffi A; den Haan, Riaan; van Zyl, Willem Heber

    2016-09-01

    Enzyme cost is a major impediment to second-generation (2G) cellulosic ethanol production. One strategy to reduce enzyme cost is to engineer enzyme production capacity in a fermentative microorganism to enable consolidated bio-processing (CBP). Ideally, a strain with a high secretory phenotype, high fermentative capacity as well as an innate robustness to bioethanol-specific stressors, including tolerance to products formed during pre-treatment and fermentation of lignocellulosic substrates should be used. Saccharomyces cerevisiae is a robust fermentative yeast but has limitations as a potential CBP host, such as low heterologous protein secretion titers. In this study, we evaluated natural S. cerevisiae isolate strains for superior secretion activity and other industrially relevant characteristics needed during the process of lignocellulosic ethanol production. Individual cellulases namely Saccharomycopsis fibuligera Cel3A (β-glucosidase), Talaromyces emersonii Cel7A (cellobiohydrolase), and Trichoderma reesei Cel5A (endoglucanase) were utilized as reporter proteins. Natural strain YI13 was identified to have a high secretory phenotype, demonstrating a 3.7- and 3.5-fold higher Cel7A and Cel5A activity, respectively, compared to the reference strain S288c. YI13 also demonstrated other industrially relevant characteristics such as growth vigor, high ethanol titer, multi-tolerance to high temperatures (37 and 40 °C), ethanol (10 % w/v), and towards various concentrations of a cocktail of inhibitory compounds commonly found in lignocellulose hydrolysates. This study accentuates the value of natural S. cerevisiae isolate strains to serve as potential robust and highly productive chassis organisms for CBP strain development. PMID:27470141

  6. Redox balancing in recombinant strains of Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Anderlund, M.

    1998-09-01

    In metabolically engineered Saccharomyces cerevisiae expressing Pichia stipitis XYL1 and XYL2 genes, encoding xylose reductase (XR) and xylitol dehydrogenase (XDH), respectively, xylitol is excreted as the major product during anaerobic xylose fermentation and only low yields of ethanol are produced. This has been interpreted as a result of the dual cofactor dependence of XR and the exclusive use of NAD{sup +} by XDH. The excretion of xylitol was completely stopped and the formation of glycerol and acetic acid were reduced in xylose utilising S. cerevisiae strains cultivated in oxygen-limited conditions by expressing lower levels of XR than of XDH. The expression level of XYL1 and XYL2 were controlled by changing the promoters and transcription directions of the genes. A new functional metabolic pathway was established when Thermus thermophilus xylA gene was expressed in S. cerevisiae. The recombinant strain was able to ferment xylose to ethanol when cultivated on a minimal medium containing xylose as only carbon source. In order to create a channeled metabolic transfer in the two first steps of the xylose metabolism, XYL1 and XYL2 were fused in-frame and expressed in S. cerevisiae. When the fusion protein, containing a linker of three amino acids, was co expressed together with native XR and XDH monomers, enzyme complexes consisting of chimeric and native subunits were formed. The total activity of these complexes exhibited 10 and 9 times higher XR and XDH activity, respectively, than the original conjugates, consisting of only chimeric subunits. This strain produced less xylitol and the xylitol yield was lower than with strains only expressing native XR and XDH monomers. In addition, more ethanol and less acetic acid were formed. A new gene encoding the cytoplasmic transhydrogenase from Azotobacter vinelandii was cloned. The enzyme showed high similarity to the family of pyridine nucleotide-disulphide oxidoreductase. To analyse the physiological effect of

  7. EasyClone 2.0:expanded toolkit of integrative vectors for stable gene expression in industrial Saccharomyces cerevisiae strains

    OpenAIRE

    Stovicek, Vratislav; Borja Zamfir, Gheorghe Manuel; Förster, Jochen; Borodina, Irina

    2015-01-01

    Saccharomyces cerevisiae is one of the key cell factories for production of chemicals and active pharmaceuticals. For large-scale fermentations, particularly in biorefinery applications, it is desirable to use stress-tolerant industrial strains. However, such strains are less amenable for metabolic engineering than the standard laboratory strains. To enable easy delivery and overexpression of genes in a wide range of industrial S. cerevisiae strains, we constructed a set of integrative vector...

  8. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains

    Science.gov (United States)

    Šuranská, Hana; Vránová, Dana; Omelková, Jiřina

    2016-01-01

    In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir) were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic) wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were employed to differentiate among indigenous S. cerevisiae strains. This combination of the methods provides a rapid and relatively simple approach for identification of yeast of S. cerevisiae at strain level. In total, 120 isolates were identified and grouped by molecular approaches and 45 of the representative strains were tested for selected important oenological properties including ethanol, sulfur dioxide and osmotic stress tolerance, intensity of flocculation and desirable enzymatic activities. Their ability to produce and utilize acetic/malic acid was examined as well; in addition, H2S production as an undesirable property was screened. The oenological characteristics of indigenous isolates were compared to a commercially available S. cerevisiae BS6 strain, which is commonly used as the starter culture. Finally, some indigenous strains coming from organically treated grape berries were chosen for their promising oenological properties and these strains will be used as the starter culture, because application of a selected indigenous S. cerevisiae strain can enhance the regional character of the wines. PMID:26887243

  9. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Šuranská, Hana; Vránová, Dana; Omelková, Jiřina

    2016-01-01

    In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir) were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic) wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were employed to differentiate among indigenous S. cerevisiae strains. This combination of the methods provides a rapid and relatively simple approach for identification of yeast of S. cerevisiae at strain level. In total, 120 isolates were identified and grouped by molecular approaches and 45 of the representative strains were tested for selected important oenological properties including ethanol, sulfur dioxide and osmotic stress tolerance, intensity of flocculation and desirable enzymatic activities. Their ability to produce and utilize acetic/malic acid was examined as well; in addition, H2S production as an undesirable property was screened. The oenological characteristics of indigenous isolates were compared to a commercially available S. cerevisiae BS6 strain, which is commonly used as the starter culture. Finally, some indigenous strains coming from organically treated grape berries were chosen for their promising oenological properties and these strains will be used as the starter culture, because application of a selected indigenous S. cerevisiae strain can enhance the regional character of the wines. PMID:26887243

  10. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Hana Šuranská

    2016-03-01

    Full Text Available Abstract In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were employed to differentiate among indigenous S. cerevisiae strains. This combination of the methods provides a rapid and relatively simple approach for identification of yeast of S. cerevisiae at strain level. In total, 120 isolates were identified and grouped by molecular approaches and 45 of the representative strains were tested for selected important oenological properties including ethanol, sulfur dioxide and osmotic stress tolerance, intensity of flocculation and desirable enzymatic activities. Their ability to produce and utilize acetic/malic acid was examined as well; in addition, H2S production as an undesirable property was screened. The oenological characteristics of indigenous isolates were compared to a commercially available S. cerevisiae BS6 strain, which is commonly used as the starter culture. Finally, some indigenous strains coming from organically treated grape berries were chosen for their promising oenological properties and these strains will be used as the starter culture, because application of a selected indigenous S. cerevisiae strain can enhance the regional character of the wines.

  11. Investigation of fatty acid accumulation in the engineered Saccharomyces cerevisiae under nitrogen limited culture condition.

    Science.gov (United States)

    Tang, Xiaoling; Chen, Wei Ning

    2014-06-01

    In this study, the Saccharomyces cerevisiae wild type strain and engineered strain with an overexpressed heterologous ATP-citrate lyase (acl) were cultured in medium with different carbon and nitrogen concentrations, and their fatty acid production levels were investigated. The results showed that when the S. cerevisiae engineered strain was cultivated under nitrogen limited culture condition, the yield of mono-unsaturated fatty acids showed higher than that under non-nitrogen limited condition; with the carbon concentration increased, the accumulation become more apparent, whereas in the wild type strain, no such correlation was found. Besides, the citrate level in the S. cerevisiae under nitrogen limited condition was found to be much higher than that under non-nitrogen limited condition, which indicated a relationship between the diminution of nitrogen and accumulation of citrate in the S. cerevisiae. The accumulated citrate could be further cleaved by acl to provide substrate for fatty acid synthesis. PMID:24755317

  12. Co-utilization of L-arabinose and D-xylose by laboratory and industrial Saccharomyces cerevisiae strains

    OpenAIRE

    Boles Eckhard; Hahn-Hägerdal Bärbel; Wiedemann Beate; Karhumaa Kaisa; Gorwa-Grauslund Marie-F

    2006-01-01

    Abstract Background Fermentation of lignocellulosic biomass is an attractive alternative for the production of bioethanol. Traditionally, the yeast Saccharomyces cerevisiae is used in industrial ethanol fermentations. However, S. cerevisiae is naturally not able to ferment the pentose sugars D-xylose and L-arabinose, which are present in high amounts in lignocellulosic raw materials. Results We describe the engineering of laboratory and industrial S. cerevisiae strains to co-ferment the pento...

  13. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains

    OpenAIRE

    Hana Šuranská; Dana Vránová; Jiřina Omelková

    2016-01-01

    Abstract In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir) were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic) wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typ...

  14. Isolation, identification and characterization of regional indigenous Saccharomyces cerevisiae strains

    OpenAIRE

    Šuranská, Hana; Vránová, Dana; Omelková, Jiřina

    2016-01-01

    In the present work we isolated and identified various indigenous Saccharomyces cerevisiae strains and screened them for the selected oenological properties. These S. cerevisiae strains were isolated from berries and spontaneously fermented musts. The grape berries (Sauvignon blanc and Pinot noir) were grown under the integrated and organic mode of farming in the South Moravia (Czech Republic) wine region. Modern genotyping techniques such as PCR-fingerprinting and interdelta PCR typing were ...

  15. Strain-engineered MOSFETs

    CERN Document Server

    Maiti, CK

    2012-01-01

    Currently strain engineering is the main technique used to enhance the performance of advanced silicon-based metal-oxide-semiconductor field-effect transistors (MOSFETs). Written from an engineering application standpoint, Strain-Engineered MOSFETs introduces promising strain techniques to fabricate strain-engineered MOSFETs and to methods to assess the applications of these techniques. The book provides the background and physical insight needed to understand new and future developments in the modeling and design of n- and p-MOSFETs at nanoscale. This book focuses on recent developments in st

  16. Malic acid production by Saccharomyces cerevisiae: engineering of pyruvate carbosylation, oxaloacetate reduction and malate export

    NARCIS (Netherlands)

    Zelle, R.M.; Hulster, de E.; Winden, van W.A.; Waard, de P.; Dijkema, C.; Winkler, A.A.; Geertman, J.M.A.

    2008-01-01

    Malic acid is a potential biomass-derivable "building block" for chemical synthesis. Since wild-type Saccharomyces cerevisiae strains produce only low levels of malate, metabolic engineering is required to achieve efficient malate production with this yeast. A promising pathway for malate production

  17. Malic Acid Production by Saccharomyces cerevisiae: Engineering of Pyruvate Carboxylation, Oxaloacetate Reduction, and Malate Export

    NARCIS (Netherlands)

    Zelle, R.M.; De Hulster, E.; Van Winden, W.A.; De Waard, P.; Dijkema, C.; Winkler, A.A.; Geertman, J.M.; Van Dijken, J.P.; Pronk, J.T.; Van Maris, A.J.A.

    2008-01-01

    Malic acid is a potential biomass-derivable "building block" for chemical synthesis. Since wild-type Saccharomyces cerevisiae strains produce only low levels of malate, metabolic engineering is required to achieve efficient malate production with this yeast. A promising pathway for malate production

  18. Evolutionary engineering of Saccharomyces cerevisiae for efficient aerobic xylose consumption

    DEFF Research Database (Denmark)

    Scalcinati, Gionata; Otero, José Manuel; Van Vleet, Jennifer R. H.;

    2012-01-01

    flux to biomass production. Such a platform may then be enhanced with complementary metabolic engineering strategies that couple biomass production with high value-added chemical. Saccharomyces cerevisiae, expressing xylose reductase, xylitol dehydrogenase and xylulose kinase, from the native xylose...

  19. Evaluation of cytochrome P-450 concentration in Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Míriam Cristina Sakuragui Matuo

    2010-09-01

    Full Text Available Saccharomyces cerevisiae has been widely used in mutagenicity tests due to the presence of a cytochrome P-450 system, capable of metabolizing promutagens to active mutagens. There are a large number of S. cerevisiae strains with varying abilities to produce cytochrome P-450. However, strain selection and ideal cultivation conditions are not well defined. We compared cytochrome P-450 levels in four different S. cerevisiae strains and evaluated the cultivation conditions necessary to obtain the highest levels. The amount of cytochrome P-450 produced by each strain varied, as did the incubation time needed to reach the maximum level. The highest cytochrome P-450 concentrations were found in media containing fermentable sugars. The NCYC 240 strain produced the highest level of cytochrome P-450 when grown in the presence of 20 % (w/v glucose. The addition of ethanol to the media also increased cytochrome P-450 synthesis in this strain. These results indicate cultivation conditions must be specific and well-established for the strain selected in order to assure high cytochrome P-450 levels and reliable mutagenicity results.Linhagens de Saccharomyces cerevisiae tem sido amplamente empregadas em testes de mutagenicidade devido à presença de um sistema citocromo P-450 capaz de metabolizar substâncias pró-mutagênicas à sua forma ativa. Devido à grande variedade de linhagens de S. cerevisiae com diferentes capacidades de produção de citocromo P-450, torna-se necessária a seleção de cepas, bem como a definição das condições ideais de cultivo. Neste trabalho, foram comparados os níveis de citocromo P-450 em quatro diferentes linhagens de S. cerevisiae e avaliadas as condições de cultivo necessárias para obtenção de altas concentrações deste sistema enzimático. O maior nível enzimático foi encontrado na linhagem NCYC 240 em presença de 20 % de glicose (p/v. A adição de etanol ao meio de cultura também produziu um aumento na s

  20. Mead production: selection and characterization assays of Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Pereira, Ana Paula; Dias, Teresa; Andrade, João; Ramalhosa, Elsa; Estevinho, Letícia M

    2009-08-01

    Mead is a traditional drink, which results from the alcoholic fermentation of diluted honey carried out by yeasts. However, when it is produced in a homemade way, mead producers find several problems, namely, the lack of uniformity in the final product, delayed and arrested fermentations, and the production of "off-flavours" by the yeasts. These problems are usually associated with the inability of yeast strains to respond and adapt to unfavourable and stressful growth conditions. The main objectives of this work were to evaluate the capacity of Saccharomyces cerevisiae strains, isolated from honey of the Trás-os-Montes (Northeast Portugal), to produce mead. Five strains from honey, as well as one laboratory strain and one commercial wine strain, were evaluated in terms of their fermentation performance under ethanol, sulphur dioxide and osmotic stress. All the strains showed similar behaviour in these conditions. Two yeasts strains isolated from honey and the commercial wine strain were further tested for mead production, using two different honey (a dark and a light honey), enriched with two supplements (one commercial and one developed by the research team), as fermentation media. The results obtained in this work show that S. cerevisiae strains isolated from honey, are appropriate for mead production. However it is of extreme importance to take into account the characteristics of the honey, and supplements used in the fermentation medium formulation, in order to achieve the best results in mead production. PMID:19481129

  1. Multiplex metabolic pathway engineering using CRISPR/Cas9 in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Jakociunas, Tadas; Bonde, Ida; Herrgard, Markus;

    2015-01-01

    CRISPR/Cas9 is a simple and efficient tool for targeted and marker-free genome engineering. Here, we report the development and successful application of a multiplex CRISPR/Cas9 system for genome engineering of up to 5 different genomic loci in one transformation step in baker's yeast Saccharomyces...... cerevisiae. To assess the specificity of the tool we employed genome re-sequencing to screen for off-target sites in all single knock-out strains targeted by different gRNAs. This extensive analysis identified no more genome variants in CRISPR/Cas9 engineered strains compared to wild-type reference strains...

  2. CRISPR-Cas9 Genome Engineering in Saccharomyces cerevisiae Cells.

    Science.gov (United States)

    Ryan, Owen W; Poddar, Snigdha; Cate, Jamie H D

    2016-01-01

    This protocol describes a method for CRISPR-Cas9-mediated genome editing that results in scarless and marker-free integrations of DNA into Saccharomyces cerevisiae genomes. DNA integration results from cotransforming (1) a single plasmid (pCAS) that coexpresses the Cas9 endonuclease and a uniquely engineered single guide RNA (sgRNA) expression cassette and (2) a linear DNA molecule that is used to repair the chromosomal DNA damage by homology-directed repair. For target specificity, the pCAS plasmid requires only a single cloning modification: replacing the 20-bp guide RNA sequence within the sgRNA cassette. This CRISPR-Cas9 protocol includes methods for (1) cloning the unique target sequence into pCAS, (2) assembly of the double-stranded DNA repair oligonucleotides, and (3) cotransformation of pCAS and linear repair DNA into yeast cells. The protocol is technically facile and requires no special equipment. It can be used in any S. cerevisiae strain, including industrial polyploid isolates. Therefore, this CRISPR-Cas9-based DNA integration protocol is achievable by virtually any yeast genetics and molecular biology laboratory. PMID:27250940

  3. Divergence in wine characteristics produced by wild and domesticated strains of Saccharomyces cerevisiae

    OpenAIRE

    Katie E Hyma; Saerens, Sofie M; Verstrepen, Kevin J.; Justin C Fay

    2011-01-01

    The budding yeast Saccharomyces cerevisiae is the primary species used by wine makers to convert sugar into alcohol during wine fermentation. Saccharomyces cerevisiae is found in vineyards, but is also found in association with oak trees and other natural sources. Although wild strains of S. cerevisiae as well as other Saccharomyces species are also capable of wine fermentation, a genetically distinct group of S. cerevisiae strains is primarily used to produce wine, consistent with the idea t...

  4. EasyClone 2.0: expanded toolkit of integrative vectors for stable gene expression in industrial Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Stovicek, Vratislav; Borja, Gheorghe M; Forster, Jochen; Borodina, Irina

    2015-11-01

    Saccharomyces cerevisiae is one of the key cell factories for production of chemicals and active pharmaceuticals. For large-scale fermentations, particularly in biorefinery applications, it is desirable to use stress-tolerant industrial strains. However, such strains are less amenable for metabolic engineering than the standard laboratory strains. To enable easy delivery and overexpression of genes in a wide range of industrial S. cerevisiae strains, we constructed a set of integrative vectors with long homology arms and dominant selection markers. The vectors integrate into previously validated chromosomal locations via double cross-over and result in homogenous stable expression of the integrated genes, as shown for several unrelated industrial strains. Cre-mediated marker rescue is possible for removing markers positioned on different chromosomes. To demonstrate the applicability of the presented vector set for metabolic engineering of industrial yeast, we constructed xylose-utilizing strains overexpressing xylose isomerase, xylose transporter and five genes of the pentose phosphate pathway. PMID:26376869

  5. Engineering of a novel cellulose-adherent cellulolytic Saccharomyces cerevisiae for cellulosic biofuel production

    OpenAIRE

    Zhuo Liu; Shih-Hsin Ho; Kengo Sasaki; Riaan den Haan; Kentaro Inokuma; Chiaki Ogino; van Zyl, Willem H; Tomohisa Hasunuma; Akihiko Kondo

    2016-01-01

    Cellulosic biofuel is the subject of increasing attention. The main obstacle toward its economic feasibility is the recalcitrance of lignocellulose requiring large amount of enzyme to break. Several engineered yeast strains have been developed with cellulolytic activities to reduce the need for enzyme addition, but exhibiting limited effect. Here, we report the successful engineering of a cellulose-adherent Saccharomyces cerevisiae displaying four different synergistic cellulases on the cell ...

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

    DEFF Research Database (Denmark)

    Borodina, Irina; Nielsen, Jens

    2014-01-01

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

  7. In vitro screening of probiotic properties of Saccharomyces cerevisiae var. boulardii and food-borne Saccharomyces cerevisiae strains

    DEFF Research Database (Denmark)

    van der Aa Kuhle, Alis; Skovgaard, Kerstin; Jespersen, Lene

    2005-01-01

    nontumorigenic porcine jejunal epithelial cell line (IPEC-J2) was investigated by incorporation of H-3-methionine into the yeast cells and use of liquid scintillation counting. Only few of the food-borne S. cerevisiae strains exhibited noteworthy adhesiveness with the strongest levels of adhesion (13...... effects hence indicating that food-borne strains of S. cerevisiae may possess probiotic properties in spite of low adhesiveness. © 2004 Elsevier B.V. All rights reserved....

  8. Draft Genome Sequence of Saccharomyces cerevisiae IR-2, a Useful Industrial Strain for Highly Efficient Production of Bioethanol

    OpenAIRE

    Sahara, Takehiko; Fujimori, Kazuhiro E.; Nezuo, Maiko; Tsukahara, Masatoshi; Tochigi, Yuki; Ohgiya, Satoru; Kamagata, Yoichi

    2014-01-01

    We sequenced the genome of Saccharomyces cerevisiae IR-2, which is a diploid industrial strain with flocculation activity and the ability to efficiently produce bioethanol. The approximately 11.4-Mb draft genome information provides useful insights into metabolic engineering for the production of bioethanol from biomass.

  9. Development Of An Efficient Glycerol Utilizing Saccharomyces Cerevisiae Strain Via Adaptive Laboratory Evolution

    DEFF Research Database (Denmark)

    Strucko, Tomas; Zirngibl, Katharina; Tharwat Tolba Mohamed, Elsayed;

    2015-01-01

    catabolism in yeast. The knowledge acquired in this study may be further applied for rational S. cerevisiae strain improvement for using glycerol as a carbon source in industrial biotechnology processes. This work is a part of the DeYeastLibrary consortium financed by ERA-IB DeYeastLibrary - Designer yeast...... fermentation processes. The most commonly known microbial cell factory, the yeast Saccharomyces cerevisiae, has been extensively applied for the production of a wide range of scientifically and industrially relevant products using saccharides (mainly glucose) as carbon source. However, it was shown...... strain library optimized for metabolic engineering applications http://www.era-ib.net/deyeast-library...

  10. Dual utilization of NADPH and NADH cofactors enhances xylitol production in engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Jo, Jung-Hyun; Oh, Sun-Young; Lee, Hyeun-Soo; Park, Yong-Cheol; Seo, Jin-Ho

    2015-12-01

    Xylitol, a natural sweetener, can be produced by hydrogenation of xylose in hemicelluloses. In microbial processes, utilization of only NADPH cofactor limited commercialization of xylitol biosynthesis. To overcome this drawback, Saccharomyces cerevisiae D452-2 was engineered to express two types of xylose reductase (XR) with either NADPH-dependence or NADH-preference. Engineered S. cerevisiae DWM expressing both the XRs exhibited higher xylitol productivity than the yeast strain expressing NADPH-dependent XR only (DWW) in both batch and glucose-limited fed-batch cultures. Furthermore, the coexpression of S. cerevisiae ZWF1 and ACS1 genes in the DWM strain increased intracellular concentrations of NADPH and NADH and improved maximum xylitol productivity by 17%, relative to that for the DWM strain. Finally, the optimized fed-batch fermentation of S. cerevisiae DWM-ZWF1-ACS1 resulted in 196.2 g/L xylitol concentration, 4.27 g/L h productivity and almost the theoretical yield. Expression of the two types of XR utilizing both NADPH and NADH is a promising strategy to meet the industrial demands for microbial xylitol production. PMID:26470683

  11. Comparative proteomics analysis of engineered Saccharomyces cerevisiae with enhanced biofuel precursor production.

    Directory of Open Access Journals (Sweden)

    Xiaoling Tang

    Full Text Available The yeast Saccharomyces cerevisiae was metabolically modified for enhanced biofuel precursor production by knocking out genes encoding mitochondrial isocitrate dehydrogenase and over-expression of a heterologous ATP-citrate lyase. A comparative iTRAQ-coupled 2D LC-MS/MS analysis was performed to obtain a global overview of ubiquitous protein expression changes in S. cerevisiae engineered strains. More than 300 proteins were identified. Among these proteins, 37 were found differentially expressed in engineered strains and they were classified into specific categories based on their enzyme functions. Most of the proteins involved in glycolytic and pyruvate branch-point pathways were found to be up-regulated and the proteins involved in respiration and glyoxylate pathway were however found to be down-regulated in engineered strains. Moreover, the metabolic modification of S. cerevisiae cells resulted in a number of up-regulated proteins involved in stress response and differentially expressed proteins involved in amino acid metabolism and protein biosynthesis pathways. These LC-MS/MS based proteomics analysis results not only offered extensive information in identifying potential protein-protein interactions, signal pathways and ubiquitous cellular changes elicited by the engineered pathways, but also provided a meaningful biological information platform serving further modification of yeast cells for enhanced biofuel production.

  12. GMAX Yeast Background Strain Made from Industrial Tolerant Saccharomyces Cerevisiae Engineered to Convert Pretreated Lignocellulosic Starch and Cellulosic Sugars Universally to Ethanol Anaerobically

    Science.gov (United States)

    Tailored GMAX yeast background strain technology for universal ethanol production industrially: Production of the stable baseline glucose, mannose, arabinose, xylose-utilizing (GMAX) yeast will be evaluated by taking the genes identified in high-throughput screening for a plasmid-based yeast to util...

  13. Mutagenic Inverted Repeats Assisted Genome Engineering (MIRAGE) in Saccharomyces cerevisiae: deletion of gal7.

    Science.gov (United States)

    Nair, Nikhil U; Zhao, Huimin

    2012-01-01

    MIRAGE is a unique in vivo genome editing technique that exploits the inherent instability of inverted repeats (palindromes) in the Saccharomyces cerevisiae chromosome. As a technique able to quickly create deletions as well as precise point mutations, it is valuable in applications that require creation of designer strains of this yeast. In particular, it has various potential applications in metabolic engineering, systems biology, synthetic biology, and molecular genetics. PMID:22144353

  14. Nanofiltration concentration of extracellular glutathione produced by engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Sasaki, Kengo; Hara, Kiyotaka Y; Kawaguchi, Hideo; Sazuka, Takashi; Ogino, Chiaki; Kondo, Akihiko

    2016-01-01

    This study aimed to optimize extracellular glutathione production by a Saccharomyces cerevisiae engineered strain and to concentrate the extracellular glutathione by membrane separation processes, including ultrafiltration (UF) and nanofiltration (NF). Synthetic defined (SD) medium containing 20 g L(-1) glucose was fermented for 48 h; the fermentation liquid was passed through an UF membrane to remove macromolecules. Glutathione in this permeate was concentrated for 48 h to 545.1 ± 33.6 mg L(-1) using the NF membrane; this was a significantly higher concentration than that obtained with yeast extract peptone dextrose (YPD) medium following 96 h NF concentration (217.9 ± 57.4 mg L(-1)). This higher glutathione concentration results from lower cellular growth in SD medium (final OD600 = 6.9 ± 0.1) than in YPD medium (final OD600 = 11.0 ± 0.6) and thus higher production of extracellular glutathione (16.0 ± 1.3 compared to 9.2 ± 2.1 mg L(-1) in YPD medium, respectively). Similar fermentation and membrane processing of sweet sorghum juice containing 20 g L(-1) total sugars provided 240.3 ± 60.6 mg L(-1) glutathione. Increased extracellular production of glutathione by this engineered strain in SD medium and subsequent UF permeation and NF concentration in shortend time may help realize industrial recovery of extracellular glutathione. PMID:26105794

  15. Production of 2,3-butanediol in Saccharomyces cerevisiae by in silico aided metabolic engineering

    Directory of Open Access Journals (Sweden)

    Ng Chiam Yu

    2012-05-01

    Full Text Available Abstract Background 2,3-Butanediol is a chemical compound of increasing interest due to its wide applications. It can be synthesized via mixed acid fermentation of pathogenic bacteria such as Enterobacter aerogenes and Klebsiella oxytoca. The non-pathogenic Saccharomyces cerevisiae possesses three different 2,3-butanediol biosynthetic pathways, but produces minute amount of 2,3-butanediol. Hence, we attempted to engineer S. cerevisiae strain to enhance 2,3-butanediol production. Results We first identified gene deletion strategy by performing in silico genome-scale metabolic analysis. Based on the best in silico strategy, in which disruption of alcohol dehydrogenase (ADH pathway is required, we then constructed gene deletion mutant strains and performed batch cultivation of the strains. Deletion of three ADH genes, ADH1, ADH3 and ADH5, increased 2,3-butanediol production by 55-fold under microaerobic condition. However, overproduction of glycerol was observed in this triple deletion strain. Additional rational design to reduce glycerol production by GPD2 deletion altered the carbon fluxes back to ethanol and significantly reduced 2,3-butanediol production. Deletion of ALD6 reduced acetate production in strains lacking major ADH isozymes, but it did not favor 2,3-butanediol production. Finally, we introduced 2,3-butanediol biosynthetic pathway from Bacillus subtilis and E. aerogenes to the engineered strain and successfully increased titer and yield. Highest 2,3-butanediol titer (2.29 g·l-1 and yield (0.113 g·g-1 were achieved by Δadh1 Δadh3 Δadh5 strain under anaerobic condition. Conclusions With the aid of in silico metabolic engineering, we have successfully designed and constructed S. cerevisiae strains with improved 2,3-butanediol production.

  16. Metabolic engineering of Saccharomyces cerevisiae for the overproduction of short branched-chain fatty acids.

    Science.gov (United States)

    Yu, Ai-Qun; Juwono, Nina Kurniasih Pratomo; Foo, Jee Loon; Leong, Susanna Su Jan; Chang, Matthew Wook

    2016-03-01

    Short branched-chain fatty acids (SBCFAs, C4-6) are versatile platform intermediates for the production of value-added products in the chemical industry. Currently, SBCFAs are mainly synthesized chemically, which can be costly and may cause environmental pollution. In order to develop an economical and environmentally friendly route for SBCFA production, we engineered Saccharomyces cerevisiae, a model eukaryotic microorganism of industrial significance, for the overproduction of SBCFAs. In particular, we employed a combinatorial metabolic engineering approach to optimize the native Ehrlich pathway in S. cerevisiae. First, chromosome-based combinatorial gene overexpression led to a 28.7-fold increase in the titer of SBCFAs. Second, deletion of key genes in competing pathways improved the production of SBCFAs to 387.4 mg/L, a 31.2-fold increase compared to the wild-type. Third, overexpression of the ATP-binding cassette (ABC) transporter PDR12 increased the secretion of SBCFAs. Taken together, we demonstrated that the combinatorial metabolic engineering approach used in this study effectively improved SBCFA biosynthesis in S. cerevisiae through the incorporation of a chromosome-based combinatorial gene overexpression strategy, elimination of genes in competitive pathways and overexpression of a native transporter. We envision that this strategy could also be applied to the production of other chemicals in S. cerevisiae and may be extended to other microbes for strain improvement. PMID:26721212

  17. Co-utilization of L-arabinose and D-xylose by laboratory and industrial Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Boles Eckhard

    2006-04-01

    Full Text Available Abstract Background Fermentation of lignocellulosic biomass is an attractive alternative for the production of bioethanol. Traditionally, the yeast Saccharomyces cerevisiae is used in industrial ethanol fermentations. However, S. cerevisiae is naturally not able to ferment the pentose sugars D-xylose and L-arabinose, which are present in high amounts in lignocellulosic raw materials. Results We describe the engineering of laboratory and industrial S. cerevisiae strains to co-ferment the pentose sugars D-xylose and L-arabinose. Introduction of a fungal xylose and a bacterial arabinose pathway resulted in strains able to grow on both pentose sugars. Introduction of a xylose pathway into an arabinose-fermenting laboratory strain resulted in nearly complete conversion of arabinose into arabitol due to the L-arabinose reductase activity of the xylose reductase. The industrial strain displayed lower arabitol yield and increased ethanol yield from xylose and arabinose. Conclusion Our work demonstrates simultaneous co-utilization of xylose and arabinose in recombinant strains of S. cerevisiae. In addition, the co-utilization of arabinose together with xylose significantly reduced formation of the by-product xylitol, which contributed to improved ethanol production.

  18. 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. PMID:26164646

  19. Improved Production of a Heterologous Amylase in Saccharomyces cerevisiae by Inverse Metabolic Engineering

    DEFF Research Database (Denmark)

    Liu, Zihe; Liu, Lifang; Osterlund, Tobias;

    2014-01-01

    The increasing demand for industrial enzymes and biopharmaceutical proteins relies on robust production hosts with high protein yield and productivity. Being one of the best-studied model organisms and capable of performing posttranslational modifications, the yeast Saccharomyces cerevisiae is...... engineering to identify novel targets for improving protein secretion. Screening that combined UV-random mutagenesis and selection for growth on starch was performed to find mutant strains producing heterologous amylase 5-fold above the level produced by the reference strain. Genomic mutations that could be...

  20. [Production of β-carotene by metabolically engineered Saccharomyces cerevisiae].

    Science.gov (United States)

    Wang, Beibei; Shi, Mingyu; Wang, Dong; Xu, Jiaoyang; Liu, Yi; Yang, Hongjiang; Dai, Zhubo; Zhang, Xueli

    2014-08-01

    β-carotene has a wide range of application in food, pharmaceutical and cosmetic industries. For microbial production of β-carotene in Saccharomyces cerevisiae, the supply of geranylgeranyl diphosphate (GGPP) was firstly increased in S. cerevisiae BY4742 to obtain strain BY4742-T2 through over-expressing truncated 3-hydroxy-3-methylglutaryl-CoA reductase (tHMGR), which is the major rate-limiting enzyme in the mevalonate (MVA) pathway, and GGPP synthase (GGPS), which is a key enzyme in the diterpenoid synthetic pathway. The β-carotene synthetic genes of Pantoea agglomerans and Xanthophyllomyces dendrorhous were further integrated into strain BY4742-T2 for comparing β-carotene production. Over-expression of tHMGR and GGPS genes led to 26.0-fold increase of β-carotene production. In addition, genes from X. dendrorhous was more efficient than those from P. agglomerans for β-carotene production in S. cerevisiae. Strain BW02 was obtained which produced 1.56 mg/g (dry cell weight) β-carotene, which could be used further for constructing cell factories for β-carotene production. PMID:25507473

  1. Metabolic engineering of Saccharomyces cerevisiae for production of carboxylic acids: current status and challenges.

    Science.gov (United States)

    Abbott, Derek A; Zelle, Rintze M; Pronk, Jack T; van Maris, Antonius J A

    2009-12-01

    To meet the demands of future generations for chemicals and energy and to reduce the environmental footprint of the chemical industry, alternatives for petrochemistry are required. Microbial conversion of renewable feedstocks has a huge potential for cleaner, sustainable industrial production of fuels and chemicals. Microbial production of organic acids is a promising approach for production of chemical building blocks that can replace their petrochemically derived equivalents. Although Saccharomyces cerevisiae does not naturally produce organic acids in large quantities, its robustness, pH tolerance, simple nutrient requirements and long history as an industrial workhorse make it an excellent candidate biocatalyst for such processes. Genetic engineering, along with evolution and selection, has been successfully used to divert carbon from ethanol, the natural endproduct of S. cerevisiae, to pyruvate. Further engineering, which included expression of heterologous enzymes and transporters, yielded strains capable of producing lactate and malate from pyruvate. Besides these metabolic engineering strategies, this review discusses the impact of transport and energetics as well as the tolerance towards these organic acids. In addition to recent progress in engineering S. cerevisiae for organic acid production, the key limitations and challenges are discussed in the context of sustainable industrial production of organic acids from renewable feedstocks. PMID:19566685

  2. Physiological and transcriptional characterization of Saccharomyces cerevisiae engineered for production of fatty acid ethyl esters.

    Science.gov (United States)

    de Jong, Bouke Wim; Siewers, Verena; Nielsen, Jens

    2016-02-01

    Saccharomyces cerevisiae has previously been engineered to become a cell factory for the production of fatty acid ethyl esters (FAEEs), molecules suitable for crude diesel replacement. To find new metabolic engineering targets for the improvement of FAEE cell factories, three different FAEE-producing strains of S. cerevisiae, constructed previously, were compared and characterized by quantification of key fluxes and genome-wide transcription analysis. From both the physiological and the transcriptional data, it was indicated that strain CB2I20, with high expression of a heterologous wax ester synthase gene (ws2) and strain BdJ15, containing disruptions of genes DGA1, LRO1, ARE1, ARE2 and POX1, which prevent the conversion of acyl-CoA to sterol esters, triacylglycerides and the degradation to acetyl-CoA, triggered oxidative stress that consequently influenced cellular growth. In the latter strain, stress was possibly triggered by disabling the buffering capacity of lipid droplets in encapsulating toxic fatty acids such as oleic acid. Additionally, it was indicated that there was an increased demand for NADPH required for the reduction steps in fatty acid biosynthesis. In conclusion, our analysis clearly shows that engineering of fatty acid biosynthesis results in transcriptional reprogramming and has a significant effect on overall cellular metabolism. PMID:26590613

  3. Lactic acid production from xylose by engineered Saccharomyces cerevisiae without PDC or ADH deletion.

    Science.gov (United States)

    Turner, Timothy L; Zhang, Guo-Chang; Kim, Soo Rin; Subramaniam, Vijay; Steffen, David; Skory, Christopher D; Jang, Ji Yeon; Yu, Byung Jo; Jin, Yong-Su

    2015-10-01

    Production of lactic acid from renewable sugars has received growing attention as lactic acid can be used for making renewable and bio-based plastics. However, most prior studies have focused on production of lactic acid from glucose despite that cellulosic hydrolysates contain xylose as well as glucose. Microbial strains capable of fermenting both glucose and xylose into lactic acid are needed for sustainable and economic lactic acid production. In this study, we introduced a lactic acid-producing pathway into an engineered Saccharomyces cerevisiae capable of fermenting xylose. Specifically, ldhA from the fungi Rhizopus oryzae was overexpressed under the control of the PGK1 promoter through integration of the expression cassette in the chromosome. The resulting strain exhibited a high lactate dehydrogenase activity and produced lactic acid from glucose or xylose. Interestingly, we observed that the engineered strain exhibited substrate-dependent product formation. When the engineered yeast was cultured on glucose, the major fermentation product was ethanol while lactic acid was a minor product. In contrast, the engineered yeast produced lactic acid almost exclusively when cultured on xylose under oxygen-limited conditions. The yields of ethanol and lactic acid from glucose were 0.31 g ethanol/g glucose and 0.22 g lactic acid/g glucose, respectively. On xylose, the yields of ethanol and lactic acid were S. cerevisiae without deleting pyruvate decarboxylase, and the formation patterns of fermentations can be altered by substrates. PMID:26043971

  4. Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems

    OpenAIRE

    DiCarlo, James; Norville, Julie; Mali, Prashant; Rios Villanueva, Xavier; Aach, John Dennis; Church, George McDonald

    2013-01-01

    Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated (Cas) systems in bacteria and archaea use RNA-guided nuclease activity to provide adaptive immunity against invading foreign nucleic acids. Here, we report the use of type II bacterial CRISPR-Cas system in Saccharomyces cerevisiae for genome engineering. The CRISPR-Cas components, Cas9 gene and a designer genome targeting CRISPR guide RNA (gRNA), show robust and specific RNA-guided endonuclease activity a...

  5. Metabolic pathway engineering for fatty acid ethyl ester production in Saccharomyces cerevisiae using stable chromosomal integration.

    Science.gov (United States)

    de Jong, Bouke Wim; Shi, Shuobo; Valle-Rodríguez, Juan Octavio; Siewers, Verena; Nielsen, Jens

    2015-03-01

    Fatty acid ethyl esters are fatty acid derived molecules similar to first generation biodiesel (fatty acid methyl esters; FAMEs) which can be produced in a microbial cell factory. Saccharomyces cerevisiae is a suitable candidate for microbial large scale and long term cultivations, which is the typical industrial production setting for biofuels. It is crucial to conserve the metabolic design of the cell factory during industrial cultivation conditions that require extensive propagation. Genetic modifications therefore have to be introduced in a stable manner. Here, several metabolic engineering strategies for improved production of fatty acid ethyl esters in S. cerevisiae were combined and the genes were stably expressed from the organisms' chromosomes. A wax ester synthase (ws2) was expressed in different yeast strains with an engineered acetyl-CoA and fatty acid metabolism. Thus, we compared expression of ws2 with and without overexpression of alcohol dehydrogenase (ADH2), acetaldehyde dehydrogenase (ALD6) and acetyl-CoA synthetase (acs SE (L641P) ) and further evaluated additional overexpression of a mutant version of acetyl-CoA decarboxylase (ACC1 (S1157A,S659A) ) and the acyl-CoA binding protein (ACB1). The combined engineering efforts of the implementation of ws2, ADH2, ALD6 and acs SE (L641P) , ACC1 (S1157A,S659A) and ACB1 in a S. cerevisiae strain lacking storage lipid formation (are1Δ, are2Δ, dga1Δ and lro1Δ) and β-oxidation (pox1Δ) resulted in a 4.1-fold improvement compared with sole expression of ws2 in S. cerevisiae. PMID:25422103

  6. Reducing alcohol levels in wines through rational and evolutionary engineering of Saccharomyces cerevisiae.

    Science.gov (United States)

    Tilloy, Valentin; Cadière, Axelle; Ehsani, Maryam; Dequin, Sylvie

    2015-11-20

    Over the past two decades, the level of ethanol in wine has increased in most wine-producing regions, raising a number of issues related to consumer health, prevention policies, the effectiveness of the fermentation and wine sensorial quality. This review focuses on metabolic challenges and recent achievements in the development of Saccharomyces cerevisiae wine strains with reduced ethanol yield. Metabolic engineering approaches that have been successfully used to optimize endogenous pathways have been gradually replaced in recent years by evolutionary engineering strategies, which can generate strains with improved phenotypes using new circuits and can be put to immediate commercial use. The power of adaptive evolutionary strategies is expected to increase with the rapid development of whole-genome sequencing, which, combined with gene expression and metabolic flux analysis, enables the identification of the genetic basis of improved phenotypes and the transfer of such phenotypes between strains. PMID:26219842

  7. Engineering of a novel cellulose-adherent cellulolytic Saccharomyces cerevisiae for cellulosic biofuel production.

    Science.gov (United States)

    Liu, Zhuo; Ho, Shih-Hsin; Sasaki, Kengo; den Haan, Riaan; Inokuma, Kentaro; Ogino, Chiaki; van Zyl, Willem H; Hasunuma, Tomohisa; Kondo, Akihiko

    2016-01-01

    Cellulosic biofuel is the subject of increasing attention. The main obstacle toward its economic feasibility is the recalcitrance of lignocellulose requiring large amount of enzyme to break. Several engineered yeast strains have been developed with cellulolytic activities to reduce the need for enzyme addition, but exhibiting limited effect. Here, we report the successful engineering of a cellulose-adherent Saccharomyces cerevisiae displaying four different synergistic cellulases on the cell surface. The cellulase-displaying yeast strain exhibited clear cell-to-cellulose adhesion and a "tearing" cellulose degradation pattern; the adhesion ability correlated with enhanced surface area and roughness of the target cellulose fibers, resulting in higher hydrolysis efficiency. The engineered yeast directly produced ethanol from rice straw despite a more than 40% decrease in the required enzyme dosage for high-density fermentation. Thus, improved cell-to-cellulose interactions provided a novel strategy for increasing cellulose hydrolysis, suggesting a mechanism for promoting the feasibility of cellulosic biofuel production. PMID:27079382

  8. Stress Tolerance Variations in Saccharomyces cerevisiae Strains from Diverse Ecological Sources and Geographical Locations.

    Directory of Open Access Journals (Sweden)

    Yan-Lin Zheng

    Full Text Available The budding yeast Saccharomyces cerevisiae is a platform organism for bioethanol production from various feedstocks and robust strains are desirable for efficient fermentation because yeast cells inevitably encounter stressors during the process. Recently, diverse S. cerevisiae lineages were identified, which provided novel resources for understanding stress tolerance variations and related shaping factors in the yeast. This study characterized the tolerance of diverse S. cerevisiae strains to the stressors of high ethanol concentrations, temperature shocks, and osmotic stress. The results showed that the isolates from human-associated environments overall presented a higher level of stress tolerance compared with those from forests spared anthropogenic influences. Statistical analyses indicated that the variations of stress tolerance were significantly correlated with both ecological sources and geographical locations of the strains. This study provides guidelines for selection of robust S. cerevisiae strains for bioethanol production from nature.

  9. A repressor activator protein1 homologue from an oleaginous strain of Candida tropicalis increases storage lipid production in Saccharomyces cerevisiae.

    Science.gov (United States)

    Chattopadhyay, Atrayee; Dey, Prabuddha; Barik, Amita; Bahadur, Ranjit P; Maiti, Mrinal K

    2015-06-01

    The repressor activator protein1 (Rap1) has been studied over the years as a multifunctional regulator in Saccharomyces cerevisiae. However, its role in storage lipid accumulation has not been investigated. This report documents the identification and isolation of a putative transcription factor CtRap1 gene from an oleaginous strain of Candida tropicalis, and establishes the direct effect of its expression on the storage lipid accumulation in S. cerevisiae, usually a non-oleaginous yeast. In silico analysis revealed that the CtRap1 polypeptide binds relatively more strongly to the promoter of fatty acid synthase1 (FAS1) gene of S. cerevisiae than ScRap1. The expression level of CtRap1 transcript in vivo was found to correlate directly with the amount of lipid produced in oleaginous native host C. tropicalis. Heterologous expression of the CtRap1 gene resulted in ∼ 4-fold enhancement of storage lipid content (57.3%) in S. cerevisiae. We also showed that the functionally active CtRap1 upregulates the endogenous ScFAS1 and ScDGAT genes of S. cerevisiae, and this, in turn, might be responsible for the increased lipid production in the transformed yeast. Our findings pave the way for the possible utility of the CtRap1 gene in suitable microorganisms to increase their storage lipid content through transcription factor engineering. PMID:25805842

  10. CRISPR–Cas system enables fast and simple genome editing of industrial Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Vratislav Stovicek

    2015-12-01

    Full Text Available There is a demand to develop 3rd generation biorefineries that integrate energy production with the production of higher value chemicals from renewable feedstocks. Here, robust and stress-tolerant industrial strains of Saccharomyces cerevisiae will be suitable production organisms. However, their genetic manipulation is challenging, as they are usually diploid or polyploid. Therefore, there is a need to develop more efficient genetic engineering tools. We applied a CRISPR–Cas9 system for genome editing of different industrial strains, and show simultaneous disruption of two alleles of a gene in several unrelated strains with the efficiency ranging between 65% and 78%. We also achieved simultaneous disruption and knock-in of a reporter gene, and demonstrate the applicability of the method by designing lactic acid-producing strains in a single transformation event, where insertion of a heterologous gene and disruption of two endogenous genes occurred simultaneously. Our study provides a foundation for efficient engineering of industrial yeast cell factories.

  11. Genome-wide transcriptional response of a Saccharomyces cerevisiae strain with an altered redox metabolism

    DEFF Research Database (Denmark)

    Bro, Christoffer; Regenberg, Birgitte; Nielsen, Jens

    2004-01-01

    The genome-wide transcriptional response of a Saccharomyces cerevisiae strain deleted in GDH1 that encodes a NADP(+)-dependent glutamate dehydrogenase was compared to a wild-type strain under anaerobic steady-state conditions. The GDH1-deleted strain has a significantly reduced NADPH requirement,...

  12. Efficient Bioethanol Production by a Recombinant Flocculent Saccharomyces cerevisiae Strain with a Genome-Integrated NADP+-Dependent Xylitol Dehydrogenase Gene▿

    Science.gov (United States)

    Matsushika, Akinori; Inoue, Hiroyuki; Watanabe, Seiya; Kodaki, Tsutomu; Makino, Keisuke; Sawayama, Shigeki

    2009-01-01

    The recombinant industrial Saccharomyces cerevisiae strain MA-R5 was engineered to express NADP+-dependent xylitol dehydrogenase using the flocculent yeast strain IR-2, which has high xylulose-fermenting ability, and both xylose consumption and ethanol production remarkably increased. Furthermore, the MA-R5 strain produced the highest ethanol yield (0.48 g/g) from nonsulfuric acid hydrolysate of wood chips. PMID:19329659

  13. Metabolic engineering of Saccharomyces cerevisiae for the production of n-butanol

    Directory of Open Access Journals (Sweden)

    Myers Samuel

    2008-12-01

    Full Text Available Abstract Background Increasing energy costs and environmental concerns have motivated engineering microbes for the production of "second generation" biofuels that have better properties than ethanol. Results and conclusion Saccharomyces cerevisiae was engineered with an n-butanol biosynthetic pathway, in which isozymes from a number of different organisms (S. cerevisiae, Escherichia coli, Clostridium beijerinckii, and Ralstonia eutropha were substituted for the Clostridial enzymes and their effect on n-butanol production was compared. By choosing the appropriate isozymes, we were able to improve production of n-butanol ten-fold to 2.5 mg/L. The most productive strains harbored the C. beijerinckii 3-hydroxybutyryl-CoA dehydrogenase, which uses NADH as a co-factor, rather than the R. eutropha isozyme, which uses NADPH, and the acetoacetyl-CoA transferase from S. cerevisiae or E. coli rather than that from R. eutropha. Surprisingly, expression of the genes encoding the butyryl-CoA dehydrogenase from C. beijerinckii (bcd and etfAB did not improve butanol production significantly as previously reported in E. coli. Using metabolite analysis, we were able to determine which steps in the n-butanol biosynthetic pathway were the most problematic and ripe for future improvement.

  14. Metabolic engineering of Saccharomyces cerevisiae for the production of n-butanol

    Energy Technology Data Exchange (ETDEWEB)

    Steen, EricJ.; Chan, Rossana; Prasad, Nilu; Myers, Samuel; Petzold, Christopher; Redding, Alyssa; Ouellet, Mario; Keasling, JayD.

    2008-11-25

    BackgroundIncreasing energy costs and environmental concerns have motivated engineering microbes for the production of ?second generation? biofuels that have better properties than ethanol.Results& ConclusionsSaccharomyces cerevisiae was engineered with an n-butanol biosynthetic pathway, in which isozymes from a number of different organisms (S. cerevisiae, Escherichia coli, Clostridium beijerinckii, and Ralstonia eutropha) were substituted for the Clostridial enzymes and their effect on n-butanol production was compared. By choosing the appropriate isozymes, we were able to improve production of n-butanol ten-fold to 2.5 mg/L. The most productive strains harbored the C. beijerinckii 3-hydroxybutyryl-CoA dehydrogenase, which uses NADH as a co-factor, rather than the R. eutropha isozyme, which uses NADPH, and the acetoacetyl-CoA transferase from S. cerevisiae or E. coli rather than that from R. eutropha. Surprisingly, expression of the genes encoding the butyryl-CoA dehydrogenase from C. beijerinckii (bcd and etfAB) did not improve butanol production significantly as previously reported in E. coli. Using metabolite analysis, we were able to determine which steps in the n-butanol biosynthetic pathway were the most problematic and ripe for future improvement.

  15. Proteomic characterization of a wild-type wine strain of Saccharomyces cerevisiae.

    Science.gov (United States)

    Trabalzini, Lorenza; Paffetti, Alessandro; Ferro, Elisa; Scaloni, Andrea; Talamo, Fabio; Millucci, Lia; Martelli, Paola; Santucci, Annalisa

    2003-12-01

    Saccharomyces cerevisiae is the optimal eukaryotic model system to study mammalian biological responses. At the same time Saccharomyces cerevisiae is also widely utilized as a biotechnological tool in the food industry. Enological Saccharomyces cerevisiae strains have been so far routinely analyzed for their microbiological aspects. Nevertheless, wine yeasts are gaining an increasing interest in the last years since they strongly affect both the vinification process and the organoleptic properties of the final product wine. The protein repertoire is responsible of such features and, consequently, 2D-PAGE can be an useful tool to evaluate and select optimal wine yeast strains. We present here the first proteomic map of a wild-type wine Saccharomyces cerevisiae strain selected for the guided fermentation of very high quality wines. PMID:15141481

  16. Lactic acid production from cellobiose and xylose by engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Turner, Timothy L; Zhang, Guo-Chang; Oh, Eun Joong; Subramaniam, Vijay; Adiputra, Andrew; Subramaniam, Vimal; Skory, Christopher D; Jang, Ji Yeon; Yu, Byung Jo; Park, In; Jin, Yong-Su

    2016-05-01

    Efficient and rapid production of value-added chemicals from lignocellulosic biomass is an important step toward a sustainable society. Lactic acid, used for synthesizing the bioplastic polylactide, has been produced by microbial fermentation using primarily glucose. Lignocellulosic hydrolysates contain high concentrations of cellobiose and xylose. Here, we constructed a recombinant Saccharomyces cerevisiae strain capable of fermenting cellobiose and xylose into lactic acid. Specifically, genes (cdt-1, gh1-1, XYL1, XYL2, XYL3, and ldhA) coding for cellobiose transporter, β-glucosidase, xylose reductase, xylitol dehydrogenase, xylulokinase, and lactate dehydrogenase were integrated into the S. cerevisiae chromosomes. The resulting strain produced lactic acid from cellobiose or xylose with high yields. When fermenting a cellulosic sugar mixture containing 10 g/L glucose, 40 g/L xylose, and 80 g/L cellobiose, the engineered strain produced 83 g/L of lactic acid with a yield of 0.66 g lactic acid/g sugar (66% theoretical maximum). This study demonstrates initial steps toward the feasibility of sustainable production of lactic acid from lignocellulosic sugars by engineered yeast. PMID:26524688

  17. Comparative proteomic analysis of engineered Saccharomyces cerevisiae with enhanced free fatty acid accumulation.

    Science.gov (United States)

    Chen, Liwei; Lee, Jaslyn Jie Lin; Zhang, Jianhua; Chen, Wei Ning

    2016-02-01

    The engineered Saccharomyces cerevisiae strain △faa1△faa4 [Acot5s] was demonstrated to accumulate more free fatty acids (FFA) previously. Here, comparative proteomic analysis was performed to get a global overview of metabolic regulation in the strain. Over 500 proteins were identified, and 82 of those proteins were found to change significantly in the engineered strains. Proteins involved in glycolysis, acetate metabolism, fatty acid synthesis, TCA cycle, glyoxylate cycle, the pentose phosphate pathway, respiration, transportation, and stress response were found to be upregulated in △faa1△faa4 [Acot5s] as compared to the wild type. On the other hand, proteins involved in glycerol, ethanol, ergosterol, and cell wall synthesis were downregulated. Taken together with our metabolite analysis, our results showed that the disruption of Faa1 and Faa4 and expression of Acot5s in the engineered strain △faa1△faa4 [Acot5s] not only relieved the feedback inhibition of fatty acyl-CoAs on fatty acid synthesis, but also caused a major metabolic rearrangement. The rearrangement redirected carbon flux toward the pathways which generate the essential substrates and cofactors for fatty acid synthesis, such as acetyl-CoA, ATP, and NADPH. Therefore, our results help shed light on the mechanism for the increased production of fatty acids in the engineered strains, which is useful in providing information for future studies in biofuel production. PMID:26450510

  18. Genome-scale consequences of cofactor balancing in engineered pentose utilization pathways in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Amit Ghosh

    Full Text Available Biofuels derived from lignocellulosic biomass offer promising alternative renewable energy sources for transportation fuels. Significant effort has been made to engineer Saccharomyces cerevisiae to efficiently ferment pentose sugars such as D-xylose and L-arabinose into biofuels such as ethanol through heterologous expression of the fungal D-xylose and L-arabinose pathways. However, one of the major bottlenecks in these fungal pathways is that the cofactors are not balanced, which contributes to inefficient utilization of pentose sugars. We utilized a genome-scale model of S. cerevisiae to predict the maximal achievable growth rate for cofactor balanced and imbalanced D-xylose and L-arabinose utilization pathways. Dynamic flux balance analysis (DFBA was used to simulate batch fermentation of glucose, D-xylose, and L-arabinose. The dynamic models and experimental results are in good agreement for the wild type and for the engineered D-xylose utilization pathway. Cofactor balancing the engineered D-xylose and L-arabinose utilization pathways simulated an increase in ethanol batch production of 24.7% while simultaneously reducing the predicted substrate utilization time by 70%. Furthermore, the effects of cofactor balancing the engineered pentose utilization pathways were evaluated throughout the genome-scale metabolic network. This work not only provides new insights to the global network effects of cofactor balancing but also provides useful guidelines for engineering a recombinant yeast strain with cofactor balanced engineered pathways that efficiently co-utilizes pentose and hexose sugars for biofuels production. Experimental switching of cofactor usage in enzymes has been demonstrated, but is a time-consuming effort. Therefore, systems biology models that can predict the likely outcome of such strain engineering efforts are highly useful for motivating which efforts are likely to be worth the significant time investment.

  19. Production of 2,3-butanediol in Saccharomyces cerevisiae by in silico aided metabolic engineering

    OpenAIRE

    Ng Chiam Yu; Jung Moo-young; Lee Jinwon; Oh Min-Kyu

    2012-01-01

    Abstract Background 2,3-Butanediol is a chemical compound of increasing interest due to its wide applications. It can be synthesized via mixed acid fermentation of pathogenic bacteria such as Enterobacter aerogenes and Klebsiella oxytoca. The non-pathogenic Saccharomyces cerevisiae possesses three different 2,3-butanediol biosynthetic pathways, but produces minute amount of 2,3-butanediol. Hence, we attempted to engineer S. cerevisiae strain to enhance 2,3-butanediol production. Results We fi...

  20. Diversity of Saccharomyces cerevisiae Strains Isolated from Two Italian Wine-Producing Regions.

    Science.gov (United States)

    Capece, Angela; Granchi, Lisa; Guerrini, Simona; Mangani, Silvia; Romaniello, Rossana; Vincenzini, Massimo; Romano, Patrizia

    2016-01-01

    Numerous studies, based on different molecular techniques analyzing DNA polymorphism, have provided evidence that indigenous Saccharomyces cerevisiae populations display biogeographic patterns. Since the differentiated populations of S. cerevisiae seem to be responsible for the regional identity of wine, the aim of this work was to assess a possible relationship between the diversity and the geographical origin of indigenous S. cerevisiae isolates from two different Italian wine-producing regions (Tuscany and Basilicata). For this purpose, sixty-three isolates from Aglianico del Vulture grape must (main cultivar in the Basilicata region) and from Sangiovese grape must (main cultivar in the Tuscany region) were characterized genotypically, by mitochondrial DNA restriction analysis and MSP-PCR by using (GTG)5 primers, and phenotypically, by determining technological properties and metabolic compounds of oenological interest after alcoholic fermentation. All the S. cerevisiae isolates from each region were inoculated both in must obtained from Aglianico grape and in must obtained from Sangiovese grape to carry out fermentations at laboratory-scale. Numerical analysis of DNA patterns resulting from both molecular methods and principal component analysis of phenotypic data demonstrated a high diversity among the S. cerevisiae strains. Moreover, a correlation between genotypic and phenotypic groups and geographical origin of the strains was found, supporting the concept that there can be a microbial aspect to terroir. Therefore, exploring the diversity of indigenous S. cerevisiae strains can allow developing tailored strategies to select wine yeast strains better adapted to each viticultural area. PMID:27446054

  1. Diversity of Saccharomyces cerevisiae Strains Isolated from Two Italian Wine-Producing Regions

    Science.gov (United States)

    Capece, Angela; Granchi, Lisa; Guerrini, Simona; Mangani, Silvia; Romaniello, Rossana; Vincenzini, Massimo; Romano, Patrizia

    2016-01-01

    Numerous studies, based on different molecular techniques analyzing DNA polymorphism, have provided evidence that indigenous Saccharomyces cerevisiae populations display biogeographic patterns. Since the differentiated populations of S. cerevisiae seem to be responsible for the regional identity of wine, the aim of this work was to assess a possible relationship between the diversity and the geographical origin of indigenous S. cerevisiae isolates from two different Italian wine-producing regions (Tuscany and Basilicata). For this purpose, sixty-three isolates from Aglianico del Vulture grape must (main cultivar in the Basilicata region) and from Sangiovese grape must (main cultivar in the Tuscany region) were characterized genotypically, by mitochondrial DNA restriction analysis and MSP-PCR by using (GTG)5 primers, and phenotypically, by determining technological properties and metabolic compounds of oenological interest after alcoholic fermentation. All the S. cerevisiae isolates from each region were inoculated both in must obtained from Aglianico grape and in must obtained from Sangiovese grape to carry out fermentations at laboratory-scale. Numerical analysis of DNA patterns resulting from both molecular methods and principal component analysis of phenotypic data demonstrated a high diversity among the S. cerevisiae strains. Moreover, a correlation between genotypic and phenotypic groups and geographical origin of the strains was found, supporting the concept that there can be a microbial aspect to terroir. Therefore, exploring the diversity of indigenous S. cerevisiae strains can allow developing tailored strategies to select wine yeast strains better adapted to each viticultural area. PMID:27446054

  2. Genome Sequence and Analysis of a Stress-Tolerant, Wild-Derived Strain of Saccharomyces cerevisiae Used in Biofuels Research

    Directory of Open Access Journals (Sweden)

    Sean J. McIlwain

    2016-06-01

    Full Text Available The genome sequences of more than 100 strains of the yeast Saccharomyces cerevisiae have been published. Unfortunately, most of these genome assemblies contain dozens to hundreds of gaps at repetitive sequences, including transposable elements, tRNAs, and subtelomeric regions, which is where novel genes generally reside. Relatively few strains have been chosen for genome sequencing based on their biofuel production potential, leaving an additional knowledge gap. Here, we describe the nearly complete genome sequence of GLBRCY22-3 (Y22-3, a strain of S. cerevisiae derived from the stress-tolerant wild strain NRRL YB-210 and subsequently engineered for xylose metabolism. After benchmarking several genome assembly approaches, we developed a pipeline to integrate Pacific Biosciences (PacBio and Illumina sequencing data and achieved one of the highest quality genome assemblies for any S. cerevisiae strain. Specifically, the contig N50 is 693 kbp, and the sequences of most chromosomes, the mitochondrial genome, and the 2-micron plasmid are complete. Our annotation predicts 92 genes that are not present in the reference genome of the laboratory strain S288c, over 70% of which were expressed. We predicted functions for 43 of these genes, 28 of which were previously uncharacterized and unnamed. Remarkably, many of these genes are predicted to be involved in stress tolerance and carbon metabolism and are shared with a Brazilian bioethanol production strain, even though the strains differ dramatically at most genetic loci. The Y22-3 genome sequence provides an exceptionally high-quality resource for basic and applied research in bioenergy and genetics.

  3. Genome Sequence and Analysis of a Stress-Tolerant, Wild-Derived Strain of Saccharomyces cerevisiae Used in Biofuels Research.

    Science.gov (United States)

    McIlwain, Sean J; Peris, David; Sardi, Maria; Moskvin, Oleg V; Zhan, Fujie; Myers, Kevin S; Riley, Nicholas M; Buzzell, Alyssa; Parreiras, Lucas S; Ong, Irene M; Landick, Robert; Coon, Joshua J; Gasch, Audrey P; Sato, Trey K; Hittinger, Chris Todd

    2016-01-01

    The genome sequences of more than 100 strains of the yeast Saccharomyces cerevisiae have been published. Unfortunately, most of these genome assemblies contain dozens to hundreds of gaps at repetitive sequences, including transposable elements, tRNAs, and subtelomeric regions, which is where novel genes generally reside. Relatively few strains have been chosen for genome sequencing based on their biofuel production potential, leaving an additional knowledge gap. Here, we describe the nearly complete genome sequence of GLBRCY22-3 (Y22-3), a strain of S. cerevisiae derived from the stress-tolerant wild strain NRRL YB-210 and subsequently engineered for xylose metabolism. After benchmarking several genome assembly approaches, we developed a pipeline to integrate Pacific Biosciences (PacBio) and Illumina sequencing data and achieved one of the highest quality genome assemblies for any S. cerevisiae strain. Specifically, the contig N50 is 693 kbp, and the sequences of most chromosomes, the mitochondrial genome, and the 2-micron plasmid are complete. Our annotation predicts 92 genes that are not present in the reference genome of the laboratory strain S288c, over 70% of which were expressed. We predicted functions for 43 of these genes, 28 of which were previously uncharacterized and unnamed. Remarkably, many of these genes are predicted to be involved in stress tolerance and carbon metabolism and are shared with a Brazilian bioethanol production strain, even though the strains differ dramatically at most genetic loci. The Y22-3 genome sequence provides an exceptionally high-quality resource for basic and applied research in bioenergy and genetics. PMID:27172212

  4. The use of genetically modified Saccharomyces cerevisiae strains in the wine industry

    OpenAIRE

    Schuller, Dorit; Casal, Margarida

    2005-01-01

    During the last decades, science and food technology have contributed at an accelerated rate to the introduction of new products to satisfy nutritional, socio-economic and quality requirements. With the emergence of modern molecular genetics, the industrial importance of Saccharomyces cerevisiae, continuously extended. The demand for suitable genetically modified (GM) S. cerevisiae strains for the biofuel, bakery and beverage industries or for the production of biotechnological products (e.g....

  5. Rapid and efficient galactose fermentation by engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Quarterman, Josh; Skerker, Jeffrey M; Feng, Xueyang; Liu, Ian Y; Zhao, Huimin; Arkin, Adam P; Jin, Yong-Su

    2016-07-10

    In the important industrial yeast Saccharomyces cerevisiae, galactose metabolism requires energy production by respiration; therefore, this yeast cannot metabolize galactose under strict anaerobic conditions. While the respiratory dependence of galactose metabolism provides benefits in terms of cell growth and population stability, it is not advantageous for producing fuels and chemicals since a substantial fraction of consumed galactose is converted to carbon dioxide. In order to force S. cerevisiae to use galactose without respiration, a subunit (COX9) of a respiratory enzyme was deleted, but the resulting deletion mutant (Δcox9) was impaired in terms of galactose assimilation. Interestingly, after serial sub-cultures on galactose, the mutant evolved rapidly and was able to use galactose via fermentation only. The evolved strain (JQ-G1) produced ethanol from galactose with a 94% increase in yield and 6.9-fold improvement in specific productivity as compared to the wild-type strain. (13)C-metabolic flux analysis demonstrated a three-fold reduction in carbon flux through the TCA cycle of the evolved mutant with redirection of flux toward the fermentation pathway. Genome sequencing of the JQ-G1 strain revealed a loss of function mutation in a master negative regulator of the Leloir pathway (Gal80p). The mutation (Glu348*) in Gal80p was found to act synergistically with deletion of COX9 for efficient galactose fermentation, and thus the double deletion mutant Δcox9Δgal80 produced ethanol 2.4 times faster and with 35% higher yield than a single knockout mutant with deletion of GAL80 alone. When we introduced a functional COX9 cassette back into the JQ-G1 strain, the JQ-G1-COX9 strain showed a 33% reduction in specific galactose uptake rate and a 49% reduction in specific ethanol production rate as compared to JQ-G1. The wild-type strain was also subjected to serial sub-cultures on galactose but we failed to isolate a mutant capable of utilizing galactose without

  6. Benchmarking two commonly used Saccharomyces cerevisiae strains for heterologous vanillin-β-glucoside production

    OpenAIRE

    Tomas Strucko; Olivera Magdenoska; Mortensen, Uffe H.

    2015-01-01

    The yeast Saccharomyces cerevisiae is a widely used eukaryotic model organism and a key cell factory for production of biofuels and wide range of chemicals. From the broad palette of available yeast strains, the most popular are those derived from laboratory strain S288c and the industrially relevant CEN.PK strain series. Importantly, in recent years these two strains have been subjected to comparative “-omics” analyzes pointing out significant genotypic and phenotypic differences. It is ther...

  7. Metabolic engineering and adaptive evolution for efficient production of D-lactic acid in Saccharomyces cerevisiae.

    Science.gov (United States)

    Baek, Seung-Ho; Kwon, Eunice Y; Kim, Yong Hwan; Hahn, Ji-Sook

    2016-03-01

    There is an increasing demand for microbial production of lactic acid (LA) as a monomer of biodegradable poly lactic acid (PLA). Both optical isomers, D-LA and L-LA, are required to produce stereocomplex PLA with improved properties. In this study, we developed Saccharomyces cerevisiae strains for efficient production of D-LA. D-LA production was achieved by expressing highly stereospecific D-lactate dehydrogenase gene (ldhA, LEUM_1756) from Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 in S. cerevisiae lacking natural LA production activity. D-LA consumption after glucose depletion was inhibited by deleting DLD1 encoding D-lactate dehydrogenase and JEN1 encoding monocarboxylate transporter. In addition, ethanol production was reduced by deleting PDC1 and ADH1 genes encoding major pyruvate decarboxylase and alcohol dehydrogenase, respectively, and glycerol production was eliminated by deleting GPD1 and GPD2 genes encoding glycerol-3-phosphate dehydrogenase. LA tolerance of the engineered D-LA-producing strain was enhanced by adaptive evolution and overexpression of HAA1 encoding a transcriptional activator involved in weak acid stress response, resulting in effective D-LA production up to 48.9 g/L without neutralization. In a flask fed-batch fermentation under neutralizing condition, our evolved strain produced 112.0 g/L D-LA with a yield of 0.80 g/g glucose and a productivity of 2.2 g/(L · h). PMID:26596574

  8. Expression of protein engineered NADP{sup +}-dependent xylitol dehydrogenase increases ethanol production from xylose in recombinant Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Matsushika, Akinori; Inoue, Hiroyuki; Murakami, Katsuji; Takimura, Osamu; Sawayama, Shigeki [National Institute of Advanced Industrial Science and Technology, Hiroshima (Japan). Biomass Technology Research Center; Watanabe, Seiya; Kodaki, Tsutomu; Makino, Keisuke [Kyoto Univ. (Japan). Inst. of Advanced Energy

    2008-11-15

    A recombinant Saccharomyces cerevisiae strain transformed with xylose reductase (XR) and xylitol dehydrogenase (XDH) genes from Pichia stipitis has the ability to convert xylose to ethanol together with the unfavorable excretion of xylitol, which may be due to cofactor imbalance between NADPH-preferring XR and NAD{sup +}-dependent XDH. To reduce xylitol formation, we have already generated several XDH mutants with a reversal of coenzyme specificity toward NADP{sup +}. In this study, we constructed a set of recombinant S. cerevisiae strains with xylose-fermenting ability, including protein-engineered NADP{sup +}-dependent XDH-expressing strains. The most positive effect on xylose-to-ethanol fermentation was found by using a strain named MA-N5, constructed by chromosomal integration of the gene for NADP{sup +}-dependent XDH along with XR and endogenous xylulokinase genes. The MA-N5 strain had an increase in ethanol production and decrease in xylitol excretion compared with the reference strain expressing wild-type XDH when fermenting not only xylose but also mixed sugars containing glucose and xylose. Furthermore, the MA-N5 strain produced ethanol with a high yield of 0.49 g of ethanol/g of total consumed sugars in the nonsulfuric acid hydrolysate of wood chips. The results demonstrate that glucose and xylose present in the lignocellulosic hydrolysate can be efficiently fermented by this redox-engineered strain. (orig.)

  9. BIOSORPTION OF CHROMIUM (VI FROM INDUSTRIAL EFFLUENT BY WILD ANDMUTANT TYPE STRAIN OF SACCHAROMYCES CEREVISIAE AND ITS IMMOBILIZED FORM

    Directory of Open Access Journals (Sweden)

    K Selvam, K Arungandhi, B Vishnupriya, T Shanmuga priya and M Yamuna

    2013-01-01

    Full Text Available Biosorption of chromium was studied by wild type Saccharomyces cerevisiae strain, mutant strain, immobilized-wild type and mutant strain. Chromium absorption pattern was observed in all experimental conditions. Hexavalent chromium (VI was analyzed by diphenyl carbazide method, by oxidizing the trivalent chromium (III. The percentage efficiency of wild type S. cerevisiae and its mutant strain, immobilized-wild type and mutant strain were 94.8%, 98.7%, 97.4% and 100% respectively. S. cerevisiae mutant strain and their immobilized form was found to be effective in biosorption of chromium (VI than the wild type forms.

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

    Directory of Open Access Journals (Sweden)

    Xiaoling Tang

    2015-12-01

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

  11. Engineering of carbon catabolite repression in recombinant xylose fermenting Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Roca, Christophe Francois Aime; Haack, Martin Brian; Olsson, Lisbeth

    2004-01-01

    Two xylose-fermenting glucose-derepressed Saccharomyces cerevisiae strains were constructed in order to investigate the influence of carbon catabolite repression on xylose metabolism. S. cerevisiae CPB.CR2 (Deltamig1, XYL1, XYL2, XKS1) and CPB.MBH2 (Deltamig1, Deltamig2, XYL1, XYL2, XKS1) were...

  12. BIOSORPTION OF CHROMIUM (VI) FROM INDUSTRIAL EFFLUENT BY WILD ANDMUTANT TYPE STRAIN OF SACCHAROMYCES CEREVISIAE AND ITS IMMOBILIZED FORM

    OpenAIRE

    K Selvam, K Arungandhi, B Vishnupriya, T Shanmuga priya and M Yamuna

    2013-01-01

    Biosorption of chromium was studied by wild type Saccharomyces cerevisiae strain, mutant strain, immobilized-wild type and mutant strain. Chromium absorption pattern was observed in all experimental conditions. Hexavalent chromium (VI) was analyzed by diphenyl carbazide method, by oxidizing the trivalent chromium (III). The percentage efficiency of wild type S. cerevisiae and its mutant strain, immobilized-wild type and mutant strain were 94.8%, 98.7%, 97.4% and 100% respectively. S. cerevisi...

  13. Engineering of Saccharomyces cerevisiae to utilize xylan as a sole carbohydrate source by co-expression of an endoxylanase, xylosidase and a bacterial xylose isomerase.

    Science.gov (United States)

    Mert, Marlin John; la Grange, Daniël Coenrad; Rose, Shaunita Hellouise; van Zyl, Willem Heber

    2016-04-01

    Xylan represents a major component of lignocellulosic biomass, and its utilization by Saccharomyces cerevisiae is crucial for the cost effective production of ethanol from plant biomass. A recombinant xylan-degrading and xylose-assimilating Saccharomyces cerevisiae strain was engineered by co-expression of the xylanase (xyn2) of Trichoderma reesei, the xylosidase (xlnD) of Aspergillus niger, the Scheffersomyces stipitis xylulose kinase (xyl3) together with the codon-optimized xylose isomerase (xylA) from Bacteroides thetaiotaomicron. Under aerobic conditions, the recombinant strain displayed a complete respiratory mode, resulting in higher yeast biomass production and consequently higher enzyme production during growth on xylose as carbohydrate source. Under oxygen limitation, the strain produced ethanol from xylose at a maximum theoretical yield of ~90 %. This study is one of only a few that demonstrates the construction of a S. cerevisiae strain capable of growth on xylan as sole carbohydrate source by means of recombinant enzymes. PMID:26749525

  14. Robust industrial Saccharomyces cerevisiae strains for very high gravity bio-ethanol fermentations

    OpenAIRE

    Pereira, Francisco B.; Guimarães, Pedro M. R.; Teixeira, J.A.; Domingues, Lucília

    2011-01-01

    The application and physiological background of two industrial Saccharomyces cerevisiae strains, isolated from harsh industrial environments, were studied in Very High Gravity (VHG) bio-ethanol fermentations. VHG laboratory fermentations, mimicking industrially relevant conditions, were performed with PE-2 and CA1185 industrial strains and the CEN.PK113-7D laboratory strain. The industrial isolates produced remarkable high ethanol titres (>19%, v/v) and accumulated an increased content of ste...

  15. Ethanol production from high-glucose industrial substrates using ethanol-tolerant Saccharomyces cerevisiae strains

    OpenAIRE

    Cunha, M. R. M.; Guimarães, Pedro M. R.; Teixeira, J.A.; Domingues, Lucília

    2008-01-01

    Ethanol is well known as a toxic metabolite for yeast cells. Thus, strains that can grow well under high ethanol stress condition are highly desirable. This work aims to select and characterize Saccharomyces cerevisiae strains with improved ethanol tolerance. Moreover, it aims to evaluate the feasibility of industrial residues as fermentation media and to optimize the composition of such media. The ethanol production and tolerance of the yeast strains have been evaluated, carrying out batc...

  16. Sensitivity to Lovastatin of Saccharomyces cerevisiae Strains Deleted for Pleiotropic Drug Resistance (PDR) Genes

    DEFF Research Database (Denmark)

    Formenti, Luca Riccardo; Kielland-Brandt, Morten

    2011-01-01

    The use of statins is well established in human therapy, and model organisms such as Saccharomyces cerevisiae are commonly used in studies of drug action at molecular and cellular levels. The investigation of the resistance mechanisms towards statins may suggest new approaches to improve therapy...... based on the use of statins. We investigated the susceptibility to lovastatin of S. cerevisiae strains deleted for PDR genes, responsible for exporting hydrophobic and amphi-philic drugs, such as lovastatin. Strains deleted for the genes tested, PDR1, PDR3, PDR5 and SNQ2, exhibited remarkably different...

  17. Evolutionary engineering of Saccharomyces cerevisiae for efficient conversion of red algal biosugars to bioethanol.

    Science.gov (United States)

    Lee, Hye-Jin; Kim, Soo-Jung; Yoon, Jeong-Jun; Kim, Kyoung Heon; Seo, Jin-Ho; Park, Yong-Cheol

    2015-09-01

    The aim of this work was to apply the evolutionary engineering to construct a mutant Saccharomyces cerevisiae HJ7-14 resistant on 2-deoxy-D-glucose and with an enhanced ability of bioethanol production from galactose, a mono-sugar in red algae. In batch and repeated-batch fermentations, HJ7-14 metabolized 5-fold more galactose and produced ethanol 2.1-fold faster than the parental D452-2 strain. Transcriptional analysis of genes involved in the galactose metabolism revealed that moderate relief from the glucose-mediated repression of the transcription of the GAL genes might enable HJ7-14 to metabolize galactose rapidly. HJ7-14 produced 7.4 g/L ethanol from hydrolysates of the red alga Gelidium amansii within 12 h, which was 1.5-times faster than that observed with D452-2. We demonstrate conclusively that evolutionary engineering is a promising tool to manipulate the complex galactose metabolism in S. cerevisiae to produce bioethanol from red alga. PMID:25804535

  18. Metabolic engineering of a synergistic pathway for n-butanol production in Saccharomyces cerevisiae.

    Science.gov (United States)

    Shi, Shuobo; Si, Tong; Liu, Zihe; Zhang, Hongfang; Ang, Ee Lui; Zhao, Huimin

    2016-01-01

    n-Butanol has several favourable properties as an advanced fuel or a platform chemical. Bio-based production of n-butanol is becoming increasingly important for sustainable chemical industry. Synthesis of n-butanol can be achieved via more than one metabolic pathway. Here we report the metabolic engineering of Saccharomyces cerevisiae to produce n-butanol through a synergistic pathway: the endogenous threonine pathway and the introduced citramalate pathway. Firstly, we characterized and optimized the endogenous threonine pathway; then, a citramalate synthase (CimA) mediated pathway was introduced to construct the synergistic pathway; next, the synergistic pathway was optimized by additional overexpression of relevant genes identified previously; meanwhile, the n-butanol production was also improved by overexpression of keto-acid decarboxylases (KDC) and alcohol dehydrogenase (ADH). After combining these strategies with co-expression of LEU1 (two copies), LEU4, LEU2 (two copies), LEU5, CimA, NFS1, ADH7 and ARO10(*), we achieved an n-butanol production of 835 mg/L in the final engineered strain, which is almost 7-fold increase compared to the initial strain. Furthermore, the production showed a 3-fold of the highest titer ever reported in yeast. Therefore, the engineered yeast strain represents a promising alternative platform for n-butanol production. PMID:27161023

  19. Metabolic engineering of a synergistic pathway for n-butanol production in Saccharomyces cerevisiae

    Science.gov (United States)

    Shi, Shuobo; Si, Tong; Liu, Zihe; Zhang, Hongfang; Ang, Ee Lui; Zhao, Huimin

    2016-01-01

    n-Butanol has several favourable properties as an advanced fuel or a platform chemical. Bio-based production of n-butanol is becoming increasingly important for sustainable chemical industry. Synthesis of n-butanol can be achieved via more than one metabolic pathway. Here we report the metabolic engineering of Saccharomyces cerevisiae to produce n-butanol through a synergistic pathway: the endogenous threonine pathway and the introduced citramalate pathway. Firstly, we characterized and optimized the endogenous threonine pathway; then, a citramalate synthase (CimA) mediated pathway was introduced to construct the synergistic pathway; next, the synergistic pathway was optimized by additional overexpression of relevant genes identified previously; meanwhile, the n-butanol production was also improved by overexpression of keto-acid decarboxylases (KDC) and alcohol dehydrogenase (ADH). After combining these strategies with co-expression of LEU1 (two copies), LEU4, LEU2 (two copies), LEU5, CimA, NFS1, ADH7 and ARO10*, we achieved an n-butanol production of 835 mg/L in the final engineered strain, which is almost 7-fold increase compared to the initial strain. Furthermore, the production showed a 3-fold of the highest titer ever reported in yeast. Therefore, the engineered yeast strain represents a promising alternative platform for n-butanol production. PMID:27161023

  20. Rational and evolutionary engineering approaches uncover a small set of genetic changes efficient for rapid xylose fermentation in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Soo Rin Kim

    Full Text Available Economic bioconversion of plant cell wall hydrolysates into fuels and chemicals has been hampered mainly due to the inability of microorganisms to efficiently co-ferment pentose and hexose sugars, especially glucose and xylose, which are the most abundant sugars in cellulosic hydrolysates. Saccharomyces cerevisiae cannot metabolize xylose due to a lack of xylose-metabolizing enzymes. We developed a rapid and efficient xylose-fermenting S. cerevisiae through rational and inverse metabolic engineering strategies, comprising the optimization of a heterologous xylose-assimilating pathway and evolutionary engineering. Strong and balanced expression levels of the XYL1, XYL2, and XYL3 genes constituting the xylose-assimilating pathway increased ethanol yields and the xylose consumption rates from a mixture of glucose and xylose with little xylitol accumulation. The engineered strain, however, still exhibited a long lag time when metabolizing xylose above 10 g/l as a sole carbon source, defined here as xylose toxicity. Through serial-subcultures on xylose, we isolated evolved strains which exhibited a shorter lag time and improved xylose-fermenting capabilities than the parental strain. Genome sequencing of the evolved strains revealed that mutations in PHO13 causing loss of the Pho13p function are associated with the improved phenotypes of the evolved strains. Crude extracts of a PHO13-overexpressing strain showed a higher phosphatase activity on xylulose-5-phosphate (X-5-P, suggesting that the dephosphorylation of X-5-P by Pho13p might generate a futile cycle with xylulokinase overexpression. While xylose consumption rates by the evolved strains improved substantially as compared to the parental strain, xylose metabolism was interrupted by accumulated acetate. Deletion of ALD6 coding for acetaldehyde dehydrogenase not only prevented acetate accumulation, but also enabled complete and efficient fermentation of xylose as well as a mixture of glucose and

  1. Metabolic engineering of Saccharomyces cerevisiae for the production of isobutanol and 3-methyl-1-butanol.

    Science.gov (United States)

    Park, Seong-Hee; Kim, Sujin; Hahn, Ji-Sook

    2014-11-01

    Saccharomyces cerevisiae naturally produces small amounts of isobutanol and 3-methyl-1-butanol via Ehrlich pathway from the catabolism of valine and leucine, respectively. In this study, we engineered CEN.PK2-1C, a leucine auxotrophic strain having a LEU2 gene mutation, for the production of isobutanol and 3-methyl-1-butanol. First, ALD6 encoding aldehyde dehydrogenase and BAT1 involved in valine synthesis were deleted to eliminate competing pathways. We also increased transcription of endogenous genes in the valine and leucine biosynthetic pathways by expressing Leu3Δ601, a constitutively active form of Leu3 transcriptional activator. For the production of isobutanol, genes involved in isobutanol production (ILV2, ILV3, ILV5, ARO10, and ADH2) were additionally overexpressed in ald6Δbat1Δ strain expressing LEU3Δ601, resulting in 376.9 mg/L isobutanol production from 100 g/L glucose. To increase 3-methyl-1-butanol production, leucine biosynthetic genes were additionally overexpressed in the final isobutanol-production strain. The resulting strain overexpressing LEU2 and LEU4 (D578Y) , a feedback inhibition-insensitive mutant of LEU4, showed a 34-fold increase in 3-methyl-1-butanol synthesis compared with CEN.PK2-1C control strain, producing 765.7 mg/L 3-methyl-1-butanol. PMID:25280745

  2. Genetic analysis of Saccharomyces cerevisiae strains isolated from Palm Wine in Eastern Nigeria. Comparison with other African strains.

    OpenAIRE

    Ezeronye, O.U.; Legras, Jean Luc

    2009-01-01

    Aims: To study the yeast diversity of Nigerian palm wines by comparison with other African strains. Methods and Results: Twenty-three Saccharomyces cerevisiae strains were obtained from palm wine samples collected at four locations in eastern Nigeria, and characterized using different molecular techniques: internal transcribed spacer restriction fragment length polymorphism and sequence analysis, pulsed field gel electrophoresis, inter delta typing and microsatellite multilocus analysis...

  3. Physiological studies in aerobic batch cultivations of Saccharomyces cerevisiae strains harboring the MEL1 gene

    DEFF Research Database (Denmark)

    Østergaard, Simon; Roca, Christophe Francois Aime; Ronnow, B.;

    2000-01-01

    Physiological studies of Saccharomyces cerevisiae strains harboring the MEL1 gene were carried out in aerobic batch cultivations on glucose-galactose mixtures and on the disaccharide melibiose, which is hydrolyzed by the enzyme melibiase (Mel1, EC 3.2.1.22) into a glucose and a galactose moiety. ...

  4. The Interaction between Saccharomyces cerevisiae and Non-Saccharomyces Yeast during Alcoholic Fermentation Is Species and Strain Specific.

    Science.gov (United States)

    Wang, Chunxiao; Mas, Albert; Esteve-Zarzoso, Braulio

    2016-01-01

    The present study analyzes the lack of culturability of different non-Saccharomyces strains due to interaction with Saccharomyces cerevisiae during alcoholic fermentation. Interaction was followed in mixed fermentations with 1:1 inoculation of S. cerevisiae and ten non-Saccharomyces strains. Starmerella bacillaris, and Torulaspora delbrueckii indicated longer coexistence in mixed fermentations compared with Hanseniaspora uvarum and Metschnikowia pulcherrima. Strain differences in culturability and nutrient consumption (glucose, alanine, ammonium, arginine, or glutamine) were found within each species in mixed fermentation with S. cerevisiae. The interaction was further analyzed using cell-free supernatant from S. cerevisiae and synthetic media mimicking both single fermentations with S. cerevisiae and using mixed fermentations with the corresponding non-Saccharomyces species. Cell-free S. cerevisiae supernatants induced faster culturability loss than synthetic media corresponding to the same fermentation stage. This demonstrated that some metabolites produced by S. cerevisiae played the main role in the decreased culturability of the other non-Saccharomyces yeasts. However, changes in the concentrations of main metabolites had also an effect. Culturability differences were observed among species and strains in culture assays and thus showed distinct tolerance to S. cerevisiae metabolites and fermentation environment. Viability kit and recovery analyses on non-culturable cells verified the existence of viable but not-culturable status. These findings are discussed in the context of interaction between non-Saccharomyces and S. cerevisiae. PMID:27148191

  5. Selection of Indigenous Saccharomyces cerevisiae Strains from Kutjevo Wine Growing Area at the Laboratoy Scale

    Directory of Open Access Journals (Sweden)

    Sandi Orlić

    2005-09-01

    Full Text Available The use of selected yeasts for winemaking has clear advantages over traditional spontaneous fermentation. Selection of wine yeasts is usually carried out within the Saccharomyces cerevisiae species. Yeast strains produce different amount of secondary compounds that impart specific characteristics to the wines. This suggests that it is necessary to isolate naturally occuring autochthone strains, which exhibit a metabolic profile that corresponds to each wine. Twenty two strains of S.cerevisiae, isolated from the Kutjevo region (Gornji and Donji Hrnjevec, Mitrovac, Graševina grapes, were tested for: fermentation vigor, ethanol resistance, volatile acidity, H2S production and β-glucosidase, polygalacturonase, and killer activity. From the results of this investigation we are able to select two yeast strains (RO 1272 and RO 1284 for more detailed fermentation trials and possible use as a starter culture in production of typical wines.

  6. Robust Metabolic Responses to Varied Carbon Sources in Natural and Laboratory Strains of Saccharomyces cerevisiae

    OpenAIRE

    Van Voorhies, Wayne A.

    2012-01-01

    Understanding factors that regulate the metabolism and growth of an organism is of fundamental biologic interest. This study compared the influence of two different carbon substrates, dextrose and galactose, on the metabolic and growth rates of the yeast Saccharomyces cerevisiae. Yeast metabolic and growth rates varied widely depending on the metabolic substrate supplied. The metabolic and growth rates of a yeast strain maintained under long-term laboratory conditions was compared to strain i...

  7. The Influence of Some Commercial Saccharomyces cerevisiae Strains on the Quality of Chardonnay Wines

    OpenAIRE

    Stanka HERJAVEC; Podgorski, Vesna; Redžepović, Sulejman; Mirošević, Nikola

    2003-01-01

    Changes in some aroma compounds and sensory properties caused by different commercial S. cerevisiae strains and by epiphyte microorganisms in Chardonnay wines were investigated. Wines fermented with the Lalvin-71 B strain contained significantly lower concentrations of alcohol, isoamyl alcohol, isobutanol and total acidity than the wines obtained from spontaneous and other inoculated fermentations. The highest concentrations of 2-phenyl ethanol and 2-phenyl ethyl acetate were found in the spo...

  8. Heterologous xylose isomerase pathway and evolutionary engineering improve xylose utilization in Saccharomyces cerevisiae.

    Science.gov (United States)

    Qi, Xin; Zha, Jian; Liu, Gao-Gang; Zhang, Weiwen; Li, Bing-Zhi; Yuan, Ying-Jin

    2015-01-01

    Xylose utilization is one key issue for the bioconversion of lignocelluloses. It is a promising approach to engineering heterologous pathway for xylose utilization in Saccharomyces cerevisiae. Here, we constructed a xylose-fermenting yeast SyBE001 through combinatorial fine-tuning the expression of XylA and endogenous XKS1. Additional overexpression of genes RKI1, RPE1, TKL1, and TAL1 in the non-oxidative pentose phosphate pathway (PPP) in SyBE001 increased the xylose consumption rate by 1.19-fold. By repetitive adaptation, the xylose utilization rate was further increased by ∼10-fold in the resultant strain SyBE003. Gene expression analysis identified a variety of genes with significantly changed expression in the PPP, glycolysis and the tricarboxylic acid cycle in SyBE003. PMID:26539187

  9. Social wasp intestines host the local phenotypic variability of Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Dapporto, Leonardo; Stefanini, Irene; Rivero, Damariz; Polsinelli, Mario; Capretti, Paolo; De Marchi, Paolo; Viola, Roberto; Turillazzi, Stefano; Cavalieri, Duccio

    2016-07-01

    Nowadays, the presence of Saccharomyces cerevisiae has been assessed in both wild and human-related environments. Social wasps have been shown to maintain and vector S. cerevisiae among different environments. The availability of strains isolated from wasp intestines represents a striking opportunity to assess whether the strains found in wasp intestines are characterized by peculiar traits. We analysed strains isolated from the intestines of social wasps and compared them with strains isolated from other sources, all collected in a restricted geographic area. We evaluated the production of volatile metabolites during grape must fermentation, the resistance to different stresses and the ability to exploit various carbon sources. Wasp strains, in addition to representing a wide range of S. cerevisiae genotypes, also represent large part of the phenotypes characterizing the sympatric set of yeast strains; their higher production of acetic acid and ethyl acetate could reflect improved ability to attract insects. Our findings suggest that the relationship between yeasts and wasps should be preserved, to safeguard not only the natural variance of this microorganism but also the interests of wine-makers, who could take advantage from the exploitation of their phenotypic variability. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27168222

  10. Automated Yeast Mating Protocol Using Open Reading Frames from Saccharomyces cerevisiae Genome to Improve Yeast Strains for Cellulosic Ethanol Production

    Science.gov (United States)

    Engineering the industrial ethanologen Saccharomyces cerevisiae to utilize pentose sugars from lignocellulosic biomass is critical for commercializing cellulosic fuel ethanol production. Approaches to engineer pentose-fermenting yeasts have required expression of additional genes. We implemented a...

  11. Engineering of a novel cellulose-adherent cellulolytic Saccharomyces cerevisiae for cellulosic biofuel production

    Science.gov (United States)

    Liu, Zhuo; Ho, Shih-Hsin; Sasaki, Kengo; den Haan, Riaan; Inokuma, Kentaro; Ogino, Chiaki; van Zyl, Willem H.; Hasunuma, Tomohisa; Kondo, Akihiko

    2016-01-01

    Cellulosic biofuel is the subject of increasing attention. The main obstacle toward its economic feasibility is the recalcitrance of lignocellulose requiring large amount of enzyme to break. Several engineered yeast strains have been developed with cellulolytic activities to reduce the need for enzyme addition, but exhibiting limited effect. Here, we report the successful engineering of a cellulose-adherent Saccharomyces cerevisiae displaying four different synergistic cellulases on the cell surface. The cellulase-displaying yeast strain exhibited clear cell-to-cellulose adhesion and a “tearing” cellulose degradation pattern; the adhesion ability correlated with enhanced surface area and roughness of the target cellulose fibers, resulting in higher hydrolysis efficiency. The engineered yeast directly produced ethanol from rice straw despite a more than 40% decrease in the required enzyme dosage for high-density fermentation. Thus, improved cell-to-cellulose interactions provided a novel strategy for increasing cellulose hydrolysis, suggesting a mechanism for promoting the feasibility of cellulosic biofuel production. PMID:27079382

  12. Historical Evolution of Laboratory Strains of Saccharomyces cerevisiae.

    Science.gov (United States)

    Louis, Edward J

    2016-01-01

    Budding yeast strains used in the laboratory have had a checkered past. Historically, the choice of strain for any particular experiment depended on the suitability of the strain for the topic of study (e.g., cell cycle vs. meiosis). Many laboratory strains had poor fermentation properties and were not representative of the robust strains used for domestic purposes. Most strains were related to each other, but investigators usually had only vague notions about the extent of their relationships. Isogenicity was difficult to confirm before the advent of molecular genetic techniques. However, their ease of growth and manipulation in laboratory conditions made them "the model" model organism, and they still provided a great deal of fundamental knowledge. Indeed, more than one Nobel Prize has been won using them. Most of these strains continue to be powerful tools, and isogenic derivatives of many of them-including entire collections of deletions, overexpression constructs, and tagged gene products-are now available. Furthermore, many of these strains are now sequenced, providing intimate knowledge of their relationships. Recent collections, new isolates, and the creation of genetically tractable derivatives have expanded the available strains for experiments. But even still, these laboratory strains represent a small fraction of the diversity of yeast. The continued development of new laboratory strains will broaden the potential questions that can be posed. We are now poised to take advantage of this diversity, rather than viewing it as a detriment to controlled experiments. PMID:27371602

  13. Research Progress of "Omics" Technologies and Its Application in Construction of Engineering Strain of Saccharomyces cerevisiae%酿酒酵母的"组学"技术研究进展及其在工程菌株构建中的应用

    Institute of Scientific and Technical Information of China (English)

    张晓阳; 李余动; 吴雪昌

    2011-01-01

    酿酒酵母是真核模式生物,已被广泛用于"组学"水平的研究."组学"技术主要由基因组学、转录组学、蛋白质组学及代谢组学构成.综述了酵母菌"组学"的研究进展,并论述了酵母菌"组学"技术在酵母菌菌株改造中的应用,包括酒类及生物燃料乙醇工业生产菌株的基因工程改造等.%The budding yeast, Saccharomyces cerevisiae, has been used as eukaryotic model organism in "ome" level research. "Omics" technologies were mainly composed of genomics,transcriptomics,proteomics and metabolomics. The recent advancements of "ome" level research in yeast were surveyed,then their application in strain improvement by genetic engineering was discussed,including the industrial yeast strains in bioethanol and winemaking processes.

  14. PHO13 deletion-induced transcriptional activation prevents sedoheptulose accumulation during xylose metabolism in engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Xu, Haiqing; Kim, Sooah; Sorek, Hagit; Lee, Youngsuk; Jeong, Deokyeol; Kim, Jungyeon; Oh, Eun Joong; Yun, Eun Ju; Wemmer, David E; Kim, Kyoung Heon; Kim, Soo Rin; Jin, Yong-Su

    2016-03-01

    The deletion of PHO13 (pho13Δ) in Saccharomyces cerevisiae, encoding a phosphatase enzyme of unknown specificity, results in the transcriptional activation of genes related to the pentose phosphate pathway (PPP) such as TAL1 encoding transaldolase. It has been also reported that the pho13Δ mutant of S. cerevisiae expressing a heterologous xylose pathway can metabolize xylose efficiently compared to its parental strain. However, the interaction between the pho13Δ-induced transcriptional changes and the phenotypes of xylose fermentation was not understood. Thus we investigated the global metabolic changes in response to pho13Δ when cells were exponentially growing on xylose. Among the 134 intracellular metabolites that we identified, the 98% reduction of sedoheptulose was found to be the most significant change in the pho13Δ mutant as compared to its parental strain. Because sedoheptulose-7-phosphate (S7P), a substrate of transaldolase, reduced significantly in the pho13Δ mutant as well, we hypothesized that limited transaldolase activity in the parental strain might cause dephosphorylation of S7P, leading to carbon loss and inefficient xylose metabolism. Mutants overexpressing TAL1 at different degrees were constructed, and their TAL1 expression levels and xylose consumption rates were positively correlated. Moreover, as TAL1 expression levels increased, intracellular sedoheptulose concentration dropped significantly. Therefore, we concluded that TAL1 upregulation, preventing the accumulation of sedoheptulose, is the most critical mechanism for the improved xylose metabolism by the pho13Δ mutant of engineered S. cerevisiae. PMID:26724864

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

    DEFF Research Database (Denmark)

    Nielsen, Jens; Jewett, Michael Christopher

    2008-01-01

    Industrial biotechnology is a rapidly growing field. With the increasing shift towards a bio-based economy, there is rising demand for developing efficient cell factories that can produce fuels, chemicals, pharmaceuticals, materials, nutraceuticals, and even food ingredients. The yeast Saccharomy...... 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.......Industrial biotechnology is a rapidly growing field. With the increasing shift towards a bio-based economy, there is rising demand for developing efficient cell factories that can produce fuels, chemicals, pharmaceuticals, materials, nutraceuticals, and even food ingredients. The yeast...... 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...

  16. An efficient xylose-fermenting recombinant Saccharomyces cerevisiae strain obtained through adaptive evolution and its global transcription profile

    Energy Technology Data Exchange (ETDEWEB)

    Shen, Yu; Chen, Xiao; Peng, Bingyin; Chen, Liyuan; Hou, Jin; Bao, Xiaoming [Shandong Univ., Jinan (China). State Key Lab. of Microbial Technology

    2012-11-15

    Factors related to ethanol production from xylose in engineered Saccharomyces cerevisiae that contain an exogenous initial metabolic pathway are still to be elucidated. In the present study, a strain that expresses the xylose isomerase gene of Piromyces sp. Pi-xylA and overexpresses XKS1, RPE1, RKI1, TAL1, and TKL1, with deleted GRE3 and COX4 genes was constructed. The xylose utilization capacity of the respiratory deficiency strain was poor but improved via adaptive evolution in xylose. The {mu}{sub max} of the evolved strain in 20 gl{sup -1} xylose is 0.11 {+-} 0.00 h{sup -1}, and the evolved strain consumed 17.83 gl{sup -1} xylose within 72 h, with an ethanol yield of 0.43 gg{sup -1} total consumed sugars during glucose-xylose cofermentation. Global transcriptional changes and effect of several specific genes were studied. The result revealed that the increased xylose isomerase activity, the upregulation of enzymes involved in glycolysis and glutamate synthesis, and the downregulation of trehalose and glycogen synthesis, may have contributed to the improved xylose utilization of the strain. Furthermore, the deletion of PHO13 decreased the xylose growth in the respiration deficiency strain although deleting PHO13 can improve the xylose metabolism in other strains. (orig.)

  17. Comparison between two selected Saccharomyces cerevisiae strains as fermentation starters in the production of traditional cachaça

    Directory of Open Access Journals (Sweden)

    Fátima de Cássia Oliveira Gomes

    2009-04-01

    Full Text Available Two Saccharomyces cerevisiae strains were tested as the starter yeasts in a traditional cachaça distillery. The strains used were S. cerevisiae UFMG-A829, isolated from a cachaça fermentation process, and S. cerevisiae K1-V1116, obtained from the wine industry. The permanence of each strain in the fermentation must was determined by RAPD (Random Amplified Polymorphic DNA-PCR, with primer M13. Both yeast strains were prevalent in the vats for approximately 30 days. Indigenous non-Saccharomyces and indigenous S. cerevisiae strains were isolated in lower counts during the fermentation period. Indigenous S. cerevisiae strains were molecularly distinct when compared to the starter yeasts. The two yeasts appeared promising starter yeasts in the fermentation process to produce traditional cachaça.Duas linhagens de Saccharomyces cerevisiae foram testadas como iniciadoras em uma destilaria de cachaça. Foram utilizadas as linhagens de S. cerevisiae UFMG-A829, isolada de fermentação de cachaça, e S. cerevisiae K1-V1116, de origem vinícola. A permanência de cada linhagem durante a fermentação foi determinada por RAPD (Random Amplified Polymorphic DNA-PCR, utilizando o iniciador M13. As duas linhagens predominaram nas dornas de fermentação por aproximadamente 30 dias. Leveduras não-Saccharomyces e S. cerevisiae indígenas foram isoladas em menor proporção durante o experimento. As linhagens de S. cerevisiae indígenas apresentaram perfis moleculares distintos em relação às linhagens iniciadoras. As duas linhagens foram promissoras para serem utilizadas como iniciadoras do processo fermentativo para a produção da cachaça.

  18. Directed evolution of pyruvate decarboxylase-negative Saccharomyces cerevisiae, yielding a C2-independent, glucose-tolerant, and pyruvate-hyperproducing yeast

    NARCIS (Netherlands)

    A.J. van Maris; J.M. Geertman; A. Vermeulen; M.K. Groothuizen; A.A. Winkler; M.D. Piper; J.P. van Dijken; J.T. Pronk

    2004-01-01

    textabstractThe absence of alcoholic fermentation makes pyruvate decarboxylase-negative (Pdc(-)) strains of Saccharomyces cerevisiae an interesting platform for further metabolic engineering of central metabolism. However, Pdc(-) S. cerevisiae strains have two growth defects:

  19. 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

  20. Selection of Saccharomyces cerevisiae strains for efficient very high gravity bio-ethanol fermentation processes

    OpenAIRE

    Pereira, Francisco B; Guimarães, Pedro M. R.; J. A. Teixeira; Domingues, Lucília

    2010-01-01

    Abstract An optimized very high gravity (VHG) glucose medium supplemented with low cost nutrient sources was used to evaluate bio-ethanol production by 11 Saccharomyces cerevisiae strains. The industrial strains PE-2 and CA1185 exhibited the best overall fermentation performance, producing an ethanol titre of 19.2% (v/v) corresponding to a batch productivity of 2.5 g l?1 h?1, while the best laboratory strain (CEN.PK 113-7D) produced 17.5% (v/v) ethanol with a productivity of 1.7 g ...

  1. Glucose and maltose metabolism in MIG1-disrupted and MAL-constitutive strains of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Klein, Christopher; Olsson, Lisbeth; Rønnow, B; Mikkelsen, Jørn Dalgaard; Nielsen, Jens Bredal

    1997-01-01

    The alleviation of glucose control of maltose metabolism brought about by MIG1 disruption was compared to that by MAL overexpression in a haploid Saccharomyces cerevisiae strain. The sugar consumption profiles during cultivation of the wild type, single transformants and a double transformant in a...... mixed glucose-maltose medium revealed that the MAL-constitutive strains were more alleviated than the single MIG1-disrupted transformant. While all transformants exhibited higher maximum specific growth rates (0.24-0.25 h(-1)) in glucose-maltose mixtures than the wild type strain (0.20 h(-1)), the MAL...

  2. Ethanol production from lignocellulosic hydrolysates using engineered Saccharomyces cerevisiae harboring xylose isomerase-based pathway.

    Science.gov (United States)

    Ko, Ja Kyong; Um, Youngsoon; Woo, Han Min; Kim, Kyoung Heon; Lee, Sun-Mi

    2016-06-01

    The efficient co-fermentation of glucose and xylose is necessary for the economically feasible bioethanol production from lignocellulosic biomass. Even with xylose utilizing Saccharomyces cerevisiae, the efficiency of the lignocellulosic ethanol production remains suboptimal mainly due to the low conversion yield of xylose to ethanol. In this study, we evaluated the co-fermentation performances of SXA-R2P-E, a recently engineered isomerase-based xylose utilizing strain, in mixed sugars and in lignocellulosic hydrolysates. In a high-sugar fermentation with 70g/L of glucose and 40g/L of xylose, SXA-R2P-E produced 50g/L of ethanol with an yield of 0.43gethanol/gsugars at 72h. From dilute acid-pretreated hydrolysates of rice straw and hardwood (oak), the strain produced 18-21g/L of ethanol with among the highest yield of 0.43-0.46gethanol/gsugars ever reported. This study shows a highly promising potential of a xylose isomerase-expressing strain as an industrially relevant ethanol producer from lignocellulosic hydrolysates. PMID:26990396

  3. Draft Genome Sequence of Saccharomyces cerevisiae Strain NCIM3186 Used in the Production of Bioethanol from Sweet Sorghum

    OpenAIRE

    Sravanthi Goud, Burragoni; Ulaganathan, Kandasamy

    2015-01-01

    Here, we report the draft genome sequence of Saccharomyces cerevisiae strain NCIM3186 used in bioethanol production from sweet sorghum. The size of the genome is approximately 11.9 Mb and contains 5,347 protein-coding genes.

  4. Production of Volatile and Sulfur Compounds by 10 Saccharomyces cerevisiae Strains Inoculated in Trebbiano Must.

    Science.gov (United States)

    Patrignani, Francesca; Chinnici, Fabio; Serrazanetti, Diana I; Vernocchi, Pamela; Ndagijimana, Maurice; Riponi, Claudio; Lanciotti, Rosalba

    2016-01-01

    In wines, the presence of sulfur compounds is the resulting of several contributions among which yeast metabolism. The characterization of the starter Saccharomyces cerevisiae needs to be performed also taking into account this ability even if evaluated together with the overall metabolic profile. In this perspective, principal aim of this experimental research was the evaluation of the volatile profiles, throughout GC/MS technique coupled with solid phase micro extraction, of wines obtained throughout the fermentation of 10 strains of S. cerevisiae. In addition, the production of sulfur compounds was further evaluated by using a gas-chromatograph coupled with a Flame Photometric Detector. Specifically, the 10 strains were inoculated in Trebbiano musts and the fermentations were monitored for 19 days. In the produced wines, volatile and sulfur compounds as well as amino acid concentrations were investigated. Also the physico-chemical characteristics of the wines and their electronic nose profiles were evaluated. PMID:26973621

  5. Production of volatile and sulfur compounds by ten Saccharomyces cerevisiae strains inoculated in Trebbiano must

    Directory of Open Access Journals (Sweden)

    Francesca ePatrignani

    2016-03-01

    Full Text Available In wines, the presence of sulphur compounds is the resulting of several contributions among which yeast metabolism. The characterization of the starter Saccharomyces cerevisiae needs to be performed also taking into account this ability even if evaluated together with the overall metabolic profile. In this perspective, principal aim of this experimental research was the evaluation of the volatile profiles, throughout GC/MS technique coupled with solid phase micro extraction, of wines obtained throughout the fermentation of 10 strains of Saccharomyces cerevisiae. In addition, the production of sulphur compounds was further evaluated by using a gas-chromatograph coupled with a Flame Photometric Detector. Specifically, the ten strains were inoculated in Trebbiano musts and the fermentations were monitored for 19 days. In the produced wines, volatile and sulphur compounds as well as amino acid concentrations were investigated. Also the physico-chemical characteristics of the wines and their electronic nose profiles were evaluated.

  6. Genome Sequencing and Comparative Analysis of Saccharomyces cerevisiae Strains of the Peterhof Genetic Collection

    Science.gov (United States)

    Drozdova, Polina B.; Tarasov, Oleg V.; Matveenko, Andrew G.; Radchenko, Elina A.; Sopova, Julia V.; Polev, Dmitrii E.; Inge-Vechtomov, Sergey G.; Dobrynin, Pavel V.

    2016-01-01

    The Peterhof genetic collection of Saccharomyces cerevisiae strains (PGC) is a large laboratory stock that has accumulated several thousands of strains for over than half a century. It originated independently of other common laboratory stocks from a distillery lineage (race XII). Several PGC strains have been extensively used in certain fields of yeast research but their genomes have not been thoroughly explored yet. Here we employed whole genome sequencing to characterize five selected PGC strains including one of the closest to the progenitor, 15V-P4, and several strains that have been used to study translation termination and prions in yeast (25-25-2V-P3982, 1B-D1606, 74-D694, and 6P-33G-D373). The genetic distance between the PGC progenitor and S288C is comparable to that between two geographically isolated populations. The PGC seems to be closer to two bakery strains than to S288C-related laboratory stocks or European wine strains. In genomes of the PGC strains, we found several loci which are absent from the S288C genome; 15V-P4 harbors a rare combination of the gene cluster characteristic for wine strains and the RTM1 cluster. We closely examined known and previously uncharacterized gene variants of particular strains and were able to establish the molecular basis for known phenotypes including phenylalanine auxotrophy, clumping behavior and galactose utilization. Finally, we made sequencing data and results of the analysis available for the yeast community. Our data widen the knowledge about genetic variation between Saccharomyces cerevisiae strains and can form the basis for planning future work in PGC-related strains and with PGC-derived alleles. PMID:27152522

  7. Fungicide residues in grapes determined the dynamics of Saccharomyces cerevisiae strains during spontaneous wine fermentation

    OpenAIRE

    Čuš Franc; Čadež Neža J.; Raspor Peter I.

    2011-01-01

    Impact of three fungicides against B. cinerea (iprodione, pyrimethanil and f ludioxonil plus cyprodinil) on the population of Saccharomyces cerevisiae strains during the spontaneous alcoholic fermentation was studied. With regard to the use of fungicides in the vineyard at two stages of the grapevine growth we followed four different spontaneous fermentations: control, iprodione, pyrimethanil and f ludioxonil plus cyprodinil. The fungicide residues in the grapes were determined by GC/MS...

  8. Adaptation of a flocculent Saccharomyces cerevisiae strain to lignocellulosic inhibitors by cell recycle batch fermentation

    OpenAIRE

    Landaeta, R.; Aroca, G.; Acevedo, F.; J. A. Teixeira; Mussatto, Solange I.

    2013-01-01

    The ethanol production from lignocellulosic feedstocks is considered a promising strategy to increase global production of biofuels without impacting food supplies. However, some compounds released during the hydrolysis of lignocellulosic materials are toxic for the microbial metabolism, causing low ethanol yield and productivity during the fermentation. As an attempt to overcome this problem, the present study evaluated the adaptation of a flocculent strain of Saccharomyces cerevisiae (NRRL ...

  9. Improving Stress Tolerance in Industrial Saccharomyces cerevisiae Strains for Ethanol Production from Lignocellulosic Biomass

    OpenAIRE

    Wallace, Valeria

    2014-01-01

    The present work was aimed at developing industrial S. cerevisiae strains with improved tolerance to two types of stressors encountered during the fermentation of lignocellulosic biomass that affect ethanol yield and productivity, namely hydrolysate-derived inhibitors and high temperature, and at understanding the response of yeast and mechanisms of adaptation to such stressors. In one part of the study, key amino acid substitutions that were responsible for the acquired ability of a mutated ...

  10. Improvement of galactose uptake in Saccharomyces cerevisiae through overexpression of phosphoglucomutase: Example of transcript analysis as a tool in inverse metabolic engineering

    DEFF Research Database (Denmark)

    Bro, Christoffer; Knudsen, S.; Regenberg, Birgitte; Olsson, Lisbeth; Nielsen, Jens

    2005-01-01

    reference strain. Based on our findings, we concluded that phosphoglucomutase plays a key role in controlling the flux through the Leloir pathway, probably due to increased conversion of glucose-l-phosphate to glucose-6-phosphate. This conclusion was supported by measurements of sugar phosphates, which......Through genome-wide transcript analysis of a reference strain and two recombinant Saccharomyces cerevisiae strains with different rates of galactose uptake, we obtained information about the global transcriptional response to metabolic engineering of the GAL gene regulatory network. One of the...... showed that there were increased concentrations of glucose-6-phosphate, galactose-6-phosphate, and fructose-6-phosphate in the strain construct overexpressing PGM2....

  11. Enhanced production of fatty alcohols by engineering the TAGs synthesis pathway in Saccharomyces cerevisiae.

    Science.gov (United States)

    Tang, Xiaoling; Chen, Wei Ning

    2015-02-01

    The production of fatty acid-derived chemicals has received a great deal of attention in recent years. In yeast cells, the main storage forms of fatty acids are TAGs. However, the conversion of TAGs into fatty acid derivatives suffers from a practical standpoint. Herein, a more direct strategy was applied to accumulate cellular fatty acyl-CoAs in Saccharomyces cerevisiae, which are the activated forms of fatty acids and used as important precursors for various converting enzymes. The dga1 gene was deleted to block the fatty acyl-CoAs dependent pathway of TAGs synthesis and a significant decrease in lipid content was observed. The FAR gene was cloned and overexpressed in the wild type strain and gene disrupted strain, to convert the fatty acyl-CoAs to the corresponding fatty acid derivatives. The metabolic engineered pathway resulted in enhanced production of fatty alcohols. Compared with the wild type strain with overexpressed FAR gene, the yield of fatty alcohols in the Δdga1 strain with FAR was dramatically increased: the intracellular fatty alcohols increased from 26 mg/L to 45 mg/L, while the extracellular fatty alcohols increased from 2.2 mg/L to 4.3 mg/L. By optimizing the culture medium with increased carbon concentration and limited nitrogen concentration, the fatty alcohols yield in the Δdga1 strain with FAR was further increased to 84 mg/L in cells and 14 mg/L secreted in broth. The results in this study demonstrated the feasibility of using the designed strategy to solve the bottleneck in utilizing TAGs for fatty acid derivatives production. PMID:25116045

  12. Metabolic engineering of Saccharomyces cerevisiae for the production of 2-phenylethanol via Ehrlich pathway.

    Science.gov (United States)

    Kim, Bosu; Cho, Bo-Ram; Hahn, Ji-Sook

    2014-01-01

    2-Phenylethanol (2-PE), a fragrance compound with a rose-like odor, is widely used in perfumery and cosmetics. Here, we report the first metabolic engineering approach for 2-PE production in Saccharomyces cerevisiae. 2-PE can be produced from the catabolism of L-phenylalanine via Ehrlich pathway, consisting of transamination to phenylpyruvate by Aro9, decarboxylation to phenylacetaldehyde by Aro10, and reduction to 2-PE by alcohol dehydrogenases. We demonstrated that Ald3 is mainly responsible for phenylacetaldehyde oxidation, competing with 2-PE production. ALD3 deletion strain overexpressing ARO9 and ARO10 both by episomal overexpression and by induction of the endogenous genes through overexpression of Aro80 transcription factor, produced 4.8 g/L 2-PE in a medium containing 10 g/L L-phenylalanine as a sole nitrogen source. Considering the cytotoxicity of 2-PE, this production titer is almost the upper limit that can be reached in batch cultures, suggesting the great potential of this yeast strain for 2-PE production. 2-PE production was further increased by applying two-phase fermentation method with polypropylene glycol 1200 as an extractant, reaching 6.1 g/L 2-PE in organic phase with the molar yield of 82.5%, which is about ninefold increase compared with wild type. PMID:23836015

  13. Improving biomass sugar utilization by engineered Saccharomyces cerevisiae

    Science.gov (United States)

    The efficient utilization of all available sugars in lignocellulosic biomass, which is more abundant than available commodity crops and starch, represents one of the most difficult technological challenges for the production of bioethanol. The well-studied yeast Saccharomyces cerevisiae has played a...

  14. Functional expression of xylose isomerase in flocculating industrial Saccharomyces cerevisiae strain for bioethanol production.

    Science.gov (United States)

    Li, Yun-Cheng; Li, Guo-Ying; Gou, Min; Xia, Zi-Yuan; Tang, Yue-Qin; Kida, Kenji

    2016-06-01

    Saccharomyces cerevisiae strains with xylose isomerase (XI) pathway were constructed using a flocculating industrial strain (YC-8) as the host. Both strains expressing wild-type xylA (coding XI) from the fungus Orpinomyces sp. and the bacterium Prevotella ruminicola, respectively, showed better growth ability and fermentation capacity when using xylose as the sole sugar than most of the reported strains expressing XI. Codon optimization of both XIs did not improve the xylose fermentation ability of the strains. Adaption significantly increased XI activity resulting in improved growth and fermentation. The strains expressing codon-optimized XI showed a higher increase in xylose consumption and ethanol production compared to strains expressing wild XI. Among all strains, the adapted strain YCPA2E expressing XI from P. ruminicola showed the best performance in the fermentation of xylose to ethanol. After 48 h of fermentation, YCPA2E assimilated 16.95 g/L xylose and produced 6.98 g/L ethanol. These results indicate that YC-8 is a suitable host strain for XI expression, especially for the codon-optimized XI originating from P. ruminicola. PMID:26645659

  15. Enhancing beta-carotene production in Saccharomyces cerevisiae by metabolic engineering.

    Science.gov (United States)

    Li, Qian; Sun, Zhiqiang; Li, Jing; Zhang, Yansheng

    2013-08-01

    Beta-carotene is known to exhibit a number of pharmacological and nutraceutical benefits to human health. Metabolic engineering of beta-carotene biosynthesis in Saccharomyces cerevisiae has been attracting the interest of many researchers. A previous work has shown that S. cerevisiae successfully integrated with phytoene synthase (crtYB) and phytoene desaturase (crtI) from Xanthophyllomyces dendrorhous could produce beta-carotene. In the present study, we achieved around 200% improvement in beta-carotene production in S. cerevisiae through specific site optimization of crtI and crtYB, in which five codons of crtI and eight codons of crtYB were rationally mutated. Furthermore, the effects of the truncated HMG-CoA reductase (tHMG1) from S. cerevisiae and HMG-CoA reductase (mva) from Staphylococcus aureus on the production of beta-carotene in S. cerevisiae were also evaluated. Our results indicated that mva from a prokaryotic organism might be more effective than tHMG1 for beta-carotene production in S. cerevisiae. PMID:23718229

  16. Consolidated bioprocessing of starchy substrates into ethanol by industrial Saccharomyces cerevisiae strains secreting fungal amylases.

    Science.gov (United States)

    Favaro, Lorenzo; Viktor, Marko J; Rose, Shaunita H; Viljoen-Bloom, Marinda; van Zyl, Willem H; Basaglia, Marina; Cagnin, Lorenzo; Casella, Sergio

    2015-09-01

    The development of a yeast strain that converts raw starch to ethanol in one step (called Consolidated Bioprocessing, CBP) could significantly reduce the commercial costs of starch-based bioethanol. An efficient amylolytic Saccharomyces cerevisiae strain suitable for industrial bioethanol production was developed in this study. Codon-optimized variants of the Thermomyces lanuginosus glucoamylase (TLG1) and Saccharomycopsis fibuligera α-amylase (SFA1) genes were δ-integrated into two S. cerevisiae yeast with promising industrial traits, i.e., strains M2n and MEL2. The recombinant M2n[TLG1-SFA1] and MEL2[TLG1-SFA1] yeast displayed high enzyme activities on soluble and raw starch (up to 8118 and 4461 nkat/g dry cell weight, respectively) and produced about 64 g/L ethanol from 200 g/L raw corn starch in a bioreactor, corresponding to 55% of the theoretical maximum ethanol yield (g of ethanol/g of available glucose equivalent). Their starch-to-ethanol conversion efficiencies were even higher on natural sorghum and triticale substrates (62 and 73% of the theoretical yield, respectively). This is the first report of direct ethanol production from natural starchy substrates (without any pre-treatment or commercial enzyme addition) using industrial yeast strains co-secreting both a glucoamylase and α-amylase. PMID:25786804

  17. Thermotolerant Kluyveromyces marxianus and Saccharomyces cerevisiae strains representing potentials for bioethanol production from Jerusalem artichoke by consolidated bioprocessing

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Nan [Agricultural Univ., Qingdao, SD (China). College of Animal Science and Technology; Chinese Academy of Sciences, Qingdao, SD (China). Key Lab. of Biofuels; Yuan, Bo; Wang, Shi-An; Li, Fu-Li [Chinese Academy of Sciences, Qingdao, SD (China). Key Lab. of Biofuels; Sun, Juan [Agricultural Univ., Qingdao, SD (China). College of Animal Science and Technology

    2012-09-15

    Thermotolerant inulin-utilizing yeast strains are desirable for ethanol production from Jerusalem artichoke tubers by consolidated bioprocessing (CBP). To obtain such strains, 21 naturally occurring yeast strains isolated by using an enrichment method and 65 previously isolated Saccharomyces cerevisiae strains were investigated in inulin utilization, extracellular inulinase activity, and ethanol fermentation from inulin and Jerusalem artichoke tuber flour at 40 C. The strains Kluyveromyces marxianus PT-1 (CGMCC AS2.4515) and S. cerevisiae JZ1C (CGMCC AS2.3878) presented the highest extracellular inulinase activity and ethanol yield in this study. The highest ethanol concentration in Jerusalem artichoke tuber flour fermentation (200 g L{sup -1}) at 40 C achieved by K. marxianus PT-1 and S. cerevisiae JZ1C was 73.6 and 65.2 g L{sup -1}, which corresponded to the theoretical ethanol yield of 90.0 and 79.7 %, respectively. In the range of 30 to 40 C, temperature did not have a significant effect on ethanol production for both strains. This study displayed the distinctive superiority of K. marxianus PT-1 and S. cerevisiae JZ1C in the thermotolerance and utilization of inulin-type oligosaccharides reserved in Jerusalem artichoke tubers. It is proposed that both K. marxianus and S. cerevisiae have considerable potential in ethanol production from Jerusalem artichoke tubers by a high temperature CBP. (orig.)

  18. Benchmarking two commonly used Saccharomyces cerevisiae strains for heterologous vanillin-β-glucoside production

    Directory of Open Access Journals (Sweden)

    Tomas Strucko

    2015-12-01

    Full Text Available The yeast Saccharomyces cerevisiae is a widely used eukaryotic model organism and a key cell factory for production of biofuels and wide range of chemicals. From the broad palette of available yeast strains, the most popular are those derived from laboratory strain S288c and the industrially relevant CEN.PK strain series. Importantly, in recent years these two strains have been subjected to comparative “-omics” analyzes pointing out significant genotypic and phenotypic differences. It is therefore possible that the two strains differ significantly with respect to their potential as cell factories for production of specific compounds. To examine this possibility, we have reconstructed a de novo vanillin-β-glucoside pathway in an identical manner in S288c and CEN.PK strains. Characterization of the two resulting strains in two standard conditions revealed that the S288c background strain produced up to 10-fold higher amounts of vanillin-β-glucoside compared to CEN.PK. This study demonstrates that yeast strain background may play a major role in the outcome of newly developed cell factories for production of a given product.

  19. Improvement of galactose uptake in Saccharomyces cerevisiae through overexpression of phosphoglucomutase: Example of transcript analysis as a tool in inverse metabolic engineering

    DEFF Research Database (Denmark)

    Bro, Christoffer; Knudsen, S.; Regenberg, Birgitte;

    2005-01-01

    Through genome-wide transcript analysis of a reference strain and two recombinant Saccharomyces cerevisiae strains with different rates of galactose uptake, we obtained information about the global transcriptional response to metabolic engineering of the GAL gene regulatory network. One of the...... reference strain. Based on our findings, we concluded that phosphoglucomutase plays a key role in controlling the flux through the Leloir pathway, probably due to increased conversion of glucose-l-phosphate to glucose-6-phosphate. This conclusion was supported by measurements of sugar phosphates, which...

  20. Engineering Saccharomyces cerevisiae toward n‐butanol production

    OpenAIRE

    Swidah, Reem

    2016-01-01

    Biobutanol represents a second generation biofuel, which can be producedfrom renewable resources by microorganisms. A Saccharomyces cerevisiae strainbearing the five butanol synthetic genes (hbd, adhe2, crt, ccr and ERG10) wasconstructed, where the hbd, adhe2, crt and ccr genes are derived from Clostridiumbeijerinckii, while ERG10 is a yeast gene. The genes were transformed individually onsingle cassettes, which integrated into specific chromosomal sites. The single integrantstrains were back...

  1. Genetic diversity and population structure of Saccharomyces cerevisiae strains isolated from different grape varieties and winemaking regions

    OpenAIRE

    Dorit Schuller; Filipa Cardoso; Susana Sousa; Paula Gomes; Gomes, Ana C.; Santos, Manuel A. S.; Margarida Casal

    2012-01-01

    We herein evaluate intraspecific genetic diversity of fermentative vineyard-associated S. cerevisiae strains and evaluate relationships between grape varieties and geographical location on populational structures. From the musts obtained from 288 grape samples, collected from two wine regions (16 vineyards, nine grape varieties), 94 spontaneous fermentations were concluded and 2820 yeast isolates were obtained that belonged mainly (92%) to the species S. cerevisiae. Isolates were classified i...

  2. Engineering the pentose phosphate pathway of Saccharomyces cerevisiae for production of ethanol and xylitol

    OpenAIRE

    Toivari, Mervi

    2007-01-01

    The baker s yeast Saccharomyces cerevisiae has a long tradition in alcohol production from D-glucose of e.g. starch. However, without genetic modifications it is unable to utilise the 5-carbon sugars D-xylose and L arabinose present in plant biomass. In this study, one key metabolic step of the catabolic D-xylose pathway in recombinant D-xylose-utilising S. cerevisiae strains was studied. This step, carried out by xylulokinase (XK), was shown to be rate-limiting, because overexpression of the...

  3. Engineering Saccharomyces cerevisiae for consolidated bioprocessing in starch and biomass conversion

    Science.gov (United States)

    The conversion of starch or biomass to biofuel is a two-stage process involving enzymatic treatment, followed by yeast fermentation. An alternative route would be to consolidate the process by engineering Saccharomyces cerevisiae capable of both saccharification and fermentation. An approach was d...

  4. Xylose-metabolizing Saccharomyces cerevisiae strains overexpressing the TKL1 and TAL1 genes encoding the pentose phosphate pathway enzymes transketolase and transaldolase.

    OpenAIRE

    Walfridsson, M; Hallborn, J; Penttilä, M.; Keränen, S; Hahn-Hägerdal, B

    1995-01-01

    Saccharomyces cerevisiae was metabolically engineered for xylose utilization. The Pichia stipitis CBS 6054 genes XYL1 and XYL2 encoding xylose reductase and xylitol dehydrogenase were cloned into S. cerevisiae. The gene products catalyze the two initial steps in xylose utilization which S. cerevisiae lacks. In order to increase the flux through the pentose phosphate pathway, the S. cerevisiae TKL1 and TAL1 genes encoding transketolase and transaldolase were overexpressed. A XYL1- and XYL2-con...

  5. Study of Saccharomyces cerevisiae wine strains for breeding through fermentation efficiency and tetrad analysis.

    Science.gov (United States)

    Fernández-González, Mónica; Úbeda, Juan F; Briones, Ana I

    2015-03-01

    One of the issues that most concerns to both winemakers and producers of active dry yeasts is the stuck and sluggish fermentations of grape musts with high levels of sugar, reflecting the inability of inoculated yeast strain to complete the fermentation process. It is difficult to obtain a wine strain that possesses both adequate oenological and technological properties; thus, the correct approach to solving these problems is the application of breeding programs primarily focused on both properties. The first step toward this process is to characterize the phenotypic diversity between potential parental strains. In the present study, we have analyzed the fermentative behavior of 26 Saccharomyces cerevisiae wine strains in high-sugar conditions at 20 °C, using a range of tests, such as sporulation ability, spore viability, and tetrad analysis to determine the tolerance of these yeasts to several stress conditions. Most tested strains were homothallic and heterozygous for more than one character. Two auxotrophic derivatives with defects in amino acid or nucleic acid metabolism were obtained, and these strains could potentially be used for the development of hybridization techniques without using laboratory strains. PMID:25447272

  6. Engineering piezoresistivity using biaxially strained silicon

    DEFF Research Database (Denmark)

    Pedersen, Jesper Goor; Richter, Jacob; Brandbyge, Mads;

    2008-01-01

    piezocoefficient on temperature and dopant density is altered qualitatively for strained silicon. In particular, we find that a vanishing temperature coefficient may result for silicon with grown-in biaxial tensile strain. These results suggest that strained silicon may be used to engineer the iezoresistivity to......We calculate the shear piezocoefficient of p-type silicon with grown-in biaxial strain using a 66 k·p method. We find a significant increase in the value of the shear piezocoefficient for compressive grown-in biaxial strain, while tensile strain decreases the piezocoefficient. The dependence of the...

  7. New integrative computational approaches unveil the Saccharomyces cerevisiae pheno-metabolomic fermentative profile and allow strain selection for winemaking.

    Science.gov (United States)

    Franco-Duarte, Ricardo; Umek, Lan; Mendes, Inês; Castro, Cristiana C; Fonseca, Nuno; Martins, Rosa; Silva-Ferreira, António C; Sampaio, Paula; Pais, Célia; Schuller, Dorit

    2016-11-15

    During must fermentation by Saccharomyces cerevisiae strains thousands of volatile aroma compounds are formed. The objective of the present work was to adapt computational approaches to analyze pheno-metabolomic diversity of a S. cerevisiae strain collection with different origins. Phenotypic and genetic characterization together with individual must fermentations were performed, and metabolites relevant to aromatic profiles were determined. Experimental results were projected onto a common coordinates system, revealing 17 statistical-relevant multi-dimensional modules, combining sets of most-correlated features of noteworthy biological importance. The present method allowed, as a breakthrough, to combine genetic, phenotypic and metabolomic data, which has not been possible so far due to difficulties in comparing different types of data. Therefore, the proposed computational approach revealed as successful to shed light into the holistic characterization of S. cerevisiae pheno-metabolome in must fermentative conditions. This will allow the identification of combined relevant features with application in selection of good winemaking strains. PMID:27283661

  8. Paclitaxel-induced microtubule stabilization causes mitotic block and apoptotic-like cell death in a paclitaxel-sensitive strain of Saccharomyces cerevisiae

    OpenAIRE

    Foland, Travis B.; Dentler, William L.; SUPRENANT, KATHY A.; Gupta, Mohan L.; Himes, Richard H.

    2005-01-01

    Wild-type Saccharomyces cerevisiae tubulin does not bind the anti-mitotic microtubule stabilizing agent paclitaxel. Previously, we introduced mutations into the S. cerevisiae gene for β-tubulin that imparted paclitaxel binding to the protein, but the mutant strain was not sensitive to paclitaxel and other microtubule-stabilizing agents, due to the multiple ABC transporters in the membranes of budding yeast. Here, we introduced the mutated β-tubulin gene into a S. cerevisiae strain with dimini...

  9. Impact of different spray-drying conditions on the viability of wine Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Aponte, Maria; Troianiello, Gabriele Danilo; Di Capua, Marika; Romano, Raffaele; Blaiotta, Giuseppe

    2016-01-01

    Spray-drying (SD) is widely considered a suitable method to preserve microorganisms, but data regarding yeasts are still scanty. In this study, the effect of growing media, process variables and carriers over viability of a wild wine Saccharomyces (S.) cerevisiae LM52 was evaluated. For biomass production, the strain was grown (batch and fed-batch fermentation) in a synthetic, as well as in a beet sugar molasses based-medium. Drying of cells resuspended in several combinations of soluble starch and maltose was performed at different inlet and outlet temperatures. Under the best conditions-suspension in soluble starch plus maltose couplet to inlet and outlet temperatures of 110 and 55 °C, respectively-the loss of viability of S. cerevisiae LM52 was 0.8 ± 0.1 and 0.5 ± 0.2 Log c.f.u. g(-1) for synthetic and molasses-based medium, respectively. Similar results were obtained when S. cerevisiae strains Zymoflore F15 and EC1118, isolated from commercial active dry yeast (ADY), were tested. Moreover, powders retained a high vitality and showed good fermentation performances up to 6 month of storage, at both 4 and -20 °C. Finally, fermentation performances of different kinds of dried formulates (SD and ADY) compared with fresh cultures did not show significant differences. The procedure proposed allowed a small-scale production of yeast in continuous operation with relatively simple equipment, and may thus represent a rapid response-on-demand for the production of autochthonous yeasts for local wine-making. PMID:26712628

  10. Engineering the Saccharomyces cerevisiae β-oxidation pathway to increase medium chain fatty acid production as potential biofuel.

    Directory of Open Access Journals (Sweden)

    Liwei Chen

    Full Text Available Fatty acid-derived biofuels and biochemicals can be produced in microbes using β-oxidation pathway engineering. In this study, the β-oxidation pathway of Saccharomyces cerevisiae was engineered to accumulate a higher ratio of medium chain fatty acids (MCFAs when cells were grown on fatty acid-rich feedstock. For this purpose, the haploid deletion strain Δpox1 was obtained, in which the sole acyl-CoA oxidase encoded by POX1 was deleted. Next, the POX2 gene from Yarrowia lipolytica, which encodes an acyl-CoA oxidase with a preference for long chain acyl-CoAs, was expressed in the Δpox1 strain. The resulting Δpox1 [pox2+] strain exhibited a growth defect because the β-oxidation pathway was blocked in peroxisomes. To unblock the β-oxidation pathway, the gene CROT, which encodes carnitine O-octanoyltransferase, was expressed in the Δpox1 [pox2+] strain to transport the accumulated medium chain acyl-coAs out of the peroxisomes. The obtained Δpox1 [pox2+, crot+] strain grew at a normal rate. The effect of these genetic modifications on fatty acid accumulation and profile was investigated when the strains were grown on oleic acids-containing medium. It was determined that the engineered strains Δpox1 [pox2+] and Δpox1 [pox2+, crot+] had increased fatty acid accumulation and an increased ratio of MCFAs. Compared to the wild-type (WT strain, the total fatty acid production of the strains Δpox1 [pox2+] and Δpox1 [pox2+, crot+] were increased 29.5% and 15.6%, respectively. The intracellular level of MCFAs in Δpox1 [pox2+] and Δpox1 [pox2+, crot+] increased 2.26- and 1.87-fold compared to the WT strain, respectively. In addition, MCFAs in the culture medium increased 3.29-fold and 3.34-fold compared to the WT strain. These results suggested that fatty acids with an increased MCFAs ratio accumulate in the engineered strains with a modified β-oxidation pathway. Our approach exhibits great potential for transforming low value fatty acid

  11. Strain engineering of graphene: a review

    Science.gov (United States)

    Si, Chen; Sun, Zhimei; Liu, Feng

    2016-02-01

    Graphene has intrigued the science community by many unique properties not found in conventional materials. In particular, it is the strongest two-dimensional material ever measured, being able to sustain reversible tensile elastic strain larger than 20%, which yields an interesting possibility to tune the properties of graphene by strain and thus opens a new field called ``straintronics''. In this article, the current progress in the strain engineering of graphene is reviewed. We first summarize the strain effects on the electronic structure and Raman spectra of graphene. We then highlight the electron-phonon coupling greatly enhanced by the biaxial strain and the strong pseudomagnetic field induced by the non-uniform strain with specific distribution. Finally, the potential application of strain-engineering in the self-assembly of foreign atoms on the graphene surface is also discussed. Given the short history of graphene straintronics research, the current progress has been notable, and many further advances in this field are expected.

  12. Fermentation of xylose to produce ethanol by recombinant Saccharomyces cerevisiae strain containing XYLA and XKS1

    Institute of Scientific and Technical Information of China (English)

    LIU Xiaolin; JIANG Ning; HE Peng; LU Dajun; SHEN An

    2005-01-01

    Fermentation of the pentose sugar xylose to produce ethanol using lignocellulosic biomass would make bioethanol production economically more competitive. Saccharomyce cerevisise, an efficient ethanol producer, cannot utilize xylose because it lacks the ability to convert xylose to its isomer xylulose. In this study, XYLA gene encoding xylose isomerase (XI) from Thermoanaerobacter tengcongensis MB4T and XKS1 gene encoding xylulokinase (XK) from Pichia stipitis were cloned and functionally coexpressed in Saccharomyces cerevisiae EF-326 to construct a recombinant xylose-utilizing strain. The resulting strain S. cerevisiae EF 1014 not only grew on xylose as sole carbon source, but also produced ethanol under anaerobic conditions. Fermentations performed with different xylose concentrations at different temperatures demonstrated that the highest ethanol productivity was 0.11 g/g xylose when xylose concentration was provided at 50 g/L. Under this condition, 28.4% of xylose was consumed and 1.54 g/L xylitol was formed. An increasing fermentation temperature from 30℃ to 37℃ did not improve ethanol yield.

  13. Construction of novel Saccharomyces cerevisiae strains for bioethanol active dry yeast (ADY production.

    Directory of Open Access Journals (Sweden)

    Daoqiong Zheng

    Full Text Available The application of active dry yeast (ADY in bioethanol production simplifies operation processes and reduces the risk of bacterial contamination. In the present study, we constructed a novel ADY strain with improved stress tolerance and ethanol fermentation performances under stressful conditions. The industrial Saccharomyces cerevisiae strain ZTW1 showed excellent properties and thus subjected to a modified whole-genome shuffling (WGS process to improve its ethanol titer, proliferation capability, and multiple stress tolerance for ADY production. The best-performing mutant, Z3-86, was obtained after three rounds of WGS, producing 4.4% more ethanol and retaining 2.15-fold higher viability than ZTW1 after drying. Proteomics and physiological analyses indicated that the altered expression patterns of genes involved in protein metabolism, plasma membrane composition, trehalose metabolism, and oxidative responses contribute to the trait improvement of Z3-86. This work not only successfully developed a novel S. cerevisiae mutant for application in commercial bioethanol production, but also enriched the current understanding of how WGS improves the complex traits of microbes.

  14. Engineering cofactor and transport mechanisms in Saccharomyces cerevisiae for enhanced acetyl-CoA and polyketide biosynthesis.

    Science.gov (United States)

    Cardenas, Javier; Da Silva, Nancy A

    2016-07-01

    Synthesis of polyketides at high titer and yield is important for producing pharmaceuticals and biorenewable chemical precursors. In this work, we engineered cofactor and transport pathways in Saccharomyces cerevisiae to increase acetyl-CoA, an important polyketide building block. The highly regulated yeast pyruvate dehydrogenase bypass pathway was supplemented by overexpressing a modified Escherichia coli pyruvate dehydrogenase complex (PDHm) that accepts NADP(+) for acetyl-CoA production. After 24h of cultivation, a 3.7-fold increase in NADPH/NADP(+) ratio was observed relative to the base strain, and a 2.2-fold increase relative to introduction of the native E. coli PDH. Both E. coli pathways increased acetyl-CoA levels approximately 2-fold relative to the yeast base strain. Combining PDHm with a ZWF1 deletion to block the major yeast NADPH biosynthesis pathway resulted in a 12-fold NADPH boost and a 2.2-fold increase in acetyl-CoA. At 48h, only this coupled approach showed increased acetyl-CoA levels, 3.0-fold higher than that of the base strain. The impact on polyketide synthesis was evaluated in a S. cerevisiae strain expressing the Gerbera hybrida 2-pyrone synthase (2-PS) for the production of the polyketide triacetic acid lactone (TAL). Titers of TAL relative to the base strain improved only 30% with the native E. coli PDH, but 3.0-fold with PDHm and 4.4-fold with PDHm in the Δzwf1 strain. Carbon was further routed toward TAL production by reducing mitochondrial transport of pyruvate and acetyl-CoA; deletions in genes POR2, MPC2, PDA1, or YAT2 each increased titer 2-3-fold over the base strain (up to 0.8g/L), and in combination to 1.4g/L. Combining the two approaches (NADPH-generating acetyl-CoA pathway plus reduced metabolite flux into the mitochondria) resulted in a final TAL titer of 1.6g/L, a 6.4-fold increase over the non-engineered yeast strain, and 35% of theoretical yield (0.16g/g glucose), the highest reported to date. These biological driving

  15. Molecular and Technological Characterization of Saccharomyces cerevisiae Strains Isolated from Natural Fermentation of Susumaniello Grape Must in Apulia, Southern Italy

    Directory of Open Access Journals (Sweden)

    Mariana Tristezza

    2014-01-01

    Full Text Available The characterization of autochthonous Saccharomyces cerevisiae strains is an important step towards the conservation and employment of microbial biodiversity. The utilization of selected autochthonous yeast strains would be a powerful tool to enhance the organoleptic and sensory properties of typical regional wines. In fact, indigenous yeasts are better tailored to a particular must and because of this they are able to praise the peculiarities of the derived wine. The present study described the biodiversity of indigenous S. cerevisiae strains isolated from natural must fermentations of an ancient and recently rediscovered Apulian grape cultivar, denoted as “Susumaniello.” The yeast strains denoted by the best oenological and technological features were identified and their fermentative performances were tested by either laboratory assay. Five yeast strains showed that they could be excellent candidates for the production of industrial starter cultures, since they dominated the fermentation process and produced wines characterized by peculiar oenological and organoleptic features.

  16. Production of fructanase by a wild strain of Saccharomyces cerevisiae on tequila agave fructan.

    Science.gov (United States)

    Corona-González, R I; Pelayo-Ortiz, C; Jacques, G; Guatemala, G; Arriola, E; Arias, J A; Toriz, G

    2015-01-01

    A new wild strain of Saccharomyces cerevisiae (CF3) isolated from tequila must was evaluated for production of fructanase on Agave tequilana Weber fructan (FT). Fructanase activity (F) was assessed by a 3(3) factorial design (substrate, temperature and pH). High enzymatic activity (31.1 U/ml) was found at 30 °C, pH 5, using FT (10 g/l) as substrate. The effect of initial substrate concentration on F (FT0, 5.7-66 g/l) was studied and it was found that F was highest (44.8 U/ml) at FT0 25 g/l. A 2(2) factorial experimental design with five central points was utilized to study the effect of stirring and aeration on fructanase activity; stirring exhibited a stronger effect on F. The ratio fructanase to invertase (F/S) was 0.57, which confirms that the enzymes are fructanase. Crude fructanase reached high substrate hydrolysis (48 wt%) in 10 h. It is shown that S. cerevisiae CF3 was able to produce large amounts of fructanase by growing it on fructan from A. tequilana. PMID:25432071

  17. Deletion of FPS1, Encoding Aquaglyceroporin Fps1p, Improves Xylose Fermentation by Engineered Saccharomyces cerevisiae

    OpenAIRE

    Wei, Na; Xu, Haiqing; Kim, Soo Rin; Jin, Yong-Su

    2013-01-01

    Accumulation of xylitol in xylose fermentation with engineered Saccharomyces cerevisiae presents a major problem that hampers economically feasible production of biofuels from cellulosic plant biomass. In particular, substantial production of xylitol due to unbalanced redox cofactor usage by xylose reductase (XR) and xylitol dehydrogenase (XDH) leads to low yields of ethanol. While previous research focused on manipulating intracellular enzymatic reactions to improve xylose metabolism, this s...

  18. IMPROVEMENT OF BORASSUS AKEASSII WINES QUALITY BY CONTROLLED FERMENTATION USING SACCHAROMYCES CEREVISIAE STRAINS

    Directory of Open Access Journals (Sweden)

    TAPSOBA François

    2016-06-01

    Full Text Available Palm wine produced traditionally and consumed by many people around the world and specifically in Burkina Faso posed health risks because of questionable quality of wine produced by mix culture fermentation and the use of antiseptics for the stabilization. In order to improve its quality, Saccharomyces cerevisiae strains isolated from Borassus akeassii wines and identified by amplification and RFLP analysis of the 5-8S-ITS region were used for in vitro fermentation of unfermented palm sap. The physicochemical characteristics of the sap were measured before and after fermentation process by High-Performance Liquid Chromatography (HPLC and the microbiological quality were also performed. HPLC analysis showed that glucose and fructose concentration in palm sap were 37.0 and 27.6 g/L respectively, ethanol content was ranged between 2.76 and 5.31 % (g/mL for controlled fermentation and 2.20 % (g/mL for spontaneous fermentation. Lactic and acetic acids were ranged between 0.1 and 0.3 g/L and 1.5 and 1.6 g/L for controlled fermentation versus 2.5 and 3.1 g/L and the spontaneous fermentation respectively. Coliforms and Staphylococcus aureus were detected only in the unfermented palm sap and the wine fermented spontaneously. Principal component analysis showed a good separation between spontaneous and controlled fermentation. Sterilization and controlled fermentation of the unfermented sap with palm wine Saccharomyces cerevisiae strains led to the improvement of palm wine quality.

  19. Biodiversity of autolytic ability in flocculent Saccharomyces cerevisiae strains suitable for traditional sparkling wine fermentation.

    Science.gov (United States)

    Perpetuini, Giorgia; Di Gianvito, Paola; Arfelli, Giuseppe; Schirone, Maria; Corsetti, Aldo; Tofalo, Rosanna; Suzzi, Giovanna

    2016-07-01

    Yeasts involved in secondary fermentation of traditional sparkling wines should show specific characteristics, such as flocculation capacity and autolysis. Recently it has been postulated that autophagy may contribute to the outcome of autolysis. In this study, 28 flocculent wine Saccahromyces cerevisiae strains characterized by different flocculation degrees were studied for their autolytic and autophagic activities. Autolysis was monitored in synthetic medium through the determination of amino acid nitrogen and total proteins released. At the same time, novel primer sets were developed to determine the expression of the genes ATG1, ATG17 and ATG29. Twelve strains were selected on the basis of their autolytic rate and ATG gene expressions in synthetic medium and were inoculated in a base wine. After 30, 60 and 180 days the autolytic process and ATG gene expressions were evaluated. The obtained data showed that autolysis and ATG gene expressions differed among strains and were independent of the degree of flocculation. This biodiversity could be exploited to select new starter stains to improve sparkling wine production. Copyright © 2016 John Wiley & Sons, Ltd. PMID:26804203

  20. A Saccharomyces cerevisiae Wine Strain Inhibits Growth and Decreases Ochratoxin A Biosynthesis by Aspergillus carbonarius and Aspergillus ochraceus

    Directory of Open Access Journals (Sweden)

    Marilena Budroni

    2012-12-01

    Full Text Available The aim of this study was to select wine yeast strains as biocontrol agents against fungal contaminants responsible for the accumulation of ochratoxin A (OTA in grape and wine and to dissect the mechanism of OTA detoxification by a Saccharomyces cerevisiae strain (DISAABA1182, which had previously been reported to reduce OTA in a synthetic must. All of the yeast strains tested displayed an ability to inhibit the growth of Aspergillus carbonarius both in vivo and in vitro and addition of culture filtrates from the tested isolates led to complete inhibition of OTA production. S. cerevisiae DISAABA1182 was selected and further tested for its capacity to inhibit OTA production and pks (polyketide synthase transcription in A. carbonarius and Aspergillus ochraceus in vitro. In order to dissect the mechanism of OTA detoxification, each of these two fungi was co-cultured with living yeast cells exposed to yeast crude or to autoclaved supernatant: S. cerevisiae DISAABA1182 was found to inhibit mycelial growth and OTA production in both Aspergilli when co-cultured in the OTA-inducing YES medium. Moreover, a decrease in pks transcription was observed in the presence of living cells of S. cerevisiae DISAABA1182 or its supernatant, while no effects were observed on transcription of either of the constitutively expressed calmodulin and β-tubulin genes. This suggests that transcriptional regulation of OTA biosynthetic genes takes place during the interaction between DISAABA1182 and OTA-producing Aspergilli.

  1. Xylose fermentation efficiency and inhibitor tolerance of the recombinant industrial Saccharomyces cerevisiae strain NAPX37.

    Science.gov (United States)

    Li, Yun-Cheng; Mitsumasu, Kanako; Gou, Zi-Xi; Gou, Min; Tang, Yue-Qin; Li, Guo-Ying; Wu, Xiao-Lei; Akamatsu, Takashi; Taguchi, Hisataka; Kida, Kenji

    2016-02-01

    Industrial yeast strains with good xylose fermentation ability and inhibitor tolerance are important for economical lignocellulosic bioethanol production. The flocculating industrial Saccharomyces cerevisiae strain NAPX37, harboring the xylose reductase-xylitol dehydrogenase (XR-XDH)-based xylose metabolic pathway, displayed efficient xylose fermentation during batch and continuous fermentation. During batch fermentation, the xylose consumption rates at the first 36 h were similar (1.37 g/L/h) when the initial xylose concentrations were 50 and 75 g/L, indicating that xylose fermentation was not inhibited even when the xylose concentration was as high as 75 g/L. The presence of glucose, at concentrations of up to 25 g/L, did not affect xylose consumption rate at the first 36 h. Strain NAPX37 showed stable xylose fermentation capacity during continuous ethanol fermentation using xylose as the sole sugar, for almost 1 year. Fermentation remained stable at a dilution rate of 0.05/h, even though the xylose concentration in the feed was as high as 100 g/L. Aeration rate, xylose concentration, and MgSO4 concentration were found to affect xylose consumption and ethanol yield. When the xylose concentration in the feed was 75 g/L, a high xylose consumption rate of 6.62 g/L/h and an ethanol yield of 0.394 were achieved under an aeration rate of 0.1 vvm, dilution rate of 0.1/h, and 5 mM MgSO4. In addition, strain NAPX37 exhibited good tolerance to inhibitors such as weak acids, furans, and phenolics during xylose fermentation. These findings indicate that strain NAPX37 is a promising candidate for application in the industrial production of lignocellulosic bioethanol. PMID:26603762

  2. Transcriptomics in human blood incubation reveals the importance of oxidative stress response in Saccharomyces cerevisiae clinical strains

    Directory of Open Access Journals (Sweden)

    Llopis Silvia

    2012-08-01

    Full Text Available Abstract Background In recent years an increasing number of yeast infections in humans have been related to certain clinical isolates of Saccharomyces cerevisiae. Some clinical strains showed in vivo and in vitro virulence traits and were able to cause death in mice whereas other clinical strains were avirulent. Results In this work, we studied the transcriptional profiles of two S. cerevisiae clinical strains showing virulent traits and two control non-virulent strains during a blood incubation model and detected a specific transcriptional response of clinical strains. This response involves an mRNA levels increase of amino acid biosynthesis genes and especially oxidative stress related genes. We observed that the clinical strains were more resistant to reactive oxygen species in vitro. In addition, blood survival of clinical isolates was high, reaching similar levels to pathogenic Candida albicans strain. Furthermore, a virulent strain mutant in the transcription factor Yap1p, unable to grow in oxidative stress conditions, presented decreased survival levels in human blood compared with the wild type or YAP1 reconstituted strain. Conclusions Our data suggest that this enhanced oxidative stress response in virulent clinical isolates, presumably induced in response to oxidative burst from host defense cells, is important to increase survival in human blood and can help to infect and even produce death in mice models.

  3. Improvement of Xylose Uptake and Ethanol Production in Recombinant Saccharomyces cerevisiae through an Inverse Metabolic Engineering Approach

    OpenAIRE

    Jin, Yong-Su; Alper, Hal; Yang, Yea-Tyng; Stephanopoulos, Gregory

    2005-01-01

    We used an inverse metabolic engineering approach to identify gene targets for improved xylose assimilation in recombinant Saccharomyces cerevisiae. Specifically, we created a genomic fragment library from Pichia stipitis and introduced it into recombinant S. cerevisiae expressing XYL1 and XYL2. Through serial subculturing enrichment of the transformant library, 16 transformants were identified and confirmed to have a higher growth rate on xylose. Sequencing of the 16 plasmids isolated from t...

  4. Kinetic Activity of Commercial Native Linamarase (CNLIN) and Engineered <&beta-glucosidase) from Saccharomyces cerevisiae on Cassava Linamarin

    OpenAIRE

    Julius Kwagh-Har Ikya; Charles Chukwuma Ariahu; James Ortese Ayatse

    2014-01-01

    The ability of Commercial Native Linamarase (CNLIN) and Engineered Linamarase (GELIN) extracts from Saccharomyces cerevisiae to hydrolyse cassava linamarin was challenged. CNLIN acting as control was used together with GELIN extracts from Saccharomyces cerevisiae to evaluate the kinetic data for test enzymes at pH 3.5, 6.8 and 10.5, respectively and ambient temperature (35°C). Data obtained from the varying activity versus substrate concentrations were fitted with the Michaelis-Menten plots a...

  5. Surfome analysis of a wild-type wine Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Braconi, Daniela; Amato, Loredana; Bernardini, Giulia; Arena, Simona; Orlandini, Maurizio; Scaloni, Andrea; Santucci, Annalisa

    2011-09-01

    The yeast Saccharomyces cerevisiae, besides being an eukaryotic cell model, plays a fundamental role in the production of fermented foods. In the winemaking industry, yeast cell walls may be involved in numerous processes and contribute substantially to the final chemical and sensorial profiles of wines. Nonetheless, apart from mannoproteins, little is known on the protein components of the yeast cell wall and their changes during the fermentation of must into wine. In this work, we performed a dynamic analysis of the cell surface proteome (surfome) of an autochthonous wine yeast strain (previously selected as a wine fermentation starter) by shaving intact cells with trypsin and identifying tryptic peptides by means of nLC-ESI-LIT-MS/MS. Out of the 42 identified proteins, 16 and 14 were found to be specifically expressed in wine yeast surfome at the beginning and at the end of fermentation, respectively. The molecular functions of these specifically expressed proteins might help in explaining their roles in the cell wall as a response to the alcoholic fermentation-related stresses. Additionally, we provided the identification of 20 new potential cell wall related proteins. Globally, our results might provide new useful data for the selection and characterization of yeast strains to be used in the winemaking industry. PMID:21645823

  6. Development of minimal fermentation media supplementation for ethanol production using two Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Tropea, Alessia; Wilson, David; Cicero, Nicola; Potortì, Angela G; La Torre, Giovanna L; Dugo, Giacomo; Richardson, David; Waldron, Keith W

    2016-05-01

    Ethanol production by fermentation is strongly dependent on media composition. Specific nutrients, such as trace elements, vitamins and nitrogen will affect the physiological state and, consequently, the fermentation performance of the micro-organism employed. The purpose of this study has been to assess the highest ethanol production by a minimal medium, instead of the more complex nutrients supplementation used during alcoholic fermentation. All fermentation tests were carried out using a microwell plate reader to monitor the processes. Two Saccharomyces cerevisiae strains (NCYC 2826 and NCYC 3445) were tested using three nitrogen sources, supplied with different vitamin and salts. The results show that solutions made of urea phosphate, KCl, MgSO4·7H2O, Ca-panthothenate, biotin allowed an ethanol yield of 22.9 and 23.4 g/L for strain NCYC 2826 and NCYC 3445, respectively, representing 90 and 92% of the theoretical yield. All tests were carried out using glucose as common reference carbon source. PMID:26469871

  7. Engineering of chromosomal wax ester synthase integrated Saccharomyces cerevisiae mutants for improved biosynthesis of fatty acid ethyl esters.

    Science.gov (United States)

    Shi, Shuobo; Valle-Rodríguez, Juan Octavio; Siewers, Verena; Nielsen, Jens

    2014-09-01

    In recent years, significant advances have been made to engineer robust microbes for overproducing biochemical products from renewable resources. These accomplishments have to a large extend been based on plasmid based methods. However, plasmid maintenance may cause a metabolic burden on the host cell and plasmid-based overexpression of genes can result in genetically unstable strains, which contributes to loss in productivity. Here, a chromosome engineering method based on delta integration was applied in Saccharomyces cerevisiae for the production of fatty acid ethyl esters (FAEEs), which can be directly used as biodiesel and would be a possible substitute for conventional petroleum-based diesel. An integration construct was designed and integrated into chromosomal delta sequences by repetitive transformation, which resulted in 1-6 copies of the integration construct per genome. The corresponding FAEE production increased up to 34 mg/L, which is an about sixfold increase compared to the equivalent plasmid-based producer. The integrated cassette in the yeast genome was stably maintained in nonselective medium after deletion of RAD52 which is essential for efficient homologous recombination. To obtain a further increase of FAEE production, genes encoding endogenous acyl-CoA binding protein (ACB1) and a bacterial NADP(+)-dependent glyceraldehyde-3-phosphate dehydrogenase (gapN) were overexpressed in the final integration strain, which resulted in another 40% percent increase in FAEE production. Our integration strategy enables easy engineering of strains with adjustable gene copy numbers integrated into the genome and this allows for an easy evaluation of the effect of the gene copy number on pathway flux. It therefore represents a valuable tool for introducing and expressing a heterologous pathway in yeast. PMID:24752598

  8. CRISPR–Cas system enables fast and simple genome editing of industrial Saccharomyces cerevisiae strains

    DEFF Research Database (Denmark)

    Stovicek, Vratislav; Borodina, Irina; Förster, Jochen

    2015-01-01

    genetic manipulation is challenging, as they are usually diploid or polyploid. Therefore, there is a need to develop more efficient genetic engineering tools. We applied a CRISPR–Cas9 system for genome editing of different industrial strains, and show simultaneous disruption of two alleles of a gene in...

  9. Ethanol from lignocellulose - Fermentation inhibitors, detoxification and genetic engineering of Saccharomyces cerevisiae for enhanced resistance

    Energy Technology Data Exchange (ETDEWEB)

    Larsson, Simona

    2000-07-01

    Ethanol can be produced from lignocellulose by first hydrolysing the material to sugars, and then fermenting the hydrolysate with the yeast Saccharomyces cerevisiae. Hydrolysis using dilute sulphuric acid has advantages over other methods, however, compounds which inhibit fermentation are generated during this kind of hydrolysis. The inhibitory effect of aliphatic acids, furans, and phenolic compounds was investigated. The generation of inhibitors during hydrolysis was studied using Norway spruce as raw material. It was concluded that the decrease in the fermentability coincided with increasing harshness of the hydrolysis conditions. The decrease in fermentability was not correlated solely to the content of aliphatic acids or furan derivatives. To increase the fermentability, detoxification is often employed. Twelve detoxification methods were compared with respect to the chemical composition of the hydrolysate and the fermentability after treatment. The most efficient detoxification methods were anion-exchange at pH 10.0, overliming and enzymatic detoxification with the phenol-oxidase laccase. Detailed analyses of ion exchange revealed that anion exchange and unspecific hydrophobic interactions greatly contributed to the detoxification effect, while cation exchange did not. The comparison of detoxification methods also showed that phenolic compounds are very important fermentation inhibitors, as their selective removal with laccase had a major positive effect on the fermentability. Selected compounds; aliphatic acids, furans and phenolic compounds, were characterised with respect to their inhibitory effect on ethanolic fermentation by S. cerevisiae. When aliphatic acids or furans were compared, the inhibitory effects were found to be in the same range, but the phenolic compounds displayed widely different inhibitory effects. The possibility of genetically engineering S. cerevisiae to achieve increased inhibitor resistance was explored by heterologous expression of

  10. Diversity of Saccharomyces cerevisiae strains isolated from Borassus akeassii palm wines from Burkina Faso in comparison to other African beverages.

    Science.gov (United States)

    Tapsoba, François; Legras, Jean-Luc; Savadogo, Aly; Dequin, Sylvie; Traore, Alfred Sababenedyo

    2015-10-15

    In South-West of Burkina Faso, palm wine is produced by spontaneous fermentation of the sap from a specific palm tree Borassus akeassii and plays an important role in people's lives. Saccharomyces cerevisiae is the main agent of this alcoholic fermentation but little is known about the diversity of the isolates from palm. In this work, 39 Saccharomyces cerevisiae strains were isolated from palm wine samples collected from 14 sites in Burkina Faso, as well as 7 isolates obtained from sorghum beer (Dolo) from 3 distant sites. Their diversity was analyzed at 12 microsatellite loci, and compared to the genotypes obtained for other African yeast populations isolated from Cocoa hulks from Ghana, sorghum beer from Ivory Coast, palm wine from Djibouti Republic, and to our database of strains from miscellaneous origins (bread, beer, wine, sake, oaks…). The ploidy of these strains has been assessed as well by flow cytometry. Our results show that B. akeassii palm wine contains a specific yeast population of diploid strains, different from Dolo produced in the same area and from other palm wine strains from Ivory Coast, Nigeria, or Djibouti Republic. In contrast, Dolo strains appeared as a group of related and mainly tetraploid strains despite being isolated from different countries. PMID:26202324

  11. De novo production of the flavonoid naringenin in engineered Saccharomyces cerevisiae

    Science.gov (United States)

    2012-01-01

    Background Flavonoids comprise a large family of secondary plant metabolic intermediates that exhibit a wide variety of antioxidant and human health-related properties. Plant production of flavonoids is limited by the low productivity and the complexity of the recovered flavonoids. Thus to overcome these limitations, metabolic engineering of specific pathway in microbial systems have been envisaged to produce high quantity of a single molecules. Result Saccharomyces cerevisiae was engineered to produce the key intermediate flavonoid, naringenin, solely from glucose. For this, specific naringenin biosynthesis genes from Arabidopsis thaliana were selected by comparative expression profiling and introduced in S. cerevisiae. The sole expression of these A. thaliana genes yielded low extracellular naringenin concentrations (Synthesis of aromatic amino acids was deregulated by alleviating feedback inhibition of 3-deoxy-d-arabinose-heptulosonate-7-phosphate synthase (Aro3, Aro4) and byproduct formation was reduced by eliminating phenylpyruvate decarboxylase (Aro10, Pdc5, Pdc6). Together with an increased copy number of the chalcone synthase gene and expression of a heterologous tyrosine ammonia lyase, these modifications resulted in a 40-fold increase of extracellular naringenin titers (to approximately 200 μM) in glucose-grown shake-flask cultures. In aerated, pH controlled batch reactors, extracellular naringenin concentrations of over 400 μM were reached. Conclusion The results reported in this study demonstrate that S. cerevisiae is capable of de novo production of naringenin by coexpressing the naringenin production genes from A. thaliana and optimization of the flux towards the naringenin pathway. The engineered yeast naringenin production host provides a metabolic chassis for production of a wide range of flavonoids and exploration of their biological functions. PMID:23216753

  12. Plasmid-mediate transfer of FLO-1 into industrial Saccharomyces cerevisiae PE-2 strain creates a strain useful for repeat-batch fermentations involving flocculation-sedimentation

    OpenAIRE

    Gomes, Daniel Gonçalves; Guimarães, Pedro M. R.; Pereira, Francisco B.; Teixeira, J.A.; Domingues, Lucília

    2012-01-01

    The flocculation gene FLO1 was transferred into the robust industrial strain Saccharomyces cerevisiae PE-2 by the lithium acetate method. The recombinant strain showed a fermentation performance similar to that of the parental strain. In 10 repeat-batch cultivations in VHG medium with 345 g glucose/L and cell recycling by flocculation–sedimentation, an average final ethanol concentration of 142 g/L and an ethanol productivity of 2.86 g/L/h were achieved. Due to the flocculent natu...

  13. Model-guided identification of gene deletion targets for metabolic engineering in Saccharomyces cerevisiae.

    Science.gov (United States)

    Brochado, Ana Rita; Patil, Kiran Raosaheb

    2014-01-01

    Identification of metabolic engineering strategies for rerouting intracellular fluxes towards a desired product is often a challenging task owing to the topological and regulatory complexity of metabolic networks. Genome-scale metabolic models help tackling this complexity through systematic consideration of mass balance and reaction directionality constraints over the entire network. Here, we describe how genome-scale metabolic models can be used for identifying gene deletion targets leading to increased production of the desired product. Vanillin production in Saccharomyces cerevisiae is used as a case study throughout this chapter. PMID:24744040

  14. Design and engineering of intracellular-metabolite-sensing/regulation gene circuits in Saccharomyces cerevisiae.

    Science.gov (United States)

    Wang, Meng; Li, Sijin; Zhao, Huimin

    2016-01-01

    The development of high-throughput phenotyping tools is lagging far behind the rapid advances of genotype generation methods. To bridge this gap, we report a new strategy for design, construction, and fine-tuning of intracellular-metabolite-sensing/regulation gene circuits by repurposing bacterial transcription factors and eukaryotic promoters. As proof of concept, we systematically investigated the design and engineering of bacterial repressor-based xylose-sensing/regulation gene circuits in Saccharomyces cerevisiae. We demonstrated that numerous properties, such as induction ratio and dose-response curve, can be fine-tuned at three different nodes, including repressor expression level, operator position, and operator sequence. By applying these gene circuits, we developed a cell sorting based, rapid and robust high-throughput screening method for xylose transporter engineering and obtained a sugar transporter HXT14 mutant with 6.5-fold improvement in xylose transportation capacity. This strategy should be generally applicable and highly useful for evolutionary engineering of proteins, pathways, and genomes in S. cerevisiae. PMID:26059511

  15. Metabolite Profiling during Fermentation of Makgeolli by the Wild Yeast Strain Saccharomyces cerevisiae Y98-5

    OpenAIRE

    Kim, Hye Ryun; Kim, Jae-Ho; Ahn, Byung Hak; Bai, Dong-Hoon

    2014-01-01

    Makgeolli is a traditional Korean alcoholic beverage. The flavor of makgeolli is primarily determined by metabolic products such as free sugars, amino acids, organic acids, and aromatic compounds, which are produced during the fermentation of raw materials by molds and yeasts present in nuruk, a Korean fermentation starter. In this study, makgeolli was brewed using the wild yeast strain Saccharomyces cerevisiae Y98-5, and temporal changes in the metabolites during fermentation were analyzed b...

  16. Systems biology approaches for the design of novel Saccharomyces cerevisiae winemaking strains for enhanced flavour compounds synthesis

    OpenAIRE

    Mendes, Inês Isabel Moreira Moutinho Vieira

    2015-01-01

    Tese de Doutoramento em Biologia Ambiental e Molecular Wine flavour and aroma are the result of yeast metabolism and must compounds interactions. During must fermentation thousands of volatile aroma compounds are formed, with higher alcohols, acetate esters and ethyl esters being the main aromatic compounds contributing to a floral and fruity aroma. The action of yeast, in particular of Saccharomyces cerevisiae strains, on the must components will build the architecture of t...

  17. Selection of autochthonous Saccharomyces cerevisiae strains as wine starters using a polyphasic approach and ochratoxin A removal.

    Science.gov (United States)

    Petruzzi, Leonardo; Bevilacqua, Antonio; Corbo, Maria Rosaria; Garofalo, Carmela; Baiano, Antonietta; Sinigaglia, Milena

    2014-07-01

    Over the last few years, the selection of autochthonous strains of Saccharomyces cerevisiae as wine starters has been studied; however, researchers have not focused on the ability to remove ochratoxin A (OTA) as a possible trait to use in oenological characterization. In this article, a polyphasic approach, including yeast genotyping, evaluation of phenotypic traits, and fermentative performance in a model system (temperature, 25 and 30°C; sugar level, 200 and 250 g liter(-1)), was proposed as a suitable approach to select wine starters of S. cerevisiae from 30 autochthonous isolates from Uva di Troia cv., a red wine grape variety grown in the Apulian region (Southern Italy). The ability to remove OTA, a desirable trait to improve the safety of wine, was also assessed using enzyme-linked immunosorbent assay. The isolates, identified by PCR-restriction fragment length polymorphism analysis of the internal transcribed spacer region and DNA sequencing, were differentiated at strain level through the amplification of the interdelta region; 11 biotypes (I to XI) were identified and further studied. Four biotypes (II, III, V, VIII) were able to reduce OTA, with the rate of toxin removal from the medium (0.6 to 42.8%, wt/vol) dependent upon the strain and the temperature, and biotypes II and VIII were promising in terms of ethanol, glycerol, and volatile acidity production, as well as for their enzymatic and stress resistance characteristics. For the first time, the ability of S. cerevisiae to remove OTA during alcoholic fermentation was used as an additional trait in the yeast-selection program; the results could have application for evaluating the potential of autochthonous S. cerevisiae strains as starter cultures for the production of typical wines with improved quality and safety. PMID:24988024

  18. Cell recycling during repeated very high gravity bio-ethanol fermentations using the industrial Saccharomyces cerevisiae strain PE-2

    OpenAIRE

    Pereira, Francisco B.; Gomes, Daniel Gonçalves; Guimarães, Pedro M. R.; Teixeira, J.A.; Domingues, Lucília

    2012-01-01

    A very high gravity (VHG) repeatedbatch fermentation system using an industrial strain of Saccharomyces cerevisiae PE-2 (isolated from sugarcane-to-ethanol distillery in Brazil) and mimicking industrially relevant conditions (high inoculation rates and low O2 availability) was successfully operated during fifteen consecutive fermentation cycles, attaining ethanol at 17.1 ± 0.2% (v/v) with a batch productivity of 3.5 ± 0.04 g l-1 h-1. Moreover, this innovative operat...

  19. Glycoprotein Hypersecretion Alters the Cell Wall in Trichoderma reesei Strains Expressing the Saccharomyces cerevisiae Dolichylphosphate Mannose Synthase Gene▿

    OpenAIRE

    Perlińska-Lenart, Urszula; Orłowski, Jacek; Laudy, Agnieszka E.; Zdebska, Ewa; Palamarczyk, Grażyna; Kruszewska, Joanna S.

    2006-01-01

    Expression of the Saccharomyces cerevisiae DPM1 gene (coding for dolichylphosphate mannose synthase) in Trichoderma reesei (Hypocrea jecorina) increases the intensity of protein glycosylation and secretion and causes ultrastructural changes in the fungal cell wall. In the present work, we undertook further biochemical and morphological characterization of the DPM1-expressing T. reesei strains. We established that the carbohydrate composition of the fungal cell wall was altered with an increas...

  20. Physicochemical characterization of pomegranate wines fermented with three different Saccharomyces cerevisiae yeast strains.

    Science.gov (United States)

    Berenguer, María; Vegara, Salud; Barrajón, Enrique; Saura, Domingo; Valero, Manuel; Martí, Nuria

    2016-01-01

    Three commercial Saccharomyces cerevisiae yeast strains: Viniferm Revelación, Viniferm SV and Viniferm PDM were evaluated for the production of pomegranate wine from a juice coupage of the two well-known varieties Mollar and Wonderfull. Further malolactic fermentation was carried out spontaneously. The same fermentation patterns were observed for pH, titratable acidity, density, sugar consumption, and ethanol and glycerol production. Glucose was exhausted while fructose residues remained at the end of alcoholic fermentation. A high ethanol concentration (10.91 ± 0.27% v/v) in combination with 1.49 g/L glycerol was achieved. Citric acid concentration increased rapidly a 31.7%, malic acid disappeared as result of malolactic fermentation and the lactic acid levels reached values between 0.40 and 0.96 g/L. The analysis of CIEa parameter and total anthocyanin content highlights a lower degradation of monomeric anthocyanins during winemaking with Viniferm PDM yeast. The resulting wine retains a 34.5% of total anthocyanin content of pomegranate juice blend. PMID:26213048

  1. Heterologous Expression of Amylase Gene from Saccharomycopsis fibuligera in an Industrial Strain of Saccharomyces cerevisiae

    Institute of Scientific and Technical Information of China (English)

    LIU Zeng-ran; ZHANG Guang-yi; LONG Zhang-fu; LIU Shi-gui

    2005-01-01

    An α-amylase encoding gene was amplified by polymerase chain reaction from Saccharomycopsis fibuligera and inserted into a shuttle vector YEp352,together with the yeast phosphoglycerate kinase 1 promoter and α-factor signal gene. The recombinant expression plasmid pLA8α was transformed into an industrial strain of Saccharomyces cerevisiae Sc-11. The activity of the α-amylase produced by the transformant Sc-11-pLA8α was 6.3 U/mL and the starch utilization rate in YPS medium was 42 %. The purified amylase was analyzed by SDS-PAGE,showing a molecular weight of 55×103 protein band. Furthermore, the residual sugar, ethanol and some volatile compounds in the fermented worts under simulating brewing conditions were determined by chromatographic analyses. The fermentation characteristics of Sc-11-pLA8α were similar to that of Sc-11 and only minor changes in the concentration of flavor compounds could be observed.

  2. Cell surface engineering of Saccharomyces cerevisiae combined with membrane separation technology for xylitol production from rice straw hydrolysate.

    Science.gov (United States)

    Guirimand, Gregory; Sasaki, Kengo; Inokuma, Kentaro; Bamba, Takahiro; Hasunuma, Tomohisa; Kondo, Akihiko

    2016-04-01

    Xylitol, a value-added polyol deriving from D-xylose, is widely used in both the food and pharmaceutical industries. Despite extensive studies aiming to streamline the production of xylitol, the manufacturing cost of this product remains high while demand is constantly growing worldwide. Biotechnological production of xylitol from lignocellulosic waste may constitute an advantageous and sustainable option to address this issue. However, to date, there have been few reports of biomass conversion to xylitol. In the present study, xylitol was directly produced from rice straw hydrolysate using a recombinant Saccharomyces cerevisiae YPH499 strain expressing cytosolic xylose reductase (XR), along with β-glucosidase (BGL), xylosidase (XYL), and xylanase (XYN) enzymes (co-)displayed on the cell surface; xylitol production by this strain did not require addition of any commercial enzymes. All of these enzymes contributed to the consolidated bioprocessing (CBP) of the lignocellulosic hydrolysate to xylitol to produce 5.8 g/L xylitol with 79.5 % of theoretical yield from xylose contained in the biomass. Furthermore, nanofiltration of the rice straw hydrolysate provided removal of fermentation inhibitors while simultaneously increasing sugar concentrations, facilitating high concentration xylitol production (37.9 g/L) in the CBP. This study is the first report (to our knowledge) of the combination of cell surface engineering approach and membrane separation technology for xylitol production, which could be extended to further industrial applications. PMID:26631184

  3. The level of glucose-6-phosphate dehydrogenase activity strongly influences xylose fermentation and inhibitor sensitivity in recombinant Saccharomyces cerevisiae strains

    DEFF Research Database (Denmark)

    Jeppsson, M.; Johansson, B.; Jensen, Peter Ruhdal;

    2003-01-01

    Disruption of the ZWF1 gene encoding glucose-6-phosphate dehydrogenase (G6PDH) has been shown to reduce the xylitol yield and the xylose consumption in the xylose-utilizing recombinant Saccharomyces cerevisiae strain TMB3255. In the present investigation we have studied the influence of different...... consumption, respectively, compared with the ZWF1-disrupted strain. Both strains exhibited decreased xylitol yields (0.13 and 0.19 g/g xylose) and enhanced ethanol yields (0.36 and 0.34 g/g xylose) compared with the control strain TMB3001 (0.29 g xylitol/g xylose, 0.31 g ethanol/g xylose). Cytoplasmic...... transhydrogenase (TH) from Azotobacter vinelandii has previously been shown to transfer NADPH and NAD(+) into NADP(+) and NADH, and TH-overproduction resulted in lower xylitol yield and enhanced glycerol yield during xylose utilization. Strains with low G6PDH-activity grew slower in a lignocellulose hydrolysate...

  4. Whole Genome Comparison Reveals High Levels of Inbreeding and Strain Redundancy Across the Spectrum of Commercial Wine Strains of Saccharomyces cerevisiae

    Science.gov (United States)

    Borneman, Anthony R.; Forgan, Angus H.; Kolouchova, Radka; Fraser, James A.; Schmidt, Simon A.

    2016-01-01

    Humans have been consuming wines for more than 7000 yr . For most of this time, fermentations were presumably performed by strains of Saccharomyces cerevisiae that naturally found their way into the fermenting must . In contrast, most commercial wines are now produced by inoculation with pure yeast monocultures, ensuring consistent, reliable and reproducible fermentations, and there are now hundreds of these yeast starter cultures commercially available. In order to thoroughly investigate the genetic diversity that has been captured by over 50 yr of commercial wine yeast development and domestication, whole genome sequencing has been performed on 212 strains of S. cerevisiae, including 119 commercial wine and brewing starter strains, and wine isolates from across seven decades. Comparative genomic analysis indicates that, despite their large numbers, commercial strains, and wine strains in general, are extremely similar genetically, possessing all of the hallmarks of a population bottle-neck, and high levels of inbreeding. In addition, many commercial strains from multiple suppliers are nearly genetically identical, suggesting that the limits of effective genetic variation within this genetically narrow group may be approaching saturation. PMID:26869621

  5. Whole Genome Comparison Reveals High Levels of Inbreeding and Strain Redundancy Across the Spectrum of Commercial Wine Strains of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Anthony R. Borneman

    2016-04-01

    Full Text Available Humans have been consuming wines for more than 7000 yr . For most of this time, fermentations were presumably performed by strains of Saccharomyces cerevisiae that naturally found their way into the fermenting must . In contrast, most commercial wines are now produced by inoculation with pure yeast monocultures, ensuring consistent, reliable and reproducible fermentations, and there are now hundreds of these yeast starter cultures commercially available. In order to thoroughly investigate the genetic diversity that has been captured by over 50 yr of commercial wine yeast development and domestication, whole genome sequencing has been performed on 212 strains of S. cerevisiae, including 119 commercial wine and brewing starter strains, and wine isolates from across seven decades. Comparative genomic analysis indicates that, despite their large numbers, commercial strains, and wine strains in general, are extremely similar genetically, possessing all of the hallmarks of a population bottle-neck, and high levels of inbreeding. In addition, many commercial strains from multiple suppliers are nearly genetically identical, suggesting that the limits of effective genetic variation within this genetically narrow group may be approaching saturation.

  6. Whole Genome Comparison Reveals High Levels of Inbreeding and Strain Redundancy Across the Spectrum of Commercial Wine Strains of Saccharomyces cerevisiae.

    Science.gov (United States)

    Borneman, Anthony R; Forgan, Angus H; Kolouchova, Radka; Fraser, James A; Schmidt, Simon A

    2016-01-01

    Humans have been consuming wines for more than 7000 yr . For most of this time, fermentations were presumably performed by strains of Saccharomyces cerevisiae that naturally found their way into the fermenting must . In contrast, most commercial wines are now produced by inoculation with pure yeast monocultures, ensuring consistent, reliable and reproducible fermentations, and there are now hundreds of these yeast starter cultures commercially available. In order to thoroughly investigate the genetic diversity that has been captured by over 50 yr of commercial wine yeast development and domestication, whole genome sequencing has been performed on 212 strains of S. cerevisiae, including 119 commercial wine and brewing starter strains, and wine isolates from across seven decades. Comparative genomic analysis indicates that, despite their large numbers, commercial strains, and wine strains in general, are extremely similar genetically, possessing all of the hallmarks of a population bottle-neck, and high levels of inbreeding. In addition, many commercial strains from multiple suppliers are nearly genetically identical, suggesting that the limits of effective genetic variation within this genetically narrow group may be approaching saturation. PMID:26869621

  7. New promoters for strain engineering of Penicillium chrysogenum.

    Science.gov (United States)

    Polli, Fabiola; Meijrink, Ben; Bovenberg, Roel A L; Driessen, Arnold J M

    2016-04-01

    Filamentous fungi such as Aspergillus and Penicillium are widely used as hosts for the industrial products such as proteins and secondary metabolites. Although filamentous fungi are versatile in recognizing transcriptional and translational elements present in genes from other filamentous fungal species, only few promoters have been applied and compared in performance so far in Penicillium chrysogenum. Therefore, a set of homologous and heterologous promoters were tested in a reporter system to obtain a set of potential different strengths. Through in vivo homologous recombination in Saccharomyces cerevisiae, twelve Aspergillus niger and P. chrysogenum promoter-reporter pathways were constructed that drive the expression of green fluorescent protein while concurrent expression of the red fluorescent protein was used as an internal standard and placed under control of the PcPAF promoter. The pathways were integrated into the genome of P. chrysogenum and tested using the BioLector system for fermentation. Reporter gene expression was monitored during growth and classified according to promoter strength and expression profile. A set of novel promoters was obtained that can be used to tune the expression of target genes in future strain engineering programs. PMID:26701309

  8. Cellulosic Ethanol Production from Saccharomyces cerevisiae Engineered for Anaerobic Conversion of Pretreated Lignocellulosic Sugars to Ethanol

    Science.gov (United States)

    Advanced high-throughput screening has resulted in the discovery of several yeast strains that are capable of anaerobically utilizing pentose, as well as hexose sugars. The growth and ethanol production of these developed strains will be described. The paradigm for using genetically engineered Sac...

  9. Comparing the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways in arabinose and xylose fermenting Saccharomyces cerevisiae strains

    Science.gov (United States)

    Bettiga, Maurizio; Hahn-Hägerdal, Bärbel; Gorwa-Grauslund, Marie F

    2008-01-01

    Background Ethanolic fermentation of lignocellulosic biomass is a sustainable option for the production of bioethanol. This process would greatly benefit from recombinant Saccharomyces cerevisiae strains also able to ferment, besides the hexose sugar fraction, the pentose sugars, arabinose and xylose. Different pathways can be introduced in S. cerevisiae to provide arabinose and xylose utilisation. In this study, the bacterial arabinose isomerase pathway was combined with two different xylose utilisation pathways: the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways, respectively, in genetically identical strains. The strains were compared with respect to aerobic growth in arabinose and xylose batch culture and in anaerobic batch fermentation of a mixture of glucose, arabinose and xylose. Results The specific aerobic arabinose growth rate was identical, 0.03 h-1, for the xylose reductase/xylitol dehydrogenase and xylose isomerase strain. The xylose reductase/xylitol dehydrogenase strain displayed higher aerobic growth rate on xylose, 0.14 h-1, and higher specific xylose consumption rate in anaerobic batch fermentation, 0.09 g (g cells)-1 h-1 than the xylose isomerase strain, which only reached 0.03 h-1 and 0.02 g (g cells)-1h-1, respectively. Whereas the xylose reductase/xylitol dehydrogenase strain produced higher ethanol yield on total sugars, 0.23 g g-1 compared with 0.18 g g-1 for the xylose isomerase strain, the xylose isomerase strain achieved higher ethanol yield on consumed sugars, 0.41 g g-1 compared with 0.32 g g-1 for the xylose reductase/xylitol dehydrogenase strain. Anaerobic fermentation of a mixture of glucose, arabinose and xylose resulted in higher final ethanol concentration, 14.7 g l-1 for the xylose reductase/xylitol dehydrogenase strain compared with 11.8 g l-1 for the xylose isomerase strain, and in higher specific ethanol productivity, 0.024 g (g cells)-1 h-1 compared with 0.01 g (g cells)-1 h-1 for the xylose reductase

  10. Comparing the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways in arabinose and xylose fermenting Saccharomyces cerevisiae strains

    Directory of Open Access Journals (Sweden)

    Hahn-Hägerdal Bärbel

    2008-10-01

    Full Text Available Abstract Background Ethanolic fermentation of lignocellulosic biomass is a sustainable option for the production of bioethanol. This process would greatly benefit from recombinant Saccharomyces cerevisiae strains also able to ferment, besides the hexose sugar fraction, the pentose sugars, arabinose and xylose. Different pathways can be introduced in S. cerevisiae to provide arabinose and xylose utilisation. In this study, the bacterial arabinose isomerase pathway was combined with two different xylose utilisation pathways: the xylose reductase/xylitol dehydrogenase and xylose isomerase pathways, respectively, in genetically identical strains. The strains were compared with respect to aerobic growth in arabinose and xylose batch culture and in anaerobic batch fermentation of a mixture of glucose, arabinose and xylose. Results The specific aerobic arabinose growth rate was identical, 0.03 h-1, for the xylose reductase/xylitol dehydrogenase and xylose isomerase strain. The xylose reductase/xylitol dehydrogenase strain displayed higher aerobic growth rate on xylose, 0.14 h-1, and higher specific xylose consumption rate in anaerobic batch fermentation, 0.09 g (g cells-1 h-1 than the xylose isomerase strain, which only reached 0.03 h-1 and 0.02 g (g cells-1h-1, respectively. Whereas the xylose reductase/xylitol dehydrogenase strain produced higher ethanol yield on total sugars, 0.23 g g-1 compared with 0.18 g g-1 for the xylose isomerase strain, the xylose isomerase strain achieved higher ethanol yield on consumed sugars, 0.41 g g-1 compared with 0.32 g g-1 for the xylose reductase/xylitol dehydrogenase strain. Anaerobic fermentation of a mixture of glucose, arabinose and xylose resulted in higher final ethanol concentration, 14.7 g l-1 for the xylose reductase/xylitol dehydrogenase strain compared with 11.8 g l-1 for the xylose isomerase strain, and in higher specific ethanol productivity, 0.024 g (g cells-1 h-1 compared with 0.01 g (g cells-1 h-1

  11. Production of 3-hydroxypropionic acid from glucose and xylose by metabolically engineered Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Kanchana R. Kildegaard

    2015-12-01

    Full Text Available Biomass, the most abundant carbon source on the planet, may in the future become the primary feedstock for production of fuels and chemicals, replacing fossil feedstocks. This will, however, require development of cell factories that can convert both C6 and C5 sugars present in lignocellulosic biomass into the products of interest. We engineered Saccharomyces cerevisiae for production of 3-hydroxypropionic acid (3HP, a potential building block for acrylates, from glucose and xylose. We introduced the 3HP biosynthetic pathways via malonyl-CoA or β-alanine intermediates into a xylose-consuming yeast. Using controlled fed-batch cultivation, we obtained 7.37±0.17 g 3HP L−1 in 120 hours with an overall yield of 29±1% Cmol 3HP Cmol−1 xylose. This study is the first demonstration of the potential of using S. cerevisiae for production of 3HP from the biomass sugar xylose.

  12. Engineering increased triacylglycerol accumulation in Saccharomyces cerevisiae using a modified type 1 plant diacylglycerol acyltransferase.

    Science.gov (United States)

    Greer, Michael S; Truksa, Martin; Deng, Wei; Lung, Shiu-Cheung; Chen, Guanqun; Weselake, Randall J

    2015-03-01

    Diacylglycerol acyltransferase (DGAT) catalyzes the acyl-CoA-dependent acylation of sn-1,2-diacylglycerol to produce triacylglycerol (TAG). This enzyme, which is critical to numerous facets of oilseed development, has been highlighted as a genetic engineering target to increase storage lipid production in microorganisms designed for biofuel applications. Here, four transcriptionally active DGAT1 genes were identified and characterized from the oil crop Brassica napus. Overexpression of each BnaDGAT1 in Saccharomyces cerevisiae increased TAG biosynthesis. Further studies showed that adding an N-terminal tag could mask the deleterious influence of the DGATs' native N-terminal sequences, resulting in increased in vivo accumulation of the polypeptides and an increase of up to about 150-fold in in vitro enzyme activity. The levels of TAG and total lipid fatty acids in S. cerevisiae producing the N-terminally tagged BnaDGAT1.b at 72 h were 53 and 28 % higher than those in cultures producing untagged BnaA.DGAT1.b, respectively. These modified DGATs catalyzed the synthesis of up to 453 mg fatty acid/L by this time point. The results will be of benefit in the biochemical analysis of recombinant DGAT1 produced through heterologous expression in yeast and offer a new approach to increase storage lipid content in yeast for industrial applications. PMID:25520169

  13. Enhancing fatty acid ethyl ester production in Saccharomyces cerevisiae through metabolic engineering and medium optimization.

    Science.gov (United States)

    Thompson, R Adam; Trinh, Cong T

    2014-11-01

    Biodiesels in the form of fatty acyl ethyl esters (FAEEs) are a promising next generation biofuel due to their chemical properties and compatibility with existing infrastructure. It has recently been shown that expression of a bacterial acyl-transferase in the established industrial workhorse Saccharomyces cerevisiae can lead to production of FAEEs by condensation of fatty acyl-CoAs and ethanol. In contrast to recent strategies to produce FAEEs in S. cerevisiae through manipulation of de novo fatty acid biosynthesis or a series of arduous genetic manipulations, we introduced a novel genetic background, which is comparable in titer to previous reports with a fraction of the genetic disruption by aiming at increasing the fatty acyl-CoA pools. In addition, we combined metabolic engineering with modification of culture conditions to produce a maximum titer of over 25 mg/L FAEEs, a 40% improvement over previous reports and a 17-fold improvement over our initial characterizations. Biotechnol. Bioeng. 2014;111: 2200-2208. © 2014 Wiley Periodicals, Inc. PMID:24895195

  14. Production of 3-hydroxypropionic acid from glucose and xylose by metabolically engineered Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kildegaard, Kanchana Rueksomtawin; Wang, Zheng; Chen, Yun;

    2015-01-01

    biomass into the products of interest. We engineered Saccharomyces cerevisiae for production of 3-hydroxypropionic acid (3HP), a potential building block for acrylates, from glucose and xylose. We introduced the 3HP biosynthetic pathways via malonyl-CoA or β-alanine intermediates into a xylose...

  15. An organic acid-tolerant HAA1-overexpression mutant of an industrial bioethanol strain of Saccharomyces cerevisiae and its application to the production of bioethanol from sugarcane molasses

    OpenAIRE

    Inaba, Takuya; Watanabe, Daisuke; Yoshiyama, Yoko; Tanaka,Koichi; Ogawa, Jun; Takagi, Hiroshi; Shimoi, Hitoshi; Shima, Jun

    2013-01-01

    Bacterial contamination is known as a major cause of the reduction in ethanol yield during bioethanol production by Saccharomyces cerevisiae. Acetate is an effective agent for the prevention of bacterial contamination, but it negatively affects the fermentation ability of S. cerevisiae. We have proposed that the combined use of organic acids including acetate and lactate and yeast strains tolerant to organic acids may be effective for the elimination of principally lactic acid bacterial (LAB)...

  16. Development of flocculent Saccharomyces cerevisiae strain GYK-10 for the selective fermentation of glucose/fructose in sugar mills.

    Science.gov (United States)

    Kato, Taku; Ohara, Satoshi; Fukushima, Yasuhiro; Sugimoto, Akira; Masuda, Takayuki; Yasuhara, Takaomi; Yamagishi, Hiromi

    2016-07-01

    Advances in glucose/fructose-selective ethanol production have successfully enhanced raw sugar extraction from sugarcane juice by converting inhibitory substances (i.e., glucose/fructose) into ethanol, which is removed by subsequent operations in cane sugar mills. However, the commercial implementation of this breakthrough process in existing cane sugar mills requires a yeast strain that (i) can be used in food production processes, (ii) exhibits stable saccharometabolic selectivity, and (iii) can be easily separated from the saccharide solution. In this study, we developed a suitable saccharometabolism-selective and flocculent strain, Saccharomyces cerevisiae GYK-10. We obtained a suitable yeast strain for selective fermentation in cane sugar mills using a yeast mating system. First, we crossed a haploid strain defective in sucrose utilization with a flocculent haploid strain. Next, we performed tetrad dissection of the resultant hybrid diploid strain and selected GYK-10 from various segregants by investigating the sucrose assimilation and flocculation capacity phenotypes. Ten consecutive fermentation tests of the GYK-10 strain using a bench-scale fermentor confirmed its suitability for the implementation of practical selective fermentation in a commercial sugar mill. The strain exhibited complete saccharometabolic selectivity and sustained flocculation, where it maintained a high ethanol yield and conversion rate throughout the test. PMID:26811219

  17. Genome Sequences of Industrially Relevant Saccharomyces cerevisiae Strain M3707, Isolated from a Sample of Distillers Yeast and Four Haploid Derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Steven D.; Klingeman, Dawn M.; Johnson, Courtney M.; Clum, Alicia; Aerts, Andrea; Salamov, Asaf; Sharma, Aditi; Zane, Matthew; Barry, Kerrie; Grigoriev, Igor V.; Davison, Brian H.; Lynd, Lee R.; Gilna, Paul; Hau, Heidi; Hogsett, David A.; Froehlich, Allan C.

    2013-04-19

    Saccharomyces cerevisiae strain M3707 was isolated from a sample of commercial distillers yeast, and its genome sequence together with the genome sequences for the four derived haploid strains M3836, M3837, M3838, and M3839 has been determined. Yeasts have potential for consolidated bioprocessing (CBP) for biofuel production, and access to these genome sequences will facilitate their development.

  18. High Level Ethanol from Sugar Cane Molasses by a New Thermotolerant Saccharomyces cerevisiae Strain in Industrial Scale

    OpenAIRE

    Fadel, M.; Abeer A. Keera; Mouafi, Foukia E.; Tarek Kahil

    2013-01-01

    A new local strain of S. cerevisiae F-514, for ethanol production during hot summer season, using Egyptian sugar cane molasses was applied in Egyptian distillery factory. The inouluum was propagated through 300 L, 3 m3, and 12 m3 fermenters charged with diluted sugar cane molasses containing 4%-5% sugars. The yeast was applied in fermentation vessels 65 m3 working volume to study the varying concentrations of urea, DAP, orthophosphoric acid (OPA), and its combinations as well as magnesium sul...

  19. Expression of the bacterial recA gene impairs genetic recombination and sporulation in a Saccharomyces cerevisiae diploid strain

    Directory of Open Access Journals (Sweden)

    Marcos Antonio de Morais Junior

    2003-01-01

    Full Text Available The Escherichia coli RecA protein (RecAp has been demonstrated to induce mutagenesis in yeast cells, although there is still little information on the role of the RecAp in yeast recombination events. We evaluated spontaneous and induced general recombination in vegetative and meiotic cells of the XS2316 strain of the yeast Saccharomyces cerevisiae bearing the recA gene. We found that RecAp decreased reciprocal recombination, gene conversion and intrachromosomal recombination and promoted an increase in error-prone processes in both vegetative and meiotic cells, while its negative effect on meiotic recombination blocked ascospore formation.

  20. Genetic engineering to enhance the Ehrlich pathway and alter carbon flux for increased isobutanol production from glucose by Saccharomyces cerevisiae.

    Science.gov (United States)

    Kondo, Takashi; Tezuka, Hironori; Ishii, Jun; Matsuda, Fumio; Ogino, Chiaki; Kondo, Akihiko

    2012-05-31

    The production of higher alcohols by engineered bacteria has received significant attention. The budding yeast, Saccharomyces cerevisiae, has considerable potential as a producer of higher alcohols because of its capacity to naturally fabricate fusel alcohols, in addition to its robustness and tolerance to low pH. However, because its natural productivity is not significant, we considered a strategy of genetic engineering to increase production of the branched-chain higher alcohol isobutanol, which is involved in valine biosynthesis. Initially, we overexpressed 2-keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH) in S. cerevisiae to enhance the endogenous activity of the Ehrlich pathway. We then overexpressed Ilv2, which catalyzes the first step in the valine synthetic pathway, and deleted the PDC1 gene encoding a major pyruvate decarboxylase with the intent of altering the abundant ethanol flux via pyruvate. Through these engineering steps, along with modification of culture conditions, the isobutanol titer of S. cerevisiae was elevated 13-fold, from 11 mg/l to 143 mg/l, and the yield was 6.6 mg/g glucose, which is higher than any previously reported value for S. cerevisiae. PMID:22342368

  1. The Improvement of SAM Accumulation by Integrating the Endogenous Methionine Adenosyltransferase Gene SAM2 in Genome of the Industrial Saccharomyces cerevisiae Strain.

    Science.gov (United States)

    Zhao, Weijun; Shi, Feng; Hang, Baojian; Huang, Lei; Cai, Jin; Xu, Zhinan

    2016-03-01

    S-Adenosyl-L-methionine (SAM) plays important roles in trans-methylation, trans-sulfuration, and polyamine synthesis in all living cells, and it is also an effective cure for liver disease, depressive syndromes, and osteoarthritis. The increased demands of SAM in pharmaceuticals industry have aroused lots of attempts to improve its production. In this study, a multiple-copy integrative plasmid pYMIKP-SAM2 was introduced into the chromosome of wild-type Saccharomyces cerevisiae strain ZJU001 to construct the recombined strain R1-ZJU001. Further studies showed that the recombinant yeast exhibited higher enzymatic activity of methionine adenosyltransferase and improved its SAM biosynthesis. With a three-phase fed-batch strategy in 15-liter bench-top fermentor, 8.81 g/L SAM was achieved after 52 h cultivation of R1-ZJU001, about 27.1 % increase over its parent strain ZJU001, whereas the SAM content was also improved from 64.6 mg/g DCW to 91.0 mg/g DCW. Our results shall provide insights into the metabolic engineering of SAM pathway in yeast for improved productivity of SAM and subsequent industrial applications. PMID:26728652

  2. Efficient fermentation of xylose to ethanol at high formic acid concentrations by metabolically engineered Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Hasunuma, Tomohisa; Yoshimura, Kazuya; Matsuda, Fumio [Kobe Univ., Hyogo (Japan). Organization of Advanced Science and Technology; Sung, Kyung-mo; Sanda, Tomoya; Kondo, Akihiko [Kobe Univ., Hyogo (Japan). Dept. of Chemical Science and Engineering

    2011-05-15

    Recombinant yeast strains highly tolerant to formic acid during xylose fermentation were constructed. Microarray analysis of xylose-fermenting Saccharomyces cerevisiae strain overexpressing endogenous xylulokinase in addition to xylose reductase and xylitol dehydrogenase from Pichia stipitis revealed that upregulation of formate dehydrogenase genes (FDH1 and FDH2) was one of the most prominent transcriptional events against excess formic acid. The quantification of formic acid in medium indicated that the innate activity of FDH was too weak to detoxify formic acid. To reinforce the capability for formic acid breakdown, the FDH1 gene was additionally overexpressed in the xylose-metabolizing recombinant yeast. This modification allowed the yeast to rapidly decompose excess formic acid. The yield and final ethanol concentration in the presence of 20 mM formic acid is as essentially same as that of control. The fermentation profile also indicated that the production of xylitol and glycerol, major by-products in xylose fermentation, was not affected by the upregulation of FDH activity. (orig.)

  3. Combined metabolic engineering of precursor and co-factor supply to increase α-santalene production by Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Scalcinati Gionata

    2012-08-01

    Full Text Available Abstract Background Sesquiterpenes are a class of natural products with a diverse range of attractive industrial proprieties. Due to economic difficulties of sesquiterpene production via extraction from plants or chemical synthesis there is interest in developing alternative and cost efficient bioprocesses. The hydrocarbon α-santalene is a precursor of sesquiterpenes with relevant commercial applications. Here, we construct an efficient Saccharomyces cerevisiae cell factory for α-santalene production. Results A multistep metabolic engineering strategy targeted to increase precursor and cofactor supply was employed to manipulate the yeast metabolic network in order to redirect carbon toward the desired product. To do so, genetic modifications were introduced acting to optimize the farnesyl diphosphate branch point, modulate the mevalonate pathway, modify the ammonium assimilation pathway and enhance the activity of a transcriptional activator. The approach employed resulted in an overall α-santalene yield of a 0.0052 Cmmol (Cmmol glucose-1 corresponding to a 4-fold improvement over the reference strain. This strategy, combined with a specifically developed continuous fermentation process, led to a final α-santalene productivity of 0.036 Cmmol (g biomass-1 h-1. Conclusions The results reported in this work illustrate how the combination of a metabolic engineering strategy with fermentation technology optimization can be used to obtain significant amounts of the high-value sesquiterpene α-santalene. This represents a starting point toward the construction of a yeast “sesquiterpene factory” and for the development of an economically viable bio-based process that has the potential to replace the current production methods.

  4. Saccharomyces cerevisiae BY4741 and W303-1A laboratory strains differ in salt tolerance

    Czech Academy of Sciences Publication Activity Database

    Petrezsélyová, Silvia; Zahrádka, Jaromír; Sychrová, Hana

    2010-01-01

    Roč. 114, 2-3 (2010), s. 144-150. ISSN 1878-6146 R&D Projects: GA MŠk(CZ) LC531; GA ČR(CZ) GA204/08/0354 Institutional research plan: CEZ:AV0Z50110509 Keywords : Saccharomyces cerevisiae * salt tolerance * potassium homeostasis Subject RIV: EB - Genetics ; Molecular Biology

  5. Comparative study of Saccharomyces cerevisiae wine strains to identify potential marker genes correlated to desiccation stress tolerance.

    Science.gov (United States)

    Capece, Angela; Votta, Sonia; Guaragnella, Nicoletta; Zambuto, Marianna; Romaniello, Rossana; Romano, Patrizia

    2016-05-01

    The most diffused formulation of starter for winemaking is active dry yeast (ADY). ADYs production process is essentially characterized by air-drying stress, a combination of several stresses, including thermal, hyperosmotic and oxidative and cell capacity to counteract such multiple stresses will determine its survival. The molecular mechanisms underlying cell stress response to desiccation have been mostly studied in laboratory and commercial yeast strains, but a growing interest is currently developing for indigenous yeast strains which represent a valuable and alternative source of genetic and molecular biodiversity to be exploited. In this work, a comparative study of different Saccharomyces cerevisiae indigenous wine strains, previously selected for their technological traits, has been carried out to identify potentially relevant genes involved in desiccation stress tolerance. Cell viability was evaluated along desiccation treatment and gene expression was analyzed by real-time PCR before and during the stress. Our data show that the observed differences in individual strain sensitivity to desiccation stress could be associated to specific gene expression over time. In particular, either the basal or the stress-induced mRNA levels of certain genes, such as HSP12, SSA3, TPS1, TPS2, CTT1 and SOD1, result tightly correlated to the strain survival advantage. This study provides a reliable and sensitive method to predict desiccation stress tolerance of indigenous wine yeast strains which could be preliminary to biotechnological applications. PMID:26882930

  6. Zearalenone and Its Derivatives α-Zearalenol and β-Zearalenol Decontamination by Saccharomyces cerevisiae Strains Isolated from Bovine Forage

    Science.gov (United States)

    Keller, Luiz; Abrunhosa, Luís; Keller, Kelly; Rosa, Carlos Alberto; Cavaglieri, Lilia; Venâncio, Armando

    2015-01-01

    Zearalenone (ZEA) and its derivatives are mycotoxins with estrogenic effects on mammals. The biotransformation for ZEA in animals involves the formation of two major metabolites, α- and β-zearalenol (α-ZOL and β-ZOL), which are subsequently conjugated with glucuronic acid. The capability of Saccharomyces cerevisiae strains isolated from silage to eliminate ZEA and its derivatives α-ZOL and β-ZOL was investigated as, also, the mechanisms involved. Strains were grown on Yeast Extract-Peptone-Dextrose medium supplemented with the mycotoxins and their elimination from medium was quantified over time by HPLC-FL. A significant effect on the concentration of ZEA was observed, as all the tested strains were able to eliminate more than 90% of the mycotoxin from the culture medium in two days. The observed elimination was mainly due to ZEA biotransformation into β-ZOL (53%) and α-ZOL (8%) rather than to its adsorption to yeast cells walls. Further, the biotransformation of α-ZOL was not observed but a small amount of β-ZOL (6%) disappeared from culture medium. ZEA biotransformation by yeasts may not be regarded as a full detoxification process because both main end-products are still estrogenic. Nonetheless, it was observed that the biotransformation favors the formation of β-ZOL which is less estrogenic than ZEA and α-ZOL. This metabolic effect is only possible if active strains are used as feed additives and may play a role in the detoxification performance of products with viable S. cerevisiae cells. PMID:26308051

  7. A strain of Saccharomyces cerevisiae evolved for fermentation of lignocellulosic biomass displays improved growth and fermentative ability in high solids concentrations and in the presence of inhibitory compounds

    Directory of Open Access Journals (Sweden)

    Hawkins Gary M

    2011-11-01

    Full Text Available Abstract Background Softwoods are the dominant source of lignocellulosic biomass in the northern hemisphere, and have been investigated worldwide as a renewable substrate for cellulosic ethanol production. One challenge to using softwoods, which is particularly acute with pine, is that the pretreatment process produces inhibitory compounds detrimental to the growth and metabolic activity of fermenting organisms. To overcome the challenge of bioconversion in the presence of inhibitory compounds, especially at high solids loading, a strain of Saccharomyces cerevisiae was subjected to evolutionary engineering and adaptation for fermentation of pretreated pine wood (Pinus taeda. Results An industrial strain of Saccharomyces, XR122N, was evolved using pretreated pine; the resulting daughter strain, AJP50, produced ethanol much more rapidly than its parent in fermentations of pretreated pine. Adaptation, by preculturing of the industrial yeast XR122N and the evolved strains in 7% dry weight per volume (w/v pretreated pine solids prior to inoculation into higher solids concentrations, improved fermentation performance of all strains compared with direct inoculation into high solids. Growth comparisons between XR122N and AJP50 in model hydrolysate media containing inhibitory compounds found in pretreated biomass showed that AJP50 exited lag phase faster under all conditions tested. This was due, in part, to the ability of AJP50 to rapidly convert furfural and hydroxymethylfurfural to their less toxic alcohol derivatives, and to recover from reactive oxygen species damage more quickly than XR122N. Under industrially relevant conditions of 17.5% w/v pretreated pine solids loading, additional evolutionary engineering was required to decrease the pronounced lag phase. Using a combination of adaptation by inoculation first into a solids loading of 7% w/v for 24 hours, followed by a 10% v/v inoculum (approximately equivalent to 1 g/L dry cell weight into 17

  8. Production of Volatile and Sulfur Compounds by 10 Saccharomyces cerevisiae Strains Inoculated in Trebbiano Must

    OpenAIRE

    Patrignani, Francesca; Chinnici, Fabio; Serrazanetti, Diana I.; Vernocchi, Pamela; Ndagijimana, Maurice; Riponi, Claudio; Lanciotti, Rosalba

    2016-01-01

    In wines, the presence of sulfur compounds is the resulting of several contributions among which yeast metabolism. The characterization of the starter Saccharomyces cerevisiae needs to be performed also taking into account this ability even if evaluated together with the overall metabolic profile. In this perspective, principal aim of this experimental research was the evaluation of the volatile profiles, throughout GC/MS technique coupled with solid phase micro extraction, of wines obtained ...

  9. Determining the fate of fluorescent quantum dots on surface of engineered budding S. cerevisiae cell molecular landscape

    OpenAIRE

    Chouhan, Raghuraj Singh; Qureshi, Anjum; Kolkar Mohammed, Javed Hussain Niazi

    2015-01-01

    In this study, we surface engineered living S. cerevisiae cells by decorating quantum dots (QDs) and traced the fate of QDs on molecular landscape of single mother cell through several generation times (progeny cells). The fate of QDs on cell-surface was tracked through the cellular division events using confocal microscopy and fluorescence emission profiles. The extent of cell-surface QDs distribution among the offspring was determined as the mother cell divides into daughter cells. Fluoresc...

  10. YeastFab: the design and construction of standard biological parts for metabolic engineering in Saccharomyces cerevisiae

    OpenAIRE

    Guo, Yakun; Dong, Junkai; Zhou, Tong; Auxillos, Jamie; Li, Tianyi; Zhang, Weimin; Wang, LiHui; Shen, Yue; Luo, Yisha; Zheng, Yijing; Lin, Jiwei; Chen, Guo-Qiang; Wu, Qingyu; Cai, Yizhi; Dai, Junbiao

    2015-01-01

    It is a routine task in metabolic engineering to introduce multicomponent pathways into a heterologous host for production of metabolites. However, this process sometimes may take weeks to months due to the lack of standardized genetic tools. Here, we present a method for the design and construction of biological parts based on the native genes and regulatory elements in Saccharomyces cerevisiae. We have developed highly efficient protocols (termed YeastFab Assembly) to synthesize these genet...

  11. Kinetic Activity of Commercial Native Linamarase (CNLIN and Engineered <&beta-glucosidase from Saccharomyces cerevisiae on Cassava Linamarin

    Directory of Open Access Journals (Sweden)

    Julius Kwagh-Har Ikya

    2014-02-01

    Full Text Available The ability of Commercial Native Linamarase (CNLIN and Engineered Linamarase (GELIN extracts from Saccharomyces cerevisiae to hydrolyse cassava linamarin was challenged. CNLIN acting as control was used together with GELIN extracts from Saccharomyces cerevisiae to evaluate the kinetic data for test enzymes at pH 3.5, 6.8 and 10.5, respectively and ambient temperature (35°C. Data obtained from the varying activity versus substrate concentrations were fitted with the Michaelis-Menten plots and Lineweaver-Burk model to obtain maximum velocity (Vmax, affinity coefficient (Km, physiological efficiency (Vmax/Km and r2 (linear regressing coefficient. The results indicated that the engineered linamarase conferred different enzyme kinetic data showing degradation of cassava linamarin by CNLIN and GELIN from Saccharomyces cerevisiae at the optimum pH and temperature. The relation was best described by the characteristic sigmoid Michaelis-Menten plots and Lineweaver-Burk model evidence from the high coefficient of linear regression (r2>0.976. Vmax and Km derived from the Lineweaver-Burk model varied from 10.0 to 13.0 µmol/min and 0.5 to 0.9 &muM respectively for engineered enzymes and 0.0-10.0 mol/min and 0.0 to 0.9 &muM respectively for CNLIN. The kinetics profiles of the studied enzymes showed their actions on cassava linamarin were influenced by degree of genetic manipulation, purification and pH at ambient temperature. The wide pH tolerance in the degradation of linamarin suggests a possible use of the engineered linamarase from Saccharomyces cerevisiae in detoxifying linamarin in cassava for the production of cyanide-free cassava-based food products.

  12. Enhancing sesquiterpene production in Saccharomyces cerevisiae through in silico driven metabolic engineering

    DEFF Research Database (Denmark)

    Asadollahi, Mohammadali; Maury, Jerome; Patil, Kiran Raosaheb;

    2009-01-01

    A genome-scale metabolic model was used to identify new target genes for enhanced biosynthesis of sesquiterpenes in the yeast Saccharomyces cerevisiae. The effect of gene deletions on the flux distributions in the metabolic model of S. cerevisiae was assessed using OptGene as the modeling framewo...

  13. Evolutionary engineering of Saccharomyces cerevisiae for enhanced tolerance to hydrolysates of lignocellulosic biomass.

    Science.gov (United States)

    Almario, María P; Reyes, Luis H; Kao, Katy C

    2013-10-01

    Lignocellulosic biomass has become an important feedstock to mitigate current ethical and economical concerns related to the bio-based production of fuels and chemicals. During the pre-treatment and hydrolysis of the lignocellulosic biomass, a complex mixture of sugars and inhibitors are formed. The inhibitors interfere with microbial growth and product yields. This study uses an adaptive laboratory evolution method called visualizing evolution in real-time (VERT) to uncover the molecular mechanisms associated with tolerance to hydrolysates of lignocellulosic biomass in Saccharomyces cerevisiae. VERT enables a more rational scheme for isolating adaptive mutants for characterization and molecular analyses. Subsequent growth kinetic analyses of the mutants in individual and combinations of common inhibitors present in hydrolysates (acetic acid, furfural, and hydroxymethylfurfural) showed differential levels of resistance to different inhibitors, with enhanced growth rates up to 57%, 12%, 22%, and 24% in hydrolysates, acetic acid, HMF and furfural, respectively. Interestingly, some of the adaptive mutants exhibited reduced fitness in the presence of individual inhibitors, but showed enhanced fitness in the presence of combinations of inhibitors compared to the parental strains. Transcriptomic analysis revealed different mechanisms for resistance to hydrolysates and a potential cross adaptation between oxidative stress and hydrolysates tolerance in several of the mutants. PMID:23613173

  14. Ribozyme-based aminoglycoside switches of gene expression engineered by genetic selection in S. cerevisiae.

    Science.gov (United States)

    Klauser, Benedikt; Atanasov, Janina; Siewert, Lena K; Hartig, Jörg S

    2015-05-15

    Systems for conditional gene expression are powerful tools in basic research as well as in biotechnology. For future applications, it is of great importance to engineer orthogonal genetic switches that function reliably in diverse contexts. RNA-based switches have the advantage that effector molecules interact immediately with regulatory modules inserted into the target RNAs, getting rid of the need of transcription factors usually mediating genetic control. Artificial riboswitches are characterized by their simplicity and small size accompanied by a high degree of modularity. We have recently reported a series of hammerhead ribozyme-based artificial riboswitches that allow for post-transcriptional regulation of gene expression via switching mRNA, tRNA, or rRNA functions. A more widespread application was so far hampered by moderate switching performances and a limited set of effector molecules available. Here, we report the re-engineering of hammerhead ribozymes in order to respond efficiently to aminoglycoside antibiotics. We first established an in vivo selection protocol in Saccharomyces cerevisiae that enabled us to search large sequence spaces for optimized switches. We then envisioned and characterized a novel strategy of attaching the aptamer to the ribozyme catalytic core, increasing the design options for rendering the ribozyme ligand-dependent. These innovations enabled the development of neomycin-dependent RNA modules that switch gene expression up to 25-fold. The presented aminoglycoside-responsive riboswitches belong to the best-performing RNA-based genetic regulators reported so far. The developed in vivo selection protocol should allow for sampling of large sequence spaces for engineering of further optimized riboswitches. PMID:24871672

  15. 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.

  16. 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. PMID:25907574

  17. Optimization of CDT-1 and XYL1 expression for balanced co-production of ethanol and xylitol from cellobiose and xylose by engineered Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Jian Zha

    Full Text Available Production of ethanol and xylitol from lignocellulosic hydrolysates is an alternative to the traditional production of ethanol in utilizing biomass. However, the conversion efficiency of xylose to xylitol is restricted by glucose repression, causing a low xylitol titer. To this end, we cloned genes CDT-1 (encoding a cellodextrin transporter and gh1-1 (encoding an intracellular β-glucosidase from Neurospora crassa and XYL1 (encoding a xylose reductase that converts xylose into xylitol from Scheffersomyces stipitis into Saccharomyces cerevisiae, enabling simultaneous production of ethanol and xylitol from a mixture of cellobiose and xylose (main components of lignocellulosic hydrolysates. We further optimized the expression levels of CDT-1 and XYL1 by manipulating their promoters and copy-numbers, and constructed an engineered S. cerevisiae strain (carrying one copy of PGK1p-CDT1 and two copies of TDH3p-XYL1, which showed an 85.7% increase in xylitol production from the mixture of cellobiose and xylose than that from the mixture of glucose and xylose. Thus, we achieved a balanced co-fermentation of cellobiose (0.165 g/L/h and xylose (0.162 g/L/h at similar rates to co-produce ethanol (0.36 g/g and xylitol (1.00 g/g.

  18. Optimization of CDT-1 and XYL1 Expression for Balanced Co-Production of Ethanol and Xylitol from Cellobiose and Xylose by Engineered Saccharomyces cerevisiae

    Science.gov (United States)

    Zha, Jian; Li, Bing-Zhi; Shen, Ming-Hua; Hu, Meng-Long; Song, Hao; Yuan, Ying-Jin

    2013-01-01

    Production of ethanol and xylitol from lignocellulosic hydrolysates is an alternative to the traditional production of ethanol in utilizing biomass. However, the conversion efficiency of xylose to xylitol is restricted by glucose repression, causing a low xylitol titer. To this end, we cloned genes CDT-1 (encoding a cellodextrin transporter) and gh1-1 (encoding an intracellular β-glucosidase) from Neurospora crassa and XYL1 (encoding a xylose reductase that converts xylose into xylitol) from Scheffersomyces stipitis into Saccharomyces cerevisiae, enabling simultaneous production of ethanol and xylitol from a mixture of cellobiose and xylose (main components of lignocellulosic hydrolysates). We further optimized the expression levels of CDT-1 and XYL1 by manipulating their promoters and copy-numbers, and constructed an engineered S. cerevisiae strain (carrying one copy of PGK1p-CDT1 and two copies of TDH3p-XYL1), which showed an 85.7% increase in xylitol production from the mixture of cellobiose and xylose than that from the mixture of glucose and xylose. Thus, we achieved a balanced co-fermentation of cellobiose (0.165 g/L/h) and xylose (0.162 g/L/h) at similar rates to co-produce ethanol (0.36 g/g) and xylitol (1.00 g/g). PMID:23844185

  19. Transcriptomes of a xylose-utilizing industrial flocculating Saccharomyces cerevisiae strain cultured in media containing different sugar sources.

    Science.gov (United States)

    Zeng, Wei-Yi; Tang, Yue-Qin; Gou, Min; Xia, Zi-Yuan; Kida, Kenji

    2016-12-01

    Lignocellulosic hydrolysates used for bioethanol production contain a mixture of sugars, with xylose being the second most abundant after glucose. Since xylose is not a natural substrate for Saccharomyces cerevisiae, recombinant S. cerevisiae strongly prefers glucose over xylose, and the fermentation rate and ethanol yield with xylose are both lower than those with glucose. To determine the molecular basis for glucose and xylose fermentation, we used microarrays to investigate the transcriptional difference of a xylose-utilizing industrial strain cultured in both single sugar media and a mixed sugar medium of glucose and xylose. The transcriptomes were nearly identical between glucose metabolizing cells in the glucose alone medium and those in the glucose fermentation phase in the mixed-sugar medium. Whereas the transcriptomes highly differed between the xylose metabolizing cells in the xylose alone medium and those in the xylose fermentation phase in the mixed sugar medium, and the differences mainly involved sulfur metabolism. When the transcriptional profiles were compared between glucose fermentation state and xylose fermentation state, we found the expression patterns of hexose transporters and glucose signaling pathway differed in response to different sugar sources, and the expression levels of the genes involved in gluconeogenesis, the glyoxylate and tricarboxylic acid cycles and respiration increased with xylose, indicating that the xylose-metabolizing cells had high requirements for maintenance energy and lacked the carbon catabolite repression capability. The effect of carbon catabolite repression by glucose lasted after glucose depletion for specific genes to different extents. PMID:27485516

  20. An organic acid-tolerant HAA1-overexpression mutant of an industrial bioethanol strain of Saccharomyces cerevisiae and its application to the production of bioethanol from sugarcane molasses.

    Science.gov (United States)

    Inaba, Takuya; Watanabe, Daisuke; Yoshiyama, Yoko; Tanaka, Koichi; Ogawa, Jun; Takagi, Hiroshi; Shimoi, Hitoshi; Shima, Jun

    2013-01-01

    Bacterial contamination is known as a major cause of the reduction in ethanol yield during bioethanol production by Saccharomyces cerevisiae. Acetate is an effective agent for the prevention of bacterial contamination, but it negatively affects the fermentation ability of S. cerevisiae. We have proposed that the combined use of organic acids including acetate and lactate and yeast strains tolerant to organic acids may be effective for the elimination of principally lactic acid bacterial (LAB) contamination. In a previous study employing laboratory S. cerevisiae strains, we showed that overexpression of the HAA1 gene, which encodes a transcriptional activator, could be a useful molecular breeding method for acetate-tolerant yeast strains. In the present study, we constructed a HAA1-overexpressing diploid strain (MATa/α, named ER HAA1-OP) derived from the industrial bioethanol strain Ethanol Red (ER). ER HAA1-OP showed tolerance not only to acetate but also to lactate, and this tolerance was dependent on the increased expression of HAA1 gene. The ethanol production ability of ER HAA1-OP was almost equivalent to that of the parent strain during the bioethanol production process from sugarcane molasses in the absence of acetate. The addition of acetate at 0.5% (w/v, pH 4.5) inhibited the fermentation ability of the parent strain, but such an inhibition was not observed in the ethanol production process using ER HAA1-OP. PMID:24373204

  1. Effect of 905 MHz microwave radiation on colony growth of the yeast Saccharomyces cerevisiae strains FF18733, FF1481 and D7

    International Nuclear Information System (INIS)

    The aim of this study was to evaluate the effect of weak radiofrequency microwave (RF/MW) radiation emitted by mobile phones on colony growth of the yeast Saccharomyces cerevisiae. S. cerevisiae strains FF18733 (wild-type), FF1481 (rad1 mutant) and D7 (commonly used to detect reciprocal and nonreciprocal mitotic recombinations) were exposed to a 905 MHz electromagnetic field that closely matched the Global System for Mobile Communication (GSM) pulse modulation signals for mobile phones at a specific absorption rate (SAR) of 0.12 W/kg. Following 15-, 30- and 60-minutes exposure to RF/MW radiation, strain FF18733 did not show statistically significant changes in colony growth compared to the control sample. The irradiated strains FF1481 and D7 demonstrated statistically significant reduction of colony growth compared to non-irradiated strains after all exposure times. Furthermore, strain FF1481 was more sensitive to RF/MW radiation than strain D7. The findings indicate that pulsed RF/MW radiation at a low SAR level can affect the rate of colony growth of different S. cerevisiae strains

  2. Magneto-strain-driven quantum engine on a graphene flake

    OpenAIRE

    Peña, Francisco J.; Muñoz, Enrique

    2015-01-01

    We propose a novel conceptual design for a graphene-based quantum engine, driven by a superposition of mechanical strain and an external magnetic field. Engineering of strain in a nanoscale graphene flake creates a gauge field with an associated uniform pseudo-magnetic field. The strain-induced pseudo-magnetic field can be combined with a real magnetic field, leading to the emergence of discrete relativistic Landau levels within the single-particle picture. The inter-level distance and hence ...

  3. Production of Rebaudioside A from Stevioside Catalyzed by the Engineered Saccharomyces cerevisiae.

    Science.gov (United States)

    Li, Yan; Li, Yangyang; Wang, Yu; Chen, Liangliang; Yan, Ming; Chen, Kequan; Xu, Lin; Ouyang, Pingkai

    2016-04-01

    Rebaudioside A has superior taste quality among the steviol glycosides extracted from Stevia rebaudiana leaves. Given its high purity as a general-purpose sweetener, rebaudioside A has received significant attention and has been widely applied in food and beverages in recent decades. Stevioside is one of the major steviol glycosides and can be converted to rebaudioside A by the uridine-diphosphate dependent glucosyltransferase UGT76G1 in S. rebaudiana. To explore the applicability of and limits in producing rebaudioside A from stevioside through whole-cell biocatalysis, the engineered Saccharomyces cerevisiae expressing UGT76G1, using a newly constructed constitutive expression vector, was used as the whole-cell biocatalyst. Citrate was added to the reaction mixture to allow metabolic regulation when glucose was fed to provide the activated sugar donor UDP-glucose for glycosylation of stevioside in vivo. In an evaluation of the whole-cell reaction parameters involving cell permeability, temperature, pH, citrate and Mg(2+) concentrations, and glucose feeding, production of 1160.5 mg/L rebaudioside A from 2 g/L stevioside was achieved after 48 h without supplementation of extracellular UDP-glucose. PMID:26733458

  4. Mitotic chromosome loss in a radiation-sensitive strain of the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Cells of Saccharomyces cerevisiae with mutations in the RAD52 gene have previously been shown to be defective in meiotic and mitotic recombination, in sporulation, and in repair of radiation-induced damage to DNA. In this study we show that diploid cells homozygous for rad52 lose chromosomes at high frequencies and that these frequencies of loss can be increased dramatically by exposure of these cells to x-rays. Genetic analyses of survivors of x-ray treatment demonstrate that chromosome loss events result in the conversion of diploid cells to cells with near haploid chromosome numbers

  5. High Level Ethanol from Sugar Cane Molasses by a New Thermotolerant Saccharomyces cerevisiae Strain in Industrial Scale.

    Science.gov (United States)

    Fadel, M; Keera, Abeer A; Mouafi, Foukia E; Kahil, Tarek

    2013-01-01

    A new local strain of S. cerevisiae F-514, for ethanol production during hot summer season, using Egyptian sugar cane molasses was applied in Egyptian distillery factory. The inouluum was propagated through 300 L, 3 m(3), and 12 m(3) fermenters charged with diluted sugar cane molasses containing 4%-5% sugars. The yeast was applied in fermentation vessels 65 m(3) working volume to study the varying concentrations of urea, DAP, orthophosphoric acid (OPA), and its combinations as well as magnesium sulfate and inoculum size. The fermenter was allowed to stay for a period of 20 hours to give time for maximum conversion of sugars into ethanol. S. cerevisiae F-514 at molasses sugar level of 18% (w/v), inoculum size of 20% (v/v) cell concentration of 3.0 × 10(8)/mL, and combinations of urea, diammonium phosphate (DAP), orthophosphoric acid (OPA), and magnesium sulfate at amounts of 20, 10, 5, and 10 kg/65 m(3) working volume fermenters, respectively, supported maximum ethanol production (9.8%, v/v), fermentation efficiency (FE) 88.1%, and remaining sugars (RS) 1.22%. The fermentation resulted 13.4 g dry yeast/L contained 34.6% crude protein and 8.2% ash. By selecting higher ethanol yielding yeast strain and optimizing, the fermentation parameters both yield and economics of the fermentation process can be improved. PMID:24363937

  6. Strain Engineering of Transition Metal Dichalcogenides

    Science.gov (United States)

    Dadgar, Ali; Pasupathy, Abhay; Herman, Irving; Wang, Dennis; Kang, Kyungnam; Yang, Eui-Hyeok

    The application of strain to materials can cause changes to bandwidth, effective masses, degeneracies and even structural phases. In the case of the transition metal dichalcogenide (TMD) semiconductors, small strain (around 1 percent) is expected to change band gaps and mobilities, while larger strains are expected to cause phase changes from the triangular 2H phase to orthorhombic 1T' phases. We will describe experimental techniques to apply small and large (around 10 percent) strains to one or few layer samples of the TMD semiconductors, and describe the effect of the strain using optical (Raman, photoluminescence) and cryogenic transport techniques.

  7. EDITORIAL: Excelling under strain: band engineering in nanomaterials Excelling under strain: band engineering in nanomaterials

    Science.gov (United States)

    Demming, Anna

    2013-08-01

    A little stress or strain has been known to improve the performance of athletes, actors and of course nanomaterials alike. In fact strain in silicon is now a major engineering tool for improving the performance of devices, and is ubiquitously used in device design and fabrication. Strain engineering alters a material's band structure, a model of electron behaviour that describes how as atoms come together in a solid, their discrete electron orbitals overlap to ultimately give rise to bands of allowed energy levels. In a strained crystal lattice of silicon or silicon germanium the distance between atoms in the lattice is greater than usual and the bands of allowed energy levels change. This July marks 100 years since Bohr submitted his paper 'On the constitution of atoms and molecules' [1] where he describes the structure of the atom in terms of discrete allowed energy levels. The paper was a seminal contribution to the development of quantum mechanics and laid the initial theoretical precepts for band gap engineering in devices. In this issue Nrauda and a collaboration of researchers in Europe and Australia study the growth of defect-free SiGe islands on pre-patterned silicon [2]. They analyse the strain in the islands and determine at what point lattice dislocations set in with a view to informing implementation of strain engineering in devices. The effects of strain on band structure in silicon and germanium were already studied and reported in the 1950s [3, 4]. Since then the increasing focus on nanoscale materials and the hunger for control of electronic properties has prompted further study of strain effects. The increased surface area to volume ratio in nanostructures changes the strain behaviour with respect to bulk materials, and this can also be exploited for handling and fine tuning strain to manipulate material properties. It is perhaps no surprise that graphene, one of the most high-profile materials in current nanotechnology research, has attracted

  8. Novel starters for old processes: use of Saccharomyces cerevisiae strains isolated from artisanal sourdough for craft beer production at a brewery scale.

    Science.gov (United States)

    Marongiu, Antonella; Zara, Giacomo; Legras, Jean-Luc; Del Caro, Alessandra; Mascia, Ilaria; Fadda, Costantino; Budroni, Marilena

    2015-01-01

    The deliberate inoculation of yeast strains isolated from food matrices such as wine or bread, could allow the transfer of novel properties to beer. In this work, the feasibility of the use of baker's yeast strains as starters for craft beer production has been evaluated at laboratory and brewery scale. Nine out of 12 Saccharomyces cerevisiae strains isolated from artisanal sourdoughs metabolized 2 % maltose, glucose and trehalose and showed growth rates and cell populations higher than those of the brewer's strain Safbrew-S33. Analysis of allelic variation at 12 microsatellite loci clustered seven baker's strains and Safbrew-S33 in the main group of bread isolates. Chemical analyses of beers produced at a brewery scale showed significant differences among the beers produced with the baker's strain S38 or Safbrew-S33, while no significant differences were observed when S38 or the brewer's strain Safbrew-F2 was used for re-fermentation. The sensory profile of beers obtained with S38 or the brewer's yeasts did not show significant differences, thus suggesting that baker's strains of S. cerevisiae could represent a reservoir of biodiversity for the selection of starter strains for craft beer production. PMID:25387611

  9. Benchmarking two commonly used Saccharomyces cerevisiae strains for heterologous vanillin-β-glucoside production

    DEFF Research Database (Denmark)

    Strucko, Tomas; Magdenoska, Olivera; Mortensen, Uffe Hasbro

    2015-01-01

    factories for production of specific compounds. To examine this possibility, we have reconstructed a de novo vanillin-β-glucoside pathway in an identical manner in S288c and CEN.PK strains. Characterization of the two resulting strains in two standard conditions revealed that the S288c background strain...... produced up to 10-fold higher amounts of vanillin-β-glucoside compared to CEN.PK. This study demonstrates that yeast strain background may play a major role in the outcome of newly developed cell factories for production of a given product....

  10. Whole Genome Analysis of 132 Clinical Saccharomyces cerevisiae Strains Reveals Extensive Ploidy Variation

    Science.gov (United States)

    Zhu, Yuan O.; Sherlock, Gavin; Petrov, Dmitri A.

    2016-01-01

    Budding yeast has undergone several independent transitions from commercial to clinical lifestyles. The frequency of such transitions suggests that clinical yeast strains are derived from environmentally available yeast populations, including commercial sources. However, despite their important role in adaptive evolution, the prevalence of polyploidy and aneuploidy has not been extensively analyzed in clinical strains. In this study, we have looked for patterns governing the transition to clinical invasion in the largest screen of clinical yeast isolates to date. In particular, we have focused on the hypothesis that ploidy changes have influenced adaptive processes. We sequenced 144 yeast strains, 132 of which are clinical isolates. We found pervasive large-scale genomic variation in both overall ploidy (34% of strains identified as 3n/4n) and individual chromosomal copy numbers (36% of strains identified as aneuploid). We also found evidence for the highly dynamic nature of yeast genomes, with 35 strains showing partial chromosomal copy number changes and eight strains showing multiple independent chromosomal events. Intriguingly, a lineage identified to be baker’s/commercial derived with a unique damaging mutation in NDC80 was particularly prone to polyploidy, with 83% of its members being triploid or tetraploid. Polyploidy was in turn associated with a >2× increase in aneuploidy rates as compared to other lineages. This dataset provides a rich source of information on the genomics of clinical yeast strains and highlights the potential importance of large-scale genomic copy variation in yeast adaptation. PMID:27317778

  11. Using mixed inocula of Saccharomyces cerevisiae killer strains to improve the quality of traditional sparkling-wine.

    Science.gov (United States)

    Velázquez, Rocío; Zamora, Emiliano; Álvarez, Manuel; Álvarez, María L; Ramírez, Manuel

    2016-10-01

    The quality of traditional sparkling-wine depends on the aging process in the presence of dead yeast cells. These cells undergo a slow autolysis process thereby releasing some compounds, mostly colloidal polymers such as polysaccharides and mannoproteins, which influence the wine's foam properties and mouthfeel. Saccharomyces cerevisiae killer yeasts were tested to increase cell death and autolysis during mixed-yeast-inoculated second fermentation and aging. These yeasts killed sensitive strains in killer plate assays done under conditions of low pH and temperature similar to those used in sparkling-wine making, although some strains showed a different killer behaviour during the second fermentation. The fast killer effect improved the foam quality and mouthfeel of the mixed-inoculated wines, while the slow killer effect gave small improvements over single-inoculated wines. The effect was faster under high-pressure than under low-pressure conditions. Wine quality improvement did not correlate with the polysaccharide, protein, mannan, or aromatic compound concentrations, suggesting that the mouthfeel and foaming quality of sparkling wine are very complex properties influenced by other wine compounds and their interactions, as well as probably by the specific chemical composition of a given wine. PMID:27375256

  12. Comparative transcriptomic analysis reveals similarities and dissimilarities in Saccharomyces cerevisiae wine strains response to nitrogen availability.

    Directory of Open Access Journals (Sweden)

    Catarina Barbosa

    Full Text Available Nitrogen levels in grape-juices are of major importance in winemaking ensuring adequate yeast growth and fermentation performance. Here we used a comparative transcriptome analysis to uncover wine yeasts responses to nitrogen availability during fermentation. Gene expression was assessed in three genetically and phenotypically divergent commercial wine strains (CEG, VL1 and QA23, under low (67 mg/L and high nitrogen (670 mg/L regimes, at three time points during fermentation (12 h, 24 h and 96 h. Two-way ANOVA analysis of each fermentation condition led to the identification of genes whose expression was dependent on strain, fermentation stage and on the interaction of both factors. The high fermenter yeast strain QA23 was more clearly distinct from the other two strains, by differential expression of genes involved in flocculation, mitochondrial functions, energy generation and protein folding and stabilization. For all strains, higher transcriptional variability due to fermentation stage was seen in the high nitrogen fermentations. A positive correlation between maximum fermentation rate and the expression of genes involved in stress response was observed. The finding of common genes correlated with both fermentation activity and nitrogen up-take underlies the role of nitrogen on yeast fermentative fitness. The comparative analysis of genes differentially expressed between both fermentation conditions at 12 h, where the main difference was the level of nitrogen available, showed the highest variability amongst strains revealing strain-specific responses. Nevertheless, we were able to identify a small set of genes whose expression profiles can quantitatively assess the common response of the yeast strains to varying nitrogen conditions. The use of three contrasting yeast strains in gene expression analysis prompts the identification of more reliable, accurate and reproducible biomarkers that will facilitate the diagnosis of deficiency of this

  13. Study on the development of high yielding alcohol resistant strain of Saccharomyces cerevisiae and the Influence of Magnetic field on Saccharomyces cerevisiae Inoculum for the production of Alcohol and Vinegar from apple juice.

    Directory of Open Access Journals (Sweden)

    Rabiul Haque

    2014-12-01

    Full Text Available Natural vinegar is one of the fermented products which has some potentiality with respect to a nutraceutical standpoint. The present study is an optimization of the fermentation conditions for apple juice vinegar production from aple juice wine, this biochemical process being aided by Acetobacter aceti.We have studied on the development of high yielding alcohol resistant strain of Saccharomyces cerevisiae for the production of alcohol. Overflow metabolism is an undesirable characteristic of aerobic cultures of Saccharomyces cerevisiae during biomassdirected processes. It results from elevated apple juice containing sugar consumption rates that cause a high substrate conversion to ethanol and other bi-products, severely affecting cell physiology, bioprocess performance, and biomass yields. Fed-batch culture, where sucrose consumption rates are controlled by the external addition of sugar aiming at its low concentrations in the fermentor, is the classical bioprocessing alternative to prevent sugar fermentation by yeasts. However, fed-batch fermentations present drawbacks that could be overcome by simpler batch cultures at relatively high initial sugar concentrations From the results it is clear that strain T2. which has been exposed to 15% alcohol for 18 hrs is the high yielding strain, as it gives 16% alcohol after distillation. We also find that as the exposure is increased, that is, with increasing exposure to 20% alcohol for 5 hrs, 18 hrs, and 20 hrs, the production of alcohol decreases. Saccharomyces cerevisiae yeast cells strain T2. which has been exposed to 15% alcohol for 18 hrs were exposed to a homogenous static magnetic field of 125 mT for periods of 24, 48 or 72 hours and then used as inoculum for the alcoholic fermentation. The exposure to the magnetic field improved the fermentation process kinetics. Biomass and ethanol yields of fermentations inoculated with treated inoculum were higher than those in the control fermentation, which

  14. Cellular and molecular engineering of yeast Saccharomyces cerevisiae for advanced biobutanol production.

    Science.gov (United States)

    Kuroda, Kouichi; Ueda, Mitsuyoshi

    2016-02-01

    Butanol is an attractive alternative energy fuel owing to several advantages over ethanol. Among the microbial hosts for biobutanol production, yeast Saccharomyces cerevisiae has a great potential as a microbial host due to its powerful genetic tools, a history of successful industrial use, and its inherent tolerance to higher alcohols. Butanol production by S. cerevisiae was first attempted by transferring the 1-butanol-producing metabolic pathway from native microorganisms or using the endogenous Ehrlich pathway for isobutanol synthesis. Utilizing alternative enzymes with higher activity, eliminating competitive pathways, and maintaining cofactor balance achieved significant improvements in butanol production. Meeting future challenges, such as enhancing butanol tolerance and implementing a comprehensive strategy by high-throughput screening, would further elevate the biobutanol-producing ability of S. cerevisiae toward an ideal microbial cell factory exhibiting high productivity of biobutanol. PMID:26712533

  15. Growth temperature exerts differential physiological and transcriptional responses in laboratory and wine strains of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Pizarra, Francisco J.; Jewett, Michael Christopher; Nielsen, Jens;

    2008-01-01

    -dependent genes. The transcriptional response was used to identify highly correlated gene expression subnetworks within yeast metabolism. We showed that temperature differences most strongly affect nitrogen metabolism and the heat shock response. A lack of stress response element-mediated gene induction, coupled...... environmental conditions and the organoleptic properties that they confer to wine. Here, we used a two-factor design to study the responses of a standard laboratory strain, CEN.PK113-7D, and an industrial wine yeast strain, EC1118, to growth temperatures of 15 degrees C and 30 degrees C in nitrogen...... exhibited higher fermentation rates. To elucidate mechanistic differences controlling the growth temperature response and underlying adaptive mechanisms between the strains, DNA microarrays and targeted metabolome analysis were used. We identified 1,007 temperature-dependent genes and 473 strain...

  16. Synergistic effects of TAL1 over-expression and PHO13 deletion on the weak acid inhibition of xylose fermentation by industrial Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Li, Yun-Cheng; Gou, Zi-Xi; Liu, Ze-Shen; Tang, Yue-Qin; Akamatsu, Takashi; Kida, Kenji

    2014-10-01

    In the industrial production of bioethanol from lignocellulosic biomass, a strain of Saccharomyces cerevisiae that can ferment xylose in the presence of inhibitors is of utmost importance. The recombinant, industrial-flocculating S. cerevisiae strain NAPX37, which can ferment xylose, was used as the parent to delete the gene encoding p-nitrophenylphosphatase (PHO13) and overexpress the gene encoding transaldolase (TAL1) to evaluate the synergistic effects of these two genes on xylose fermentation in the presence of weak acid inhibitors, including formic, acetic, or levulinic acids. TAL1 over-expression or PHO13 deletion improved xylose fermentation as well as the tolerance of NAPX37 to all three weak acids. The simultaneous deletion of PHO13 and the over-expression of TAL1 had synergistic effects and improved ethanol production and reduction of xylitol accumulation in the absence and presence of weak acid inhibitors. PMID:24966040

  17. Nanoscale strain engineering of graphene and graphene-based devices

    Institute of Scientific and Technical Information of China (English)

    N-C Yeh; C-C Hsu; M L Teague; J-Q Wang; D A Boyd; C-C Chen

    2016-01-01

    Structural distortions in nano-materials can induce dramatic changes in their electronic properties. This situation is well manifested in graphene, a two-dimensional honeycomb structure of carbon atoms with only one atomic layer thickness. In particular, strained graphene can result in both charging effects and pseudo-magnetic fields, so that controlled strain on a perfect graphene lattice can be tailored to yield desirable electronic properties. Here, we describe the theoretical foundation for strain-engineering of the electronic properties of graphene, and then provide experimental evidence for strain-induced pseudo-magnetic fields and charging effects in monolayer graphene. We further demonstrate the feasibility of nano-scale strain engineering for graphene-based devices by means of theoretical simula-tions and nano-fabrication technology.

  18. Nanoscale strain engineering of graphene and graphene-based devices

    Science.gov (United States)

    Yeh, N.-C.; Hsu, C.-C.; Teague, M. L.; Wang, J.-Q.; Boyd, D. A.; Chen, C.-C.

    2016-06-01

    Structural distortions in nano-materials can induce dramatic changes in their electronic properties. This situation is well manifested in graphene, a two-dimensional honeycomb structure of carbon atoms with only one atomic layer thickness. In particular, strained graphene can result in both charging effects and pseudo-magnetic fields, so that controlled strain on a perfect graphene lattice can be tailored to yield desirable electronic properties. Here, we describe the theoretical foundation for strain-engineering of the electronic properties of graphene, and then provide experimental evidence for strain-induced pseudo-magnetic fields and charging effects in monolayer graphene. We further demonstrate the feasibility of nano-scale strain engineering for graphene-based devices by means of theoretical simulations and nano-fabrication technology.

  19. Contribution of PRS3, RPB4 and ZWF1 to the resistance of industrial Saccharomyces cerevisiae CCUG53310 and PE-2 strains to lignocellulosic hydrolysate-derived inhibitors

    OpenAIRE

    Cunha, J. T.; Aguiar, Tatiana Quinta; Romaní, Aloia; Oliveira, Carla Cristina Marques de; Domingues, Lucília

    2015-01-01

    PRS3, RPB4 and ZWF1 were previously identified as key genes for yeast tolerance to lignocellulose-derived inhibitors. To better understand their contribution to yeast resistance to the multiple stresses occurring during lignocellulosic hydrolysate fermentations, we overexpressed these genes in two industrial Saccharomyces cerevisiae strains, CCUG53310 and PE-2, and evaluated their impact on the fermentation of Eucalyptus globulus wood and corn cob hydrolysates. PRS3 overexpression improved...

  20. Consolidated bioprocessing of transgenic switchgrass by an engineered and evolved Clostridium thermocellum strain

    Energy Technology Data Exchange (ETDEWEB)

    Yee, Kelsey L [ORNL; Rodriguez Jr, Miguel [ORNL; Thompson, Olivia A [ORNL; Fu, Chunxiang [Noble Foundation; Wang, Zeng-Yu [Noble Foundation; Davison, Brian H [ORNL; Mielenz, Jonathan R [ORNL

    2014-01-01

    Background: Switchgrass is an abundant and dedicated bioenergy feedstock however its inherent recalcitrance is one of the economic hurdles for producing biofuels. The down-regulation of the caffeic acid O-methyl transferase (COMT) gene in the lignin pathway of switchgrass reduced lignin content and S/G ratio, and the transgenic lines showed improved fermentation yield with S. cerevisiae and C. thermocellum (ATCC 27405) in comparison to the wild-type switchgrass. Results: Here we examine the fermentation potential of the COMT transgenic switchgrass and its wild-type line, with an engineered and evolved Clostridium thermocellum (M1570) strain. The fermentation of the transgenic switchgrass had superior conversion relative to the control line with an increase of 20% and ethanol was the primary metabolite accounting for 90% of the total metabolites measured by HPLC. Conclusions: The down-regulation of the COMT gene in switchgrass reduced recalcitrance and improved microbial bioconversion yield. Moreover, these results showed ethanol as the main fermentation metabolite produced by an engineered and evolved C. thermocellum strain grown on a transgenic switchgrass.

  1. Engineering Saccharomyces cerevisiae to produce feruloyl esterase for the release of ferulic acid from switchgrass

    Science.gov (United States)

    The Aspergillus niger ferulic acid esterase gene (faeA) was cloned into Saccharomyces cerevisiae via a yeast expression vector, resulting in efficient expression and secretion of the enzyme in the medium. The recombinant enzyme was purified to homogeneity by anion-exchange and hydrophobic interactio...

  2. Strain Engineering to Modify the Electrochemistry of Energy Storage Electrodes

    Science.gov (United States)

    Muralidharan, Nitin; Carter, Rachel; Oakes, Landon; Cohn, Adam P.; Pint, Cary L.

    2016-01-01

    Strain engineering has been a critical aspect of device design in semiconductor manufacturing for the past decade, but remains relatively unexplored for other applications, such as energy storage. Using mechanical strain as an input parameter to modulate electrochemical potentials of metal oxides opens new opportunities intersecting fields of electrochemistry and mechanics. Here we demonstrate that less than 0.1% strain on a Ni-Ti-O based metal-oxide formed on superelastic shape memory NiTi alloys leads to anodic and cathodic peak potential shifts by up to ~30 mV in an electrochemical cell. Moreover, using the superelastic properties of NiTi to enable strain recovery also recovers the electrochemical potential of the metal oxide, providing mechanistic evidence of strain-modified electrochemistry. These results indicate that mechanical energy can be coupled with electrochemical systems to efficiently design and optimize a new class of strain-modulated energy storage materials. PMID:27283872

  3. Strain Engineering to Modify the Electrochemistry of Energy Storage Electrodes

    Science.gov (United States)

    Muralidharan, Nitin; Carter, Rachel; Oakes, Landon; Cohn, Adam P.; Pint, Cary L.

    2016-06-01

    Strain engineering has been a critical aspect of device design in semiconductor manufacturing for the past decade, but remains relatively unexplored for other applications, such as energy storage. Using mechanical strain as an input parameter to modulate electrochemical potentials of metal oxides opens new opportunities intersecting fields of electrochemistry and mechanics. Here we demonstrate that less than 0.1% strain on a Ni-Ti-O based metal-oxide formed on superelastic shape memory NiTi alloys leads to anodic and cathodic peak potential shifts by up to ~30 mV in an electrochemical cell. Moreover, using the superelastic properties of NiTi to enable strain recovery also recovers the electrochemical potential of the metal oxide, providing mechanistic evidence of strain-modified electrochemistry. These results indicate that mechanical energy can be coupled with electrochemical systems to efficiently design and optimize a new class of strain-modulated energy storage materials.

  4. Comparative transcriptomic analysis reveals similarities and dissimilarities in saccharomyces cerevisiae wine strains response to nitrogen availability

    OpenAIRE

    Barbosa, Catarina; García Martínez, José; Pérez Ortín, José Enrique; Mendes Ferreira, A.

    2015-01-01

    Nitrogen levels in grape-juices are of major importance in winemaking ensuring adequate yeast growth and fermentation performance. Here we used a comparative transcriptome analysis to uncover wine yeasts responses to nitrogen availability during fermentation. Gene expression was assessed in three genetically and phenotypically divergent commercial wine strains (CEG, VL1 and QA23), under low (67 mg/L) and high nitrogen (670 mg/L) regimes, at three time points during fermentation (12h, 24h and ...

  5. Promoters inducible by aromatic amino acids and γ-aminobutyrate (GABA) for metabolic engineering applications in Saccharomyces cerevisiae.

    Science.gov (United States)

    Kim, Sujin; Lee, Kyusung; Bae, Sang-Jeong; Hahn, Ji-Sook

    2015-03-01

    A wide range of promoters with different strengths and regulatory mechanisms are valuable tools in metabolic engineering and synthetic biology. While there are many constitutive promoters available, the number of inducible promoters is still limited for pathway engineering in Saccharomyces cerevisiae. Here, we constructed aromatic amino-acid-inducible promoters based on the binding sites of Aro80 transcription factor, which is involved in the catabolism of aromatic amino acids through transcriptional activation of ARO9 and ARO10 genes in response to aromatic amino acids. A dynamic range of tryptophan-inducible promoter strengths can be obtained by modulating the number of Aro80 binding sites, plasmid copy numbers, and tryptophan concentrations. Using low and high copy number plasmid vectors and different tryptophan concentrations, a 29-fold range of fluorescence intensities of enhanced green fluorescent protein (EGFP) reporter could be achieved from a synthetic U4C ARO9 promoter, which is composed of four repeats of Aro80 binding half site (CCG) and ARO9 core promoter element. The U4C ARO9 promoter was applied to express alsS and alsD genes from Bacillus subtilis for acetoin production in S. cerevisiae, resulting in a gradual increase in acetoin titers depending on tryptophan concentrations. Furthermore, we demonstrated that γ-aminobutyrate (GABA)-inducible UGA4 promoter, regulated by Uga3, can also be used in metabolic engineering as a dose-dependent inducible promoter. The wide range of controllable expression levels provided by these tryptophan- and GABA-inducible promoters might contribute to fine-tuning gene expression levels and timing for the optimization of pathways in metabolic engineering. PMID:25573467

  6. BioREFINE-2G project – Engineering of industrial yeast strains for production of dicarboxylic acids from side and waste streams

    DEFF Research Database (Denmark)

    Stovicek, Vratislav; Chen, Xiao; Borodina, Irina;

    2014-01-01

    compounds can be polymerised to biodegradable polymersthat can find application as plastics, coatings or adhesives. To reach the goals, the identification of relevant metabolic routes, strain engineering and the development of a toolbox for manipulation of industrial S. cerevisiae strains are required. Here......, we present advanced genetic engineering tools applicable for generally hardly amenable strains with industrial background. This involves tools forstable heterologous gene (over-)expression and a strategy for fast performance of gene disruption inmultiple ploidy strains. The use of the developed...... generation biorefineries utilize less than 20% of the biomass feedstock for ethanol production. Major side-streams are produced such as pentose and lignin waste streams that are used for biogas and energy production. Converting the carbon from these waste streams into added-value products would improve the...

  7. Strain Engineering Water Transport in Graphene Nano-channels

    OpenAIRE

    Xiong, Wei; Liu, Jefferson Zhe; Ma, Ming; Xu, Zhiping; Sheridan, John; Zheng, Quanshui

    2011-01-01

    Using equilibrium and non-equilibrium molecular dynamic (MD) simulations, we found that engineering the strain on the graphene planes forming a channel can drastically change the interfacial friction of water transport through it. There is a sixfold change of interfacial friction stress when the strain changes from -10% to 10%. Stretching the graphene walls increases the interfacial shear stress, while compressing the graphene walls reduces it. Detailed analysis of the molecular structure rev...

  8. Strain Engineering of Kapitza Resistance in Few-Layer Graphene

    DEFF Research Database (Denmark)

    Chen, Jie; Walther, Jens Honore; Koumoutsakos, Petros

    2014-01-01

    We demonstrate through molecular dynamics simulations that the Kapitza resistance in few-layer graphene (FLG) can be controlled by applying mechanical strain. For unstrained FLG, the Kapitza resistance decreases with the increase of thickness and reaches an asymptotic value of 6 × 10–10 m2K/W at ...... is applied. Our study suggests that graphene can be exploited for both heat dissipation and insulation through strain engineering....

  9. Breaking phylogenetic barriers for fine and bulk chemical products in engineered Saccharomyces cerevisiae

    OpenAIRE

    Codazzi,

    2011-01-01

    Industrial biotechnologies allow today to obtain both fine and bulk chemicals and yeasts as cell factories can produce many products belonging to both field (Branduardi et al., 2008, Porro and Branduardi, 2009). Among yeasts, Saccharomyces cerevisiae still represents the microorganism of election to develop such cell factories. As regard bioethanol production, yeasts utilization is well established for its natural fermentation ability, but new generation biofuels require ...

  10. Metabolic Engineering of Saccharomyces cerevisiae for Astaxanthin Production and Oxidative Stress Tolerance▿

    OpenAIRE

    Ukibe, Ken; Hashida, Keisuke; Yoshida, Nobuyuki; Takagi, Hiroshi

    2009-01-01

    The red carotenoid astaxanthin possesses higher antioxidant activity than other carotenoids and has great commercial potential for use in the aquaculture, pharmaceutical, and food industries. In this study, we produced astaxanthin in the budding yeast Saccharomyces cerevisiae by introducing the genes involved in astaxanthin biosynthesis of carotenogenic microorganisms. In particular, expression of genes of the red yeast Xanthophyllomyces dendrorhous encoding phytoene desaturase (crtI product)...

  11. Engineering of core Pentose Metabolism in Saccharomyces cerevisiae for Bio-ethanol Production

    OpenAIRE

    Pereira, Filipa Alexandra Barroso

    2013-01-01

    Tese de Doutoramento em Ciências (Especialidade em Biologia) Renewable fuels that do not contribute to atmospheric carbon dioxide have gained increased attention due to peak oil and the possibility of carbon dioxide induced climate change. Bioethanol is the currently largest biofuel in terms of annual production and is mainly produce by fermentation of hexose sugars in sucrose or starch from sugarcane or corn by the yeast Saccharomyces cerevisiae. Second generation biofuel is ...

  12. Strain engineering in semiconducting two-dimensional crystals

    International Nuclear Information System (INIS)

    One of the fascinating properties of the new families of two-dimensional crystals is their high stretchability and the possibility to use external strain to manipulate, in a controlled manner, their optical and electronic properties. Strain engineering, understood as the field that study how the physical properties of materials can be tuned by controlling the elastic strain fields applied to it, has a perfect platform for its implementation in the atomically thin semiconducting materials. The object of this review is to give an overview of the recent progress to control the optical and electronics properties of 2D crystals, by means of strain engineering. We will concentrate on semiconducting layered materials, with especial emphasis in transition metal dichalcogenides (MoS2, WS2, MoSe2 and WSe2). The effect of strain in other atomically thin materials like black phosphorus, silicene, etc, is also considered. The benefits of strain engineering in 2D crystals for applications in nanoelectronics and optoelectronics will be revised, and the open problems in the field will be discussed. (topical review)

  13. Enhanced 3-Sulfanylhexan-1-ol Production in Sequential Mixed Fermentation with Torulaspora delbrueckii/Saccharomyces cerevisiae Reveals a Situation of Synergistic Interaction between Two Industrial Strains.

    Science.gov (United States)

    Renault, Philippe; Coulon, Joana; Moine, Virginie; Thibon, Cécile; Bely, Marina

    2016-01-01

    The aim of this work was to study the volatile thiol productions of two industrial strains of Torulaspora delbrueckii and Saccharomyces cerevisiae during alcoholic fermentation (AF) of Sauvignon Blanc must. In order to evaluate the influence of the inoculation procedure, sequential and simultaneous mixed cultures were carried out and compared to pure cultures of T. delbrueckii and S. cerevisiae. The results confirmed the inability of T. delbrueckii to release 4-methyl-4-sulfanylpentan-2-one (4MSP) and its low capacity to produce 3-sulfanylhexyl acetate (3SHA), as already reported in previous studies. A synergistic interaction was observed between the two species, resulting in higher levels of 3SH (3-sulfanylhexan-1-ol) and its acetate when S. cerevisiae was inoculated 24 h after T. delbrueckii, compared to the pure cultures. To elucidate the nature of the interactions between these two species, the yeast population kinetics were examined and monitored, as well as the production of 3SH, its acetate and their related non-odorous precursors: Glut-3SH (glutathionylated conjugate precursor) and Cys-3SH (cysteinylated conjugate precursor). For the first time, it was suggested that, unlike S. cerevisiae, which is able to metabolize the two precursor forms, T. delbrueckii was only able to metabolize the glutathionylated precursor. Consequently, the presence of T. delbrueckii during mixed fermentation led to an increase in Glut-3SH degradation and Cys-3SH production. This overproduction was dependent on the T. delbrueckii biomass. In sequential culture, thus favoring T. delbrueckii development, the higher availability of Cys-3SH throughout AF resulted in more abundant 3SH and 3SHA production by S. cerevisiae. PMID:27014216

  14. Enhanced 3-sulfanylhexan-1-ol production in sequential mixed fermentation with Torulaspora delbrueckii/Saccharomyces cerevisiae reveals a situation of synergistic interaction between two industrial strains

    Directory of Open Access Journals (Sweden)

    Philippe eRenault

    2016-03-01

    Full Text Available The aim of this work was to study the volatile thiol productions of 2 industrial strains of Torulaspora delbrueckii and Saccharomyces cerevisiae during alcoholic fermentation (AF of Sauvignon Blanc must. In order to evaluate the influence of the inoculation procedure, sequential and simultaneous mixed cultures were carried out and compared to pure cultures of T. delbrueckii and S. cerevisiae. The results confirmed the inability of T. delbrueckii to release 4-methyl-4-sulfanylpentan-2-one (4MSP and its low capacity to produce 3-sulfanylhexyl acetate (3SHA, as already reported in previous studies. A synergistic interaction was observed between the two species, resulting in higher levels of 3SH (3-sulfanylhexan-1-ol and its acetate when S. cerevisiae was inoculated 24 hours after T. delbrueckii, compared to the pure cultures. To elucidate the nature of the interactions between these 2 species, the yeast population kinetics were examined and monitored, as well as the production of 3SH, its acetate and their related non-odorous precursors: Glut-3SH (glutathionylated conjugate precursor and Cys-3SH (cysteinylated conjugate precursor. For the first time, it was suggested that, unlike, S. cerevisiae, which is able to metabolize the two precursor forms, T. delbrueckii was only able to metabolize the glutathionylated precursor. Consequently, the presence of T. delbrueckii during mixed fermentation led to an increase in Glut-3SH degradation and Cys-3SH production. This overproduction was dependent on the T. delbrueckii biomass. In sequential culture, thus favouring T. delbrueckii development, the higher availability of Cys-3SH throughout AF (alcoholic fermentation resulted in more abundant 3SH and 3SHA production by S. cerevisiae

  15. An event-specific method for the detection and quantification of ML01, a genetically modified Saccharomyces cerevisiae wine strain, using quantitative PCR.

    Science.gov (United States)

    Vaudano, Enrico; Costantini, Antonella; Garcia-Moruno, Emilia

    2016-10-01

    The availability of genetically modified (GM) yeasts for winemaking and, in particular, transgenic strains based on the integration of genetic constructs deriving from other organisms into the genome of Saccharomyces cerevisiae, has been a reality for several years. Despite this, their use is only authorized in a few countries and limited to two strains: ML01, able to convert malic acid into lactic acid during alcoholic fermentation, and ECMo01 suitable for reducing the risk of carbamate production. In this work we propose a quali-quantitative culture-independent method for the detection of GM yeast ML01 in commercial preparations of ADY (Active Dry Yeast) consisting of efficient extraction of DNA and qPCR (quantitative PCR) analysis based on event-specific assay targeting MLC (malolactic cassette), and a taxon-specific S. cerevisiae assay detecting the MRP2 gene. The ADY DNA extraction methodology has been shown to provide good purity DNA suitable for subsequent qPCR. The MLC and MRP2 qPCR assay showed characteristics of specificity, dynamic range, limit of quantification (LOQ) limit of detection (LOD), precision and trueness, which were fully compliant with international reference guidelines. The method has been shown to reliably detect 0.005% (mass/mass) of GM ML01 S. cerevisiae in commercial preparations of ADY. PMID:27367966

  16. Field performance of a genetically engineered strain of pink bollworm.

    Directory of Open Access Journals (Sweden)

    Gregory S Simmons

    Full Text Available Pest insects harm crops, livestock and human health, either directly or by acting as vectors of disease. The Sterile Insect Technique (SIT--mass-release of sterile insects to mate with, and thereby control, their wild counterparts--has been used successfully for decades to control several pest species, including pink bollworm, a lepidopteran pest of cotton. Although it has been suggested that genetic engineering of pest insects provides potential improvements, there is uncertainty regarding its impact on their field performance. Discrimination between released and wild moths caught in monitoring traps is essential for estimating wild population levels. To address concerns about the reliability of current marking methods, we developed a genetically engineered strain of pink bollworm with a heritable fluorescent marker, to improve discrimination of sterile from wild moths. Here, we report the results of field trials showing that this engineered strain performed well under field conditions. Our data show that attributes critical to SIT in the field--ability to find a mate and to initiate copulation, as well as dispersal and persistence in the release area--were comparable between the genetically engineered strain and a standard strain. To our knowledge, these represent the first open-field experiments with a genetically engineered insect. The results described here provide encouragement for the genetic control of insect pests.

  17. Ultrafast strain engineering in complex oxide heterostructures

    Energy Technology Data Exchange (ETDEWEB)

    Popovich, Paul; Caviglia, Andrea; Hu, Wanzheng; Bromberger, Hubertus; Singla, Rashmi; Mitrano, Matteo; Hoffmann, Matthias C.; Kaiser, Stefan; Foerst, Michael [Max-Planck Research Group for Structural Dynamics - Center for Free Electron Laser Science, University of Hamburg (Germany); Scherwitzl, Raoul; Zubko, Pavlo; Gariglio, Sergio; Triscone, Jean-Marc [Departement de Physique de la Matiere Condensee, University of Geneva, 24 Quai Ernest-Ansermet, 1211 Geneve 4, Geneva (Switzerland); Cavalleri, Andrea [Max-Planck Research Group for Structural Dynamics - Center for Free Electron Laser Science, University of Hamburg (Germany); Department of Physics, Clarendon Laboratory, University of Oxford (United Kingdom)

    2012-07-01

    The mechanical coupling between the substrate and the thin film is expected to be effective on the ultrafast timescale, and could be exploited for the dynamic control of materials properties. Here, we demonstrate that a large-amplitude mid-infrared field, made resonant with a stretching mode of the substrate, can switch the electronic properties of a thin film across an interface. Exploiting dynamic strain propagation between different components of a heterostructure, insulating antiferromagnetic NdNiO{sub 3} is driven through a prompt, five-order-of-magnitude increase of the electrical conductivity, with resonant frequency and susceptibility that is controlled by choice of the substrate material. Vibrational phase control, extended here to a wide class of heterostructures and interfaces, may be conductive to new strategies for electronic phase control at THz repetition rates.

  18. Strain engineered barium strontium titanate for tunable thin film resonators

    Energy Technology Data Exchange (ETDEWEB)

    Khassaf, H.; Khakpash, N. [Department of Materials Science and Engineering and Institute of Materials Science, University of Connecticut, Storrs, Connecticut 06269 (United States); Sun, F. [Department of Physics, University of Connecticut, Storrs, Connecticut 06269 (United States); Sbrockey, N. M.; Tompa, G. S. [Structured Materials Industries, Inc., Piscataway, New Jersey 08854 (United States); Kalkur, T. S. [Department of Electrical and Computer Engineering, University of Colorado at Colorado Springs, Colorado Springs, Colorado 80918 (United States); Alpay, S. P., E-mail: p.alpay@ims.uconn.edu [Department of Materials Science and Engineering and Institute of Materials Science, University of Connecticut, Storrs, Connecticut 06269 (United States); Department of Physics, University of Connecticut, Storrs, Connecticut 06269 (United States)

    2014-05-19

    Piezoelectric properties of epitaxial (001) barium strontium titanate (BST) films are computed as functions of composition, misfit strain, and temperature using a non-linear thermodynamic model. Results show that through adjusting in-plane strains, a highly adaptive rhombohedral ferroelectric phase can be stabilized at room temperature with outstanding piezoelectric response exceeding those of lead based piezoceramics. Furthermore, by adjusting the composition and the in-plane misfit, an electrically tunable piezoelectric response can be obtained in the paraelectric state. These findings indicate that strain engineered BST films can be utilized in the development of electrically tunable and switchable surface and bulk acoustic wave resonators.

  19. Alcoholic chestnut fermentation in mixed culture. Compatibility criteria between Aspergillus oryzae and Saccharomyces cerevisiae strains.

    Science.gov (United States)

    Murado, Miguel Anxo; Pastrana, Lorenzo; Vázquez, José Antonio; Mirón, Jesús; González, María Pilar

    2008-10-01

    The main objective of the present work consisted in the transfer to the case of the chestnut of a rice fermentative process that carried out to the Japanese traditional way to lead to an alcoholic bagasse, the moromi, capable of obtaining distilled. This way, selection assays of amylolitic Aspergillus oryzae strains and studies of compatibility between microfungi and yeast were carried out. These mixed cultivations were performed operating in batch submerged culture. Later on, using solid state system (chestnut, microfungi, yeast), a fermentative fed-batch process (koji, moto, moromi) was defined. By means of this approach a yield of 70% was reached in the conversion of total carbohydrates in ethanol. Also, the time required by the traditional operation was reduced in half. PMID:18289846

  20. Ethanol production from kitchen waste using the flocculating yeast Saccharomyces cerevisiae strain KF-7

    International Nuclear Information System (INIS)

    A process for producing ethanol from kitchen waste was developed in this study. The process consists of freshness preservation of the waste, saccharification of the sugars in the waste, continuous ethanol fermentation of the saccharified liquid, and anaerobic treatment of the saccharification residue and the stillage. Spraying lactic acid bacteria (LCB) on the kitchen waste kept the waste fresh for over 1 week. High glucose recovery (85.5%) from LCB-sprayed waste was achieved after saccharification using Nagase N-40 glucoamylase. The resulting saccharified liquid was used directly for ethanol fermentation, without the addition of any nutrients. High ethanol productivity (24.0 g l-1 h-1) was obtained when the flocculating yeast strain KF-7 was used in a continuous ethanol fermentation process at a dilution rate of 0.8 h-1. The saccharification residue was mixed with stillage and treated in a thermophilic anaerobic continuous stirred tank reactor (CSTR); a VTS loading rate of 6 g l-1 d-1 with 72% VTS digestion efficiency was achieved. Using this process, 30.9 g ethanol, and 65.2 l biogas with 50% methane, was produced from 1 kg of kitchen waste containing 118.0 g total sugar. Thus, energy in kitchen waste can be converted to ethanol and methane, which can then be used as fuels, while simultaneously treating kitchen waste

  1. De novo production of resveratrol from glucose or ethanol by engineered Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Li, Mingji; Kildegaard, Kanchana Rueksomtawin; Chen, Yun;

    2015-01-01

    Resveratrol is a natural antioxidant compound, used as food supplement and cosmetic ingredient. Microbial production of resveratrol has until now been achieved by supplementation of expensive substrates, p-coumaric acid or aromatic amino acids. Here we engineered the yeast Saccharomyces cerevisiae...... to produce resveratrol directly from glucose or ethanol via tyrosine intermediate. First we introduced the biosynthetic pathway, consisting of tyrosine ammonia-lyase from Herpetosiphon aurantiacus, 4-coumaryl-CoA ligase from Arabidopsis thaliana and resveratrol synthase from Vitis vinifera, and...... obtained 2.73±0.05 mg L−1 resveratrol from glucose. Then we over-expressed feedback-insensitive alleles of ARO4 encoding 3-deoxy-D-arabino-heptulosonate-7-phosphate and ARO7 encoding chorismate mutase, resulting in production of 4.85±0.31 mg L−1 resveratrol from glucose as the sole carbon source. Next we...

  2. Saccharomyces cerevisiae strain improvement using selection, mutation, and adaptation for the resistance to lignocellulose-derived fermentation inhibitor for ethanol production.

    Science.gov (United States)

    Jang, Youri; Lim, Younghoon; Kim, Keun

    2014-05-01

    Twenty-five Saccharomyces cerevisiae strains were screened for the highest sugar tolerance, ethanol-tolerance, ethanol production, and inhibitor resistance, and S. cerevisiae KL5 was selected as the best strain. Inhibitor cocktail (100%) was composed of 75 mM formic acid, 75 mM acetic acid, 30 mM furfural, 30 mM hydroxymethyl furfural (HMF), and 2.7 mM vanillin. The cells of strain KL5 were treated with γ-irradiation, and among the survivals, KL5- G2 with improved inhibitor resistance and the highest ethanol yield in the presence of inhibitor cocktail was selected. The KL5-G2 strain was adapted to inhibitor cocktail by sequential transfer of cultures to a minimal YNB medium containing increasing concentrations of inhibitor cocktail. After 10 times of adaptation, most of the isolated colonies could grow in YNB with 80% inhibitor cocktail, whereas the parental KL5 strain could not grow at all. Among the various adapted strains, the best strain (KL5-G2-A9) producing the highest ethanol yield in the presence of inhibitor cocktail was selected. In a complex YP medium containing 60% inhibitor cocktail and 5% glucose, the theoretical yield and productivity (at 48 h) of KL5- G2-A9 were 81.3% and 0.304 g/l/h, respectively, whereas those of KL5 were 20.8% and 0.072 g/l/h, respectively. KL5-G2-A9 reduced the concentrations of HMF, furfural, and vanillin in the medium in much faster rates than KL5. PMID:24608567

  3. Origin of co-expression patterns in E. coli and S. cerevisiae emerging from reverse engineering algorithms.

    Directory of Open Access Journals (Sweden)

    Mattia Zampieri

    Full Text Available BACKGROUND: The concept of reverse engineering a gene network, i.e., of inferring a genome-wide graph of putative gene-gene interactions from compendia of high throughput microarray data has been extensively used in the last few years to deduce/integrate/validate various types of "physical" networks of interactions among genes or gene products. RESULTS: This paper gives a comprehensive overview of which of these networks emerge significantly when reverse engineering large collections of gene expression data for two model organisms, E. coli and S. cerevisiae, without any prior information. For the first organism the pattern of co-expression is shown to reflect in fine detail both the operonal structure of the DNA and the regulatory effects exerted by the gene products when co-participating in a protein complex. For the second organism we find that direct transcriptional control (e.g., transcription factor-binding site interactions has little statistical significance in comparison to the other regulatory mechanisms (such as co-sharing a protein complex, co-localization on a metabolic pathway or compartment, which are however resolved at a lower level of detail than in E. coli. CONCLUSION: The gene co-expression patterns deduced from compendia of profiling experiments tend to unveil functional categories that are mainly associated to stable bindings rather than transient interactions. The inference power of this systematic analysis is substantially reduced when passing from E. coli to S. cerevisiae. This extensive analysis provides a way to describe the different complexity between the two organisms and discusses the critical limitations affecting this type of methodologies.

  4. The effects of dietary administration with chemical treated Saccharomyces cerevisiae strain YG3-1 on the growth of aquatic invertebrates in Artemia

    Directory of Open Access Journals (Sweden)

    Behnam Shekarchi

    2016-02-01

    Full Text Available Objective: To investigate the biological effects of β-glucans in cell wall of new identified strain Saccharomyces cerevisiae strain YG3-1 on the growth of aquatic invertebrates, in Artemia as model organism. Methods: All yeasts used in the present study were isolated from Rainbow trout intestine and then cultured in yeast extract-peptone-glycerol medium. Activation of β-glucan in yeasts was performed by chemical treatment with 2-mercaptoethanol (2ME (3.5% v/v. Then nauplii and larvae individuals of Artemia urmiana and Artemia franciscana (two different species of Artemia as test organisms were fed with 2ME-treated yeasts during the culture. At the end of experiment, after feeding individual length (total length and growth rate in adult individuals of Artemia was measured. Results: Following this administration, growth in both species of Artemia was improved (P < 0.05. So, the results showed that Artemia urmiana adults individuals that fed with 2MEtreated yeasts had the highest growth and total length. These results were confirmed with growth measurement in adult individuals of Artemia. Conclusions: This study suggested that 2ME-treated Saccharomyces cerevisiae strain YG3-1 yeasts can be used for enhancing the growth of other aquatic invertebrates like shrimps as probiotic supplement and growth promoter.

  5. The effects of dietary administration with chemical treated Saccharomyces cerevisiae strain YG3-1 on the growth of aquatic invertebrates in Artemia

    Institute of Scientific and Technical Information of China (English)

    Behnam Shekarchi; Ali Nekuiefard; Ramin Manaffar

    2016-01-01

    Objective: To investigate the biological effects of β-glucans in cell wall of new identified strain Saccharomyces cerevisiae strain YG3-1 on the growth of aquatic invertebrates, in Artemia as model organism. Methods: All yeasts used in the present study were isolated from Rainbow trout intestine and then cultured in yeast extract-peptone-glycerol medium. Activation of β-glucan in yeasts was performed by chemical treatment with 2-mercaptoethanol (2ME) (3.5% v/v). Then nauplii and larvae individuals of Artemia urmiana and Artemia franciscana (two different species of Artemia as test organisms) were fed with 2ME-treated yeasts during the culture. At the end of experiment, after feeding individual length (total length and growth rate) in adult individuals of Artemia was measured. Results: Following this administration, growth in both species of Artemia was improved (P Conclusions: This study suggested that 2ME-treated Saccharomyces cerevisiae strain YG3-1 yeasts can be used for enhancing the growth of other aquatic invertebrates like shrimps as probiotic supplement and growth promoter.

  6. Engineering related neutron diffraction measurements probing strains, texture and microstructure

    Energy Technology Data Exchange (ETDEWEB)

    Clausen, Bjorn [Los Alamos National Laboratory; Brown, Donald W [Los Alamos National Laboratory; Tome, Carlos N [Los Alamos National Laboratory; Balogh, Levente [Los Alamos National Laboratory; Vogel, Sven C [Los Alamos National Laboratory

    2010-01-01

    Neutron diffraction has been used for engineering applications for nearly three decades. The basis of the technique is powder diffraction following Bragg's Law. From the measured diffraction patterns information about internal, or residual, strain can be deduced from the peak positions, texture information can be extracted from the peak intensities, and finally the peak widths can provide information about the microstructure, e.g. dislocation densities and grain sizes. The strains are measured directly from changes in lattice parameters, however, in many cases it is non-trivial to determine macroscopic values of stress or strain from the measured data. The effects of intergranular strains must be considered, and combining the neutron diffraction measurements with polycrystal deformation modeling has proven invaluable in determining the overall stress and strain values of interest in designing and dimensioning engineering components. Furthelmore, the combined use of measurements and modeling has provided a tool for elucidating basic material properties, such as critical resolved shear stresses for the active deformation modes and their evolution as a function of applied deformation.

  7. Differential Azole Antifungal Efficacies Contrasted Using a Saccharomyces cerevisiae Strain Humanized for Sterol 14α-Demethylase at the Homologous Locus▿

    OpenAIRE

    Parker, J. E.; Merkamm, M.; Manning, N J; Pompon, D; Kelly, S. L.; Kelly, D. E.

    2008-01-01

    Inhibition of sterol-14α-demethylase, a cytochrome P450 (CYP51, Erg11p), is the mode of action of azole antifungal drugs, and with high frequencies of fungal infections new agents are required. New drugs that target fungal CYP51 should not inhibit human CYP51, although selective inhibitors of the human target are also of interest as anticholesterol agents. A strain of Saccharomyces cerevisiae that was humanized with respect to the amino acids encoded at the CYP51 (ERG11) yeast locus (BY4741:h...

  8. THE EFFECT INDUCED BY MILLIMETER WAVES WITH THE FREQUENCY 53.33 GHZ ON SACCHAROMYCES CEREVISIAE CNMN-Y-18 YEAST STRAIN

    OpenAIRE

    Agafia Usatîi; Elena Molodoi; Nadejda Efromova; Ludmila Fulga

    2015-01-01

    The effect of extremely high frequency electromagnetic waves on the biosynthetic activity of Saccharomyces cerevisiae CNMN-Y-18 yeast strain in dependence on the duration of irradiation was studied. The maximum amount of biomass, protein, carbohydrates, mannoproteins and catalase has been showed to accumulate when the yeast cells were irradiated with a frequency f = 53.33 GHz for 10 minutes. High degree of dependence between the content of cellular components (a correlation coefficient betwee...

  9. Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Bojsen, Rasmus K; Andersen, Kaj Scherz; Regenberg, Birgitte

    2012-01-01

    Microbial biofilms can be defined as multi-cellular aggregates adhering to a surface and embedded in an extracellular matrix (ECM). The nonpathogenic yeast, Saccharomyces cerevisiae, follows the common traits of microbial biofilms with cell-cell and cell-surface adhesion. S. cerevisiae is shown t...... cues, cell-to-cell variation and niches in S. cerevisiae biofilm. Being closely related to Candida species, S. cerevisiae is a model to investigate biofilms of pathogenic yeast....

  10. Fiber Bragg grating strain sensors for marine engineering

    Science.gov (United States)

    Wang, Tingting; Yuan, Zilin; Gong, Yuan; Wu, Yu; Rao, Yunjiang; Wei, Lili; Guo, Peng; Wang, Junpu; Wan, Fu

    2013-09-01

    For the health monitoring of the offshore drilling platform in the salt-fog environment, three nonmetallic materials, i.e., silica, FR-4 epoxy board and sheet molding compound (SMC), with the good anti-corrosion ability were chosen as the packaging materials for the fiber Bragg grating (FBG) strain sensors. By selecting a highly sensitive structure of the fiber Bragg grating strain sensor, the performances of the sensors with three materials were investigated both numerically and experimentally. The strain sensitivities were 3.76 pm/μɛ, 3.02 pm/μɛ and 3.03 pm/μɛ, respectively. The linearity was better than 0.998. It provides useful information for developing sensors for the marine engineering.

  11. Tunable thermoelectric transport in nanomeshes via elastic strain engineering

    Energy Technology Data Exchange (ETDEWEB)

    Piccione, Brian; Gianola, Daniel S., E-mail: gianola@seas.upenn.edu [Department of Materials Science and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104 (United States)

    2015-03-16

    Recent experimental explorations of silicon nanomeshes have shown that the unique metastructures exhibit reduced thermal conductivity while preserving bulk electrical conductivity via feature sizes between relevant phonon and electron mean free paths, aiding in the continued promise that nanometer-scale engineering may further enhance thermoelectric behavior. Here, we introduce a strategy for tuning thermoelectric transport phenomena in semiconductor nanomeshes via heterogeneous elastic strain engineering, using silicon as a model material for demonstration of the concept. By combining analytical models for electron mobility in uniformly stressed silicon with finite element analysis of strained silicon nanomeshes in a lumped physical model, we show that the nonuniform and multiaxial strain fields defined by the nanomesh geometry give rise to spatially varying band shifts and warping, which in aggregate accelerate electron transport along directions of applied stress. This allows for global electrical conductivity and Seebeck enhancements beyond those of homogenous samples under equivalent far-field stresses, ultimately increasing thermoelectric power factor nearly 50% over unstrained samples. The proposed concept and structures—generic to a wide class of materials with large dynamic ranges of elastic strain in nanoscale volumes—may enable a new pathway for active and tunable control of transport properties relevant to waste heat scavenging and thermal management.

  12. Tunable thermoelectric transport in nanomeshes via elastic strain engineering

    International Nuclear Information System (INIS)

    Recent experimental explorations of silicon nanomeshes have shown that the unique metastructures exhibit reduced thermal conductivity while preserving bulk electrical conductivity via feature sizes between relevant phonon and electron mean free paths, aiding in the continued promise that nanometer-scale engineering may further enhance thermoelectric behavior. Here, we introduce a strategy for tuning thermoelectric transport phenomena in semiconductor nanomeshes via heterogeneous elastic strain engineering, using silicon as a model material for demonstration of the concept. By combining analytical models for electron mobility in uniformly stressed silicon with finite element analysis of strained silicon nanomeshes in a lumped physical model, we show that the nonuniform and multiaxial strain fields defined by the nanomesh geometry give rise to spatially varying band shifts and warping, which in aggregate accelerate electron transport along directions of applied stress. This allows for global electrical conductivity and Seebeck enhancements beyond those of homogenous samples under equivalent far-field stresses, ultimately increasing thermoelectric power factor nearly 50% over unstrained samples. The proposed concept and structures—generic to a wide class of materials with large dynamic ranges of elastic strain in nanoscale volumes—may enable a new pathway for active and tunable control of transport properties relevant to waste heat scavenging and thermal management

  13. Anaerobic and sequential aerobic production of high-titer ethanol and single cell protein from NaOH-pretreated corn stover by a genome shuffling-modified Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Ren, Xueliang; Wang, Juncong; Yu, Hui; Peng, Chunlan; Hu, Jinlong; Ruan, Zhiyong; Zhao, Shumiao; Liang, Yunxiang; Peng, Nan

    2016-10-01

    In this study, a Saccharomyces cerevisiae recombinant strain 14 was constructed through genome shuffling method by transferring the whole genomic DNA of Candida intermedia strain 23 into a thermo-tolerant S. cerevisiae strain. The recombinant strain 14 combined the good natures of both parent strains that efficiently produced ethanol from glucose and single cell protein from xylose with 54.6% crude protein and all essential amino acids except cysteine at 35°C. Importantly, the recombinant strain 14 produced 64.07g/L ethanol from 25%(w/v) NaOH-pretreated and washed corn stover with the ethanol yield of 0.26g/g total stover by fed-batch simultaneous saccharification and fermentation and produced 66.50g/L dry cell mass subsequently from the residual hydrolysate and ethanol. Therefore, this study represents a feasible method to comprehensively utilize hexose and pentose in lignocellulosic materials. PMID:27416512

  14. POTENCIALIDADES DE LINHAGENS DE LEVEDURA Saccharomyces cerevisiae PARA A FERMENTAÇÃO DO CALDO DE CANA POTENTIALITIES OF YEAST STRAINS OF Saccharomyces cerevisiae FOR SUGAR CANE JUICE FERMENTATION

    Directory of Open Access Journals (Sweden)

    Carlos Alberto França Ribeiro

    1999-01-01

    Full Text Available Três linhagens de levedura da espécie Saccharomyces cerevisiae, sendo duas floculantes, das quais uma não produtora de sulfeto de hidrogênio, foram avaliadas para se verificar seus desempenhos, sob parâmetros cinéticos, bem como a formação de compostos secundários na fermentação do caldo de cana-de-açúcar, destinado à produção de aguardente. O acompanhamento da cinética fermentativa mostrou melhores resultados de eficiência de fermentação, fator de conversão de substrato em etanol e velocidade específica de crescimento pela linhagem floculante IZ 987, que foram de 89,9%, 0,46 g.g-1 e 0,0996 h-1 respectivamente. Esta linhagem foi também responsável pela maior produção de álcoois superiores, 185 mg.L-1, inerente à sua característica de não produzir H2S. A melhor produtividade de fermentação, de 3,40 g.L-1.h-1, foi a obtida pela linhagem floculante LF. A linhagem FP, não floculante, isolada à partir do fermento prensado, proporcionou os menores valores dos parâmetros cinéticos estudados.Three yeast strains of Saccharomyces cerevisiae species, two flocculants, one of them non hydrogen sulfide producer, were evaluated for their behavior on kinetic parameters and production of volatile compounds, during sugar cane juice fermentation. The fermentation kinetics presented better performance in terms of fermentation efficiency, ethanol yield and specific growth rate, 89.9%; 0.46 g.g-1 and 0.0996 h-1, respectively, for the IZ 987 strain, that also produced larger amounts of higher alcohols, 185 mg.L-1, inherent to its H2S negative character. Higher ethanol productivity, about 3.40 g.L-1.h-1, was achieved by the flocculant strain. The strain isolated from baker's yeast promoted the poorest results.

  15. Engineering Saccharomyces cerevisiae to produce odd chain-length fatty alcohols.

    Science.gov (United States)

    Jin, Zhu; Wong, Adison; Foo, Jee Loon; Ng, Joey; Cao, Ying-Xiu; Chang, Matthew Wook; Yuan, Ying-Jin

    2016-04-01

    Fatty aldehydes and alcohols are valuable precursors used in the industrial manufacturing of a myriad of specialty products. Herein, we demonstrate the de novo production of odd chain-length fatty aldehydes and fatty alcohols in Saccharomyces cerevisiae by expressing a novel biosynthetic pathway involving cytosolic thioesterase, rice α-dioxygenase and endogenous aldehyde reductases. We attained production titers of ∼20 mg/l fatty aldehydes and ∼20 mg/l fatty alcohols in shake flask cultures after 48 and 60 h respectively without extensive fine-tuning of metabolic fluxes. In contrast to prior studies which relied on bi-functional fatty acyl-CoA reductase to produce even chain-length fatty alcohols, our biosynthetic route exploits α-oxidation reaction to produce odd chain-length fatty aldehyde intermediates without using NAD(P)H cofactor, thereby conserving cellular resource during the overall synthesis of odd chain-length fatty alcohols. The biosynthetic pathway presented in this study has the potential to enable sustainable and efficient synthesis of fatty acid-derived chemicals from processed biomass. PMID:26461930

  16. Enhancement of Farnesyl Diphosphate Pool as Direct Precursor of Sesquiterpenes Through Metabolic Engineering of the Mevalonate Pathway in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Asadollahi, Mohammadali; Maury, Jerome; Schalk, M.;

    2010-01-01

    , resulted in higher production of cubebol, a plant originating sesquiterpene, and increased squalene accumulation. Down-regulation of ERG9 by replacing its native promoter with the regulatable MET3 promoter, enhanced cubebol titers but simultaneous overexpression of tHMG1 and repression of ERG9 did not...... further improve cubebol production. Furtheremore, the concentrations of squalene and ergosterol were measured in the engineered strains. Unexpectedly, significant accumulation of squalene and restoring the ergosterol biosynthesis were observed in the ERG9 repressed strains transformed with the plasmids...

  17. Mutation Breeding of Salt-tolerant and Ethanol-producing Strain S. cerevisiae H058 by Low-energy Ion Implantation

    Directory of Open Access Journals (Sweden)

    Shoubao Yan

    2014-07-01

    Full Text Available To obtain an industrial strain with high ethanol fermentation efficiency under salted conditions, the wild strain H058 of Saccharomyces cerevisiae was mutated by means of nitrogen ions implantation. Mutagenic effects of strain H058 by low energy N+ ion implantation were studied. A similar “saddle shape” survival curve due to ion beam irradiation appeared again in this study. By repeated screening, a high salt-tolerant and ethanol-producing strain M158 was obtained. Results showed that in medium contained 0, 1.5, 3.0, 4.5, 6.0% NaCl, M158 produced maximal ethanol of 98.3, 97.2, 96.4, 95.6 and 78.3 g/L at 54, 54, 54 and 72 h, respectively. However, the original strain H058 maximal ethanol of 95.2, 90.9, 84.8, 79.4 and 67.5 g/L at 60, 60, 66 and 72 h, respectively. In addition, the ethanol yield (g/g in all of the NaCl concentrations for M158 is 0.492, 0.486, 0.482, 0.48 and 0.392 g/g, respectively, which were higher than those (0.476, 0.455, 0.424, 0.397 and 0.338 g/g, respectively of the original strain H058. The higher production and shorter fermentation period suggest that strain M158 is a good salt-tolerant and ethanol-producing strain.

  18. Characterization of plasmid burden and copy number in Saccharomyces cerevisiae for optimization of metabolic engineering applications

    OpenAIRE

    Karim, Ashty S.; Curran, Kathleen A.; Alper, Hal S.

    2012-01-01

    Many metabolic engineering and genetic engineering applications in yeast rely on the use of plasmids. Despite their pervasive use and the diverse collections available, there is a fundamental lack of understanding of how commonly used DNA plasmids affect the cell’s ability to grow and how the choice of plasmid components can influence plasmid load and burden. In this study, we characterized the major attributes of the 2μ and centromeric plasmids typically used in yeast by examining the impact...

  19. The dual mechanism of the antifungal effect of new lysosomotropic agents on the saccharomyces cerevisiae RXII strain

    Czech Academy of Sciences Publication Activity Database

    Krasowska, A.; Chmielewská, L.; Luczynski, J.; Witek, S.; Sigler, Karel

    2003-01-01

    Roč. 8, č. 1 (2003), s. 111-120. ISSN 1425-8153 R&D Projects: GA MŠk ME 577; GA AV ČR IBS5020202 Grant ostatní: GA-(PL) 23/40 of the Czech-Polish Trety on Scientific-Technical Cooperation Institutional research plan: CEZ:AV0Z5020903 Keywords : saccharomyces cerevisiae, lysosomotropic * drugs Subject RIV: EE - Microbiology, Virology Impact factor: 0.455, year: 2003

  20. Increasing cell biomass in Saccharomyces cerevisiae increases recombinant protein yield: the use of a respiratory strain as a microbial cell factory

    Directory of Open Access Journals (Sweden)

    Hedfalk Kristina

    2010-06-01

    Full Text Available Abstract Background Recombinant protein production is universally employed as a solution to obtain the milligram to gram quantities of a given protein required for applications as diverse as structural genomics and biopharmaceutical manufacture. Yeast is a well-established recombinant host cell for these purposes. In this study we wanted to investigate whether our respiratory Saccharomyces cerevisiae strain, TM6*, could be used to enhance the productivity of recombinant proteins over that obtained from corresponding wild type, respiro-fermentative strains when cultured under the same laboratory conditions. Results Here we demonstrate at least a doubling in productivity over wild-type strains for three recombinant membrane proteins and one recombinant soluble protein produced in TM6* cells. In all cases, this was attributed to the improved biomass properties of the strain. The yield profile across the growth curve was also more stable than in a wild-type strain, and was not further improved by lowering culture temperatures. This has the added benefit that improved yields can be attained rapidly at the yeast's optimal growth conditions. Importantly, improved productivity could not be reproduced in wild-type strains by culturing them under glucose fed-batch conditions: despite having achieved very similar biomass yields to those achieved by TM6* cultures, the total volumetric yields were not concomitantly increased. Furthermore, the productivity of TM6* was unaffected by growing cultures in the presence of ethanol. These findings support the unique properties of TM6* as a microbial cell factory. Conclusions The accumulation of biomass in yeast cell factories is not necessarily correlated with a proportional increase in the functional yield of the recombinant protein being produced. The respiratory S. cerevisiae strain reported here is therefore a useful addition to the matrix of production hosts currently available as its improved biomass

  1. Crystallization engineering as a route to epitaxial strain control

    Energy Technology Data Exchange (ETDEWEB)

    Akbashev, Andrew R.; Plokhikh, Aleksandr V.; Barbash, Dmitri [Department of Materials Science and Engineering, Drexel University, Philadelphia, Pennsylvania 19104 (United States); Lofland, Samuel E. [Department of Physics, Rowan University, Glassboro, New Jersey 08028 (United States); Spanier, Jonathan E., E-mail: spanier@drexel.edu [Department of Materials Science and Engineering, Drexel University, Philadelphia, Pennsylvania 19104 (United States); Department of Physics, Drexel University, Philadelphia, Pennsylvania 19104 (United States)

    2015-10-01

    The controlled synthesis of epitaxial thin films offers opportunities for tuning their functional properties via enabling or suppressing strain relaxation. Examining differences in the epitaxial crystallization of amorphous oxide films, we report on an alternate, low-temperature route for strain engineering. Thin films of amorphous Bi–Fe–O were grown on (001)SrTiO{sub 3} and (001)LaAlO{sub 3} substrates via atomic layer deposition. In situ X-ray diffraction and X-ray photoelectron spectroscopy studies of the crystallization of the amorphous films into the epitaxial (001)BiFeO{sub 3} phase reveal distinct evolution profiles of crystallinity with temperature. While growth on (001)SrTiO{sub 3} results in a coherently strained film, the same films obtained on (001)LaAlO{sub 3} showed an unstrained, dislocation-rich interface, with an even lower temperature onset of the perovskite phase crystallization than in the case of (001)SrTiO{sub 3}. Our results demonstrate how the strain control in an epitaxial film can be accomplished via its crystallization from the amorphous state.

  2. Crystallization engineering as a route to epitaxial strain control

    Directory of Open Access Journals (Sweden)

    Andrew R. Akbashev

    2015-10-01

    Full Text Available The controlled synthesis of epitaxial thin films offers opportunities for tuning their functional properties via enabling or suppressing strain relaxation. Examining differences in the epitaxial crystallization of amorphous oxide films, we report on an alternate, low-temperature route for strain engineering. Thin films of amorphous Bi–Fe–O were grown on (001SrTiO3 and (001LaAlO3 substrates via atomic layer deposition. In situ X-ray diffraction and X-ray photoelectron spectroscopy studies of the crystallization of the amorphous films into the epitaxial (001BiFeO3 phase reveal distinct evolution profiles of crystallinity with temperature. While growth on (001SrTiO3 results in a coherently strained film, the same films obtained on (001LaAlO3 showed an unstrained, dislocation-rich interface, with an even lower temperature onset of the perovskite phase crystallization than in the case of (001SrTiO3. Our results demonstrate how the strain control in an epitaxial film can be accomplished via its crystallization from the amorphous state.

  3. Identification of yeasts isolated from raffia wine (Raphia hookeri) produced in Côte d'Ivoire and genotyping of Saccharomyces cerevisiae strains by PCR inter-delta.

    Science.gov (United States)

    Tra Bi, Charles Y; N'guessan, Florent K; Kouakou, Clémentine A; Jacques, Noemie; Casaregola, Serge; Djè, Marcellin K

    2016-08-01

    Raffia wine is a traditional alcoholic beverage produced in several African countries where it plays a significant role in traditional customs and population diet. Alcoholic fermentation of this beverage is ensured by a complex natural yeast flora which plays a decisive role in the quality of the final product. This present study aims to evaluate the distribution and the diversity of the yeast strains isolated in raffia wine from four sampling areas (Abengourou, Alépé, Grand-Lahou and Adzopé) in Côte d'Ivoire. Based on the D1/D2 domain of the LSU rDNA sequence analysis, nine species belonging to six genera were distinguished. With a percentage of 69.5 % out of 171 yeast isolates, Saccharomyces cerevisiae was the predominant species in the raffia wine, followed by Kodamaea ohmeri (20.4 %). The other species isolated were Candida haemulonii (4.1 %), Candida phangngensis (1.8 %), Pichia kudriavzevii (1.2 %), Hanseniaspora jakobsenii (1.2 %), Candida silvae (0.6 %), Hanseniaspora guilliermondii (0.6 %) and Meyerozyma caribbica (0.6 %). The molecular characterization of S. cerevisiae isolates at the strain level using the PCR-interdelta method revealed the presence of 21 profiles (named I to XXI) within 115 isolates. Only four profiles (I, III, V and XI) were shared by the four areas under study. Phenotypic characterization of K. ohmeri strains showed two subgroups for sugar fermentation and no diversity for the nitrogen compound assimilations and the growth at different temperatures. PMID:27339306

  4. Bio-Engineering High Performance Microbial Strains for MEOR

    Energy Technology Data Exchange (ETDEWEB)

    Xiangdong Fang; Qinghong Wang; Patrick Shuler

    2007-12-30

    The main objectives of this three-year research project are: (1) to employ the latest advances in genetics and bioengineering, especially Directed Protein Evolution technology, to improve the effectiveness of the microbial enhanced oil recovery (MEOR) process. (2) to improve the surfactant activity and the thermal stability of bio-surfactant systems for MEOR; and (3) to develop improved laboratory methods and tools that screen quickly candidate bio-systems for EOR. Biosurfactants have been receiving increasing attention as Enhanced Oil Recovery (EOR) agents because of their unique properties (i.e., mild production conditions, lower toxicity, and higher biodegradability) compared to their synthetic chemical counterparts. Rhamnolipid as a potent natural biosurfactant has a wide range of potential applications, including EOR and bioremediation. During the three-year of the project period, we have successfully cloned the genes involved in the rhamnolipid bio-synthesis. And by using the Transposon containing Rhamnosyltransferase gene rhlAB, we engineered the new mutant strains P. aeruginosa PEER02 and E. coli TnERAB so they can produce rhamnolipid biosurfactans. We were able to produce rhamnolipds in both P. aeroginosa PAO1-RhlA- strain and P. fluorescens ATCC15453 strain, with the increase of 55 to 175 fold in rhamnolipid production comparing with wild type bacteria strain. We have also completed the first round direct evolution studies using Error-prone PCR technique and have constructed the library of RhlAB-containing Transposon to express mutant gene in heterologous hosts. Several methods, such as colorimetric agar plate assay, colorimetric spectrophotometer assay, bioactive assay and oil spreading assay have been established to detect and screen rhamnolipid production. Our engineered P. aeruginosa PEER02 strain can produce rhamnolipids with different carbon sources as substrate. Interfacial tension analysis (IFT) showed that different rhamnolipids from different

  5. Strain-engineered manufacturing of freeform carbon nanotube microstructures

    Science.gov (United States)

    de Volder, M.; Park, S.; Tawfick, S.; Hart, A. J.

    2014-07-01

    The skins of many plants and animals have intricate microscale surface features that give rise to properties such as directed water repellency and adhesion, camouflage, and resistance to fouling. However, engineered mimicry of these designs has been restrained by the limited capabilities of top-down fabrication processes. Here we demonstrate a new technique for scalable manufacturing of freeform microstructures via strain-engineered growth of aligned carbon nanotubes (CNTs). Offset patterning of the CNT growth catalyst is used to locally modulate the CNT growth rate. This causes the CNTs to collectively bend during growth, with exceptional uniformity over large areas. The final shape of the curved CNT microstructures can be designed via finite element modeling, and compound catalyst shapes produce microstructures with multidirectional curvature and unusual self-organized patterns. Conformal coating of the CNTs enables tuning of the mechanical properties independently from the microstructure geometry, representing a versatile principle for design and manufacturing of complex microstructured surfaces.

  6. 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...... 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...

  7. Systematic strain construction and process development: Xylitol production by Saccharomyces cerevisiae expressing Candida tenuis xylose reductase in wild-type or mutant form.

    Science.gov (United States)

    Pratter, S M; Eixelsberger, T; Nidetzky, B

    2015-12-01

    A novel Saccharomyces cerevisiae whole-cell biocatalyst for xylitol production based on Candida tenuis xylose reductase (CtXR) is presented. Six recombinant strains expressing wild-type CtXR or an NADH-specific mutant were constructed and evaluated regarding effects of expression mode, promoter strength, biocatalyst concentration and medium composition. Intracellular XR activities ranged from 0.09 U mgProt(-1) to 1.05 U mgProt(-1) but did not correlate with the strains' xylitol productivities, indicating that other factors limited xylose conversion in the high-activity strains. The CtXR mutant decreased the biocatalyst's performance, suggesting use of the NADPH-preferring wild-type enzyme when (semi-)aerobic conditions are applied. In a bioreactor process, the best-performing strain converted 40 g L(-1) xylose with an initial productivity of 1.16 g L(-1)h(-1) and a xylitol yield of 100%. The obtained results underline the potential of CtXR wild-type for xylose reduction and point out parameters to improve "green" xylitol production. PMID:26452180

  8. Metabolic opportunities offered by wild-type and engineered Saccharomyces cerevisiae strains for biofuels production

    OpenAIRE

    Rossi,, A.

    2011-01-01

    Human society has always been dependent on biomass-derived carbonand energy for nutrition and survival. Recently, we have also become dependent on petroleum-derived carbon and energy for commodity chemicals and fuels. However, the nonrenewable nature of petroleum stands in stark contrast to the renewable carbon and energy present in biomass. Thus, there is an increasing demand to develop and implement strategies for production of commodity chemicals and fuels from biomass instead of petroleum...

  9. Fermentation performance of engineered and evolved xylose-fermenting Saccharomyces cerevisiae strains

    DEFF Research Database (Denmark)

    Sonderegger, M.; Jeppsson, M.; Larsson, C.;

    2004-01-01

    Lignocellulose hydrolysate is an abundant substrate for bioethanol production. The ideal microorganism for such a fermentation process should combine rapid and efficient conversion of the available carbon sources to ethanol with high tolerance to ethanol and to inhibitory components in the hydrol...

  10. Metabolic Engineering of Saccharomyces cerevisiae for Conversion of d-Glucose to Xylitol and Other Five-Carbon Sugars and Sugar Alcohols▿

    Science.gov (United States)

    Toivari, Mervi H.; Ruohonen, Laura; Miasnikov, Andrei N.; Richard, Peter; Penttilä, Merja

    2007-01-01

    Recombinant Saccharomyces cerevisiae strains that produce the sugar alcohols xylitol and ribitol and the pentose sugar d-ribose from d-glucose in a single fermentation step are described. A transketolase-deficient S. cerevisiae strain accumulated d-xylulose 5-phosphate intracellularly and released ribitol and pentose sugars (d-ribose, d-ribulose, and d-xylulose) into the growth medium. Expression of the xylitol dehydrogenase-encoding gene XYL2 of Pichia stipitis in the transketolase-deficient strain resulted in an 8.5-fold enhancement of the total amount of the excreted sugar alcohols ribitol and xylitol. The additional introduction of the 2-deoxy-glucose 6-phosphate phosphatase-encoding gene DOG1 into the transketolase-deficient strain expressing the XYL2 gene resulted in a further 1.6-fold increase in ribitol production. Finally, deletion of the endogenous xylulokinase-encoding gene XKS1 was necessary to increase the amount of xylitol to 50% of the 5-carbon sugar alcohols excreted. PMID:17630301

  11. Evaluation of microbial qPCR workflows using engineered Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    S.M. Da Silva

    2016-03-01

    Significance and impact of the study: The engineered yeast has potential to support measurement assurance for the analytical process of qPCR, encompassing the method, equipment, and operator, to increase confidence in results and better inform decision-making in areas of applied microbiology. This material can also support process assessment for other DNA-based detection technologies.

  12. Engineering precursor supply in Saccharomyces cerevisiae: new strategies for cytosolic acetyl-CoA formation

    NARCIS (Netherlands)

    Kozak, B.U.

    2015-01-01

    Metabolic engineering – the improvement and addition, by genetic modification, of industrially relevant properties of microorganisms with respect to catalysis, transport and regulatory functions – is a well-established method for development of more cost-effective and ‘green’ industrial processes. R

  13. 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

  14. Comparative analysis of the effects of locally used herbicides and their active ingredients on a wild-type wine Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Braconi, Daniela; Sotgiu, Michele; Millucci, Lia; Paffetti, Alessandro; Tasso, Flavia; Alisi, Chiara; Martini, Silvia; Rappuoli, Roberto; Lusini, Paola; Sprocati, Anna Rosa; Rossi, Claudio; Santucci, Annalisa

    2006-04-19

    Herbicides are released to the environment with potential ecotoxicological risks for mammals. Yeast is a good model to elucidate toxicity mechanisms. We investigated how three commercial herbicides (Proper Energy, Pointer, and Silglif) and their active ingredients (respectively, fenoxaprop-P-ethyl, tribenuron methyl, and glyphosate) can affect biological activities of an oenological Saccharomyces cerevisiae strain, which may be resident on grape vineyards of the same geographical areas where herbicides are used. The use of commercial grade herbicides employed in Italy allowed us to reproduce the same conditions applied in crops; at the same time, assaying pure single active compounds made it possible to compare the effects obtained with commercial formulations. Interestingly, we found that while pure active compounds affect cell growth and metabolism at a lower extent, commercial preparations have a significant major negative influence on yeast biology. PMID:16608247

  15. THE EFFECT INDUCED BY MILLIMETER WAVES WITH THE FREQUENCY 53.33 GHZ ON SACCHAROMYCES CEREVISIAE CNMN-Y-18 YEAST STRAIN

    Directory of Open Access Journals (Sweden)

    Agafia Usatîi

    2015-04-01

    Full Text Available The effect of extremely high frequency electromagnetic waves on the biosynthetic activity of Saccharomyces cerevisiae CNMN-Y-18 yeast strain in dependence on the duration of irradiation was studied. The maximum amount of biomass, protein, carbohydrates, mannoproteins and catalase has been showed to accumulate when the yeast cells were irradiated with a frequency f = 53.33 GHz for 10 minutes. High degree of dependence between the content of cellular components (a correlation coefficient between R2 = 0.875 and 0.926 it has been shown which demonstrates that biosynthetic processes were influenced by the same phenomenon - millimeter waves. A procedure for increasing of mannoprotein content in yeasts with the utilization of extremely high frequency waves has been proposed in this study.

  16. Engineering precursor supply in Saccharomyces cerevisiae: new strategies for cytosolic acetyl-CoA formation

    OpenAIRE

    Kozak, B.U.

    2015-01-01

    Metabolic engineering – the improvement and addition, by genetic modification, of industrially relevant properties of microorganisms with respect to catalysis, transport and regulatory functions – is a well-established method for development of more cost-effective and ‘green’ industrial processes. Rapid depletion of oil reserves and a growing demand for sustainable, environmentally friendly processes provide incentives for efficient exploitation of new, renewable resources for the production ...

  17. Directed evolution of a cellobiose utilization pathway in Saccharomyces cerevisiae by simultaneously engineering multiple proteins

    OpenAIRE

    Eriksen, Dawn T.; Hsieh, Pei Chiun Helen; Lynn, Patrick; Zhao, Huimin

    2013-01-01

    Background The optimization of metabolic pathways is critical for efficient and economical production of biofuels and specialty chemicals. One such significant pathway is the cellobiose utilization pathway, identified as a promising route in biomass utilization. Here we describe the optimization of cellobiose consumption and ethanol productivity by simultaneously engineering both proteins of the pathway, the β-glucosidase (gh1-1) and the cellodextrin transporter (cdt-1), in an example of path...

  18. Coordinated transcription factor and promoter engineering to establish strong expression elements in Saccharomyces cerevisiae.

    Science.gov (United States)

    Leavitt, John M; Tong, Alice; Tong, Joyce; Pattie, Jonathan; Alper, Hal S

    2016-07-01

    Gene expression requires the coordination of trans-acting factors and cis-DNA elements to initiate transcription. Here we present a coordinated approach that combines cis-acting element engineering with mutant trans-acting factors to engineer yeast promoters. Specifically, we first construct a hybrid promoter based on the ARO9 upstream region that exhibits high constitutive and inducible expression with respect to exogenous tryptophan. Next, we perform protein engineering to identify a mutant Aro80p that affords both high constitutive expression while retaining inducible traits. We then use this mutant trans-acting factor to drive expression and generate ultra-strong promoters with transcriptional output roughly 2 fold higher than TDH3 (GPD), one of the strongest promoters to-date. Finally, we used this element to construct a modular expression system capable of staged outputs resulting in a system with nearly 6-fold, 12-fold and 15-fold expression relative to the off-state. This work further highlights the potential of using endogenous transcription factors (including mutant factors) along with hybrid promoters to expand the yeast synthetic biology toolbox. PMID:27152757

  19. Switching the mode of sucrose utilization by Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Miletti Luiz C

    2008-02-01

    Full Text Available Abstract Background Overflow metabolism is an undesirable characteristic of aerobic cultures of Saccharomyces cerevisiae during biomass-directed processes. It results from elevated sugar consumption rates that cause a high substrate conversion to ethanol and other bi-products, severely affecting cell physiology, bioprocess performance, and biomass yields. Fed-batch culture, where sucrose consumption rates are controlled by the external addition of sugar aiming at its low concentrations in the fermentor, is the classical bioprocessing alternative to prevent sugar fermentation by yeasts. However, fed-batch fermentations present drawbacks that could be overcome by simpler batch cultures at relatively high (e.g. 20 g/L initial sugar concentrations. In this study, a S. cerevisiae strain lacking invertase activity was engineered to transport sucrose into the cells through a low-affinity and low-capacity sucrose-H+ symport activity, and the growth kinetics and biomass yields on sucrose analyzed using simple batch cultures. Results We have deleted from the genome of a S. cerevisiae strain lacking invertase the high-affinity sucrose-H+ symporter encoded by the AGT1 gene. This strain could still grow efficiently on sucrose due to a low-affinity and low-capacity sucrose-H+ symport activity mediated by the MALx1 maltose permeases, and its further intracellular hydrolysis by cytoplasmic maltases. Although sucrose consumption by this engineered yeast strain was slower than with the parental yeast strain, the cells grew efficiently on sucrose due to an increased respiration of the carbon source. Consequently, this engineered yeast strain produced less ethanol and 1.5 to 2 times more biomass when cultivated in simple batch mode using 20 g/L sucrose as the carbon source. Conclusion Higher cell densities during batch cultures on 20 g/L sucrose were achieved by using a S. cerevisiae strain engineered in the sucrose uptake system. Such result was accomplished by

  20. Global mRNA expression analysis in myosin II deficient strains of Saccharomyces cerevisiae reveals an impairment of cell integrity functions

    Directory of Open Access Journals (Sweden)

    Rivera-Molina Félix E

    2008-01-01

    Full Text Available Abstract Background The Saccharomyces cerevisiae MYO1 gene encodes the myosin II heavy chain (Myo1p, a protein required for normal cytokinesis in budding yeast. Myo1p deficiency in yeast (myo1Δ causes a cell separation defect characterized by the formation of attached cells, yet it also causes abnormal budding patterns, formation of enlarged and elongated cells, increased osmotic sensitivity, delocalized chitin deposition, increased chitin synthesis, and hypersensitivity to the chitin synthase III inhibitor Nikkomycin Z. To determine how differential expression of genes is related to these diverse cell wall phenotypes, we analyzed the global mRNA expression profile of myo1Δ strains. Results Global mRNA expression profiles of myo1Δ strains and their corresponding wild type controls were obtained by hybridization to yeast oligonucleotide microarrays. Results for selected genes were confirmed by real time RT-PCR. A total of 547 differentially expressed genes (p ≤ 0.01 were identified with 263 up regulated and 284 down regulated genes in the myo1Δ strains. Gene set enrichment analysis revealed the significant over-representation of genes in the protein biosynthesis and stress response categories. The SLT2/MPK1 gene was up regulated in the microarray, and a myo1Δslt2Δ double mutant was non-viable. Overexpression of ribosomal protein genes RPL30 and RPS31 suppressed the hypersensitivity to Nikkomycin Z and increased the levels of phosphorylated Slt2p in myo1Δ strains. Increased levels of phosphorylated Slt2p were also observed in wild type strains under these conditions. Conclusion Following this analysis of global mRNA expression in yeast myo1Δ strains, we conclude that 547 genes were differentially regulated in myo1Δ strains and that the stress response and protein biosynthesis gene categories were coordinately regulated in this mutant. The SLT2/MPK1 gene was confirmed to be essential for myo1Δ strain viability, supporting that the up

  1. A shuttle vector series for precise genetic engineering of Saccharomyces cerevisiae.

    Science.gov (United States)

    Gnügge, Robert; Liphardt, Thomas; Rudolf, Fabian

    2016-03-01

    Shuttle vectors allow for an efficient transfer of recombinant DNA into yeast cells and are widely used in fundamental research and biotechnology. While available shuttle vectors are applicable in many experimental settings, their use in quantitative biology is hampered by insufficient copy number control. Moreover, they often have practical constraints, such as limited modularity and few unique restriction sites. We constructed the pRG shuttle vector series, consisting of single- and multi-copy integrative, centromeric and episomal plasmids with marker genes for the selection in all commonly used auxotrophic yeast strains. The vectors feature a modular design and a large number of unique restriction sites, enabling an efficient exchange of every vector part and expansion of the series. Integration into the host genome is achieved using a double-crossover recombination mechanism, resulting in stable single- and multi-copy modifications. As centromeric and episomal plasmids give rise to a heterogeneous cell population, an analysis of their copy number distribution and loss behaviour was performed. Overall, the shuttle vector series supports the efficient cloning of genes and their maintenance in yeast cells with improved copy number control. PMID:26647923

  2. Construction of an environmental safe Bacillus thuringiensis engineered strain against Coleoptera.

    Science.gov (United States)

    Yu, Yajun; Yuan, Yihui; Gao, Meiying

    2016-05-01

    Cloning of new toxic genes from Bacillus thuringiensis (Bt) and construction of Bt engineered strains are two key strategies for bio-control of coleopteran pests in agriculture and forestry. In this study, we cloned a new cry3Aa-type gene, cry3Aa8, from wild Bt strain YC-03 against coleopteran, and constructed a Bt engineered strain, ACE-38, containing insecticidal protein-encoding gene cry3Aa8. The engineered strain, with almost four times of Cry3Aa yield compared with strain YC-03, was an antibiotic marker-free strain. Though no selective pressure was presented in the medium, cry3Aa8 in the engineered strain ACE-38 remained stable. The yield of Cry3Aa by strain ACE-38 reached 2.09 mg/ml in the optimized fermentation medium. The activity of strain ACE-38 against Plagiodera versicolora was tested, and the LC50 of ACE-38 cultures in the optimized fermentation medium was 1.13 μl/ml. Strain ACE-38 is a non-antibiotic Bt engineered strain with high Chrysomelidae toxicity and exhibits good fermentation property. The modified indigenous site-specific recombination system constructed in this study might be useful for the construction of Bt engineered strains containing genes that cannot be expressed in the indigenous site-specific recombination system using plasmid pBMB1205R. PMID:26767987

  3. Bioprospecting and evolving alternative xylose and arabinose pathway enzymes for use in Saccharomyces cerevisiae.

    Science.gov (United States)

    Lee, Sun-Mi; Jellison, Taylor; Alper, Hal S

    2016-03-01

    Bioprospecting is an effective way to find novel enzymes from strains with desirable phenotypes. Such bioprospecting has enabled organisms such as Saccharomyces cerevisiae to utilize nonnative pentose sugars. Yet, the efficiency of this pentose catabolism (especially for the case of arabinose) remains suboptimal. Thus, further pathway optimization or identification of novel, optimal pathways is needed. Previously, we identified a novel set of xylan catabolic pathway enzymes from a superior pentose-utilizing strain of Ustilago bevomyces. These enzymes were used to successfully engineer a xylan-utilizing S. cerevisiae through a blended approach of bioprospecting and evolutionary engineering. Here, we expanded this approach to xylose and arabinose catabolic pathway engineering and demonstrated that bioprospected xylose and arabinose catabolic pathways from U. bevomyces offer alternative choices for enabling efficient pentose catabolism in S. cerevisiae. By introducing a novel set of xylose catabolic genes from U. bevomyces, growth rates were improved up to 85 % over a set of traditional Scheffersomyces stipitis pathway genes. In addition, we suggested an alternative arabinose catabolic pathway which, after directed evolution and pathway engineering, enabled S. cerevisiae to grow on arabinose as a sole carbon source in minimal medium with growth rates upwards of 0.05 h(-1). This pathway represents the most efficient growth of yeast on pure arabinose minimal medium. These pathways provide great starting points for further strain development and demonstrate the utility of bioprospecting from U. bevomyces. PMID:26671616

  4. Identification of MET10-932 and characterization as an allele reducing hydrogen sulfide formation in wine strains of Saccharomyces cerevisiae.

    Science.gov (United States)

    Linderholm, Angela; Dietzel, Kevin; Hirst, Marissa; Bisson, Linda F

    2010-12-01

    A vineyard isolate of the yeast Saccharomyces cerevisiae, UCD932, was identified as a strain producing little or no detectable hydrogen sulfide during wine fermentation. Genetic analysis revealed that this trait segregated as a single genetic determinant. The gene also conferred a white colony phenotype on BiGGY agar (bismuth-glucose-glycine-yeast agar), which is thought to indicate low basal levels of sulfite reductase activity. However, this isolate does not display a requirement for S-containing amino acids, indicating that the sulfate reduction pathway is fully operational. Genetic crosses against known mutations conferring white colony color on BiGGY agar identified the gene leading to reduced H(2)S formation as an allele of MET10 (MET10-932), which encodes a catalytic subunit of sulfite reductase. Sequence analysis of MET10-932 revealed several corresponding amino acid differences in relation to laboratory strain S288C. Allele differences for other genes of the sulfate reduction pathway were also detected in UCD932. The MET10 allele of UCD932 was found to be unique in comparison to the sequences of several other vineyard isolates with differing levels of production of H(2)S. Replacing the MET10 allele of high-H(2)S-producing strains with MET10-932 prevented H(2)S formation by those strains. A single mutative change, corresponding to T662K, in MET10-932 resulted in a loss of H(2)S production. The role of site 662 in sulfide reduction was further analyzed by changing the encoded amino acid at this position. A change back to threonine or to the conservative serine fully restored the H(2)S formation conferred by this allele. In addition to T662K, arginine, tryptophan, and glutamic acid substitutions similarly reduced sulfide formation. PMID:20889780

  5. A novel wild-type Saccharomyces cerevisiae strain TSH1 in scaling-up of solid-state fermentation of ethanol from sweet sorghum stalks.

    Directory of Open Access Journals (Sweden)

    Ran Du

    Full Text Available The rising demand for bioethanol, the most common alternative to petroleum-derived fuel used worldwide, has encouraged a feedstock shift to non-food crops to reduce the competition for resources between food and energy production. Sweet sorghum has become one of the most promising non-food energy crops because of its high output and strong adaptive ability. However, the means by which sweet sorghum stalks can be cost-effectively utilized for ethanol fermentation in large-scale industrial production and commercialization remains unclear. In this study, we identified a novel Saccharomyces cerevisiae strain, TSH1, from the soil in which sweet sorghum stalks were stored. This strain exhibited excellent ethanol fermentative capacity and ability to withstand stressful solid-state fermentation conditions. Furthermore, we gradually scaled up from a 500-mL flask to a 127-m3 rotary-drum fermenter and eventually constructed a 550-m3 rotary-drum fermentation system to establish an efficient industrial fermentation platform based on TSH1. The batch fermentations were completed in less than 20 hours, with up to 96 tons of crushed sweet sorghum stalks in the 550-m3 fermenter reaching 88% of relative theoretical ethanol yield (RTEY. These results collectively demonstrate that ethanol solid-state fermentation technology can be a highly efficient and low-cost solution for utilizing sweet sorghum, providing a feasible and economical means of developing non-food bioethanol.

  6. CasEMBLR: Cas9-Facilitated Multiloci Genomic Integration of in Vivo Assembled DNA Parts in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Jakociunas, Tadas; Rajkumar, Arun Stephen; Zhang, Jie;

    2015-01-01

    Homologous recombination (HR) in Saccharomyces cerevisiae has been harnessed for both plasmid construction and chromosomal integration of foreign DNA. Still, native HR machinery is not efficient enough for complex and marker-free genome engineering required for modern metabolic engineering. Here...... method CasEMBLR and validate its applicability for genome engineering and cell factory development in two ways: (i) introduction of the carotenoid pathway from 15 DNA parts into three targeted loci, and (ii) creation of a tyrosine production strain using ten parts into two loci, simultaneously knocking...... out two genes. This method complements and improves the current set of tools available for genome engineering in S. cerevisiae....

  7. Analysis of the secondary compounds produced by Saccharomyces cerevisiae and wild yeast strains during the production of "cachaça" Análise dos componentes secundários produzidos por Saccharomyces cerevisiae e leveduras selvagens durante a produção de cachaça

    OpenAIRE

    Maria Cecília Fachine Dato; João Martins Pizauro Júnior; Márcia Justino Rossini Mutton

    2005-01-01

    The aim of this study is to compare the composition of "cachaças" produced in 10 fermentation cycles by Saccharomyces cerevisiae (Sc) and wild yeast strains [Pichia silvicola (Ps), Pichia anomala 1 (Pa1), Pichia anomala 2 (Pa2) and Dekkera bruxelensis (Db)], isolated from distilleries in Jaboticabal - SP, Brazil. The secondary components of the heart fraction were determined by gas chromatography. The levels of secondary components were influenced by the wine pH, which varied among yeast stra...

  8. Analysis of the secondary compounds produced by Saccharomyces cerevisiae and wild yeast strains during the production of "cachaça" Análise dos componentes secundários produzidos por Saccharomyces cerevisiae e leveduras selvagens durante a produção de cachaça

    Directory of Open Access Journals (Sweden)

    Maria Cecília Fachine Dato

    2005-03-01

    Full Text Available The aim of this study is to compare the composition of "cachaças" produced in 10 fermentation cycles by Saccharomyces cerevisiae (Sc and wild yeast strains [Pichia silvicola (Ps, Pichia anomala 1 (Pa1, Pichia anomala 2 (Pa2 and Dekkera bruxelensis (Db], isolated from distilleries in Jaboticabal - SP, Brazil. The secondary components of the heart fraction were determined by gas chromatography. The levels of secondary components were influenced by the wine pH, which varied among yeast strains. S. cerevisiae showed slightly more secondary components, whereas wild strains produced more higher alcohols. Wild yeast strains were shown to be adequate for the production of a high quality "cachaça".O presente trabalho visou estabelecer uma comparação entre composição de cachaças produzidas por Saccharomyces cerevisiae (Sc e estirpes de leveduras selvagens [Pichia silvicola (Ps, Pichia anomala 1 (Pa1, Pichia anomala 2 (Pa2 e Dekkera bruxelensis (Db], isoladas em destilarias da região de Jaboticabal-SP. Os componentes secundários da fração denominada coração foram determinados por cromatografia gasosa. Os níveis dos componentes secundários foram influenciados pelo pH dos respectivos vinhos, os quais dependem da estirpe de levedura empregada no processo fermentativo. A Saccharomyces cerevisiae apresentou valores ligeiramente superiores de componentes secundários, enquanto as estirpes selvagens produziram maiores teores de álcoois superiores. As estirpes selvagens de leveduras mostraram-se adequadas para obtenção de uma cachaça de boa qualidade.

  9. Engineering cytosolic acetyl-coenzyme A supply in Saccharomyces cerevisiae: Pathway stoichiometry, free-energy conservation and redox-cofactor balancing.

    Science.gov (United States)

    van Rossum, Harmen M; Kozak, Barbara U; Pronk, Jack T; van Maris, Antonius J A

    2016-07-01

    Saccharomyces cerevisiae is an important industrial cell factory and an attractive experimental model for evaluating novel metabolic engineering strategies. Many current and potential products of this yeast require acetyl coenzyme A (acetyl-CoA) as a precursor and pathways towards these products are generally expressed in its cytosol. The native S. cerevisiae pathway for production of cytosolic acetyl-CoA consumes 2 ATP equivalents in the acetyl-CoA synthetase reaction. Catabolism of additional sugar substrate, which may be required to generate this ATP, negatively affects product yields. Here, we review alternative pathways that can be engineered into yeast to optimize supply of cytosolic acetyl-CoA as a precursor for product formation. Particular attention is paid to reaction stoichiometry, free-energy conservation and redox-cofactor balancing of alternative pathways for acetyl-CoA synthesis from glucose. A theoretical analysis of maximally attainable yields on glucose of four compounds (n-butanol, citric acid, palmitic acid and farnesene) showed a strong product dependency of the optimal pathway configuration for acetyl-CoA synthesis. Moreover, this analysis showed that combination of different acetyl-CoA production pathways may be required to achieve optimal product yields. This review underlines that an integral analysis of energy coupling and redox-cofactor balancing in precursor-supply and product-formation pathways is crucial for the design of efficient cell factories. PMID:27016336

  10. Conclusion on the peer review of the pesticide risk assessment of the active substance cerevisane (cell walls of Saccharomyces cerevisiae strain LAS117

    Directory of Open Access Journals (Sweden)

    European Food Safety Authority

    2014-06-01

    Full Text Available The conclusions of the European Food Safety Authority (EFSA following the peer review of the initial risk assessments carried out by the competent authority of the rapporteur Member State France for the pesticide active substance cerevisane (cell walls of Saccharomyces cerevisiae strain LAS117, and the assessment of the proposal for inclusion of the substance in Annex IV of Regulation (EC No 396/2005, are reported. The context of the peer review was that required by Regulation (EC No 1107/2009 of the European Parliament and of the Council. The conclusions were reached on the basis of the evaluation of the representative uses of the active substance as a systemic resistance inducer against fungi and bacteria in lettuce and other salad crops. The reliable endpoints concluded as being appropriate for use in regulatory risk assessment, derived from the available studies and literature in the dossier peer reviewed, are presented. Missing information identified as being required by the regulatory framework is listed. No concerns are identified.

  11. Enhanced ethanol production, volatile compound biosynthesis and fungicide removal during growth of a newly isolated Saccharomyces cerevisiae strain on enriched pasteurized grape musts

    Energy Technology Data Exchange (ETDEWEB)

    Sarris, Dimitris; Kotseridis, Yorgos; Galiotou-Panayotou, Maria; Papanikolaou, Seraphim [Department of Food Science and Technology, Agricultural University of Athens (Greece); Linga, Maria [Oinognosia, Wine analysis and consulting, Kiato (Greece)

    2009-02-15

    The kinetic behavior of a newly isolated Saccharomyces cerevisiae strain, grown on pasteurized grape musts enriched with industrial sugars, was studied after the addition of various concentrations [0.0 (reference), 0.4 and 2.4 mg/L] of the fungicide quinoxyfen to the medium. Batch-flask cultures were carried out. Significant quantities of biomass (10.0{+-}0.8 g/L) were produced regardless of quinoxyfen addition to the medium; therefore, the addition of the fungicide did not seriously inhibit biomass production. Ethanol was synthesized in very high quantities in all trials (highest concentrations 106.4-119.2 g/L). A slight decrease of ethanol production in terms of both absolute value and conversion yield of ethanol produced per sugar consumed was, however, observed when the quinoxyfen concentration was increased. The addition of quinoxyfen led to significantly lower ethylic ester levels, which also pertains to the acetates analyzed in this study. Fusel alcohol synthesis seemed to be activated when 0.4 mg/L quinoxyfen was added, but at 2.4 mg/L of added fungicide, no statistically significant differences were observed compared with the control trial. Volatile acid levels did not present a uniform trend in relation with the added fungicide. Finally, the fermentation was accompanied by a significant reduction of the fungicide concentration (79-82 wt% fungicide removal). (Abstract Copyright [2009], Wiley Periodicals, Inc.)

  12. Increased mannosylphosphorylation of N-glycans by heterologous expression of YlMPO1 in glyco-engineered Saccharomyces cerevisiae for mannose-6-phosphate modification.

    Science.gov (United States)

    Gil, Jin Young; Park, Jeong-Nam; Lee, Kyung Jin; Kang, Ji-Yeon; Kim, Yeong Hun; Kim, Seonghun; Kim, Sang-Yoon; Kwon, Ohsuk; Lim, Yong Taik; Kang, Hyun Ah; Oh, Doo-Byoung

    2015-07-20

    Mannosylphosphorylated N-glycans found in yeasts can be converted to those containing mannose-6-phosphate, which is a key factor for lysosomal targeting. In the traditional yeast Saccharomyces cerevisiae, both ScMNN4 and ScMNN6 genes are required for efficient mannosylphosphorylation. ScMnn4 protein has been known to be a positive regulator of ScMnn6p, a real enzyme for mannosylphosphorylation. On the other hand, YlMpo1p, a ScMnn4p homologue, mediates mannosylphosphorylation in Yarrowia lypolytica without the involvement of ScMnn6p homologues. In this study, we show that heterologous expression of YlMpo1p can perform and enhance mannosylphosphorylation in S. cerevisiae in the absence of ScMnn4p and ScMnn6p. Moreover, the level of mannosylphosphorylation of N-glycans enhanced by YlMpo1p overexpression is much higher than that with ScMnn4p overexpression, and this is highlighted further in Scmnn4- and Scmnn6-disrupted mutants. When YlMpo1p overexpression is applied to glyco-engineered S. cerevisiae in which the synthesis of immunogenic glycans is abolished, a great increase of bi-mannosylphosphorylated glycan is observed. Through an in vitro process involving the uncapping of the outer mannose residue, this bi-mannosylphosphorylated structure is changed to a bi-phosphorylated structure with high affinity for mannose-6-phosphate receptor. The superior ability of YlMpo1p to increase bi-mannosylphosphorylated glycan in yeast shows promise for the production of therapeutic enzymes with improved lysosomal targeting capability. PMID:25907834

  13. Efeitos do cádmio sobre o crescimento das leveduras Saccharomyces cerevisiae PE-2 e Saccharomyces cerevisiae IZ-1904, e a capacidade da vinhaça em atenuar a toxicidade Effect of cadmium on the growth of two Saccharomyces cerevisiae strains, and the vinasse capacity to atenuate the toxicity

    Directory of Open Access Journals (Sweden)

    Samuel Mariano-da-Silva

    2004-03-01

    , minimizando os efeitos deletérios do metal.The present study was carried out in order to evaluate the capability of different cadmium concentration (0; 0,05, 0,10 and 0,50mM to affect the growth of two S. cerevisiae strains (PE-2 and IZ-1904 in YED (yeast extract 1% and dextrose 2% medium, and to evaluate the three vinasse concentration capability (0,15 and 30% to attenuate the two cadmium concentration toxicity (0,1 and 0,5mM, using S. cerevisiae PE-2 strain in YED medium. In the first assay, the medium was inoculated in aseptic conditions with 1mL of 1% yeast suspension (PE-2 or IZ-1904 and incubated at 30ºC, 70 RPM for 18 hours. During anaerobic growth (0, 2, 4, 6, 8, 10, 12, 14, 16 and 18 hours, portions of cell suspension were taken out and biomass concentration was determined. At the end of fermentation, yeast viability, budding rate and bacterial contamination were determined. Both, initial and final trehalose, was measured. In the second assay, the medium was inoculated in aseptic conditions with 2mL of 1% PE-2 suspension and incubated at 30ºC, 120 rpm for 18 hours. During the anaerobic growth (0, 2, 4, 6, 8, 10, 12,14, 16 and 18 hours portions of cell suspension were taken out and biomass concentration was determined. At the end of fermentation, alcohol production, yeast viability, budding rate and bacterial contamination were determined. Both, initial and final trehalose, was measured. The increase of cadmium levels showed a reduction on yeast growth and cell viability. Vinasse showed low toxicity, but protected yeast cells very effectively against the toxic effects of cadmium.

  14. Strain engineering of diamond silicon vacancy centers in MEMS cantilevers

    Science.gov (United States)

    Meesala, Srujan; Sohn, Young-Ik; Atikian, Haig; Holzgrafe, Jeffrey; Zhang, Mian; Burek, Michael; Loncar, Marko

    2016-05-01

    The silicon vacancy (SiV) center in diamond has recently attracted attention as a solid state quantum emitter due to its attractive optical properties. We fabricate diamond MEMS cantilevers, and use electrostatic actuation to apply controlled strain fields to single SiV centers implanted in these devices. The strain response of the four electronic transitions of the SiV at 737 nm is measured via cryogenic (4 K) photoluminescence excitation. We demonstrate over 300 GHz of tuning for the mean transition frequency between the ground and excited states, and over 100 GHz of tuning for the orbital splittings within the ground and excited states. The interaction Hamiltonian for strain fields is inferred, and large strain susceptibilities of the order 1 PHz/strain are measured. We discuss prospects to utilize our device to reduce phonon-induced decoherence in SiV spin qubits, and to exploit the large strain susceptibilities for hybrid quantum systems based on nanomechanical resonators.

  15. Mechanical Control of Graphene on Engineered Pyramidal Strain Arrays

    OpenAIRE

    Gill, Stephen T.; Hinnefeld, John H.; Zhu, Shuze; Swanson, William T.; Li, Teng; Mason, Nadya

    2015-01-01

    Strain can tune desirable electronic behavior in graphene, but there has been limited progress in controlling strain in graphene devices. In this paper, we study the mechanical response of graphene on substrates patterned with arrays of mesoscale pyramids. Using atomic force microscopy, we demonstrate that the morphology of graphene can be controlled from conformal to suspended depending on the arrangement of pyramids and the aspect ratio of the array. Non-uniform strains in graphene suspende...

  16. Strain engineering of electronic properties of transition metal dichalcogenide monolayers

    Science.gov (United States)

    Maniadaki, Aristea E.; Kopidakis, Georgios; Remediakis, Ioannis N.

    2016-02-01

    We present Density Functional Theory (DFT) results for the electronic and dielectric properties of single-layer (2D) semiconducting transition metal dichalcogenides MX2 (M=Mo, W; X=S, Se, Te) under isotropic, uniaxial (along the zigzag and armchair directions), and shear strain. Electronic band gaps decrease while dielectric constants increase for heavier chalcogens X. The direct gaps of equilibrium structures often become indirect under certain types of strain, depending on the material. The effects of strain and of broken symmetry on the band structure are discussed. Gaps reach maximum values at small compressive strains or in equilibrium, and decrease with larger strains. In-plane dielectric constants generally increase with strain, reaching a minimum value at small compressive strains. The out-of-plane constants exhibit a similar behavior under shear strain but under isotropic and uniaxial strain they increase with compression and decrease with tension, thus exhibiting a monotonic behavior. These DFT results are theoretically explained using only structural parameters and equilibrium dielectric constants. Our findings are consistent with available experimental data.

  17. Effects on Anthocyanin Composition in Red Wines Fermented by Different Saccharomyces cerevisiae Strains%不同酿酒酵母发酵对红葡萄酒中花色苷组成的影响

    Institute of Scientific and Technical Information of China (English)

    李双石; 苏宁; 吴志明; 李淳; 兰蓉

    2012-01-01

    To explore wild yeast resources in China, the anthocyanin composition and content in the red wines fermented by different Saccharomyces cerevisiae strains were compared. Two different Saccharomyces cerevisiae strains (indigenous Saccharomyces cerevisiae strain Y17, commercial active dry yeast F15) were separately used in red wines production. The anthocyanin components in wines were studied by high performance liquid chromatography-mass spec- trometry (HPLC-MS). The results showed that the diversities of anthocyanins in the wines fermented by different yeast strains were similar, but the contents of different anthocyanins in the wines were quite different. The indigenous yeast strain Y17 produced wines with higher concentrations of anthocyanins.%比较了不同酿酒酵母菌株发酵对红葡萄酒中花色苷组分的影响,为本土酵母的开发应用提供依据。采用本土自筛酿酒酵母菌株Y17和进口商业酵母菌株F15分别酿造干红葡萄酒,并利用高效液相色谱-质谱联用技术(HPLC-MS)对葡萄酒中的花色苷成分进行检测。结果表明,不同酵母发酵葡萄酒中的花色苷成分种类大体相同,但各种花色苷的含量具有较大差别,本土酵母菌株Y17在发酵生产高花色苷含量葡萄酒方面具有优势。

  18. Sensitivities to monochromatic 254-nm and 365-nm radiation of closely related strains of Saccharomyces cerevisiae with differing repair capabilities

    International Nuclear Information System (INIS)

    Sensitivity to monochromatic 254- and 365-nm radiation was compared in closely related yeast strains with defects in one or more of the excision-repair (rad1), error-prone repair (rad18), or recombinational-repair (rad51) pathways. At 254 nm, mutants defective in a single repair pathway exhibited slight to moderate UV sensitivity;,those defective in two separate pathways were somewhat more UV sensitive, while triple mutants defective in all three pathways exhibited extreme UV sensitivity with a lethal event corresponding to 0.05 J m-2. Repair defects also rendered mutants sensitive to 365-nm radiation: strains with single defects exhibited slight sensitivity, mutants with two defective pathways were more sensitive, and triple mutants exhibited maximal sensitivity with a lethal event corresponding to 2.4 x 104 J m-2. Evidence for dimer involvement in the yeast mutant was obtained by demonstrating that lethality at both 254 and 365 nm was photoreactivated by light at 405 nm. (author)

  19. Proteomics and redox-proteomics of the effects of herbicides on a wild-type wine Saccharomyces cerevisiae strain.

    Science.gov (United States)

    Braconi, Daniela; Bernardini, Giulia; Possenti, Silvia; Laschi, Marcella; Arena, Simona; Scaloni, Andrea; Geminiani, Michela; Sotgiu, Michele; Santucci, Annalisa

    2009-01-01

    Several toxicological and environmental problems are associated with the extensive use of agricultural pesticides, such as herbicides. Nevertheless, little is known about the toxic effects of formulated herbicides, since many studies have been carried out using pure active molecules alone. In this work, we used as an eukaryotic model system an autochthonous wine yeast strain to investigate the effects of three commercial herbicides, currently used in the same geographical area from where this strain had been isolated. We carried out a comparative proteomic analysis to study the effects at the protein level of the herbicide-related stress, and found that the herbicides tested can alter the yeast proteome producing responses that share homologies with those observed treating yeast cells with the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) or with well-known oxidizing agents. We evaluated, through redox-proteomic techniques, protein carbonylation as a biomarker of oxidative stress. This analysis showed that herbicide-induced carbonylation is a dynamic phenomenon with degrees of selectivity. PMID:19032026

  20. Strain engineering of WS2, WSe2, and WTe2

    KAUST Repository

    Amin, Bin

    2014-01-01

    We perform first-principles calculations to investigate the structural, electronic, and vibrational properties of WS2, WSe2, and WTe2 monolayers, taking into account the strong spin orbit coupling. A transition from a direct to an indirect band gap is achieved for compressive strain of 1% for WS2, 1.5% for WSe2, and 2% for WTe 2, while the nature of the band gap remains direct in the case of tensile strain. The size of the band gap passes through a maximum under compressive strain and decreases monotonically under tensile strain. A strong spin splitting is found for the valence band in all three compounds, which is further enhanced by tensile strain. The mobility of the electrons grows along the series WS2 < WSe2 < WTe2. This journal is © the Partner Organisations 2014.

  1. Performance of the auxotrophic Saccharomyces cerevisiae BY4741 as host for the production of IL-1β in aerated fed-batch reactor: role of ACA supplementation, strain viability, and maintenance energy

    Directory of Open Access Journals (Sweden)

    Zueco Jesus

    2009-12-01

    Full Text Available Abstract Background Saccharomyces cerevisiae BY4741 is an auxotrophic commonly used strain. In this work it has been used as host for the expression and secretion of human interleukin-1β (IL1β, using the cell wall protein Pir4 as fusion partner. To achieve high cell density and, consequently, high product yield, BY4741 [PIR4-IL1β] was cultured in an aerated fed-batch reactor, using a defined mineral medium supplemented with casamino acids as ACA (auxotrophy-complementing amino acid source. Also the S. cerevisiae mutant BY4741 Δyca1 [PIR4-IL1β], carrying the deletion of the YCA1 gene coding for a caspase-like protein involved in the apoptotic response, was cultured in aerated fed-batch reactor and compared to the parental strain, to test the effect of this mutation on strain robustness. Viability of the producer strains was examined during the runs and a mathematical model, which took into consideration the viable biomass present in the reactor and the glucose consumption for both growth and maintenance, was developed to describe and explain the time-course evolution of the process for both, the BY4741 parental and the BY4741 Δyca1 mutant strain. Results Our results show that the concentrations of ACA in the feeding solution, corresponding to those routinely used in the literature, are limiting for the growth of S. cerevisiae BY4741 [PIR4-IL1β] in fed-batch reactor. Even in the presence of a proper ACA supplementation, S. cerevisiae BY4741 [PIR4-IL1β] did not achieve a high cell density. The Δyca1 deletion did not have a beneficial effect on the overall performance of the strain, but it had a clear effect on its viability, which was not impaired during fed-batch operations, as shown by the kd value (0.0045 h-1, negligible if compared to that of the parental strain (0.028 h-1. However, independently of their robustness, both the parental and the Δyca1 mutant ceased to grow early during fed-batch runs, both strains using most of the

  2. Effect of Engineering Character on Stress-Strain Relationship in Post-Peak Area

    Institute of Scientific and Technical Information of China (English)

    TANG Lei; KE Min-yong; YAN Jian-hua

    2003-01-01

    Constitutive experiments are the base of all rock mechanics works. The effect of engineering character on constitutive law is a new problem of rock mechanics. The results of series specimens based on the uniaxial and plane strain compression experiments were presented and discussed. It is found that engineering or experiment character has obvious effects on stress-strain relationship and especially on mechanic parameters in post-peak area. And the law of size effect of softening materials was also discussed.

  3. Modulation of Chaperone Gene Expression in Mutagenized Saccharomyces cerevisiae Strains Developed for Recombinant Human Albumin Production Results in Increased Production of Multiple Heterologous Proteins▿

    OpenAIRE

    Payne, T.; Finnis, C.; Evans, L R; Mead, D. J.; Avery, S V; Archer, D. B.; Sleep, D.

    2008-01-01

    The yeast Saccharomyces cerevisiae has been successfully established as a commercially viable system for the production of recombinant proteins. Manipulation of chaperone gene expression has been utilized extensively to increase recombinant protein production from S. cerevisiae, focusing predominantly on the products of the protein disulfide isomerase gene PDI1 and the hsp70 gene KAR2. Here we show that the expression of the genes SIL1, LHS1, JEM1, and SCJ1, all of which are involved in regul...

  4. Strain-displacement relations for strain engineering in single-layer 2d materials

    Science.gov (United States)

    Midtvedt, Daniel; Lewenkopf, Caio H.; Croy, Alexander

    2016-03-01

    We investigate the electromechanical coupling in single-layer 2d materials. For non-Bravais lattices, we find important corrections to the standard macroscopic strain-microscopic atomic-displacement theory. We put forward a general and systematic approach to calculate strain-displacement relations for several classes of 2d materials. We apply our findings to graphene as a study case, by combining a tight binding and a valence force-field model to calculate electronic and mechanical properties of graphene nanoribbons under strain. The results show good agreement with the predictions of the Dirac equation coupled to continuum mechanics. For this long wave-limit effective theory, we find that the strain-displacement relations lead to a renormalization correction to the strain-induced pseudo-magnetic fields. A similar renormalization is found for the strain-induced band-gap of black phosphorous. Implications for nanomechanical properties and electromechanical coupling in 2d materials are discussed.

  5. Protein engineering of cyclodextrin glycosyltransferase from Bacillus circulans strain 251

    OpenAIRE

    Penninga, Dirk

    1996-01-01

    An enormous diversity of molecular functions in living organisms is carried out by proteins. Our studies have focussed on the functional analysis of a starch-converting enzyme, cyclodextrin glycosyltransferase (CGTase) from Bacillus circulans strain 251. Zie: Summary

  6. Evolved hexose transporter enhances xylose uptake and glucose/xylose co-utilization in Saccharomyces cerevisiae.

    Science.gov (United States)

    Reider Apel, Amanda; Ouellet, Mario; Szmidt-Middleton, Heather; Keasling, Jay D; Mukhopadhyay, Aindrila

    2016-01-01

    Enhancing xylose utilization has been a major focus in Saccharomyces cerevisiae strain-engineering efforts. The incentive for these studies arises from the need to use all sugars in the typical carbon mixtures that comprise standard renewable plant-biomass-based carbon sources. While major advances have been made in developing utilization pathways, the efficient import of five carbon sugars into the cell remains an important bottleneck in this endeavor. Here we use an engineered S. cerevisiae BY4742 strain, containing an established heterologous xylose utilization pathway, and imposed a laboratory evolution regime with xylose as the sole carbon source. We obtained several evolved strains with improved growth phenotypes and evaluated the best candidate using genome resequencing. We observed remarkably few single nucleotide polymorphisms in the evolved strain, among which we confirmed a single amino acid change in the hexose transporter HXT7 coding sequence to be responsible for the evolved phenotype. The mutant HXT7(F79S) shows improved xylose uptake rates (Vmax = 186.4 ± 20.1 nmol•min(-1)•mg(-1)) that allows the S. cerevisiae strain to show significant growth with xylose as the sole carbon source, as well as partial co-utilization of glucose and xylose in a mixed sugar cultivation. PMID:26781725

  7. Nanoscale strain engineering of graphene and graphene-based devices

    OpenAIRE

    Yeh, N. -C.; Hsu, C.-C.; Teague, M. L.; Wang, J.-Q.; Boyd, D A; Chen, C.-C.

    2016-01-01

    Structural distortions in nano-materials can induce dramatic changes in their electronic properties. This situation is well manifested in graphene, a two-dimensional honeycomb structure of carbon atoms with only one atomic layer thickness. In particular, strained graphene can result in both charging effects and pseudo-magnetic fields, so that controlled strain on a perfect graphene lattice can be tailored to yield desirable electronic properties. Here, we describe the theoretical foundation f...

  8. Strain engineered optoelectronic properties of transition metal dichalcogenides lateral heterostructures

    Science.gov (United States)

    Lee, Jaekwang; Yoon, Mina

    2015-03-01

    Most three-dimensional bulk-scale materials rarely survive beyond 1% strain, while recently spotlighted two-dimensional (2-D) materials can sustain a high elastic strain (up to 10%) to optimize optical quantities such as band gaps and absorption spectra governing optoelectronic device performance. Despite the enormous interest in strained 2-D materials, most researches are focused on single materials or vertical heterostructures where precise control of stacking orientation is challenging. Here, using first-principles density-functional calculations, we explore how uniaxial tensile strains modify overall electronic and optical properties of transition metal dichalcogenides lateral heterostructures, such as MoX2/WX2 (X =S, Se). Based on the detailed optoelectronic information, we predict the optimal strain condition for maximal power efficiency. Furthermore, we find that uniaxial tensile strain readily develops a continuously varying direct-bandgap across the lateral heterojunctions, which results in the broad range absorption of solar spectrum useful for future optoelectronic devices. This research was conducted at the CNMS, which is sponsored at Oak Ridge National Laboratory (ORNL) by the Office of Basic Energy Sciences, U.S. Department of Energy; a portion of theory work was supported by the LDRD Program of ORNL.

  9. Strain engineering the work function in monolayer metal dichalcogenides

    International Nuclear Information System (INIS)

    We use first-principles density functional theory to investigate the effect of both tensile and compressive strain on the work functions of various metal dichalcogenide monolayers. We find that for all six species considered, including MoS2, WS2, SnS2, VS2, MoSe2 and MoTe2, that compressive strain of up to 10% decreases the work function continuously by as much as 1.0 eV. Large enough tensile strain is also found to decrease the work function, although in some cases we observe an increase in the work function for intermediate values of tensile strain. This work function modulation is attributed to a weakening of the chalcogenide-metal bonds and an increase in total energy of each system as a function of strain. Values of strain which bring the metal atoms closer together lead to an increase in electrostatic potential energy, which in turn results in an increase in the vacuum potential level. The net effect on the work function can be explained in terms of the balance between the increases in the vacuum potential levels and Fermi energy. (paper)

  10. Bulk segregant analysis by high-throughput sequencing reveals a novel xylose utilization gene from Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Jared W Wenger

    2010-05-01

    Full Text Available Fermentation of xylose is a fundamental requirement for the efficient production of ethanol from lignocellulosic biomass sources. Although they aggressively ferment hexoses, it has long been thought that native Saccharomyces cerevisiae strains cannot grow fermentatively or non-fermentatively on xylose. Population surveys have uncovered a few naturally occurring strains that are weakly xylose-positive, and some S. cerevisiae have been genetically engineered to ferment xylose, but no strain, either natural or engineered, has yet been reported to ferment xylose as efficiently as glucose. Here, we used a medium-throughput screen to identify Saccharomyces strains that can increase in optical density when xylose is presented as the sole carbon source. We identified 38 strains that have this xylose utilization phenotype, including strains of S. cerevisiae, other sensu stricto members, and hybrids between them. All the S. cerevisiae xylose-utilizing strains we identified are wine yeasts, and for those that could produce meiotic progeny, the xylose phenotype segregates as a single gene trait. We mapped this gene by Bulk Segregant Analysis (BSA using tiling microarrays and high-throughput sequencing. The gene is a putative xylitol dehydrogenase, which we name XDH1, and is located in the subtelomeric region of the right end of chromosome XV in a region not present in the S288c reference genome. We further characterized the xylose phenotype by performing gene expression microarrays and by genetically dissecting the endogenous Saccharomyces xylose pathway. We have demonstrated that natural S. cerevisiae yeasts are capable of utilizing xylose as the sole carbon source, characterized the genetic basis for this trait as well as the endogenous xylose utilization pathway, and demonstrated the feasibility of BSA using high-throughput sequencing.

  11. Cystathionine accumulation in Saccharomyces cerevisiae.

    OpenAIRE

    Ono, B; Suruga, T; Yamamoto, M.; Yamamoto, S.; Murata, K; Kimura, A; Shinoda, S; Ohmori, S.

    1984-01-01

    A cysteine-dependent strain of Saccharomyces cerevisiae and its prototrophic revertants accumulated cystathionine in cells. The cystathionine accumulation was caused by a single mutation having a high incidence of gene conversion. The mutation was designated cys3 and was shown to cause loss of gamma-cystathionase activity. Cysteine dependence of the initial strain was determined by two linked and interacting mutations, cys3 and cys1 . Since cys1 mutations cause a loss of serine acetyltransfer...

  12. Investigating xylose metabolism in recombinant Saccharomyces cerevisiae via 13C metabolic flux analysis

    OpenAIRE

    Feng, Xueyang; Zhao, Huimin

    2013-01-01

    Background To engineer Saccharomyces cerevisiae for efficient xylose utilization, a fungal pathway consisting of xylose reductase, xylitol dehydrogenase, and xylulose kinase is often introduced to the host strain. Despite extensive in vitro studies on the xylose pathway, the intracellular metabolism rewiring in response to the heterologous xylose pathway remains largely unknown. In this study, we applied 13C metabolic flux analysis and stoichiometric modeling to systemically investigate the f...

  13. Use of Nonionic Surfactants for Improvement of Terpene Production in Saccharomyces cerevisiae

    OpenAIRE

    Kirby, James; Nishimoto, Minobu; Chow, Ruthie W. N.; Pasumarthi, Venkata N.; Chan, Rossana; Chan, Leanne Jade G.; Petzold, Christopher J.; Keasling, Jay D.

    2014-01-01

    To facilitate enzyme and pathway engineering, a selection was developed for improved sesquiterpene titers in Saccharomyces cerevisiae. α-Bisabolene, a candidate advanced biofuel, was found to protect yeast against the disruptive action of nonionic surfactants such as Tween 20 (T20). An experiment employing competition between two strains of yeast, one of which makes twice as much bisabolene as the other, demonstrated that growth in the presence of T20 provided sufficient selective pressure to...

  14. Electronic structure and magnetism in g-C4N3 controlled by strain engineering

    International Nuclear Information System (INIS)

    Regulation of magnetism and half-metallicity has attracted much attention because of its potential in spintronics. The magnetic properties and electronic structure of graphitic carbon nitride (g-C4N3) with external strain are determined theoretically based on the density function theory and many-body perturbation theory (G0W0). Asymmetric deformation induced by uniaxial strain not only regulates the magnetic characteristics but also leads to a transformation from half-metallicity to metallicity. However, this transition cannot occur in the structure with symmetric deformation induced by biaxial strain. Our results suggest the use of strain engineering in metal-free spintronics applications

  15. Fermentation of deproteinized cheese whey powder solutions to ethanol by engineered Saccharomyces cerevisiae : effect of supplementation with corn steep liquor and repeated-batch operation with biomass recycling by flocculation

    OpenAIRE

    Silva, Ana Carina; Guimarães, Pedro M. R.; Teixeira, J.A.; Domingues, Lucília

    2010-01-01

    The lactose in cheese whey is an interesting substrate for the production of bulk commodities such as bio-ethanol, due to the large amounts of whey surplus generated globally. In this work, we studied the performance of a recombinant Saccharomyces cerevisiae strain expressing the lactose permease and intracellular ß-galactosidase from Kluyveromyces lactis in fermentations of deproteinized concentrated cheese whey powder solutions. Supplementation with 10 g/l of corn st...

  16. Strain engineered nanomembranes as anodes for lithium ion batteries

    OpenAIRE

    Deng, Junwen

    2015-01-01

    Lithium ion batteries (LIBs) have attracted considerable interest due to their wide range of applications, such as portable electronics, electric vehicles (EVs) and aerospace applications. Particularly, the emergence of a variety of nanostructured materials has driven the development of LIBs towards the next generation, which is featured with high specific energy and large power density. Herein, rolled-up nanotechnology is introduced for the design of strain-released materials as anodes of...

  17. Electrostatic engineering of strained ferroelectric perovskites from first-principles

    OpenAIRE

    Cazorla, Claudio; Stengel, Massimiliano

    2015-01-01

    Design of novel artificial materials based on ferroelectric perovskites relies on the basic principles of electrostatic coupling and in-plane lattice matching. These rules state that the out-of-plane component of the electric displacement field and the in-plane components of the strain are preserved across a layered superlattice, provided that certain growth conditions are respected. Intense research is currently directed at optimizing materials functionalities based on these guidelines, ofte...

  18. Strain-Engineering of Graphene Based Topological Quantum Devices

    Science.gov (United States)

    Diniz, Ginetom S.; Guassi, Marcos R.; Qu, Fanyao

    2015-03-01

    We have investigated the spin-charge transport in quantum devices based on graphene nanoribbons (GNR). Our calculation is based on the surface Green's function technique, considering the presence of an uniform uniaxial strain, spin-orbit interactions (SOIs), exchange field and a smooth staggered potential. We propose the use of uniaxial strain as an efficient mechanism to tune the conductance profiles of GNR with different edge terminations. Our results show that distinct behaviors can be achieved: for armchair GNR there is a complete suppression of the conductance close to the Fermi level with the formation of a band gap that depends on the direction and strength of the strain deformation, while for zigzag GNR there is only a small conductance suppression. We also discuss the effects of SOIs and the appearance of spin-resolved conductance oscillations, and the local density of states of these GNR devices in the quantum anomalous Hall regime. Furthermore, we demonstrate that the local density of states show that depending on the smoothness of the staggered potential, the edge states of AGNR can either emerge or be suppressed. These emerging states can be probed by scanning tunneling microscope. Our findings can be potentially used in novel GNR based topological quantum devices. Supported by FAP-DF, CNPq and CAPES.

  19. Straining mode-dependent collagen remodeling in engineered cardiovascular tissue

    NARCIS (Netherlands)

    Rubbens, M.P.; Mol, A.; Marion, M.H. van; Hanemaaijer, R.; Bank, R.A.; Baaijens, F.P.T.; Bouten, C.V.C.

    2009-01-01

    Similar to native cardiovascular tissues, the mechanical properties of engineered cardiovascular constructs depend on the composition and quality of the extracellular matrix, which is a net result of matrix remodeling processes within the tissue. To improve tissue remodeling, and hence tissue mechan

  20. Tunable gaps and enhanced mobilities in strain-engineered silicane

    International Nuclear Information System (INIS)

    The recent demonstration of single-atom thick, sp3-hybridized group 14 analogues of graphene enables the creation of materials with electronic structures that are manipulated by the nature of the covalently bound substituents above and below the sheet. These analogues can be electronically derived from isolated (111) layers of the bulk diamond lattice. Here, we perform systematic Density Functional Theory calculations to understand how the band dispersions, effective masses, and band gaps change as the bulk silicon (111) layers are continuously separated from each other until they are electronically isolated, and then passivated with hydrogen. High-level calculations based on HSE06 hybrid functionals were performed on each endpoint to compare directly with experimental values. We find that the change in the electronic structure due to variations in the Si-H bond length, Si-Si-Si bond angle, and most significantly the Si-Si bond length can tune the nature of the band gap from indirect to direct with dramatic effects on the transport properties. First-principles calculations of the phonon-limited electron mobility predict a value of 464 cm2/Vs for relaxed indirect band gap Si-H monolayers at room temperature. However, for 1.6% tensile strain, the band gap becomes direct, which increases the mobility significantly (8 551 cm2/Vs at 4% tensile strain). In total, this analysis of Si-based monolayers suggests that strain can change the nature of the band gap from indirect to direct and increase the electron mobility more than 18-fold

  1. Phonon Bandgap Engineering of Strained Monolayer MoS2

    OpenAIRE

    Jiang, Jin-Wu

    2014-01-01

    The phonon band structure of monolayer MoS2 is characteristic for a large energy gap between acoustic and optical branches, which protects the vibration of acoustic modes from being scattered by optical phonon modes. Therefore, the phonon bandgap engineering is of practical significance for the manipulation of phonon-related mechanical or thermal properties in monolayer MoS2. We perform both phonon analysis and molecular dynamics simulations to investigate the tension effect on the phonon ban...

  2. New phenotypes of functional expression of the mKir2.1 channel in potassium efflux-deficient Saccharomyces cerevisiae strains

    Czech Academy of Sciences Publication Activity Database

    Koláčná, Lucie; Zimmermannová, Olga; Hasenbrink, G.; Schwarzer, S.; Ludwig, J.; Lichtenberg-Fraté, H.; Sychrová, Hana

    2005-01-01

    Roč. 22, č. 16 (2005), s. 1315-1323. ISSN 0749-503X Grant ostatní: GA-(XE) Multiplex; GA-(XE) QLK3-CT2001-00401 Institutional research plan: CEZ:AV0Z50110509 Keywords : potassium channel * Saccharomyces cerevisiae * salt sensitivity Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.301, year: 2005

  3. 2-DE based proteomic analysis of Saccharomyces cerevisiae wild and K+ transport-affected mutant (trk1,2) strains at the growth exponential and stationary phases

    Czech Academy of Sciences Publication Activity Database

    Curto, M.; Valledor, L.; Navarrete, C.; Gutiérrez, D.; Sychrová, Hana; Ramos, J.; Jorrin, J.

    2010-01-01

    Roč. 73, č. 12 (2010), s. 2316-2335. ISSN 1874-3919 Institutional research plan: CEZ:AV0Z50110509 Keywords : potassium starvation * S. cerevisiae * proteom Subject RIV: CE - Biochemistry Impact factor: 5.074, year: 2010

  4. Strain- and twist-engineered optical absorption of few-layer black phosphorus

    Science.gov (United States)

    Jia, Qian; Kong, XiangHua; Qiao, JingSi; Ji, Wei

    2016-09-01

    Density functional and many-body perturbation theories calculations were carried out to investigate fundamental and optical bandgap, exciton binding energy and optical absorption property of normal and strain- and twist-engineered few-layer black phosphorus (BP). We found that the fundamental bandgaps of few layer BP can be engineered by layer stacking and in-plane strain, with linear relationships to their associated exciton binding energies. The strain-dependent optical absorption behaviors are also anisotropic that the position of the first absorption peak monotonically blue-shifts as the strain applies to either direction for incident light polarized along the armchair direction, but this is not the case for that along the zigzag direction. Given those striking properties, we proposed two prototype devices for building potentially more balanced light absorbers and light filter passes, which promotes further applications and investigations of BP in nanoelectronics and optoelectronics.

  5. Impact of strain engineering on nanoscale strained InGaAs MOSFET devices.

    Science.gov (United States)

    Lee, Chang-Chun; Chang, Shu-Tong; Sun, P-H; Huang, C-X

    2011-07-01

    The strain distributions in the In(0.53)Ga(0.47)As channel regions of the In(0.4)Ga(0.6)As source/drain (S/D) with various lengths and widths were studied via 3D process simulations. The resulting mobility improvement was analyzed. The tensile strain along the transport direction was found to dominate the mobility improvement. The strain along the vertical direction perpendicular to the gate oxide was found to affect the mobility the least, while the strain along the width direction was slightly degraded. The impact of the channel width and length on the performance improvement, such as on the mobility gain, was analyzed via TCAD simulations. The novelty of this paper stems from its study of the impact of the channel width and length on the performance of InGaAs NMOSFETs, such as on their mobility gain, and from its exploration of physical insights for scaling the future III-V MOS devices. PMID:22121581

  6. Tuning Optical Conductivity of Large-Scale CVD Graphene by Strain Engineering

    OpenAIRE

    Ni, Guang-Xin; Yang, Hong-Zhi; Ji, Wei; Baeck, Seung-Jae; Toh, Chee-Tat; Ahn, Jong-Hyun; Pereira, Vitor M.; Özyilmaz, Barbaros

    2013-01-01

    Strain engineering has been recently recognized as an effective way to tailor the electrical properties of graphene. In the optical domain, effects such as strain-induced anisotropic absorption add an appealing functionality to graphene, opening the prospect for atomically thin optical elements. Indeed, graphene is currently one of the notable players in the intense drive towards bendable, thin, and portable electronic displays, where its intrinsically metallic, optically transparent, and mec...

  7. Biosynthesis of poly(3-hydroxypropionate) from glycerol using engineered Klebsiella pneumoniae strain without vitamin B12

    OpenAIRE

    Feng, Xinjun; Xian, Mo; Liu, Wei; Xu, Chao; Zhang, Haibo; Zhao, Guang

    2015-01-01

    Poly(3-hydroxypropionate) (P3HP) is a biodegradable and biocompatible thermoplastic. Previous studies demonstrated that engineered Escherichia coli strains can produce P3HP with supplementation of expensive vitamin B12. The present study examined the production of P3HP from glycerol in the recombinant Klebsiella pneumoniae strain, which naturally synthesizes vitamin B12. The genes glycerol dehydratase and its reactivation factor (dhaB123, gdrA, and gdrB from K. pneumoniae), aldehyde dehydroge...

  8. L-Histidine Inhibits Biofilm Formation and FLO11-Associated Phenotypes in Saccharomyces cerevisiae Flor Yeasts

    OpenAIRE

    Marc Bou Zeidan; Giacomo Zara; Carlo Viti; Francesca Decorosi; Ilaria Mannazzu; Marilena Budroni; Luciana Giovannetti; Severino Zara

    2014-01-01

    Flor yeasts of Saccharomyces cerevisiae have an innate diversity of FLO11 which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling FLO11 alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce FLO11p. The flor strains generally metabolized amino acids and ...

  9. L-Histidine Inhibits Biofilm Formation and FLO11-Associated Phenotypes in Saccharomyces cerevisiae Flor Yeasts

    OpenAIRE

    Marc Bou Zeidan; Giacomo Zara; Carlo Viti; Francesca Decorosi; Ilaria Mannazzu; Marilena Budroni; Luciana Giovannetti; Severino Zara

    2014-01-01

    Flor yeasts of Saccharomyces cerevisiae have an innate diversity of Flo11p which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling Flo11p alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce Flo11p. The flor strains generally metabolized amino acids and dipeptides...

  10. 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

  11. Effects of spaceflight on polysaccharides of Saccharomyces cerevisiae cell wall.

    Science.gov (United States)

    Liu, Hong-Zhi; Wang, Qiang; Liu, Xiao-Yong; Tan, Sze-Sze

    2008-12-01

    Freeze-dried samples of four Saccharomyces cerevisiae strains, namely, FL01, FL03, 2.0016, and 2.1424, were subjected to spaceflight. After the satellite's landing on Earth, the samples were recovered and changes in yeast cell wall were analyzed. Spaceflight strains of all S. cerevisiae strains showed significant changes in cell wall thickness (P growth curve analysis showed spaceflight S. cerevisiae 2.0016 had a faster growth rate, shorter lag phase periods, higher final biomass, and higher content of beta-glucan. Genetic stability analysis showed that prolonged subculturing of spaceflight strain S. cerevisiae 2.0016 did not lead to the appearance of variants, indicating that the genetic stability of S. cerevisiae 2.0016 mutant could be sufficient for its exploitation of beta-glucan production. PMID:18797865

  12. Strain engineering on structures and properties in ferroelectric thin films with perovskite structures

    Directory of Open Access Journals (Sweden)

    TANG Yanxue

    2015-08-01

    Full Text Available Ferroelectric thin films possess ferroelectric,piezoelectric,pyroelectric and photovoltaic properties,which have bright prospect for transducers,actuators,sensors,energy harvesting and solar cells.The properties of ferroelectric films are closely related to their strain due to films constrained by substrates.Therefore,the key to improve the properties of ferroelectric films is how to use substrates to regulate and control their strain,and then regulate their polarized state.This paper review the research progress of regulating the properties of ferroelectric films with perovskite structure by strain engineering and the problems needed to be resolved.

  13. Analysis of strain gage reliability in F-100 jet engine testing at NASA Lewis Research Center

    Science.gov (United States)

    Holanda, R.

    1983-01-01

    A reliability analysis was performed on 64 strain gage systems mounted on the 3 rotor stages of the fan of a YF-100 engine. The strain gages were used in a 65 hour fan flutter research program which included about 5 hours of blade flutter. The analysis was part of a reliability improvement program. Eighty-four percent of the strain gages survived the test and performed satisfactorily. A post test analysis determined most failure causes. Five failures were caused by open circuits, three failed gages showed elevated circuit resistance, and one gage circuit was grounded. One failure was undetermined.

  14. Analysis and prediction of the physiological effects of altered coenzyme specificity in xylose reductase and xylitol dehydrogenase during xylose fermentation by Saccharomyces cerevisiae

    OpenAIRE

    Krahulec, Stefan; Klimacek, Mario; Nidetzky, Bernd

    2012-01-01

    An advanced strategy of Saccharomyces cerevisiae strain development for fermentation of xylose applies tailored enzymes in the process of metabolic engineering. The coenzyme specificities of the NADPH-preferring xylose reductase (XR) and the NAD+-dependent xylitol dehydrogenase (XDH) have been targeted in previous studies by protein design or evolution with the aim of improving the recycling of NADH or NADPH in their two-step pathway, converting xylose to xylulose. Yeast strains expressing va...

  15. Cell Surface Display of Four Types of Solanum nigrum Metallothionein on Saccharomyces cerevisiae for Biosorption of Cadmium.

    Science.gov (United States)

    Wei, Qinguo; Zhang, Honghai; Guo, Dongge; Ma, Shisheng

    2016-05-28

    We displayed four types of Solanum nigrum metallothionein (SMT) for the first time on the surface of Saccharomyces cerevisiae using an α-agglutinin-based display system. The SMT genes were amplified by RT-PCR. The plasmid pYES2 was used to construct the expression vector. Transformed yeast strains were confirmed by PCR amplification and custom sequencing. Surface-expressed metallothioneins were indirectly indicated by the enhanced cadmium sorption capacity. Flame atomic absorption spectrophotometry was used to examine the concentration of Cd(2+) in this study. The transformed yeast strains showed much higher resistance ability to Cd(2+) compared with the control. Strikingly, their Cd(2+) accumulation was almost twice as much as that of the wild-type yeast cells. Furthermore, surface-engineered yeast strains could effectively adsorb ultra-trace cadmium and accumulate Cd(2+) under a wide range of pH levels, from 3 to 7, without disturbing the Cu(2+) and Hg(2+). Four types of surfaceengineered Saccharomyces cerevisiae strains were constructed and they could be used to purify Cd(2+)-contaminated water and adsorb ultra-trace cadmium effectively. The surface-engineered Saccharomyces cerevisiae strains would be useful tools for the bioremediation and biosorption of environmental cadmium contaminants. PMID:26838339

  16. Characterizing Strain Variation in Engineered E. coli Using a Multi-Omics-Based Workflow.

    Science.gov (United States)

    Brunk, Elizabeth; George, Kevin W; Alonso-Gutierrez, Jorge; Thompson, Mitchell; Baidoo, Edward; Wang, George; Petzold, Christopher J; McCloskey, Douglas; Monk, Jonathan; Yang, Laurence; O'Brien, Edward J; Batth, Tanveer S; Martin, Hector Garcia; Feist, Adam; Adams, Paul D; Keasling, Jay D; Palsson, Bernhard O; Lee, Taek Soon

    2016-05-25

    Understanding the complex interactions that occur between heterologous and native biochemical pathways represents a major challenge in metabolic engineering and synthetic biology. We present a workflow that integrates metabolomics, proteomics, and genome-scale models of Escherichia coli metabolism to study the effects of introducing a heterologous pathway into a microbial host. This workflow incorporates complementary approaches from computational systems biology, metabolic engineering, and synthetic biology; provides molecular insight into how the host organism microenvironment changes due to pathway engineering; and demonstrates how biological mechanisms underlying strain variation can be exploited as an engineering strategy to increase product yield. As a proof of concept, we present the analysis of eight engineered strains producing three biofuels: isopentenol, limonene, and bisabolene. Application of this workflow identified the roles of candidate genes, pathways, and biochemical reactions in observed experimental phenomena and facilitated the construction of a mutant strain with improved productivity. The contributed workflow is available as an open-source tool in the form of iPython notebooks. PMID:27211860

  17. Genetic determinants for enhanced glycerol growth of Saccharomyces cerevisiae.

    Science.gov (United States)

    Swinnen, Steve; Ho, Ping-Wei; Klein, Mathias; Nevoigt, Elke

    2016-07-01

    The yeast Saccharomyces cerevisiae generally shows a low natural capability to utilize glycerol as the sole source of carbon, particularly when synthetic medium is used and complex supplements are omitted. Nevertheless, wild type isolates have been identified that show a moderate growth under these conditions. In the current study we made use of intraspecies diversity to identify targets suitable for reverse metabolic engineering of the non-growing laboratory strain CEN.PK113-1A. A genome-wide genetic mapping experiment using pooled-segregant whole-genome sequence analysis was conducted, and one major and several minor genetic loci were identified responsible for the superior glycerol growth phenotype of the previously selected S. cerevisiae strain CBS 6412-13A. Downscaling of the major locus by fine-mapping and reciprocal hemizygosity analysis allowed the parallel identification of two superior alleles (UBR2CBS 6412-13A and SSK1CBS 6412-13A). These alleles together with the previously identified GUT1CBS 6412-13A allele were used to replace the corresponding alleles in the strain CEN.PK113-1A. In this way, glycerol growth could be established reaching a maximum specific growth rate of 0.08h(-1). Further improvement to a maximum specific growth rate of 0.11h(-1) could be achieved by heterologous expression of the glycerol facilitator FPS1 from Cyberlindnera jadinii. PMID:26971668

  18. A vaccine grade of yeast Saccharomyces cerevisiae expressing mammalian myostatin

    Directory of Open Access Journals (Sweden)

    Zhang Tingting

    2012-12-01

    Full Text Available Abstract Background Yeast Saccharomyces cerevisiae is a widely-used system for protein expression. We previously showed that heat-killed whole recombinant yeast vaccine expressing mammalian myostatin can modulate myostatin function in mice, resulting in increase of body weight and muscle composition in these animals. Foreign DNA introduced into yeast cells can be lost soon unless cells are continuously cultured in selection media, which usually contain antibiotics. For cost and safety concerns, it is essential to optimize conditions to produce quality food and pharmaceutical products. Results We developed a simple but effective method to engineer a yeast strain stably expressing mammalian myostatin. This method utilized high-copy-number integration of myostatin gene into the ribosomal DNA of Saccharomyces cerevisiae. In the final step, antibiotic selection marker was removed using the Cre-LoxP system to minimize any possible side-effects for animals. The resulting yeast strain can be maintained in rich culture media and stably express mammalian myostatin for two years. Oral administration of the recombinant yeast was able to induce immune response to myostatin and modulated the body weight of mice. Conclusions Establishment of such yeast strain is a step further toward transformation of yeast cells into edible vaccine to improve meat production in farm animals and treat human muscle-wasting diseases in the future.

  19. Mechanical Self-Assembly of a Strain-Engineered Flexible Layer: Wrinkling, Rolling, and Twisting

    Science.gov (United States)

    Chen, Zi; Huang, Gaoshan; Trase, Ian; Han, Xiaomin; Mei, Yongfeng

    2016-01-01

    Self-shaping of curved structures, especially those involving flexible thin layers, is attracting increasing attention because of their broad potential applications in, e.g., nanoelectromechanical andmicroelectromechanical systems, sensors, artificial skins, stretchable electronics, robotics, and drug delivery. Here, we provide an overview of recent experimental, theoretical, and computational studies on the mechanical self-assembly of strain-engineered thin layers, with an emphasis on systems in which the competition between bending and stretching energy gives rise to a variety of deformations, such as wrinkling, rolling, and twisting. We address the principle of mechanical instabilities, which is often manifested in wrinkling or multistability of strain-engineered thin layers. The principles of shape selection and transition in helical ribbons are also systematically examined. We hope that a more comprehensive understanding of the mechanical principles underlying these rich phenomena can foster the development of techniques for manufacturing functional three-dimensional structures on demand for a broad spectrum of engineering applications.

  20. Shuffling of Promoters for Multiple Genes To Optimize Xylose Fermentation in an Engineered Saccharomyces cerevisiae Strain▿ †

    OpenAIRE

    Lu, Chenfeng; Jeffries, Thomas

    2007-01-01

    We describe here a useful metabolic engineering tool, multiple-gene-promoter shuffling (MGPS), to optimize expression levels for multiple genes. This method approaches an optimized gene overexpression level by fusing promoters of various strengths to genes of interest for a particular pathway. Selection of these promoters is based on the expression levels of the native genes under the same physiological conditions intended for the application. MGPS was implemented in a yeast xylose fermentati...

  1. Electrostatic actuated strain engineering in monolithically integrated VLS grown silicon nanowires

    Science.gov (United States)

    Wagesreither, Stefan; Bertagnolli, Emmerich; Kawase, Shinya; Isono, Yoshitada; Lugstein, Alois

    2014-11-01

    In this paper we demonstrate the fabrication and application of an electrostatic actuated tensile straining test (EATEST) device enabling strain engineering in individual suspended nanowires (NWs). Contrary to previously reported approaches, this special setup guarantees the application of pure uniaxial tensile strain with no shear component of the stress while e.g. simultaneously measuring the resistance change of the NW. To demonstrate the potential of this approach we investigated the piezoresistivity of about 3 μm long and 100 nm thick SiNWs but in the same way one can think about the application of such a device on other geometries, other materials beyond Si as well as the use of other characterization techniques beyond electrical measurements. Therefore single-crystal SiNWs were monolithically integrated in a comb drive actuated MEMS device based on a silicon-on-insulator (SOI) wafer using the vapor-liquid-solid (VLS) growth technique. Strain values were verified by a precise measurement of the NW elongation with scanning electron microscopy (SEM). Further we employed confocal μ-Raman microscopy for in situ, high spatial resolution measurements of the strain in individual SiNWs during electrical characterization. A giant piezoresistive effect was observed, resulting in a fivefold increase in conductivity for 3% uniaxially strained SiNWs. As the EATEST approach can be easily integrated into an existing Si technology platform this architecture may pave the way toward a new generation of nonconventional devices by leveraging the strain degree of freedom.

  2. Electrostatic actuated strain engineering in monolithically integrated VLS grown silicon nanowires

    International Nuclear Information System (INIS)

    In this paper we demonstrate the fabrication and application of an electrostatic actuated tensile straining test (EATEST) device enabling strain engineering in individual suspended nanowires (NWs). Contrary to previously reported approaches, this special setup guarantees the application of pure uniaxial tensile strain with no shear component of the stress while e.g. simultaneously measuring the resistance change of the NW. To demonstrate the potential of this approach we investigated the piezoresistivity of about 3 μm long and 100 nm thick SiNWs but in the same way one can think about the application of such a device on other geometries, other materials beyond Si as well as the use of other characterization techniques beyond electrical measurements. Therefore single-crystal SiNWs were monolithically integrated in a comb drive actuated MEMS device based on a silicon-on-insulator (SOI) wafer using the vapor–liquid–solid (VLS) growth technique. Strain values were verified by a precise measurement of the NW elongation with scanning electron microscopy (SEM). Further we employed confocal μ-Raman microscopy for in situ, high spatial resolution measurements of the strain in individual SiNWs during electrical characterization. A giant piezoresistive effect was observed, resulting in a fivefold increase in conductivity for 3% uniaxially strained SiNWs. As the EATEST approach can be easily integrated into an existing Si technology platform this architecture may pave the way toward a new generation of nonconventional devices by leveraging the strain degree of freedom. (paper)

  3. Development of new USER-based cloning vectors for multiple genes expression in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kildegaard, Kanchana Rueksomtawin; Jensen, Niels Bjerg; Maury, Jerome;

    2013-01-01

    the production strain with the proper phenotype and product yield. However, the sequential number of metabolic engineering is time-consuming. Furthermore, the number of available selectable markers is also limiting the number of genetic modifications. To overcome these limitations, we have developed a......Saccharomyces cerevisiae is one of the most widely used cell factory in industrial biotechnology and it is used for the production of fuels, chemicals, food ingredients, food and beverages, and pharmaceuticals. Such bioprocesses frequently require multiple rounds of metabolic engineering to obtain...

  4. Improving Engineered Escherichia coli strains for High-level Biosynthesis of Isobutyrate

    Directory of Open Access Journals (Sweden)

    Mingyong Xiong

    2015-05-01

    Full Text Available Isobutyrate is an important platform chemical with various industrial applications. Previously, a synthetic metabolic pathway was constructed in E. coli to produce isobutyrate from glucose. However, isobutanol was found to be a major byproduct. Herein, gene knockouts and enzyme overexpressions were performed to optimize further the engineered E. coli strain. Besides yqhD, the knockouts of three genes eutG, yiaY and ygjB increased isobutyrate production in shake flasks. Furthermore, the introduction of an additional padA on a medium copy number plasmid under the constitutive promoter significantly reduced isobutanol formation. The IBA15-2C strain (BW25113, DyqhD, DygjB; carrying two copies of padA produced 39.2% more isobutyrate (0.39 g/glucose yield, 80% of the theoretical maximum yield than IBA1-1C strain (BW25113, DyqhD; carrying one copy of padA. A scale-up process was also investigated for IBA15-2C strain to optimize the conditions for the production of isobutyrate in the fermentor. With Ca(OH2 as the base for pH control and 10% dissolved oxygen level, IBA15-2C strain produced 90 g/L isobutyrate after 144 h. This study has engineered E. coli to achieve biosynthesis of a nonnative compound with the highest titer and opened up the possibility of the industrial production of isobutyrate.

  5. Combining Protein and Strain Engineering for the Production of Glyco-Engineered Horseradish Peroxidase C1A in Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Simona Capone

    2015-09-01

    Full Text Available Horseradish peroxidase (HRP, conjugated to antibodies and lectins, is widely used in medical diagnostics. Since recombinant production of the enzyme is difficult, HRP isolated from plant is used for these applications. Production in the yeast Pichia pastoris (P. pastoris, the most promising recombinant production platform to date, causes hyperglycosylation of HRP, which in turn complicates conjugation to antibodies and lectins. In this study we combined protein and strain engineering to obtain an active and stable HRP variant with reduced surface glycosylation. We combined four mutations, each being beneficial for either catalytic activity or thermal stability, and expressed this enzyme variant as well as the unmutated wildtype enzyme in both a P. pastoris benchmark strain and a strain where the native α-1,6-mannosyltransferase (OCH1 was knocked out. Considering productivity in the bioreactor as well as enzyme activity and thermal stability, the mutated HRP variant produced in the P. pastoris benchmark strain turned out to be interesting for medical diagnostics. This variant shows considerable catalytic activity and thermal stability and is less glycosylated, which might allow more controlled and efficient conjugation to antibodies and lectins.

  6. Growth-inhibitory activity of the D-mannan of Saccharomyces cerevisiae X2180-1A-5 mutant strain against mouse-implanted sarcoma 180 and Ehrlich-carcinoma solid tumor.

    Science.gov (United States)

    Matsumoto, T; Takanohashi, M; Okubo, Y; Suzuki, M; Suzuki, S

    1980-08-15

    The D-mannan of Saccharomyces cerevisiae X2180-1A-5 mutant strain, which possesses a main chain composed of alpha-(1 yields 6) linked D-mannopyranosyl residues and a small proportion of branches composed of alpha-(1 yields 2)- and alpha-(1 yields 3)-linked D-mannopyranosyl residues, showed strong growth-inhibitory activity against mouse-implanted Sarcoma 180 and Ehrlich-carcinoma solid tumor. The observation that the level of this activity was nearly identical with that of the D-mannan of a wild-type strain of bakers' yeast, which possesses a high proportion of branches composed of alpha-(1 yields 2)-and alpha-(1 yields 3)-linked D-mannopyranosyl residues, suggests that the branches are not essential for antitumor activity. The partial acid-degradation products of both D-mannans, the molecular weight of which was one-third of that of each parent D-mannan, had only one half of the antitumor activity of the parent D-mannans. This suggests that molecular size is the most important factor for the differences in acitvity of the polysaccharides of wild and mutant strains. PMID:6996813

  7. Evidence for Domesticated and Wild Populations of Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    2005-07-01

    Full Text Available Saccharomyces cerevisiae is predominantly found in association with human activities, particularly the production of alcoholic beverages. S. paradoxus, the closest known relative of S. cerevisiae, is commonly found on exudates and bark of deciduous trees and in associated soils. This has lead to the idea that S. cerevisiae is a domesticated species, specialized for the fermentation of alcoholic beverages, and isolates of S. cerevisiae from other sources simply represent migrants from these fermentations. We have surveyed DNA sequence diversity at five loci in 81 strains of S. cerevisiae that were isolated from a variety of human and natural fermentations as well as sources unrelated to alcoholic beverage production, such as tree exudates and immunocompromised patients. Diversity within vineyard strains and within saké strains is low, consistent with their status as domesticated stocks. The oldest lineages and the majority of variation are found in strains from sources unrelated to wine production. We propose a model whereby two specialized breeds of S. cerevisiae have been created, one for the production of grape wine and one for the production of saké wine. We estimate that these two breeds have remained isolated from one another for thousands of years, consistent with the earliest archeological evidence for winemaking. We conclude that although there are clearly strains of S. cerevisiae specialized for the production of alcoholic beverages, these have been derived from natural populations unassociated with alcoholic beverage production, rather than the opposite.

  8. Hybrid orientation technology and strain engineering for ultra-high speed MOSFETs

    Indian Academy of Sciences (India)

    T K Maiti; C K Maiti

    2012-10-01

    We report here RF MOSFET performance in sub-45-nm hybrid orientation CMOS technology. Based on the combination of hybrid orientation technology (HOT) and process-induced local strain engineering,MOSFET RF performance is investigated using CAD (TCAD) technology. Transistor optimization on (100) substrate via silicon nitride (Si3N4) cap layer thickness for -MOSFETs, Ge mole fraction optimization for -MOSFETs on (110) substrates and channel length scaling have resulted in record RF performance, viz. the cut-off frequency, T.

  9. An internal deletion in MTH1 enables growth on glucose of pyruvate-decarboxylase negative, non-fermentative Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Oud Bart

    2012-09-01

    Full Text Available Abstract Background Pyruvate-decarboxylase negative (Pdc- strains of Saccharomyces cerevisiae combine the robustness and high glycolytic capacity of this yeast with the absence of alcoholic fermentation. This makes Pdc-S. cerevisiae an interesting platform for efficient conversion of glucose towards pyruvate-derived products without formation of ethanol as a by-product. However, Pdc- strains cannot grow on high glucose concentrations and require C2-compounds (ethanol or acetate for growth under conditions with low glucose concentrations, which hitherto has limited application in industry. Results Genetic analysis of a Pdc- strain previously evolved to overcome these deficiencies revealed a 225bp in-frame internal deletion in MTH1, encoding a transcriptional regulator involved in glucose sensing. This internal deletion contains a phosphorylation site required for degradation, thereby hypothetically resulting in increased stability of the protein. Reverse engineering of this alternative MTH1 allele into a non-evolved Pdc- strain enabled growth on 20 g l-1 glucose and 0.3% (v/v ethanol at a maximum specific growth rate (0.24 h-1 similar to that of the evolved Pdc- strain (0.23 h-1. Furthermore, the reverse engineered Pdc- strain grew on glucose as sole carbon source, albeit at a lower specific growth rate (0.10 h-1 than the evolved strain (0.20 h-1. The observation that overexpression of the wild-type MTH1 allele also restored growth of Pdc-S. cerevisiae on glucose is consistent with the hypothesis that the internal deletion results in decreased degradation of Mth1. Reduced degradation of Mth1 has been shown to result in deregulation of hexose transport. In Pdc- strains, reduced glucose uptake may prevent intracellular accumulation of pyruvate and/or redox problems, while release of glucose repression due to the MTH1 internal deletion may contribute to alleviation of the C2-compound auxotrophy. Conclusions In this study we have discovered and

  10. Synthesis and accumulation of aromatic aldehydes in an engineered strain of Escherichia coli.

    Science.gov (United States)

    Kunjapur, Aditya M; Tarasova, Yekaterina; Prather, Kristala L J

    2014-08-20

    Aromatic aldehydes are useful in numerous applications, especially as flavors, fragrances, and pharmaceutical precursors. However, microbial synthesis of aldehydes is hindered by rapid, endogenous, and redundant conversion of aldehydes to their corresponding alcohols. We report the construction of an Escherichia coli K-12 MG1655 strain with reduced aromatic aldehyde reduction (RARE) that serves as a platform for aromatic aldehyde biosynthesis. Six genes with reported activity on the model substrate benzaldehyde were rationally targeted for deletion: three genes that encode aldo-keto reductases and three genes that encode alcohol dehydrogenases. Upon expression of a recombinant carboxylic acid reductase in the RARE strain and addition of benzoate during growth, benzaldehyde remained in the culture after 24 h, with less than 12% conversion of benzaldehyde to benzyl alcohol. Although individual overexpression results demonstrated that all six genes could contribute to benzaldehyde reduction in vivo, additional experiments featuring subset deletion strains revealed that two of the gene deletions were dispensable under the conditions tested. The engineered strain was next investigated for the production of vanillin from vanillate and succeeded in preventing formation of the byproduct vanillyl alcohol. A pathway for the biosynthesis of vanillin directly from glucose was introduced and resulted in a 55-fold improvement in vanillin titer when using the RARE strain versus the wild-type strain. Finally, synthesis of the chiral pharmaceutical intermediate L-phenylacetylcarbinol (L-PAC) was demonstrated from benzaldehyde and glucose upon expression of a recombinant mutant pyruvate decarboxylase in the RARE strain. Beyond allowing accumulation of aromatic aldehydes as end products in E. coli, the RARE strain expands the classes of chemicals that can be produced microbially via aldehyde intermediates. PMID:25076127

  11. Novel Strategies for Genomic Manipulation of Trichoderma reesei with the Purpose of Strain Engineering.

    Science.gov (United States)

    Derntl, Christian; Kiesenhofer, Daniel P; Mach, Robert L; Mach-Aigner, Astrid R

    2015-09-01

    The state-of-the-art procedure for gene insertions into Trichoderma reesei is a cotransformation of two plasmids, one bearing the gene of interest and the other a marker gene. This procedure yields up to 80% transformation efficiency, but both the number of integrated copies and the loci of insertion are unpredictable. This can lead to tremendous pleiotropic effects. This study describes the development of a novel transformation system for site-directed gene insertion based on auxotrophic markers. For this purpose, we tested the applicability of the genes asl1 (encoding an enzyme of the l-arginine biosynthesis pathway), the hah1 (encoding an enzyme of the l-lysine biosynthesis pathway), and the pyr4 (encoding an enzyme of the uridine biosynthesis pathway). The developed transformation system yields strains with an additional gene at a defined locus that are prototrophic and ostensibly isogenic compared to their parental strain. A positive transformation rate of 100% was achieved due to the developed split-marker system. Additionally, a double-auxotrophic strain that allows multiple genomic manipulations was constructed, which facilitates metabolic engineering purposes in T. reesei. By employing goxA of Aspergillus niger as a reporter system, the influence on the expression of an inserted gene caused by the orientation of the insertion and the transformation strategy used could be demonstrated. Both are important aspects to be considered during strain engineering. PMID:26150462

  12. High level secretion of cellobiohydrolases by Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Ahlgren Simon

    2011-09-01

    Full Text Available Abstract Background The main technological impediment to widespread utilization of lignocellulose for the production of fuels and chemicals is the lack of low-cost technologies to overcome its recalcitrance. Organisms that hydrolyze lignocellulose and produce a valuable product such as ethanol at a high rate and titer could significantly reduce the costs of biomass conversion technologies, and will allow separate conversion steps to be combined in a consolidated bioprocess (CBP. Development of Saccharomyces cerevisiae for CBP requires the high level secretion of cellulases, particularly cellobiohydrolases. Results We expressed various cellobiohydrolases to identify enzymes that were efficiently secreted by S. cerevisiae. For enhanced cellulose hydrolysis, we engineered bimodular derivatives of a well secreted enzyme that naturally lacks the carbohydrate-binding module, and constructed strains expressing combinations of cbh1 and cbh2 genes. Though there was significant variability in the enzyme levels produced, up to approximately 0.3 g/L CBH1 and approximately 1 g/L CBH2 could be produced in high cell density fermentations. Furthermore, we could show activation of the unfolded protein response as a result of cellobiohydrolase production. Finally, we report fermentation of microcrystalline cellulose (Avicel™ to ethanol by CBH-producing S. cerevisiae strains with the addition of beta-glucosidase. Conclusions Gene or protein specific features and compatibility with the host are important for efficient cellobiohydrolase secretion in yeast. The present work demonstrated that production of both CBH1 and CBH2 could be improved to levels where the barrier to CBH sufficiency in the hydrolysis of cellulose was overcome.

  13. Xylose Fermentation by Saccharomyces cerevisiae: Challenges and Prospects

    Directory of Open Access Journals (Sweden)

    Danuza Nogueira Moysés

    2016-02-01

    Full Text Available Many years have passed since the first genetically modified Saccharomyces cerevisiae strains capable of fermenting xylose were obtained with the promise of an environmentally sustainable solution for the conversion of the abundant lignocellulosic biomass to ethanol. Several challenges emerged from these first experiences, most of them related to solving redox imbalances, discovering new pathways for xylose utilization, modulation of the expression of genes of the non-oxidative pentose phosphate pathway, and reduction of xylitol formation. Strategies on evolutionary engineering were used to improve fermentation kinetics, but the resulting strains were still far from industrial application. Lignocellulosic hydrolysates proved to have different inhibitors derived from lignin and sugar degradation, along with significant amounts of acetic acid, intrinsically related with biomass deconstruction. This, associated with pH, temperature, high ethanol, and other stress fluctuations presented on large scale fermentations led the search for yeasts with more robust backgrounds, like industrial strains, as engineering targets. Some promising yeasts were obtained both from studies of stress tolerance genes and adaptation on hydrolysates. Since fermentation times on mixed-substrate hydrolysates were still not cost-effective, the more selective search for new or engineered sugar transporters for xylose are still the focus of many recent studies. These challenges, as well as under-appreciated process strategies, will be discussed in this review.

  14. Xylose Fermentation by Saccharomyces cerevisiae: Challenges and Prospects.

    Science.gov (United States)

    Moysés, Danuza Nogueira; Reis, Viviane Castelo Branco; de Almeida, João Ricardo Moreira; de Moraes, Lidia Maria Pepe; Torres, Fernando Araripe Gonçalves

    2016-01-01

    Many years have passed since the first genetically modified Saccharomyces cerevisiae strains capable of fermenting xylose were obtained with the promise of an environmentally sustainable solution for the conversion of the abundant lignocellulosic biomass to ethanol. Several challenges emerged from these first experiences, most of them related to solving redox imbalances, discovering new pathways for xylose utilization, modulation of the expression of genes of the non-oxidative pentose phosphate pathway, and reduction of xylitol formation. Strategies on evolutionary engineering were used to improve fermentation kinetics, but the resulting strains were still far from industrial application. Lignocellulosic hydrolysates proved to have different inhibitors derived from lignin and sugar degradation, along with significant amounts of acetic acid, intrinsically related with biomass deconstruction. This, associated with pH, temperature, high ethanol, and other stress fluctuations presented on large scale fermentations led the search for yeasts with more robust backgrounds, like industrial strains, as engineering targets. Some promising yeasts were obtained both from studies of stress tolerance genes and adaptation on hydrolysates. Since fermentation times on mixed-substrate hydrolysates were still not cost-effective, the more selective search for new or engineered sugar transporters for xylose are still the focus of many recent studies. These challenges, as well as under-appreciated process strategies, will be discussed in this review. PMID:26927067

  15. Xylose Fermentation by Saccharomyces cerevisiae: Challenges and Prospects

    Science.gov (United States)

    Moysés, Danuza Nogueira; Reis, Viviane Castelo Branco; de Almeida, João Ricardo Moreira; de Moraes, Lidia Maria Pepe; Torres, Fernando Araripe Gonçalves

    2016-01-01

    Many years have passed since the first genetically modified Saccharomyces cerevisiae strains capable of fermenting xylose were obtained with the promise of an environmentally sustainable solution for the conversion of the abundant lignocellulosic biomass to ethanol. Several challenges emerged from these first experiences, most of them related to solving redox imbalances, discovering new pathways for xylose utilization, modulation of the expression of genes of the non-oxidative pentose phosphate pathway, and reduction of xylitol formation. Strategies on evolutionary engineering were used to improve fermentation kinetics, but the resulting strains were still far from industrial application. Lignocellulosic hydrolysates proved to have different inhibitors derived from lignin and sugar degradation, along with significant amounts of acetic acid, intrinsically related with biomass deconstruction. This, associated with pH, temperature, high ethanol, and other stress fluctuations presented on large scale fermentations led the search for yeasts with more robust backgrounds, like industrial strains, as engineering targets. Some promising yeasts were obtained both from studies of stress tolerance genes and adaptation on hydrolysates. Since fermentation times on mixed-substrate hydrolysates were still not cost-effective, the more selective search for new or engineered sugar transporters for xylose are still the focus of many recent studies. These challenges, as well as under-appreciated process strategies, will be discussed in this review. PMID:26927067

  16. Systems biology and pathway engineering enable Saccharomyces cerevisiae to utilize C-5 and C-6 sugars simultaneously for cellulosic ethanol production

    Science.gov (United States)

    Saccharomyces cerevisiae is a traditional industrial workhorse for ethanol production. However, conventional ethanologenic yeast is superior in fermentation of hexose sugars (C-6) such as glucose but unable to utilize pentose sugars (C-5) such as xylose richly embedded in lignocellulosic biomass. In...

  17. Overexpressing enzymes of the Ehrlich pathway and deleting genes of the competing pathway in Saccharomyces cerevisiae for increasing 2-phenylethanol production from glucose.

    Science.gov (United States)

    Shen, Li; Nishimura, Yuya; Matsuda, Fumio; Ishii, Jun; Kondo, Akihiko

    2016-07-01

    2-Phenylethanol (2-PE) is a higher aromatic alcohol that is used in the cosmetics and food industries. The budding yeast Saccharomyces cerevisiae is considered to be a suitable host for the industrial production of higher alcohols, including 2-PE. To produce 2-PE from glucose in S. cerevisiae, we searched for suitable 2-keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH) enzymes of the Ehrlich pathway for overexpression in strain YPH499, and found that overexpression of the ARO10 and/or ADH1 genes increased 2-PE production from glucose. Further, we screened ten BY4741 single-deletion mutants of genes involved in the competing pathways for 2-PE production, and found that strains aro8Δ and aat2Δ displayed increased 2-PE production. Based on these results, we engineered a BY4741 strain that overexpressed ARO10 and contained an aro8Δ deletion, and demonstrated that the strain produced 96 mg/L 2-PE from glucose as the sole carbon source. As this engineered S. cerevisiae strain showed a significant increase in 2-PE production from glucose without the addition of an intermediate carbon substrate, it is a promising candidate for the large-scale production of 2-PE. PMID:26975754

  18. N-hypermannose glycosylation disruption enhances recombinant protein production by regulating secretory pathway and cell wall integrity in Saccharomyces cerevisiae.

    Science.gov (United States)

    Tang, Hongting; Wang, Shenghuan; Wang, Jiajing; Song, Meihui; Xu, Mengyang; Zhang, Mengying; Shen, Yu; Hou, Jin; Bao, Xiaoming

    2016-01-01

    Saccharomyces cerevisiae is a robust host for heterologous protein expression. The efficient expression of cellulases in S. cerevisiae is important for the consolidated bioprocess that directly converts lignocellulose into valuable products. However, heterologous proteins are often N-hyperglycosylated in S. cerevisiae, which may affect protein activity. In this study, the expression of three heterologous proteins, β-glucosidase, endoglucanase and cellobiohydrolase, was found to be N-hyperglycosylated in S. cerevisiae. To block the formation of hypermannose glycan, these proteins were expressed in strains with deletions in key Golgi mannosyltransferases (Och1p, Mnn9p and Mnn1p), respectively. Their extracellular activities improved markedly in the OCH1 and MNN9 deletion strains. Interestingly, truncation of the N-hypermannose glycan did not increase the specific activity of these proteins, but improved the secretion yield. Further analysis showed OCH1 and MNN9 deletion up-regulated genes in the secretory pathway, such as protein folding and vesicular trafficking, but did not induce the unfolded protein response. The cell wall integrity was also affected by OCH1 and MNN9 deletion, which contributed to the release of secretory protein extracellularly. This study demonstrated that mannosyltransferases disruption improved protein secretion through up-regulating secretory pathway and affecting cell wall integrity and provided new insights into glycosylation engineering for protein secretion. PMID:27156860

  19. [Construction and evaluation of an engineered bacterial strain for producing lipopeptide under anoxic conditions].

    Science.gov (United States)

    Liang, Xiao-long; Zhao, Feng; Shi, Rong-jiu; Ban, Yun-he; Zhou, Ji-dong; Han, Si-qin; Zhang, Ying

    2015-08-01

    Biosurfactant-facilitated oil recovery is one of the most important aspects of microbial enhanced oil recovery (MEOR). However, the biosurfactant production by biosurfactant-producing microorganisms, most of which are aerobes, is severely suppressed due to the in-situ anoxic conditions within oil reservoirs. In this research, we successfully engineered a strain JD-3, which could grow rapidly and produce lipopeptide under anoxic conditions, by protoplast confusion using a Bacillus amyloliquefaciens strain BQ-2 which produces biosurfactant aerobically, and a facultative anaerobic Pseudomonas stutzeri strain DQ-1 as parent strains. The alignment of 16S rDNA sequence (99% similarity) and comparisons of cell colony morphology showed that fusant JD-3 was closer to the parental strain B. amyloliquefaciens BQ-2. The surface tension of culture broth of fusant JD-3, after 36-hour cultivation under anaerobic conditions, decreased from initially 63.0 to 32.5 mN · m(-1). The results of thin layer chromatography and infrared spectrum analysis demonstrated that the biosurfactant produced by JD-3 was lipopeptide. The surface-active lipopeptide had a low critical micelle concentration (CMC) of 90 mg · L(-1) and presented a good ability to emulsify various hydrocarbons such as crude oil, liquid paraffin, and kerosene. Strain JD-3 could utilize peptone as nitrogen source and sucrose, glucose, glycerin or other common organics as carbon sources for anaerobic lipopeptide synthesis. The subculture of fusant JD-3 showed a stable lipopeptide-producing ability even after ten serial passages. All these results indicated that fusant JD-3 holds a great potential to microbially enhance oil recovery under anoxic conditions. PMID:26685621

  20. Evaluation results of the 700 deg C Chinese strain gauges. [for gas turbine engine

    Science.gov (United States)

    Hobart, H. F.

    1985-01-01

    Gauges fabricated from specially developed Fe-Cr-Al-V-Ti-Y alloy wire in the Republic of China were evaluated for use in static strain measurement of hot gas turbine engines. Gauge factor variation with temperature, apparent strain, and drift were included. Results of gauge factor versus temperature tests show gauge factor decreasing with increasing temperature. The average slope is -3-1/2 percent/100 K, with an uncertainty band of + or - 8 percent. Values of room temperature gauge factor for the Chinese and Kanthal A-1 gauges averaged 2.73 and 2.12, respectively. The room temperature gauge factor of the Chinese gauges was specified to be 2.62. The apparent strain data for both the Chinese alloy and Kanthal A-1 showed large cycle to cycle nonrepeatability. All apparent strain curves had a similar S-shape, first going negative and then rising to positive value with increasing temperatures. The mean curve for the Chinese gauges between room temperature and 100 K had a total apparent strain of 1500 microstrain. The equivalent value for Kanthal A-1 was about 9000 microstrain. Drift tests at 950 K for 50 hr show an average drift rate of about -9 microstrain/hr. Short-term (1 hr) rates are higher, averaging about -40 microstrain for the first hour. In the temperature range 700 to 870 K, however, short-term drift rates can be as high as 1700 microstrain for the first hour. Therefore, static strain measurements in this temperature range should be avoided.

  1. Increase of betulinic acid production in Saccharomyces cerevisiae by balancing fatty acids and betulinic acid forming pathways.

    Science.gov (United States)

    Li, Jing; Zhang, Yansheng

    2014-04-01

    Betulinic acid is a plant-based triterpenoid that has been recognized for its antitumor and anti-HIV activities. The level of betulinic acid in its natural hosts is extremely low. In the present study, we constructed betulinic acid biosynthetic pathway in Saccharomyces cerevisiae by metabolic engineering. Given the betulinic acid forming pathways sharing the common substrate acetyl-CoA with fatty acid synthesis, the metabolic fluxes between the two pathways were varied by changing gene expressions, and their effects on betulinic acid production were investigated. We constructed nine S. cerevisiae strains representing nine combinations of the flux distributions between betulinic acid and fatty acid pathways. Our results demonstrated that it was possible to improve the betulinic acid production in S. cerevisiae while keeping a desirable growth phenotype by optimally balancing the carbon fluxes of the two pathways. Through modulating the expressions of the key genes on betulinic acid and fatty acid pathways, the difference in betulinic acid yield varied largely in the range of 0.01-1.92 mg L(-1) OD(-1). The metabolic engineering approach used in this study could be extended for synthesizing other triterpenoids in S. cerevisiae. PMID:24389702

  2. Strain-engineered diffusive atomic switching in two-dimensional crystals

    Science.gov (United States)

    Kalikka, Janne; Zhou, Xilin; Dilcher, Eric; Wall, Simon; Li, Ju; Simpson, Robert E.

    2016-01-01

    Strain engineering is an emerging route for tuning the bandgap, carrier mobility, chemical reactivity and diffusivity of materials. Here we show how strain can be used to control atomic diffusion in van der Waals heterostructures of two-dimensional (2D) crystals. We use strain to increase the diffusivity of Ge and Te atoms that are confined to 5 Å thick 2D planes within an Sb2Te3–GeTe van der Waals superlattice. The number of quintuple Sb2Te3 2D crystal layers dictates the strain in the GeTe layers and consequently its diffusive atomic disordering. By identifying four critical rules for the superlattice configuration we lay the foundation for a generalizable approach to the design of switchable van der Waals heterostructures. As Sb2Te3–GeTe is a topological insulator, we envision these rules enabling methods to control spin and topological properties of materials in reversible and energy efficient ways. PMID:27329563

  3. Metabolic engineering of a reduced-genome strain of Escherichia coli for L-threonine production

    Directory of Open Access Journals (Sweden)

    Yoon Byoung

    2009-01-01

    Full Text Available Abstract Background Deletion of large blocks of nonessential genes that are not needed for metabolic pathways of interest can reduce the production of unwanted by-products, increase genome stability, and streamline metabolism without physiological compromise. Researchers have recently constructed a reduced-genome Escherichia coli strain MDS42 that lacks 14.3% of its chromosome. Results Here we describe the reengineering of the MDS42 genome to increase the production of the essential amino acid L-threonine. To this end, we over-expressed a feedback-resistant threonine operon (thrA*BC, deleted the genes that encode threonine dehydrogenase (tdh and threonine transporters (tdcC and sstT, and introduced a mutant threonine exporter (rhtA23 in MDS42. The resulting strain, MDS-205, shows an ~83% increase in L-threonine production when cells are grown by flask fermentation, compared to a wild-type E. coli strain MG1655 engineered with the same threonine-specific modifications described above. And transcriptional analysis revealed the effect of the deletion of non-essential genes on the central metabolism and threonine pathways in MDS-205. Conclusion This result demonstrates that the elimination of genes unnecessary for cell growth can increase the productivity of an industrial strain, most likely by reducing the metabolic burden and improving the metabolic efficiency of cells.

  4. Engineering Pichia pastoris for improved NADH regeneration: A novel chassis strain for whole-cell catalysis.

    Science.gov (United States)

    Geier, Martina; Brandner, Christoph; Strohmeier, Gernot A; Hall, Mélanie; Hartner, Franz S; Glieder, Anton

    2015-01-01

    Many synthetically useful reactions are catalyzed by cofactor-dependent enzymes. As cofactors represent a major cost factor, methods for efficient cofactor regeneration are required especially for large-scale synthetic applications. In order to generate a novel and efficient host chassis for bioreductions, we engineered the methanol utilization pathway of Pichia pastoris for improved NADH regeneration. By deleting the genes coding for dihydroxyacetone synthase isoform 1 and 2 (DAS1 and DAS2), NADH regeneration via methanol oxidation (dissimilation) was increased significantly. The resulting Δdas1 Δdas2 strain performed better in butanediol dehydrogenase (BDH1) based whole-cell conversions. While the BDH1 catalyzed acetoin reduction stopped after 2 h reaching ~50% substrate conversion when performed in the wild type strain, full conversion after 6 h was obtained by employing the knock-out strain. These results suggest that the P. pastoris Δdas1 Δdas2 strain is capable of supplying the actual biocatalyst with the cofactor over a longer reaction period without the over-expression of an additional cofactor regeneration system. Thus, focusing the intrinsic carbon flux of this methylotrophic yeast on methanol oxidation to CO2 represents an efficient and easy-to-use strategy for NADH-dependent whole-cell conversions. At the same time methanol serves as co-solvent, inductor for catalyst and cofactor regeneration pathway expression and source of energy. PMID:26664594

  5. Understanding and strain-engineering wrinkle networks in supported graphene through simulations

    Science.gov (United States)

    Zhang, Kuan; Arroyo, Marino

    2014-12-01

    Wrinkle networks are ubiquitous buckle-induced delaminations in supported graphene, which locally modify the electronic structure and degrade device performance. Although the strong property-deformation coupling of graphene can be potentially harnessed by strain engineering, it has not been possible to precisely control the geometry of wrinkle networks. Through numerical simulations based on an atomistically informed continuum theory, we understand how strain anisotropy, adhesion and friction govern spontaneous wrinkling. We then propose a strategy to control the location of wrinkles through patterns of weaker adhesion. This strategy is deceptively simple, and can in fact fail in several ways, particularly under biaxial compression. However, within bounds set by the physics of wrinkling, it is possible to robustly create by strain a variety of wrinkle network geometries and junction configurations. Graphene is nearly unstrained in the planar regions bounded by wrinkles, highly curved along wrinkles, and highly stretched and curved at junctions, which can either locally attenuate or amplify the applied strain depending on their configuration. These mechanically self-assembled networks are stable under the pressure produced by an enclosed fluid and form continuous channels, opening the door to nano-fluidic applications.

  6. Exopolysaccharide production by a genetically engineered Enterobacter cloacae strain for microbial enhanced oil recovery.

    Science.gov (United States)

    Sun, Shanshan; Zhang, Zhongzhi; Luo, Yijing; Zhong, Weizhang; Xiao, Meng; Yi, Wenjing; Yu, Li; Fu, Pengcheng

    2011-05-01

    Microbial enhanced oil recovery (MEOR) is a petroleum biotechnology for manipulating function and/or structure of microbial environments existing in oil reservoirs for prolonged exploitation of the largest source of energy. In this study, an Enterobacter cloacae which is capable of producing water-insoluble biopolymers at 37°C and a thermophilic Geobacillus strain were used to construct an engineered strain for exopolysaccharide production at higher temperature. The resultant transformants, GW3-3.0, could produce exopolysaccharide up to 8.83 g l(-1) in molasses medium at 54°C. This elevated temperature was within the same temperature range as that for many oil reservoirs. The transformants had stable genetic phenotype which was genetically fingerprinted by RAPD analysis. Core flooding experiments were carried out to ensure effective controlled profile for the simulation of oil recovery. The results have demonstrated that this approach has a promising application potential in MEOR. PMID:21444201

  7. EVALUATION OF ETHANOL PRODUCTION FROM TWO RECOMBINANT AND A COMMERCIAL STRAINS OF SACCHAROMYCES CEREVISIAE (FUNGI: ASCOMYCOTA IN SUGAR-CANE MOLASSES AND REJECTED-BANANA JUICE FROM URABÁ, COLOMBIA EVALUACIÓN DE LA PRODUCCIÓN DE ETANOL POR DOS CEPAS RECOMBINANTES Y UNA COMERCIAL DE SACCHAROMYCES CEREVISIAE (FUNGI: ASCOMYCOTA EN MELAZA DE CAÑA DE AZÚCAR Y MOSTOS DE BANANO DE RECHAZO DE URABÁ, COLOMBIA

    Directory of Open Access Journals (Sweden)

    Carolina Peña-Serna

    2011-12-01

    Full Text Available The production of bioethanol using Saccharomyces cerevisiae (Fungi: Ascomycota is influenced by sugar concentrations and the fermentation substrate. For that reason, in this study the kinetics of biomass production, residual sugar and ethanol production of four S. cerevisiae strains were evaluated in two fermentation media (sugar-cane molasses and rejected-banana juice at two sugar concentrations (100 and 170 g/l. The Ethanol Red® and GG570- CIBII strains exhibited the greatest ethanol production, with peak values of 119.74 (35 h and 62 g/l (15 h, Yps 0.75 and 0.43 g/g, and Qp 3.42 and 2.61 g/l/h, respectively, at 170 g/l of sugar in the sugar-cane molasses broth. In additional, the GG570-CIBII strain showed an increase of 37.1 g/l ethanol with respect to the control strain.La producción de bioetanol a partir de Saccharomyces cerevisiae (Fungi: Ascomycota está influenciada por la concentración de azúcares y el sustrato de fermentación. Por ello, en este trabajo se evaluaron las cinéticas de producción de biomasa, azúcares residuales y producción de etanol de cuatro cepas de S. cerevisiae en dos medios de fermentación (melaza de caña de azúcar y banano de rechazo a dos concentraciones de azúcares (100 y 170 g/l. Las cepas Ethanol Red® y GG570-CIBII presentaron mayor producción de etanol con pico de producción de 119,74 (35 h y 62 g/l (15 h, Yps 0,75 y 0,43 g/g y Qp 3,42 y 2,61 g/l/h, respectivamente a 170 g/l de azúcares en melaza de caña de azúcar. Adicionalmente, la cepa GG570-CIBII mostró un incremento de 37,1 g/l de etanol con respecto a la cepa control.

  8. EVALUATION OF ETHANOL PRODUCTION FROM TWO RECOMBINANT AND A COMMERCIAL STRAINS OF SACCHAROMYCES CEREVISIAE (FUNGI: ASCOMYCOTA IN SUGAR-CANE MOLASSES AND REJECTED-BANANA JUICE FROM URABÁ (ANTIOQUIA, COLOMBIA EVALUACIÓN DE LA PRODUCCIÓN DE ETANOL POR DOS CEPAS RECOMBINANTES Y UNA COMERCIAL DE SACCHAROMYCES CEREVISIAE (FUNGI: ASCOMYCOTA EN MELAZA DE CAÑA DE AZÚCAR Y MOSTOS DE BANANO DE RECHAZO DE URABÁ (ANTIOQUIA, COLOMBIA

    Directory of Open Access Journals (Sweden)

    Carolina Peña-Serna

    2012-06-01

    Full Text Available The production of bioethanol using Saccharomyces cerevisiae (Fungi: Ascomycota is influenced by sugar concentration and fermentation substrate. For that reason, the kinetics of biomass production, residual sugars, and ethanol production of four of S. cerevisiae strains were evaluated in two different fermentation media (sugar-cane molasses and rejected-banana juice at two different sugar concentrations (100 and 170 g/l. The EthanolRed® and GG570-CIBII strains exhibited the greatest ethanol production, with peak values of 119.74 (35 h and 62 g/l (15 h, Yps 0.75 and 0.43 g/g, and Qp 3.42 and 2.61 g/l/h, respectively, at 170 g/l of sugar in sugar-cane molasses broth. Additionally, the strain GG570-CIBII showed an increase of 37.1 g/l ethanol with respect to the control strain.La producción de bioetanol a partir de Saccharomyces cerevisiae (Fungi: Ascomycota está influenciada por la concentración de azúcares y el sustrato de fermentación. Por ello en este trabajo se evaluaron las cinéticas de producción de biomasa, azúcares residuales y producción de etanol de cuatro cepas de S. cerevisiae en dos medios de fermentación (melaza de caña de azúcar y banano de rechazo a dos concentraciones de azúcares (100 y 170 g/l. Las cepas EthanolRed® y GG570-CIBII presentaron mayor producción de etanol con pico de producción de 119,74 (35 h y 62 g/l (15 h, Yps 0,75 y 0,43 g/g y, Qp 3,42 y 2,61 g/l/h, respectivamente a 170 g/l de azúcares en melaza de caña de azúcar. Adicionalmente, la cepa GG570-CIBII mostró un incremento de 37,1 g/l de etanol con respecto a la cepa control.

  9. Characterization of global yeast quantitative proteome data generated from the wild-type and glucose repression Saccharomyces cerevisiae strains: The comparison of two quantitative methods

    DEFF Research Database (Denmark)

    Usaite, Renata; Wohlschlegel, James; Venable, John D.;

    2008-01-01

    -type yeast and yeast strains lacking key components of the Snf1 kinase complex. Four different strains were grown under well-controlled chemostat conditions. Multidimensional protein identification technology followed by quantitation using either spectral counting or stable isotope labeling approaches was...... labeling strategy. The stable isotope labeling based quantitative approach was found to be highly reproducible among biological replicates when complex protein mixtures containing small expression changes were analyzed. Where poor correlation between stable isotope labeling and spectral counting was found...

  10. Engineering of a plasmid-free Escherichia coli strain for improved in vivo biosynthesis of astaxanthin

    Directory of Open Access Journals (Sweden)

    Steuer Kristin

    2011-04-01

    Full Text Available Abstract Background The xanthophyll astaxanthin is a high-value compound with applications in the nutraceutical, cosmetic, food, and animal feed industries. Besides chemical synthesis and extraction from naturally producing organisms like Haematococcus pluvialis, heterologous biosynthesis in non-carotenogenic microorganisms like Escherichia coli, is a promising alternative for sustainable production of natural astaxanthin. Recent achievements in the metabolic engineering of E. coli strains have led to a significant increase in the productivity of carotenoids like lycopene or β-carotene by increasing the metabolic flux towards the isoprenoid precursors. For the heterologous biosynthesis of astaxanthin in E. coli, however, the conversion of β-carotene to astaxanthin is obviously the most critical step towards an efficient biosynthesis of astaxanthin. Results Here we report the construction of the first plasmid-free E. coli strain that produces astaxanthin as the sole carotenoid compound with a yield of 1.4 mg/g cdw (E. coli BW-ASTA. This engineered E. coli strain harbors xanthophyll biosynthetic genes from Pantoea ananatis and Nostoc punctiforme as individual expression cassettes on the chromosome and is based on a β-carotene-producing strain (E. coli BW-CARO recently developed in our lab. E. coli BW-CARO has an enhanced biosynthesis of the isoprenoid precursor isopentenyl diphosphate (IPP and produces β-carotene in a concentration of 6.2 mg/g cdw. The expression of crtEBIY along with the β-carotene-ketolase gene crtW148 (NpF4798 and the β-carotene-hydroxylase gene (crtZ under controlled expression conditions in E. coli BW-ASTA directed the pathway exclusively towards the desired product astaxanthin (1.4 mg/g cdw. Conclusions By using the λ-Red recombineering technique, genes encoding for the astaxanthin biosynthesis pathway were stably integrated into the chromosome of E. coli. The expression levels of chromosomal integrated recombinant

  11. Direct Production of Ethanol from Raw Corn Starch via Fermentation by Use of a Novel Surface-Engineered Yeast Strain Codisplaying Glucoamylase and α-Amylase

    OpenAIRE

    Shigechi, Hisayori; Koh, Jun; Fujita, Yasuya; Matsumoto, Takeshi; Bito, Yohei; Ueda, Mitsuyoshi; SATOH, Eiichi; Fukuda, Hideki; Kondo, Akihiko

    2004-01-01

    Direct and efficient production of ethanol by fermentation from raw corn starch was achieved by using the yeast Saccharomyces cerevisiae codisplaying Rhizopus oryzae glucoamylase and Streptococcus bovis α-amylase by using the C-terminal-half region of α-agglutinin and the flocculation functional domain of Flo1p as the respective anchor proteins. In 72-h fermentation, this strain produced 61.8 g of ethanol/liter, with 86.5% of theoretical yield from raw corn starch.

  12. A dual approach for improving homogeneity of a human-type N-glycan structure in Saccharomyces cerevisiae.

    Science.gov (United States)

    Piirainen, Mari A; Boer, Harry; de Ruijter, Jorg C; Frey, Alexander D

    2016-04-01

    N-glycosylation is an important feature of therapeutic and other industrially relevant proteins, and engineering of the N-glycosylation pathway provides opportunities for developing alternative, non-mammalian glycoprotein expression systems. Among yeasts, Saccharomyces cerevisiae is the most established host organism used in therapeutic protein production and therefore an interesting host for glycoengineering. In this work, we present further improvements in the humanization of the N-glycans in a recently developed S. cerevisiae strain. In this strain, a tailored trimannosyl lipid-linked oligosaccharide is formed and transferred to the protein, followed by complex-type glycan formation by Golgi apparatus-targeted human N-acetylglucosamine transferases. We improved the glycan pattern of the glycoengineered strain both in terms of glycoform homogeneity and the efficiency of complex-type glycosylation. Most of the interfering structures present in the glycoengineered strain were eliminated by deletion of the MNN1 gene. The relative abundance of the complex-type target glycan was increased by the expression of a UDP-N-acetylglucosamine transporter from Kluyveromyces lactis, indicating that the import of UDP-N-acetylglucosamine into the Golgi apparatus is a limiting factor for efficient complex-type N-glycosylation in S. cerevisiae. By a combination of the MNN1 deletion and the expression of a UDP-N-acetylglucosamine transporter, a strain forming complex-type glycans with a significantly improved homogeneity was obtained. Our results represent a further step towards obtaining humanized glycoproteins with a high homogeneity in S. cerevisiae. PMID:26983412

  13. The Reacquisition of Biotin Prototrophy in Saccharomyces cerevisiae Involved Horizontal Gene Transfer, Gene Duplication and Gene Clustering

    OpenAIRE

    Hall, Charles; Dietrich, Fred S

    2007-01-01

    The synthesis of biotin, a vitamin required for many carboxylation reactions, is a variable trait in Saccharomyces cerevisiae. Many S. cerevisiae strains, including common laboratory strains, contain only a partial biotin synthesis pathway. We here report the identification of the first step necessary for the biotin synthesis pathway in S. cerevisiae. The biotin auxotroph strain S288c was able to grow on media lacking biotin when BIO1 and the known biotin synthesis gene BIO6 were introduced t...

  14. Metabolic and bioprocess engineering for production of selenized yeast with increased content of seleno-methylselenocysteine

    DEFF Research Database (Denmark)

    Mapelli, Valeria; Hillestrøm, Peter René; Kápolna, Emese;

    2011-01-01

    Specific Se-metabolites have been recognized to be the main elements responsible for beneficial effects of Se-enriched diet, and Se-methylselenocysteine (SeMCys) is thought to be among the most effective ones. Here we show that an engineered Saccharomyces cerevisiae strain, expressing a codon opt...

  15. Optimization of ethanol, citric acid, and α-amylase production from date wastes by strains of Saccharomyces cerevisiae, Aspergillus niger, and Candida guilliermondii.

    Science.gov (United States)

    Acourene, S; Ammouche, A

    2012-05-01

    The present study deals with submerged ethanol, citric acid, and α-amylase fermentation by Saccharomyces cerevisiae SDB, Aspergillus niger ANSS-B5, and Candida guilliermondii CGL-A10, using date wastes as the basal fermentation medium. The physical and chemical parameters influencing the production of these metabolites were optimized. As for the ethanol production, the optimum yield obtained was 136.00 ± 0.66 g/l under optimum conditions of an incubation period of 72 h, inoculum content of 4% (w/v), sugars concentration of 180.0 g/l, and ammonium phosphate concentration of 1.0 g/l. Concerning citric acid production, the cumulative effect of temperature (30°C), sugars concentration of 150.0 g/l, methanol concentration of 3.0%, initial pH of 3.5, ammonium nitrate concentration of 2.5 g/l, and potassium phosphate concentration of 2.5 g/l during the fermentation process of date wastes syrup did increase the citric acid production to 98.42 ± 1.41 g/l. For the production of α-amylase, the obtained result shows that the presence of starch strongly induces the production of α-amylase with a maximum at 5.0 g/l. Among the various nitrogen sources tested, urea at 5.0 g/l gave the maximum biomass and α-amylase estimated at 5.76 ± 0.56 g/l and 2,304.19 ± 31.08 μmol/l/min, respectively after 72 h incubation at 30°C, with an initial pH of 6.0 and potassium phosphate concentration of 6.0 g/l. PMID:22193823

  16. Engineered measles virus Edmonston strain used as a novel oncolytic viral system against human hepatoblastoma

    International Nuclear Information System (INIS)

    Hepatoblastoma (HB) is the most common primary, malignant pediatric liver tumor in children. The treatment results for affected children have markedly improved in recent decades. However, the prognosis for high-risk patients who have extrahepatic extensions, invasion of the large hepatic veins, distant metastases and very high alpha-fetoprotein (AFP) serum levels remains poor. There is an urgent need for the development of novel therapeutic approaches. An attenuated strain of measles virus, derived from the Edmonston vaccine lineage, was genetically engineered to produce carcinoembryonic antigen (CEA). We investigated the antitumor potential of this novel viral agent against human HB both in vitro and in vivo. Infection of the Hep2G and HUH6 HB cell lines, at multiplicities of infection (MOIs) ranging from 0.01 to 1, resulted in a significant cytopathic effect consisting of extensive syncytia formation and massive cell death at 72–96 h after infection. Both of the HB lines overexpressed the measles virus receptor CD46 and supported robust viral replication, which correlated with CEA production. The efficacy of this approach in vivo was examined in murine Hep2G xenograft models. Flow cytometry assays indicated an apoptotic mechanism of cell death. Intratumoral administration of MV-CEA resulted in statistically significant delay of tumor growth and prolongation of survival. The engineered measles virus Edmonston strain MV-CEA has potent therapeutic efficacy against HB cell lines and xenografts. Trackable measles virus derivatives merit further exploration in HB treatment

  17. Display of phytase on the cell surface of Saccharomyces cerevisiae to degrade phytate phosphorus and improve bioethanol production.

    Science.gov (United States)

    Chen, Xianzhong; Xiao, Yan; Shen, Wei; Govender, Algasan; Zhang, Liang; Fan, You; Wang, Zhengxiang

    2016-03-01

    Currently, development of biofuels as an alternative fuel has gained much attention due to resource and environmental challenges. Bioethanol is one of most important and dominant biofuels, and production using corn or cassava as raw materials has become a prominent technology. However, phytate contained in the raw material not only decreases the efficiency of ethanol production, but also leads to an increase in the discharge of phosphorus, thus impacting on the environment. In this study, to decrease phytate and its phosphorus content in an ethanol fermentation process, Saccharomyces cerevisiae was engineered through a surface-displaying system utilizing the C-terminal half of the yeast α-agglutinin protein. The recombinant yeast strain, PHY, was constructed by successfully displaying phytase on the surface of cells, and enzyme activity reached 6.4 U/g wet biomass weight. Ethanol productions using various strains were compared, and the results demonstrated that the specific growth rate and average fermentation rate of the PHY strain were higher 20 and 18 %, respectively, compared to the control strain S. cerevisiae CICIMY0086, in a 5-L bioreactor process by simultaneous saccharification and fermentation. More importantly, the phytate phosphorus concentration decreased by 89.8 % and free phosphorus concentration increased by 142.9 % in dry vinasse compared to the control in a 5-L bioreactor. In summary, we constructed a recombinant S. cerevisiae strain displaying phytase on the cell surface, which could improve ethanol production performance and effectively reduce the discharge of phosphorus. The strain reported here represents a useful novel engineering platform for developing an environment-friendly system for bioethanol production from a corn substrate. PMID:26610799

  18. Controlling the thermal conductance of graphene/h -BN lateral interface with strain and structure engineering

    Science.gov (United States)

    Ong, Zhun-Yong; Zhang, Gang; Zhang, Yong-Wei

    2016-02-01

    Although phonon-mediated thermal conduction in pristine graphene and hexagonal boron nitride is well understood, less is known about phonon transport in single-sheet graphene-hexagonal boron nitride (Gr /h -BN ) lateral heterostructures, where the thermal resistance of the interfaces plays an important role in the overall thermal conductivity. We apply the newly developed extended atomistic Green's function method to analyze with detail the effect of strain and structure engineering on the thermal conductance Gint of the Gr /h -BN interface. Our calculations show that longitudinal tensile strain leads to significant Gint enhancement (up to 25 % at 300 K) primarily through the improved alignment of the flexural acoustic phonon bands, despite the reduction in the longitudinal acoustic (LA) and transverse acoustic phonon velocities. In addition, we find that alternating C-N zigzag bonds along the zigzag interface lead to a greater Gint than C-B bonds through more effective transmission of high-frequency LA and transverse optical phonons, especially at high strain levels. We also demonstrate how the interfacial structure dramatically affects the orientation of the transmitted optical phonons, a phenomenon that is neither seen for acoustic phonons nor predictable from conventional acoustic wave scattering theory. Insights from our paper can provide the basis for manipulating the interfacial thermal conductance in other two-dimensional heterostructures.

  19. Strong Modulation of Optical Properties in Black Phosphorus through Strain-Engineered Rippling

    Science.gov (United States)

    Quereda, Jorge; San-Jose, Pablo; Parente, Vincenzo; Vaquero-Garzon, Luis; Molina-Mendoza, Aday J.; Agraït, Nicolás; Rubio-Bollinger, Gabino; Guinea, Francisco; Roldán, Rafael; Castellanos-Gomez, Andres

    2016-05-01

    Controlling the bandgap through local-strain engineering is an exciting avenue for tailoring optoelectronic materials. Two-dimensional crystals are particularly suited for this purpose because they can withstand unprecedented non-homogeneous deformations before rupture: one can literally bend them and fold them up almost like a piece of paper. Here, we study multi-layer black phosphorus sheets subjected to periodic stress to modulate their optoelectronic properties. We find a remarkable shift of the optical absorption band-edge of up to ~0.7 eV between the regions under tensile and compressive stress, greatly exceeding the strain tunability reported for transition metal dichalcogenides. This observation is supported by theoretical models which also predict that this periodic stress modulation can yield to quantum confinement of carriers at low temperatures. The possibility of generating large strain-induced variations in the local density of charge carriers opens the door for a variety of applications including photovoltaics, quantum optics and two-dimensional optoelectronic devices.

  20. Strong Modulation of Optical Properties in Black Phosphorus through Strain-Engineered Rippling.

    Science.gov (United States)

    Quereda, Jorge; San-Jose, Pablo; Parente, Vincenzo; Vaquero-Garzon, Luis; Molina-Mendoza, Aday J; Agraït, Nicolás; Rubio-Bollinger, Gabino; Guinea, Francisco; Roldán, Rafael; Castellanos-Gomez, Andres

    2016-05-11

    Controlling the bandgap through local-strain engineering is an exciting avenue for tailoring optoelectronic materials. Two-dimensional crystals are particularly suited for this purpose because they can withstand unprecedented nonhomogeneous deformations before rupture; one can literally bend them and fold them up almost like a piece of paper. Here, we study multilayer black phosphorus sheets subjected to periodic stress to modulate their optoelectronic properties. We find a remarkable shift of the optical absorption band-edge of up to ∼0.7 eV between the regions under tensile and compressive stress, greatly exceeding the strain tunability reported for transition metal dichalcogenides. This observation is supported by theoretical models that also predict that this periodic stress modulation can yield to quantum confinement of carriers at low temperatures. The possibility of generating large strain-induced variations in the local density of charge carriers opens the door for a variety of applications including photovoltaics, quantum optics, and two-dimensional optoelectronic devices. PMID:27042865

  1. Comparison of process configurations for ethanol production from acid- and alkali-pretreated corncob by Saccharomyces cerevisiae strains with and without β-glucosidase expression.

    Science.gov (United States)

    Wang, Guoqiang; Liu, Cheng; Hong, Jiefang; Ma, Yuanyuan; Zhang, Kun; Huang, Xinyu; Zou, Shaolan; Zhang, Minhua

    2013-08-01

    β-Glucosidase was shown to have synergistic effects with commercial cellulase in the hydrolysis of acid- and alkali-pretreated corncob, especially at the dose of 5 U/g biomass and 5 or 10 FPU/g biomass. An integrating yeast strain 45# expressing β-glucosidase was constructed that utilized cellobiose quickly and efficiently. Process configurations were compared under conditions of 10% solid content, 10 FPU cellulase/g biomass, 5 U β-glucosidase/g biomass (only used for parental strain W303-1A), 1g/kg yeast loading and 3.3g/kg urea supplementation. While separate hydrolysis and fermentation was optimal for W303-1A and the ethanol titer and yield reached 3.22 g/100g and 75.6% (expressed as a percentage of the theoretical yield), respectively, simultaneous saccharification and fermentation was optimal for strain 45# and the ethanol titer and yield reached 3.31 g/100g and 77.7%, respectively. These results are valuable in optimization of the process configuration and improving the yeast strain selected for cellulosic ethanol production. PMID:23735797

  2. Genomic landscapes of endogenous retroviruses unveil intricate genetics of conventional and genetically-engineered laboratory mouse strains.

    Science.gov (United States)

    Lee, Kang-Hoon; Lim, Debora; Chiu, Sophia; Greenhalgh, David; Cho, Kiho

    2016-04-01

    Laboratory strains of mice, both conventional and genetically engineered, have been introduced as critical components of a broad range of studies investigating normal and disease biology. Currently, the genetic identity of laboratory mice is primarily confirmed by surveying polymorphisms in selected sets of "conventional" genes and/or microsatellites in the absence of a single completely sequenced mouse genome. First, we examined variations in the genomic landscapes of transposable repetitive elements, named the TREome, in conventional and genetically engineered mouse strains using murine leukemia virus-type endogenous retroviruses (MLV-ERVs) as a probe. A survey of the genomes from 56 conventional strains revealed strain-specific TREome landscapes, and certain families (e.g., C57BL) of strains were discernible with defined patterns. Interestingly, the TREome landscapes of C3H/HeJ (toll-like receptor-4 [TLR4] mutant) inbred mice were different from its control C3H/HeOuJ (TLR4 wild-type) strain. In addition, a CD14 knock-out strain had a distinct TREome landscape compared to its control/backcross C57BL/6J strain. Second, an examination of superantigen (SAg, a "TREome gene") coding sequences of mouse mammary tumor virus-type ERVs in the genomes of the 46 conventional strains revealed a high diversity, suggesting a potential role of SAgs in strain-specific immune phenotypes. The findings from this study indicate that unexplored and intricate genomic variations exist in laboratory mouse strains, both conventional and genetically engineered. The TREome-based high-resolution genetics surveillance system for laboratory mice would contribute to efficient study design with quality control and accurate data interpretation. This genetics system can be easily adapted to other species ranging from plants to humans. PMID:26779669

  3. Highly efficient biosynthesis of astaxanthin in Saccharomyces cerevisiae by integration and tuning of algal crtZ and bkt.

    Science.gov (United States)

    Zhou, Pingping; Ye, Lidan; Xie, Wenping; Lv, Xiaomei; Yu, Hongwei

    2015-10-01

    Astaxanthin is a highly valued carotenoid with strong antioxidant activity and has wide applications in aquaculture, food, cosmetic, and pharmaceutical industries. The market demand for natural astaxanthin promotes research in metabolic engineering of heterologous hosts for astaxanthin production. In this study, an astaxanthin-producing Saccharomyces cerevisiae strain was created by successively introducing the Haematococcus pluvialis β-carotenoid hydroxylase (crtZ) and ketolase (bkt) genes into a previously constructed β-carotene hyperproducer. Further integration of strategies including codon optimization, gene copy number adjustment, and iron cofactor supplementation led to significant increase in the astaxanthin production, reaching up to 4.7 mg/g DCW in the shake-flask cultures which is the highest astaxanthin content in S. cerevisiae reported to date. Besides, the substrate specificity of H. pluvialis CrtZ and BKT and the probable formation route of astaxanthin from β-carotene in S. cerevisiae were figured out by expressing the genes separately and in combination. The yeast strains engineered in this work provide a basis for further improving biotechnological production of astaxanthin and might offer a useful general approach to the construction of heterologous biosynthetic pathways for other natural products. PMID:26156241

  4. Genetically engineered immunomodulatory Streptococcus thermophilus strains producing antioxidant enzymes exhibit enhanced anti-inflammatory activities.

    Science.gov (United States)

    Del Carmen, Silvina; de Moreno de LeBlanc, Alejandra; Martin, Rebeca; Chain, Florian; Langella, Philippe; Bermúdez-Humarán, Luis G; LeBlanc, Jean Guy

    2014-02-01

    The aims of this study were to develop strains of lactic acid bacteria (LAB) having both immunomodulatory and antioxidant properties and to evaluate their anti-inflammatory effects both in vitro, in different cellular models, and in vivo, in a mouse model of colitis. Different Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains were cocultured with primary cultures of mononuclear cells. Analysis of the pro- and anti-inflammatory cytokines secreted by these cells after coincubation with candidate bacteria revealed that L. delbrueckii subsp. bulgaricus CRL 864 and S. thermophilus CRL 807 display the highest anti-inflammatory profiles in vitro. Moreover, these results were confirmed in vivo by the determination of the cytokine profiles in large intestine samples of mice fed with these strains. S. thermophilus CRL 807 was then transformed with two different plasmids harboring the genes encoding catalase (CAT) or superoxide dismutase (SOD) antioxidant enzymes, and the anti-inflammatory effects of recombinant streptococci were evaluated in a mouse model of colitis induced by trinitrobenzenesulfonic acid (TNBS). Our results showed a decrease in weight loss, lower liver microbial translocation, lower macroscopic and microscopic damage scores, and modulation of the cytokine production in the large intestines of mice treated with either CAT- or SOD-producing streptococci compared to those in mice treated with the wild-type strain or control mice without any treatment. Furthermore, the greatest anti-inflammatory activity was observed in mice receiving a mixture of both CAT- and SOD-producing streptococci. The addition of L. delbrueckii subsp. bulgaricus CRL 864 to this mixture did not improve their beneficial effects. These findings show that genetically engineering a candidate bacterium (e.g., S. thermophilus CRL 807) with intrinsic immunomodulatory properties by introducing a gene expressing an antioxidant enzyme enhances its anti

  5. Enhanced Bioconversion of Cellobiose by Industrial Saccharomyces cerevisiae Used for Cellulose Utilization.

    Science.gov (United States)

    Hu, Meng-Long; Zha, Jian; He, Lin-Wei; Lv, Ya-Jin; Shen, Ming-Hua; Zhong, Cheng; Li, Bing-Zhi; Yuan, Ying-Jin

    2016-01-01

    Cellobiose accumulation and the compromised temperature for yeast fermentation are the main limiting factors of enzymatic hydrolysis process during simultaneous saccharification and fermentation (SSF). In this study, genes encoding cellobiose transporter and β-glucosidase were introduced into an industrial Saccharomyces cerevisiae strain, and evolution engineering was carried out to improve the cellobiose utilization of the engineered yeast strain. The evolved strain exhibited significantly higher cellobiose consumption rate (2.8-fold) and ethanol productivity (4.9-fold) compared with its parent strain. Besides, the evolved strain showed a high cellobiose consumption rate of 3.67 g/L/h at 34°C and 3.04 g/L/h at 38°C. Moreover, little cellobiose was accumulated during SSF of Avicel using the evolved strain at 38°C, and the ethanol yield from Avicel increased by 23% from 0.34 to 0.42 g ethanol/g cellulose. Overexpression of the genes encoding cellobiose transporter and β-glucosidase accelerated cellobiose utilization, and the improvement depended on the strain background. The results proved that fast cellobiose utilization enhanced ethanol production by reducing cellobiose accumulation during SSF at high temperature. PMID:26973619

  6. Fermentation of deproteinized cheese whey powder solutions to ethanol by engineered Saccharomyces cerevisiae: effect of supplementation with corn steep liquor and repeated-batch operation with biomass recycling by flocculation.

    Science.gov (United States)

    Silva, Ana Carina; Guimarães, Pedro M R; Teixeira, José A; Domingues, Lucília

    2010-09-01

    The lactose in cheese whey is an interesting substrate for the production of bulk commodities such as bio-ethanol, due to the large amounts of whey surplus generated globally. In this work, we studied the performance of a recombinant Saccharomyces cerevisiae strain expressing the lactose permease and intracellular beta-galactosidase from Kluyveromyces lactis in fermentations of deproteinized concentrated cheese whey powder solutions. Supplementation with 10 g/l of corn steep liquor significantly enhanced whey fermentation, resulting in the production of 7.4% (v/v) ethanol from 150 g/l initial lactose in shake-flask fermentations, with a corresponding productivity of 1.2 g/l/h. The flocculation capacity of the yeast strain enabled stable operation of a repeated-batch process in a 5.5-l air-lift bioreactor, with simple biomass recycling by sedimentation of the yeast flocs. During five consecutive batches, the average ethanol productivity was 0.65 g/l/h and ethanol accumulated up to 8% (v/v) with lactose-to-ethanol conversion yields over 80% of theoretical. Yeast viability (>97%) and plasmid retention (>84%) remained high throughout the operation, demonstrating the stability and robustness of the strain. In addition, the easy and inexpensive recycle of the yeast biomass for repeated utilization makes this process economically attractive for industrial implementation. PMID:20535525

  7. Industrial systems biology of Saccharomyces cerevisiae enables novel succinic acid cell factory.

    Directory of Open Access Journals (Sweden)

    José Manuel Otero

    Full Text Available Saccharomyces cerevisiae is the most well characterized eukaryote, the preferred microbial cell factory for the largest industrial biotechnology product (bioethanol, and a robust commerically compatible scaffold to be exploitted for diverse chemical production. Succinic acid is a highly sought after added-value chemical for which there is no native pre-disposition for production and accmulation in S. cerevisiae. The genome-scale metabolic network reconstruction of S. cerevisiae enabled in silico gene deletion predictions using an evolutionary programming method to couple biomass and succinate production. Glycine and serine, both essential amino acids required for biomass formation, are formed from both glycolytic and TCA cycle intermediates. Succinate formation results from the isocitrate lyase catalyzed conversion of isocitrate, and from the α-keto-glutarate dehydrogenase catalyzed conversion of α-keto-glutarate. Succinate is subsequently depleted by the succinate dehydrogenase complex. The metabolic engineering strategy identified included deletion of the primary succinate consuming reaction, Sdh3p, and interruption of glycolysis derived serine by deletion of 3-phosphoglycerate dehydrogenase, Ser3p/Ser33p. Pursuing these targets, a multi-gene deletion strain was constructed, and directed evolution with selection used to identify a succinate producing mutant. Physiological characterization coupled with integrated data analysis of transcriptome data in the metabolically engineered strain were used to identify 2(nd-round metabolic engineering targets. The resulting strain represents a 30-fold improvement in succinate titer, and a 43-fold improvement in succinate yield on biomass, with only a 2.8-fold decrease in the specific growth rate compared to the reference strain. Intuitive genetic targets for either over-expression or interruption of succinate producing or consuming pathways, respectively, do not lead to increased succinate. Rather, we

  8. Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Zhao, Jianzhi; Bao, Xiaoming; Li, Chen; Shen, Yu; Hou, Jin

    2016-05-01

    Monoterpenes have wide applications in the food, cosmetics, and medicine industries and have recently received increased attention as advanced biofuels. However, compared with sesquiterpenes, monoterpene production is still lagging in Saccharomyces cerevisiae. In this study, geraniol, a valuable acyclic monoterpene alcohol, was synthesized in S. cerevisiae. We evaluated three geraniol synthases in S. cerevisiae, and the geraniol synthase Valeriana officinalis (tVoGES), which lacked a plastid-targeting peptide, yielded the highest geraniol production. To improve geraniol production, synthesis of the precursor geranyl diphosphate (GPP) was regulated by comparing three specific GPP synthase genes derived from different plants and the endogenous farnesyl diphosphate synthase gene variants ERG20 (G) (ERG20 (K197G) ) and ERG20 (WW) (ERG20 (F96W-N127W) ), and controlling endogenous ERG20 expression, coupled with increasing the expression of the mevalonate pathway by co-overexpressing IDI1, tHMG1, and UPC2-1. The results showed that overexpressing ERG20 (WW) and strengthening the mevalonate pathway significantly improved geraniol production, while expressing heterologous GPP synthase genes or down-regulating endogenous ERG20 expression did not show positive effect. In addition, we constructed an Erg20p(F96W-N127W)-tVoGES fusion protein, and geraniol production reached 66.2 mg/L after optimizing the amino acid linker and the order of the proteins. The best strain yielded 293 mg/L geraniol in a fed-batch cultivation, a sevenfold improvement over the highest titer previously reported in an engineered S. cerevisiae strain. Finally, we showed that the toxicity of geraniol limited its production. The platform developed here can be readily used to synthesize other monoterpenes. PMID:26883346

  9. Cofactor modification analysis: a computational framework to identify cofactor specificity engineering targets for strain improvement.

    Science.gov (United States)

    Lakshmanan, Meiyappan; Chung, Bevan Kai-Sheng; Liu, Chengcheng; Kim, Seon-Won; Lee, Dong-Yup

    2013-12-01

    Cofactors, such as NAD(H) and NADP(H), play important roles in energy transfer within the cells by providing the necessary redox carriers for a myriad of metabolic reactions, both anabolic and catabolic. Thus, it is crucial to establish the overall cellular redox balance for achieving the desired cellular physiology. Of several methods to manipulate the intracellular cofactor regeneration rates, altering the cofactor specificity of a particular enzyme is a promising one. However, the identification of relevant enzyme targets for such cofactor specificity engineering (CSE) is often very difficult and labor intensive. Therefore, it is necessary to develop more systematic approaches to find the cofactor engineering targets for strain improvement. Presented herein is a novel mathematical framework, cofactor modification analysis (CMA), developed based on the well-established constraints-based flux analysis, for the systematic identification of suitable CSE targets while exploring the global metabolic effects. The CMA algorithm was applied to E. coli using its genome-scale metabolic model, iJO1366, thereby identifying the growth-coupled cofactor engineering targets for overproducing four of its native products: acetate, formate, ethanol, and lactate, and three non-native products: 1-butanol, 1,4-butanediol, and 1,3-propanediol. Notably, among several target candidates for cofactor engineering, glyceraldehyde-3-phosphate dehydrogenase (GAPD) is the most promising enzyme; its cofactor modification enhanced both the desired product and biomass yields significantly. Finally, given the identified target, we further discussed potential mutational strategies for modifying cofactor specificity of GAPD in E. coli as suggested by in silico protein docking experiments. PMID:24372035

  10. L-Histidine inhibits biofilm formation and FLO11- associated phenotypes in Saccharomyces cerevisiae flor yeasts

    OpenAIRE

    Bou Zeidan, Marc; Zara, Giacomo; Viti, Carlo; Decorosi, Francesca; Mannazzu, Ilaria Maria; Budroni, Marilena; Giovannetti, Luciana; Zara, Severino

    2014-01-01

    Flor yeasts of Saccharomyces cerevisiae have an innate diversity of FLO11 which codes for a highly hydrophobic and anionic cell-wall glycoprotein with a fundamental role in biofilm formation. In this study, 380 nitrogen compounds were administered to three S. cerevisiae flor strains handling FLO11 alleles with different expression levels. S. cerevisiae strain S288c was used as the reference strain as it cannot produce FLO11p. The flor strains generally metabolized amino acids and dipeptides a...

  11. Strain-induced band-gap engineering of graphene monoxide and its effect on graphene

    Science.gov (United States)

    Pu, H. H.; Rhim, S. H.; Hirschmugl, C. J.; Gajdardziska-Josifovska, M.; Weinert, M.; Chen, J. H.

    2013-02-01

    Using first-principles calculations we demonstrate the feasibility of band-gap engineering in two-dimensional crystalline graphene monoxide (GMO), a recently reported graphene-based material with a 1:1 carbon/oxygen ratio. The band gap of GMO, which can be switched between direct and indirect, is tunable over a large range (0-1.35 eV) for accessible strains. Electron and hole transport occurs predominantly along the zigzag and armchair directions (armchair for both) when GMO is a direct- (indirect-) gap semiconductor. A band gap of ˜0.5 eV is also induced in graphene at the K' points for GMO/graphene hybrid systems.

  12. Band gap engineering in polymers through chemical doping and applied mechanical strain.

    Science.gov (United States)

    Lanzillo, Nicholas A; Breneman, Curt M

    2016-08-17

    We report simulations based on density functional theory and many-body perturbation theory exploring the band gaps of common crystalline polymers including polyethylene, polypropylene and polystyrene. Our reported band gaps of 8.6 eV for single-chain polyethylene and 9.1 eV for bulk crystalline polyethylene are in excellent agreement with experiment. The effects of chemical doping along the polymer backbone and side-groups are explored, and the use mechanical strain as a means to modify the band gaps of these polymers over a range of several eV while leaving the dielectric constant unchanged is discussed. This work highlights some of the opportunities available to engineer the electronic properties of polymers with wide-reaching implications for polymeric dielectric materials used for capacitive energy storage. PMID:27324304

  13. Strain-Engineered Low-Density InAs Bilayer Quantum Dots for Single Photon Emission

    Institute of Scientific and Technical Information of China (English)

    LI Zhan-Guo; LIU Guo-Jun; LI Lin; FENG Ming; LI Mei; LU Peng; ZOU Yong-Gang; LI Lian-He; GAO Xin

    2010-01-01

    @@ We investigate the growth of strain-engineered low-density InAs bilayer quantum dots(BQDs)on GaAs bymolecular beam epitaxy.Owing to increasing dot size and In composition of the upper QDs,low-density BQDs in a GaAs matrix with an emission wavelength up to 1.4 μm at room temperature are achieved.Such a wavelength is larger than that of conventional QDs in a GaAs matrix(generally of about 1.3 μm).The optical properties of the BQDs are sensitive to annealing temperature used after spacer layer growth.Significant decrease of integrated PL intensity is observed as the annealing temperature increases.At IO K,single photon emission from the BQDs with wavelength around 1.3μm is observed.

  14. Strain-Engineered Low-Density InAs Bilayer Quantum Dots for Single Photon Emission

    International Nuclear Information System (INIS)

    We investigate the growth of strain-engineered low-density InAs bilayer quantum dots (BQDs) on GaAs by molecular beam epitaxy. Owing to increasing dot size and In composition of the upper QDs, low-density BQDs in a GaAs matrix with an emission wavelength up to 1.4 μm at room temperature are achieved. Such a wavelength is larger than that of conventional QDs in a GaAs matrix (generally of about 1.3 μm). The optical properties of the BQDs are sensitive to annealing temperature used after spacer layer growth. Significant decrease of integrated PL intensity is observed as the annealing temperature increases. At 10K, single photon emission from the BQDs with wavelength around 1.3 μm is observed. (condensed matter: structure, mechanical and thermal properties)

  15. Band gap engineering in polymers through chemical doping and applied mechanical strain

    Science.gov (United States)

    Lanzillo, Nicholas A.; Breneman, Curt M.

    2016-08-01

    We report simulations based on density functional theory and many-body perturbation theory exploring the band gaps of common crystalline polymers including polyethylene, polypropylene and polystyrene. Our reported band gaps of 8.6 eV for single-chain polyethylene and 9.1 eV for bulk crystalline polyethylene are in excellent agreement with experiment. The effects of chemical doping along the polymer backbone and side-groups are explored, and the use mechanical strain as a means to modify the band gaps of these polymers over a range of several eV while leaving the dielectric constant unchanged is discussed. This work highlights some of the opportunities available to engineer the electronic properties of polymers with wide-reaching implications for polymeric dielectric materials used for capacitive energy storage.

  16. Engineering control of bacterial cellulose production using a genetic toolkit and a new cellulose-producing strain

    OpenAIRE

    Florea, Michael; Hagemann, Henrik; Santosa, Gabriella; Abbott, James; Micklem, Chris N.; Spencer-Milnes, Xenia; de Arroyo Garcia, Laura; Paschou, Despoina; Lazenbatt, Christopher; Kong, Deze; Chughtai, Haroon; Jensen, Kirsten; Freemont, Paul S.; Kitney, Richard; Reeve, Benjamin

    2016-01-01

    Bacterial cellulose is a remarkable material that is malleable, biocompatible, and over 10-times stronger than plant-based cellulose. It is currently used to create materials for tissue engineering, medicine, defense, electronics, acoustics, and fabrics. We describe here a bacterial strain that is readily amenable to genetic engineering and produces high quantities of bacterial cellulose in low-cost media. To reprogram this organism for biotechnology applications, we created a set of genetic ...

  17. Elastic strain engineering of quantum dot excitonic emission in nanomembranes and optical resonators

    International Nuclear Information System (INIS)

    We study the effect of an external biaxial stress on the light emission of single InGaAs/GaAs(001) quantum dots (QD) embedded in a 200 nm-thick-membrane. Reversible and bi-directional spectral tuning of QD excitonic emission is demonstrated via a simple electro-mechanical device. The most intriguing finding is that biaxial strain is a reliable tool to engineer the QD electronic structure and reach color coincidence between exciton and biexciton emission, providing a vital prerequisite for the generation of polarization entangled photon pairs through a time reordering strategy. The physical origin of this new phenomenon is discussed based on the empirical pseudopotential calculations. With similar technique we study the effect of biaxial stress on single QDs embedded in microring resonators. The microrings can be reversibly stretched or squeezed, resulting in a controllable engineering of both QD emissions and optical modes. Our results open up a new tuning strategy to study cQED with semiconductor quantum dots.

  18. Nicotinamide cofactor ratios in engineered strains of Clostridium thermocellum and Thermoanaerobacterium saccharolyticum.

    Science.gov (United States)

    Beri, Dhananjay; Olson, Daniel G; Holwerda, Evert K; Lynd, Lee R

    2016-06-01

    Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are bacteria under investigation for production of biofuels from plant biomass. Thermoanaerobacterium saccharolyticum has been engineered to produce ethanol at high yield (>90% of theoretical) and titer (>70 g/l). Efforts to engineer C. thermocellum have not, to date, been as successful, and efforts are underway to transfer the ethanol production pathway from T. saccharolyticum to C. thermocellum One potential challenge in transferring metabolic pathways is the possibility of incompatible levels of nicotinamide cofactors. These cofactors (NAD(+), NADH, NADP(+) and NADPH) and their oxidation state are important in the context of microbial redox metabolism. In this study we directly measured the concentrations and reduced oxidized ratios of these cofactors in a number of strains of C. thermocellum and T. saccharolyticum by using acid/base extraction and enzymatic assays. We found that cofactor ratios are maintained in a fairly narrow range, regardless of the metabolic network modifications considered. We have found that the ratios are similar in both organisms, which is a relevant observation in the context of transferring the T. saccharolyticum ethanol production pathway to C. thermocellum. PMID:27190292

  19. TOTAL ANTIOXIDANT ACTIVITY OF YEAST SACCHAROMYCES CEREVISIAE

    Directory of Open Access Journals (Sweden)

    Blažena Lavová

    2013-02-01

    Full Text Available Antioxidants are health beneficial compounds that can protect cells and macromolecules (e.g. fats, lipids, proteins and DNA from the damage of reactive oxygen species (ROS. Sacchamomyces cerevisiae are know as organisms with very important antioxidative enzyme systems such as superoxide dismutase or catalase. The total antioxidant activity (mmol Trolox equivalent – TE.g-1 d.w. of Saccharomyces cerevisiae was measured by 2,2´-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid during the yeast cultivation. It was found that the total antioxidant activity was the highest (1.08 mmol TE.g-1 d.w. in the strain Kolín after 32 hours of cultivation and the lowest (0.26 mmol TE.g-1 d.w. in the strain Gyöng after 12 hours of cultivation.

  20. III-V strain layer superlattice based band engineered avalanche photodiodes (Presentation Recording)

    Science.gov (United States)

    Ghosh, Sid

    2015-08-01

    Laser detection and ranging (LADAR)-based systems operating in the Near Infrared (NIR) and Short Wave Infrared (SWIR) have become popular optical sensors for remote sensing, medical, and environmental applications. Sophisticated laser-based radar and weapon systems used for long-range military and astronomical applications need to detect, recognize, and track a variety of targets under a wide spectrum of atmospheric conditions. Infrared APDs play an important role in LADAR systems by integrating the detection and gain stages in a single device. Robust silicon-APDs are limited to visible and very near infrared region ( 3um) infrared photon detection applications. Recently, various research groups (including Ghosh et. al.) have reported SWIR and MWIR HgCdTe APDs on CdZnTe and Si substrates. However, HgCdTe APDs suffer from low breakdown fields due to material defects, and excess noise increases significantly at high electric fields. During the past decade, InAs/GaSb Strain Layer Superlattice (SLS) material system has emerged as a potential material for the entire infrared spectrum because of relatively easier growth, comparable absorption coefficients, lower tunneling currents and longer Auger lifetimes resulting in enhanced detectivities (D*). Band engineering in type II SLS allows us to engineer avalanche properties of electrons and holes. This is a great advantage over bulk InGaAs and HgCdTe APDs where engineering avalanche properties is not possible. The talk will discuss the evolution of superlattice based avalanche photodiodes and some of the recent results on the work being done at Raytheon on SWIR avalanche photodiodes.

  1. Isolation of the catalase T structural gene of Saccharomyces cerevisiae by functional complementation.

    OpenAIRE

    Spevak, W; Fessl, F; Rytka, J; Traczyk, A; Skoneczny, M; Ruis, H

    1983-01-01

    The catalase T structural gene of Saccharomyces cerevisiae was cloned by functional complementation of a mutation causing specific lack of the enzyme (cttl). Catalase T-deficient mutants were obtained by UV mutagenesis of an S. cerevisiae strain bearing the cas1 mutation, which causes insensitivity of catalase T to glucose repression. Since the second catalase protein of S. cerevisiae, catalase A, is completely repressed on 10% glucose, catalase T-deficient mutant colonies could be detected u...

  2. Construction of Killer Industrial Yeast Saccharomyces Cerevisiae Hau-1 and its Fermentation Performance

    OpenAIRE

    Bajaj, Bijender K.; S Sharma

    2010-01-01

    Saccharomyces cerevisiae HAU-1, a time tested industrial yeast possesses most of the desirable fermentation characteristics like fast growth and fermentation rate, osmotolerance, high ethanol tolerance, ability to ferment molasses, and to ferment at elevated temperatures etc. However, this yeast was found to be sensitive against the killer strains of Saccharomyces cerevisiae. In the present study, killer trait was introduced into Saccharomyces cerevisiae HAU-1 by protoplast fusion with Saccha...

  3. Construction of killer industrial yeast Saccharomyces cerevisiae HAU-1 and its fermentation performance

    OpenAIRE

    Bajaj, Bijender K.; S Sharma

    2010-01-01

    Saccharomyces cerevisiae HAU-1, a time tested industrial yeast possesses most of the desirable fermentation characteristics like fast growth and fermentation rate, osmotolerance, high ethanol tolerance, ability to ferment molasses, and to ferment at elevated temperatures etc. However, this yeast was found to be sensitive against the killer strains of Saccharomyces cerevisiae. In the present study, killer trait was introduced into Saccharomyces cerevisiae HAU-1 by protoplast fusion with Saccha...

  4. Phosphate transport and sensing in Saccharomyces cerevisiae.

    OpenAIRE

    Wykoff, D D; O'Shea, E K

    2001-01-01

    Cellular metabolism depends on the appropriate concentration of intracellular inorganic phosphate; however, little is known about how phosphate concentrations are sensed. The similarity of Pho84p, a high-affinity phosphate transporter in Saccharomyces cerevisiae, to the glucose sensors Snf3p and Rgt2p has led to the hypothesis that Pho84p is an inorganic phosphate sensor. Furthermore, pho84Delta strains have defects in phosphate signaling; they constitutively express PHO5, a phosphate starvat...

  5. Converting Carbon Dioxide to Butyrate with an Engineered Strain of Clostridium ljungdahlii

    Energy Technology Data Exchange (ETDEWEB)

    Ueki, T; Nevin, KP; Woodard, TL; Lovley, DR

    2014-08-26

    Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70% of the carbon and electron flow was diverted to the production of butyrate with H-2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. IMPORTANCE The development of a microbial chassis for efficient conversion of carbon dioxide directly to desired organic products would greatly advance the environmentally sustainable production of biofuels and other commodities. Clostridium ljungdahlii is an effective catalyst for microbial electrosynthesis, a technology in which electricity generated with renewable technologies, such as solar or wind, powers the conversion of carbon dioxide and water to organic products. Other electron donors

  6. Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii.

    Science.gov (United States)

    Ueki, Toshiyuki; Nevin, Kelly P; Woodard, Trevor L; Lovley, Derek R

    2014-01-01

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

  7. Phenotypical signs and chemical composition of Saccharomyces cerevisiae – mannoprotein producers

    Directory of Open Access Journals (Sweden)

    Agafia USATII

    2012-11-01

    Full Text Available Phenotypical signs and chemical composition of Saccharomyces cerevisiae CNMN-Y-18 and Saccharomyces cerevisiae CNMN-Y-19 yeast strains are described in this article. The presence of protein complexes with high content of irreplaceable amino acids and antioxidant enzymes, as well as polysaccharides with predominance of mannoproteins allow to recommend these yeast strains for the utilization in biotechnology. Results are of interest for the standard description of yeast strains offered as object for industrial appointment.

  8. Surface display of malolactic enzyme from Oenococcus oeni on Saccharomyces cerevisiae.

    Science.gov (United States)

    Zhang, Xiuyan; Hou, Xiaoyan; Liang, Fang; Chen, Fusheng; Wang, Xiaohong

    2013-04-01

    In order to display malolactic enzyme (MLE) on the cell surface of Saccharomyces cerevisiae, a yeast cell surface display plasmid pADH1-AGG was constructed by fusing the α-factor signal encoding sequence (267 bp) and the C-terminal half of α-agglutinin encoding sequence (1,645 bp) into the plasmid pADH1. The pADH1-AGG could successfully express and anchor the enhanced green fluorescent protein (EGFP) onto the yeast cell surface when the EGFP was used to verify its function. Then the pADH1-MLE was constructed by inserting the MLE encoding sequence (1,600 bp) into the pADH1-AGG and introduced into S. cerevisiae cells. The positive strain carrying pADH1-MLE was confirmed by use of the 6× His monoclonal antibody and fluorescein isothiocyanate-conjugated goat anti-mouse IgG. All results indicated that the MLE was displayed successfully on the cell surface of positive transformant. The MLE activity of genetically engineered yeast strain could turn 21.11 % L-malate into lactic acid after 12 h reaction with L-malate. The constructed yeast strain might be used to conduct malolactic fermentation (MLF) in wine to solve the important issues of sluggish MLF, microbial spoilage, and adverse metabolic substances produced by the lactic acid bacteria. PMID:23446978

  9. Advances in metabolic pathway and strain engineering paving the way for sustainable production of chemical building blocks

    DEFF Research Database (Denmark)

    Chen, Yun; Nielsen, Jens

    2013-01-01

    Bio-based production of chemical building blocks from renewable resources is an attractive alternative to petroleum-based platform chemicals. Metabolic pathway and strain engineering is the key element in constructing robust microbial chemical factories within the constraints of cost effective...

  10. Selection of indigenous Saccharomyces cerevisiae strains in Tianjin and their oenological characterization of low-alcohol white wine%本土酿酒酵母的筛选及其低醇葡萄酒特性研究

    Institute of Scientific and Technical Information of China (English)

    崔艳; 吕文; 王伟; 蒋珍珍; 刘金福

    2011-01-01

    以天津宝坻产区的三种葡萄为原料,分别从自然发酵的葡萄汁中筛选出340株酵母菌,经生理生化,产酸、酯、尿酶、H2S,发酵能力测定及26SrRNA测序,确定4株为酿酒酵母,用于贵人香低醇白葡萄酒的酿造。以商用酵母为对照,定量分析了不同酵母所酿酒中的香气成分、氨基甲酸乙酯含量,并进行了理化指标和感官测定,发现酵母C508和G611酿造的低醇酒具有更独特的香气特点,氨基甲酸乙酯的含量较低,微生物稳定性较高,更适合低醇酒的酿造。%In order to identify the Saccharomyces spp.associated with spontaneous fermentation of three different grape varieties(Italian Riesling,Muscat and Cabernet Sauvignon grapes)from Tianjin Baodi wine region and to evaluate the micro-fermentation performance of low-alcohol white wine.The indigeous yeast flora isolated during fermentation was studied and analyzed.Firstly,yeasts were identified to genus level by growth on WL nutrient agar and the tests of assimilation carbon and nitrogen source and so on.Later,formation of amyloid,acids,esters,H2S and urease and fermentation power were studied.Three hundred and forty strains were isolated,four of which were identified as Saccharomyces cerevisiae by PCR of D1/D2 regions of the 26S rRNA gene and tested as starters in low-alcohol white wine fermentations for their satisfactory oenological properties.The quality of the produced wines was evaluated after determination of their physical-chemical parameters,volatile compounds and ethyl carbamate produced in wine fermentation.The results showed the potential of employing indigenous yeast strains for the production low-alcohol white wine with improved stability,a richer aromatic bouquet,less ethyl carbamate concentration.

  11. Activation of futile cycles as an approach to increase ethanol yield during glucose fermentation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Semkiv, Marta V; Dmytruk, Kostyantyn V; Abbas, Charles A; Sibirny, Andriy A

    2016-04-01

    An increase in ethanol yield by yeast from the fermentation of conventional sugars such as glucose and sucrose is possible by reducing the production of a key byproduct such as cellular biomass. Previously we have reported that overexpression of PHO8 gene encoding non-specific ATP-hydrolyzing alkaline phosphatase can lead to a decrease in cellular ATP content and to an increase in ethanol yield during glucose fermentation by Saccharomyces cerevisiae. In this work we further report on 2 new successful approaches to reduce cellular levels of ATP that increase ethanol yield and productivity. The first approach is based on the overexpression of the heterologous Escherichia coli apy gene encoding apyrase or SSB1 part of the chaperon that exhibit ATPase activity in yeast. In the second approach we constructed a futile cycle by the overexpression of S. cerevisiae genes encoding pyruvate carboxylase and phosphoenolpyruvate carboxykinase in S. cerevisiae. These genetically engineered strains accumulated more ethanol compared to the wild-type strain during alcoholic fermentation. PMID:26890808

  12. Electrospun polycaprolactone scaffolds under strain and their application in cartilage tissue engineering

    Science.gov (United States)

    Nam, Jin

    Electrospinning is a promising fabrication method for three dimensional tissue engineering scaffolds due to its ability to produce a nano-/micro-sized non-woven fibrous structure which resembles the natural extracellular matrix. We investigated the mechanical behavior of two different electrospun microstructures. Polycaprolactone (PCL) fibers with or without "point-bonding" exhibited different deformation behaviors having significant biomedical consequences. While fibers with point-bonded structure failed due to the generation of voids by the fracture of fiber interconnections under strain, fibers without point-bonds produced a 'bamboo' structure with fiber joining visible at higher levels of strain. In addition, gelatin and PCL were electrospun and the residual solvent contents were systematically investigated. A simple and effective means of reducing residual solvent content was developed. The interaction between these electrospun matrices and chondrocytic cells were compared to other topographies having the same chemistry. Electrospun polycaprolactone fibers supported better proliferation and extracellular matrix production than the corresponding semi-porous and dense surfaces and even, at some time points, glass surfaces. The intrinsic capability of electrospinning to produce high porosity appears to offset the relative hydrophobicity of polycaprolactone resulting in a more uniform cell seeding. Electrospun fibers induced a higher level of glycosaminoglycans (GAG) production by providing a 'dynamic scaffold' in which chondrocytes are able to maintain a morphology associated with the appropriate phenotype. Finally, based on this study, a method producing macro-pores within an electrospun scaffold was developed. With this method, not only can cellular infiltration into a thick electrospun scaffold be facilitated, but scaffolds having designed, anisotropic structures can be produced that better approximate the final tissue.

  13. Methionine catabolism in Saccharomyces cerevisiae.

    Science.gov (United States)

    Perpète, Philippe; Duthoit, Olivier; De Maeyer, Simon; Imray, Louise; Lawton, Andrew I; Stavropoulos, Konstantinos E; Gitonga, Virginia W; Hewlins, Michael J E; Dickinson, J Richard

    2006-01-01

    The catabolism of methionine to methionol and methanethiol in Saccharomyces cerevisiae was studied using (13)C NMR spectroscopy, GC-MS, enzyme assays and a number of mutants. Methionine is first transaminated to alpha-keto-gamma-(methylthio)butyrate. Methionol is formed by a decarboxylation reaction, which yields methional, followed by reduction. The decarboxylation is effected specifically by Ydr380wp. Methanethiol is formed from both methionine and alpha-keto-gamma-(methylthio)butyrate by a demethiolase activity. In all except one strain examined, demethiolase was induced by the presence of methionine in the growth medium. This pathway results in the production of alpha-ketobutyrate, a carbon skeleton, which can be re-utilized. Hence, methionine catabolism is more complex and economical than the other amino acid catabolic pathways in yeast, which use the Ehrlich pathway and result solely in the formation of a fusel alcohol. PMID:16423070

  14. Metabolic engineering of bacteria for environmental applications: construction of Pseudomonas strains for biodegradation of 2-chlorotoluene.

    Science.gov (United States)

    Haro, M A; de Lorenzo, V

    2001-02-13

    In this article, we illustrate the challenges and bottlenecks in the metabolic engineering of bacteria destined for environmental bioremediation, by reporting current efforts to construct Pseudomonas strains genetically designed for degradation of the recalcitrant compound 2-chlorotoluene. The assembled pathway includes one catabolic segment encoding the toluene dioxygenase of the TOD system of Pseudomonas putida F1 (todC1C2BA), which affords the bioconversion of 2-chlorotoluene into 2-chlorobenzaldehyde by virtue of its residual methyl-monooxygenase activity on o-substituted substrates. A second catabolic segment encoded the entire upper TOL pathway from pWW0 plasmid of P. putida mt-2. The enzymes, benzyl alcohol dehydrogenase (encoded by xylB) and benzaldehyde dehydrogenase (xylC) of this segment accept o-chloro-substituted substrates all the way down to 2-chlorobenzoate. These TOL and TOD segments were assembled in separate mini-Tn5 transposon vectors, such that expression of the encoded genes was dependent on the toluene-responsive Pu promoter of the TOL plasmid and the cognate XylR regulator. Such gene cassettes (mini-Tn5 [UPP2] and mini-Tn5 [TOD2]) were inserted in the chromosome of the 2-chlorobenzoate degraders Pseudomonas aeruginosa PA142 and P. aeruginosa JB2. GC-MS analysis of the metabolic intermediates present in the culture media of the resulting strains verified that these possessed, not only the genetic information, but also the functional ability to mineralise 2-chlorotoluene. However, although these strains did convert the substrate into 2-chlorobenzoate, they failed to grow on 2-chlorotoluene as the only carbon source. These results pinpoint the rate of the metabolic fluxes, the non-productive spill of side-metabolites and the physiological control of degradative pathways as the real bottlenecks for degradation of certain pollutants, rather than the theoretical enzymatic and genetic fitness of the recombinant bacteria to the process. Choices to

  15. Construction and evaluation of an exopolysaccharide-producing engineered bacterial strain by protoplast fusion for microbial enhanced oil recovery.

    Science.gov (United States)

    Sun, Shanshan; Luo, Yijing; Cao, Siyuan; Li, Wenhong; Zhang, Zhongzhi; Jiang, Lingxi; Dong, Hanping; Yu, Li; Wu, Wei-Min

    2013-09-01

    Enterobacter cloacae strain JD, which produces water-insoluble biopolymers at optimal temperature of 30°C, and a thermophilic Geobacillus strain were used to construct an engineered strain for exopolysaccharide production at high temperatures by protoplast fusion. The obtained fusant strain ZR3 produced exopolysaccharides at up to 45°C with optimal growth temperature at 35°C. The fusant produced exopolysaccharides of approximately 7.5 g/L or more at pH between 7.0 and 9.0. The feasibility of the enhancement of crude oil recovery with the fusant was tested in a sand-packed column at 40°C. The results demonstrated that bioaugmentation of the fusant was promising approach for MEOR. Mass growth of the fusant was confirmed in fermentor tests. PMID:23856587

  16. Electronic structure and magnetism in g-C{sub 4}N{sub 3} controlled by strain engineering

    Energy Technology Data Exchange (ETDEWEB)

    Liu, L. Z.; Liu, X. X. [Key Laboratory of Modern Acoustics, MOE, Institute of Acoustics, Collaborative Innovation Center of Advanced Microstructures, National Laboratory of Solid State Microstructures, Nanjing University, Nanjing 210093 (China); Wu, X. L., E-mail: hkxlwu@nju.edu.cn, E-mail: paul.chu@cityu.edu.hk [Key Laboratory of Modern Acoustics, MOE, Institute of Acoustics, Collaborative Innovation Center of Advanced Microstructures, National Laboratory of Solid State Microstructures, Nanjing University, Nanjing 210093 (China); Department of Physics, NingBo University, NingBo 315001 (China); Chu, Paul K., E-mail: hkxlwu@nju.edu.cn, E-mail: paul.chu@cityu.edu.hk [Department of Physics and Materials Science, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong (China)

    2015-03-30

    Regulation of magnetism and half-metallicity has attracted much attention because of its potential in spintronics. The magnetic properties and electronic structure of graphitic carbon nitride (g-C{sub 4}N{sub 3}) with external strain are determined theoretically based on the density function theory and many-body perturbation theory (G{sub 0}W{sub 0}). Asymmetric deformation induced by uniaxial strain not only regulates the magnetic characteristics but also leads to a transformation from half-metallicity to metallicity. However, this transition cannot occur in the structure with symmetric deformation induced by biaxial strain. Our results suggest the use of strain engineering in metal-free spintronics applications.

  17. Functional co-operation between the nuclei of Saccharomyces cerevisiae and mitochondria from other yeast species

    DEFF Research Database (Denmark)

    Spirek, M.; Horvath, A.; Piskur, Jure;

    2000-01-01

    We elaborated a simple method that allows the transfer of mitochondria from collection yeasts to Saccharomyces cerevisiae. Protoplasts prepared from different yeasts were fused to the protoplasts of the ade2-1, ura3-52, kar1-1, rho (0) strain of S. cerevisiae and were selected for respiring cybrids...

  18. EasyClone: method for iterative chromosomal integration of multiple genes in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Jensen, Niels Bjerg; Strucko, Tomas; Kildegaard, Kanchana Rueksomtawin;

    2014-01-01

    Development of strains for efficient production of chemicals and pharmaceuticals requires multiple rounds of genetic engineering. In this study, we describe construction and characterization of EasyClone vector set for baker's yeast Saccharomyces cerevisiae, which enables simultaneous expression of...... multiple genes with an option of recycling selection markers. The vectors combine the advantage of efficient uracil excision reaction-based cloning and Cre-LoxP-mediated marker recycling system. The episomal and integrative vector sets were tested by inserting genes encoding cyan, yellow, and red...

  19. Multiple gene mediated aldehyde reduction is a mechanism of in situ detoxification of furfural and 5-hydroxymethylfurfural by Saccharomyces cerevisiae

    Science.gov (United States)

    Furfural and HMF (5-hydroxymethylfurfural) are representative inhibitors to ethanologenic yeast generated from biomass pretreatment using dilute acid hydrolysis. Few yeast strains tolerant to inhibitors are available. We have developed tolerant strains of Saccharomyces cerevisiae with enhanced bio...

  20. Adaption of Saccharomyces cerevisiae expressing a heterologous protein

    DEFF Research Database (Denmark)

    Krogh, Astrid Mørkeberg; Beck, Vibe; Højlund Christensen, Lars;

    2008-01-01

    Production of the heterologous protein, bovine aprotinin, in Saccharomyces cerevisiae was shown to affect the metabolism of the host cell to various extent depending on the strain genotype. Strains with different genotypes, industrial and laboroatory, respectively, were investigated. The maximal ...... result of the adaptation. Determination of the level of mRNA encoding aprotinin and the plasmid copy number pointed to different mechanisms responsible for the decline in aprotinin yield in the different strains. (C) 2008 Elsevier B.V. All rights reserved....

  1. Direct production of ethanol from raw corn starch via fermentation by use of a novel surface-engineered yeast strain codisplaying glucoamylase and alpha-amylase.

    Science.gov (United States)

    Shigechi, Hisayori; Koh, Jun; Fujita, Yasuya; Matsumoto, Takeshi; Bito, Yohei; Ueda, Mitsuyoshi; Satoh, Eiichi; Fukuda, Hideki; Kondo, Akihiko

    2004-08-01

    Direct and efficient production of ethanol by fermentation from raw corn starch was achieved by using the yeast Saccharomyces cerevisiae codisplaying Rhizopus oryzae glucoamylase and Streptococcus bovis alpha-amylase by using the C-terminal-half region of alpha-agglutinin and the flocculation functional domain of Flo1p as the respective anchor proteins. In 72-h fermentation, this strain produced 61.8 g of ethanol/liter, with 86.5% of theoretical yield from raw corn starch. PMID:15294847

  2. Functional replacement of the Saccharomyces cerevisiae fatty acid synthase with a bacterial type II system allows flexible product profiles.

    Science.gov (United States)

    Fernandez-Moya, Ruben; Leber, Christopher; Cardenas, Javier; Da Silva, Nancy A

    2015-12-01

    The native yeast type I fatty acid synthase (FAS) is a complex, rigid enzyme, and challenging to engineer for the production of medium- or short-chain fatty acids. Introduction of a type II FAS is a promising alternative as it allows expression control for each discrete enzyme and the addition of heterologous thioesterases. In this study, the native Saccharomyces cerevisiae FAS was functionally replaced by the Escherichia coli type II FAS (eFAS) system. The E. coli acpS + acpP (together), fabB, fabD, fabG, fabH, fabI, fabZ, and tesA were expressed in individual S. cerevisiae strains, and enzyme activity was confirmed by in vitro activity assays. Eight genes were then integrated into the yeast genome, while tesA or an alternate thioesterase gene, fatB from Ricinus communis or TEII from Rattus novergicus, was expressed from a multi-copy plasmid. Native FAS activity was eliminated by knocking out the yeast FAS2 gene. The strains expressing only the eFAS as de novo fatty acid source grew without fatty acid supplementation demonstrating that this type II FAS is able to functionally replace the native yeast FAS. The engineered strain expressing the R. communis fatB thioesterase increased total fatty acid titer 1.7-fold and shifted the fatty acid profile towards C14 production, increasing it from fatty acids, and reducing C18 production from 39% to 8%. PMID:26084339

  3. [Construction of Saccharomyces cerevisiae cell factories for lycopene production].

    Science.gov (United States)

    Shi, Ming-Yu; Liu Yi; Wang, Dong; Lu, Fu-Ping; Huang, Lu-Qi; Dai, Zhu-Bo; Zhang, Xue-Li

    2014-10-01

    For microbial production of lycopene, the lycopene synthetic genes from Pantoea agglomerans were integrated into Saccharomyces cerevisiae strain BY4742, to obtain strain ZD-L-000 for production of 0.17 mg · L(-1) lycopene. Improving supplies of isoprenoid precursors was then investigated for increasing lycopene production. Four key genes were chosen to be overexpressed, inclu- ding truncated 3-hydroxy-3-methylglutaryl-CoA reductase gene (tHMG1), which is the major rate-limiting enzyme in the mevalonate (MVA) pathway, a mutated global regulatory factor gene (upc2.1), a fusion gene of FPP synthase (ERG20) and endogenous GGPP synthase (BTS1), which is a key enzyme in the diterpenoid synthetic pathway, and GGPP synthase gene (SaGGPS) from Sulfolobus acidocaldarius. Over-expression of upc2.1 could not improve lycopene production, while over-expression of tHMGI , BTS1-ERG20 and SaGGPS genes led to 2-, 16. 9- and20. 5-fold increase of lycopene production, respectively. In addition, three effective genes, tHMG1, BTS1-ERG20 and SaGGPS, were integrated into rDNA sites of ZD-L-000, resulting in strain ZD-L-201 for production of 13.23 mg · L(-1) lycopene, which was 77-fold higher than that of the parent strain. Finally, two-phase extractive fermentation was performed. The titer of lycopene increased 10-fold to 135.21 mg · L(-1). The engineered yeast strains obtained in this work provided the basis for fermentative production of lycopene. PMID:25751950

  4. Expression of native and mutant extracellular lipases fromYarrowia lipolytica in Saccharomyces cerevisiae

    OpenAIRE

    Darvishi, Farshad

    2012-01-01

    Summary Saccharomyces cerevisiae cannot produce extracellular lipase and utilize low‐cost lipid substrates. This study aimed to express extracellular lipase from Yarrowia lipolytica in S. cerevisiae, construct recombinant oily substrate consumer strains, and compare the roles of native and mutant Y. lipolytica extracellular lipases in S. cerevisiae. The LIP2 gene of Y. lipolytica DSM3286 and its mutant Y. lipolytica U6 were isolated and cloned by expression vector in S. cerevisiae. New recomb...

  5. Engineering Cofactor Preference of Ketone Reducing Biocatalysts: A Mutagenesis Study on a γ-Diketone Reductase from the Yeast Saccharomyces cerevisiae Serving as an Example

    Directory of Open Access Journals (Sweden)

    Michael Katzberg

    2010-04-01

    Full Text Available The synthesis of pharmaceuticals and catalysts more and more relies on enantiopure chiral building blocks. These can be produced in an environmentally benign and efficient way via bioreduction of prochiral ketones catalyzed by dehydrogenases. A productive source of these biocatalysts is the yeast Saccharomyces cerevisiae, whose genome also encodes a reductase catalyzing the sequential reduction of the γ-diketone 2,5-hexanedione furnishing the diol (2S,5S-hexanediol and the γ-hydroxyketone (5S-hydroxy-2-hexanone in high enantio- as well as diastereoselectivity (ee and de >99.5%. This enzyme prefers NADPH as the hydrogen donating cofactor. As NADH is more stable and cheaper than NADPH it would be more effective if NADH could be used in cell-free bioreduction systems. To achieve this, the cofactor binding site of the dehydrogenase was altered by site-directed mutagenesis. The results show that the rational approach based on a homology model of the enzyme allowed us to generate a mutant enzyme having a relaxed cofactor preference and thus is able to use both NADPH and NADH. Results obtained from other mutants are discussed and point towards the limits of rationally designed mutants.

  6. Band gap tuning of epitaxial SrTiO{sub 3-δ}/Si(001) thin films through strain engineering

    Energy Technology Data Exchange (ETDEWEB)

    Cottier, Ryan J.; Steinle, Nathan A.; Currie, Daniel A.; Theodoropoulou, Nikoleta, E-mail: ntheo@txstate.edu [Physics Department, Texas State University, San Marcos, Texas 78666 (United States)

    2015-11-30

    We investigate the effect of strain and oxygen vacancies (V{sub O}) on the crystal and optical properties of oxygen deficient, ultra-thin (4–30 nm) films of SrTiO{sub 3-δ} (STO) grown heteroepitaxially on p-Si(001) substrates by molecular beam epitaxy. We demonstrate that STO band gap tuning can be achieved through strain engineering and show that the energy shift of the direct energy gap transition of SrTiO{sub 3-δ}/Si films has a quantifiable dimensional and doping dependence that correlates well with the changes in crystal structure.

  7. Construction of killer industrial yeast Saccharomyces cerevisiae HAU-1 and its fermentation performance

    Directory of Open Access Journals (Sweden)

    Bijender K. Bajaj

    2010-06-01

    Full Text Available Saccharomyces cerevisiae HAU-1, a time tested industrial yeast possesses most of the desirable fermentation characteristics like fast growth and fermentation rate, osmotolerance, high ethanol tolerance, ability to ferment molasses, and to ferment at elevated temperatures etc. However, this yeast was found to be sensitive against the killer strains of Saccharomyces cerevisiae. In the present study, killer trait was introduced into Saccharomyces cerevisiae HAU-1 by protoplast fusion with Saccharomyces cerevisiae MTCC 475, a killer strain. The resultant fusants were characterized for desirable fermentation characteristics. All the technologically important characteristics of distillery yeast Saccharomyces cerevisiae HAU-1 were retained in the fusants, and in addition the killer trait was also introduced into them. Further, the killer activity was found to be stably maintained during hostile conditions of ethanol fermentations in dextrose or molasses, and even during biomass recycling.

  8. High-level production of animal-free recombinant transferrin from saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Goldenberg Hans

    2010-11-01

    Full Text Available Abstract Background Animal-free recombinant proteins provide a safe and effective alternative to tissue or serum-derived products for both therapeutic and biomanufacturing applications. While recombinant insulin and albumin already exist to replace their human counterparts in cell culture media, until recently there has been no equivalent for serum transferrin. Results The first microbial system for the high-level secretion of a recombinant transferrin (rTf has been developed from Saccharomyces cerevisiae strains originally engineered for the commercial production of recombinant human albumin (Novozymes' Recombumin® USP-NF and albumin fusion proteins (Novozymes' albufuse®. A full-length non-N-linked glycosylated rTf was secreted at levels around ten-fold higher than from commonly used laboratory strains. Modification of the yeast 2 μm-based expression vector to allow overexpression of the ER chaperone, protein disulphide isomerase, further increased the secretion of rTf approximately twelve-fold in high cell density fermentation. The rTf produced was functionally equivalent to plasma-derived transferrin. Conclusions A Saccharomyces cerevisiae expression system has enabled the cGMP manufacture of an animal-free rTf for industrial cell culture application without the risk of prion and viral contamination, and provides a high-quality platform for the development of transferrin-based therapeutics.

  9. Flux Balance Analysis Inspired Bioprocess Upgrading for Lycopene Production by a Metabolically Engineered Strain of Yarrowia lipolytica

    OpenAIRE

    Komi Nambou; Xingxing Jian; Xinkai Zhang; Liujing Wei; Jiajia Lou; Catherine Madzak; Qiang Hua

    2015-01-01

    Genome-scale metabolic models embody a significant advantage of systems biology since their applications as metabolic flux simulation models enable predictions for the production of industrially-interesting metabolites. The biotechnological production of lycopene from Yarrowia lipolytica is an emerging scope that has not been fully scrutinized, especially for what concerns cultivation conditions of newly generated engineered strains. In this study, by combining flux balance analysis (FBA) and...

  10. 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

    2016-08-01

    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. PMID:27493184

  11. Optimization of feeding strategy for the ergosterol production by yeasts saccharomyces cerevisiae

    OpenAIRE

    Mojmir Rychtera; Josef Cermak; Jaroslav Votruba; Jan Nahlik; Karel Melzoch; Christopher A. Kent; Estela Escalante, Waldir D.

    2010-01-01

    Objective of this study was to optimize ergosterol production by yeast strain Saccharomyces cerevisiae with the use of computer controlled feeding of cultivation medium. Baker´s yeasts strain of Saccharomyces cerevisiae originally modified and selected as mutant D7 was further applied in an industrial scale and also in this investigation. Composition of cultivation medium was optimized with the use of a modified Rosenbrock´s method with regard to following components: glucose, yeast extract, ...

  12. The Bioconversion of Red Ginseng Ethanol Extract into Compound K by Saccharomyces cerevisiae HJ-014

    OpenAIRE

    Choi, Hak Joo; Kim, Eun A.; Kim, Dong Hee; Shin, Kwang-Soo

    2014-01-01

    A β-glucosidase producing yeast strain was isolated from Korean traditional rice wine. Based on the sequence of the YCL008c gene and analysis of the fatty acid composition, the isolate was identified as Saccharomyces cerevisiae strain HJ-014. S. cerevisiae HJ-014 produced ginsenoside Rd, F2, and compound K from the ethanol extract of red ginseng. The production was increased by shaking culture, where the bioconversion efficiency was increased 2-fold compared to standing culture. The productio...

  13. Enhancement of free fatty acid production in Saccharomyces cerevisiae by control of fatty acyl-CoA metabolism.

    Science.gov (United States)

    Chen, Liwei; Zhang, Jianhua; Lee, Jaslyn; Chen, Wei Ning

    2014-08-01

    Production of biofuels derived from microbial fatty acids has attracted great attention in recent years owing to their potential to replace petroleum-derived fuels. To be cost competitive with current petroleum fuel, flux toward the direct precursor fatty acids needs to be enhanced to approach high yields. Herein, fatty acyl-CoA metabolism in Saccharomyces cerevisiae was engineered to accumulate more free fatty acids (FFA). For this purpose, firstly, haploid S. cerevisiae double deletion strain △faa1△faa4 was constructed, in which the genes FAA1 and FAA4 encoding two acyl-CoA synthetases were deleted. Then the truncated version of acyl-CoA thioesterase ACOT5 (Acot5s) encoding Mus musculus peroxisomal acyl-CoA thioesterase 5 was expressed in the cytoplasm of the strain △faa1△faa4. The resulting strain △faa1△faa4 [Acot5s] accumulated more extracellular FFA with higher unsaturated fatty acid (UFA) ratio as compared to the wild-type strain and double deletion strain △faa1△faa4. The extracellular total fatty acids (TFA) in the strain △faa1△faa4 [Acot5s] increased to 6.43-fold as compared to the wild-type strain during the stationary phase. UFA accounted for 42 % of TFA in the strain △faa1△faa4 [Acot5s], while no UFA was detected in the wild-type strain. In addition, the expression of Acot5s in △faa1△faa4 restored the growth, which indicates that FFA may not be the reason for growth inhibition in the strain △faa1△faa4. RT-PCR results demonstrated that the de-repression of fatty acid synthesis genes led to the increase of extracellular fatty acids. The study presented here showed that through control of the acyl-CoA metabolism by deleting acyl-CoA synthetase and expressing thioesterase, more FFA could be produced in S. cerevisiae, demonstrating great potential for exploitation in the platform of microbial fatty acid-derived biofuels. PMID:24769906

  14. Construction of engineering Trichoderma strains and their characteristics against tomato gray mold

    Institute of Scientific and Technical Information of China (English)

    LIU Xian; ZHUANG Jing-hua; GAO Zeng-gui; YANG Chang-cheng; CHEN Jie

    2004-01-01

    @@ The transformed Trichoderma strains Ttrm31, Ttrm34 and Ttrm55 were obtained from Trichoderma wild strain T21 mutated by REMI technique for more effective biocontrol of tomato gray mold (Botrytis cinerea) with Trichoderma agent. Those transformants appeared much better in biocontrol activity in vitro or in vivo against tomato gray mold were better than that of wild strain T21. The main results were as follow:

  15. Reconstitution of an efficient thymidine salvage pathway in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Vernis, L.; Piskur, Jure; Diffley, J.F.X.

    2003-01-01

    The budding yeast Saccharomyces cerevisiae is unable to incorporate exogenous nucleosides into DNA. We have made a number of improvements to existing strategies to reconstitute an efficient thymidine salvage pathway in yeast. We have constructed strains that express both a nucleoside kinase as well...

  16. Roles of human sulfotransferases in genotoxicity of carcinogens using genetically engineered umu test strains.

    Science.gov (United States)

    Oda, Yoshimitsu; Zhang, Yu; Buchinger, Sebastian; Reifferscheid, Georg; Yang, Min

    2012-03-01

    Human sulfotransferase (SULT) 1A1, 1A2, and 1A3 cDNA genes were subcloned separately into the pTrc99A(KM) vector. The generated plasmids were introduced into the Salmonella typhimurium O-acetyltransferase-deficient strain NM6000 (TA1538/1,8-DNP/pSK1002), resulting in the new strains NM7001, NM7002, and NM7003. We compared the sensitivities of these three strains with the parental strain NM7000 against 51 chemicals including aromatic amines, nitroarenes, alkenylbenzenes, estrogens-like chemicals, and other compounds with and without S9 mix by making use of the umu test system that is based on the bacterial SOS induction. 2-Amino-6-methyl-dipyrido[1,2-α:3',2'-d]imidazole, 3-methoxy-4-aminoazobenzene, 3-nitrobenzanthrone, 5-nitroacenaphthene, and 3,9-dinitrofluoranthene caused high genotoxicity in the NM7001 strain. The genotoxic effects of 2-aminofluorene, 2-acetylaminofluorene, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, 2-nitrofluorene, 1-nitropyrene, and 2-nitropropane were stronger in the NM7002 strain compared with the NM7001 and NM7003 strains. Among the tested benzylic and allylic compounds, 1-hydroxymethylpyrene was detected in the NM7001 strain with the highest sensitivity. Estragole and 1'-hydroxysafrole exhibited strong genotoxicity in the NM7003 strain. The estrogen-like chemicals such as bisphenol A, genistein, p,n-nonylphenol, and 4-hydroxytamoxifen were not detected as genotoxins in any strain used. Collectively, the present results suggest that the generated test strains are valuable tools in order to elucidate the role of SULT enzymes in the bioactivation of chemicals to environmental carcinogens. PMID:22072630

  17. Genetically engineered Pseudomonas putida X3 strain and its potential ability to bioremediate soil microcosms contaminated with methyl parathion and cadmium.

    Science.gov (United States)

    Zhang, Rong; Xu, Xingjian; Chen, Wenli; Huang, Qiaoyun

    2016-02-01

    A multifunctional Pseudomonas putida X3 strain was successfully engineered by introducing methyl parathion (MP)-degrading gene and enhanced green fluorescent protein (EGFP) gene in P. putida X4 (CCTCC: 209319). In liquid cultures, the engineered X3 strain utilized MP as sole carbon source for growth and degraded 100 mg L(-1) of MP within 24 h; however, this strain did not further metabolize p-nitrophenol (PNP), an intermediate metabolite of MP. No discrepancy in minimum inhibitory concentrations (MICs) to cadmium (Cd), copper (Cu), zinc (Zn), and cobalt (Co) was observed between the engineered X3 strain and its host strain. The inoculated X3 strain accelerated MP degradation in different polluted soil microcosms with 100 mg MP kg(-1) dry soil and/or 5 mg Cd kg(-1) dry soil; MP was completely eliminated within 40 h. However, the presence of Cd in the early stage of remediation slightly delayed MP degradation. The application of X3 strain in Cd-contaminated soil strongly affected the distribution of Cd fractions and immobilized Cd by reducing bioavailable Cd concentrations with lower soluble/exchangeable Cd and organic-bound Cd. The inoculated X3 strain also colonized and proliferated in various contaminated microcosms. Our results suggested that the engineered X3 strain is a potential bioremediation agent showing competitive advantage in complex contaminated environments. PMID:26521245

  18. Bandgap engineering of monolayer MoS2 under strain: A DFT study

    Science.gov (United States)

    Li, Can; Fan, Bowen; Li, Weiyi; Wen, Luowei; Liu, Yan; Wang, Tao; Sheng, Kuang; Yin, You

    2015-06-01

    In this paper, density functional theory calculations are used to investigate the monolayer MoS2 in terms of the strain by analyzing the structure parameters: the bandgap, the density of states (DOS) and the Milliken charges. The calculations indicate that an increasing external stain tends to depress the ripple structure with a shorter S-S interlayer spacing and to enlarge the length of the Mo-S bond. Tensile strain dramatically alteres the bandgap; however, compressive strain almost does not. The change in the bandgap is explained by an analysis of the DOS, the partial density of states (PDOS), the structure parameters and the Mulliken charge distribution. The effects of strain on the Mulliken charge and the length of the Mo-S band cause bandgap differences under tensile and compressive strain.

  19. Mead production: selection and characterization assays of Saccharomyces cerevisiae

    OpenAIRE

    de Pereira, Ana Paula; Dias, Teresa; Andrade, João Verdial; Ramalhosa, Elsa; Mendes-Ferreira, Ana; Mendes-Faia, Arlete; Leticia M. Estevinho

    2009-01-01

    Mead is a traditional alcoholic drink which results from the fermentation of diluted honey. Yeasts used in mead production are, usually, wine Saccharomyces cerevisiae strains. Most of these yeasts are not adapted to the conditions of mead production namely, high sugar levels, low pH values and reduced nitrogen concentrations. The inability of yeast strains to respond and adapt to unfavorable stressful growth conditions, leads to several problems, such as lack of uniformity of the final ...

  20. Heterooligomeric phosphoribosyl diphosphate synthase of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Hove-Jensen, Bjarne

    2004-01-01

    The yeast Saccharomyces cerevisiae contains five phosphoribosyl diphosphate (PRPP) synthase-homologous genes (PRS1-5), which specify PRPP synthase subunits 1-5. Expression of the five S. cerevisiae PRS genes individually in an Escherichia coli PRPP-less strain (Deltaprs) showed that a single PRS...... gene product had no PRPP synthase activity. In contrast, expression of five pairwise combinations of PRS genes resulted in the formation of active PRPP synthase. These combinations were PRS1 PRS2, PRS1 PRS3, and PRS1 PRS4, as well as PRS5 PRS2 and PRS5 PRS4. None of the remaining five possible pairwise...... combinations of PRS genes appeared to produce active enzyme. Extract of an E. coli strain containing a plasmid-borne PRS1 gene and a chromosome-borne PRS3 gene contained detectable PRPP synthase activity, whereas extracts of strains containing PRS1 PRS2, PRS1 PRS4, PRS5 PRS2, or PRS5 PRS4 contained no...

  1. Flux Balance Analysis Inspired Bioprocess Upgrading for Lycopene Production by a Metabolically Engineered Strain of Yarrowia lipolytica

    Directory of Open Access Journals (Sweden)

    Komi Nambou

    2015-12-01

    Full Text Available Genome-scale metabolic models embody a significant advantage of systems biology since their applications as metabolic flux simulation models enable predictions for the production of industrially-interesting metabolites. The biotechnological production of lycopene from Yarrowia lipolytica is an emerging scope that has not been fully scrutinized, especially for what concerns cultivation conditions of newly generated engineered strains. In this study, by combining flux balance analysis (FBA and Plackett-Burman design, we screened chemicals for lycopene production from a metabolically engineered strain of Y. lipolytica. Lycopene concentrations of 126 and 242 mg/L were achieved correspondingly from the FBA-independent and the FBA-assisted designed media in fed-batch cultivation mode. Transcriptional studies revealed upregulations of heterologous genes in media designed according to FBA, thus implying the efficiency of model predictions. Our study will potentially support upgraded lycopene and other terpenoids production from existing or prospect bioengineered strains of Y. lipolytica and/or closely related yeast species.

  2. Flux Balance Analysis Inspired Bioprocess Upgrading for Lycopene Production by a Metabolically Engineered Strain of Yarrowia lipolytica.

    Science.gov (United States)

    Nambou, Komi; Jian, Xingxing; Zhang, Xinkai; Wei, Liujing; Lou, Jiajia; Madzak, Catherine; Hua, Qiang

    2015-01-01

    Genome-scale metabolic models embody a significant advantage of systems biology since their applications as metabolic flux simulation models enable predictions for the production of industrially-interesting metabolites. The biotechnological production of lycopene from Yarrowia lipolytica is an emerging scope that has not been fully scrutinized, especially for what concerns cultivation conditions of newly generated engineered strains. In this study, by combining flux balance analysis (FBA) and Plackett-Burman design, we screened chemicals for lycopene production from a metabolically engineered strain of Y. lipolytica. Lycopene concentrations of 126 and 242 mg/L were achieved correspondingly from the FBA-independent and the FBA-assisted designed media in fed-batch cultivation mode. Transcriptional studies revealed upregulations of heterologous genes in media designed according to FBA, thus implying the efficiency of model predictions. Our study will potentially support upgraded lycopene and other terpenoids production from existing or prospect bioengineered strains of Y. lipolytica and/or closely related yeast species. PMID:26703753

  3. Ferromagnetic (Ga,Mn)As layers and nanostructures: control of magnetic anisotropy by strain engineering

    Energy Technology Data Exchange (ETDEWEB)

    Wenisch, Jan

    2008-07-01

    This work studies the fundamental connection between lattice strain and magnetic anisotropy in the ferromagnetic semiconductor (Ga,Mn)As. The first chapters provide a general introduction into the material system and a detailed description of the growth process by molecular beam epitaxy. A finite element simulation formalism is developed to model the strain distribution in (Ga,Mn)As nanostructures is introduced and its predictions verified by high-resolution X-ray diffraction methods. The influence of lattice strain on the magnetic anisotropy is explained by an magnetostatic model. A possible device application is described in the closing chapter. (orig.)

  4. [Construction of the flavinogenic yeast Candida famata strains with high riboflavin kinase activity using gene engineering].

    Science.gov (United States)

    Ishchuk, O P; Iatsyshyn, V Iu; Dmytruk, K V; Voronovs'kyĭ, A Ia; Fedorovych, D V; Sybirnyĭ, A A

    2006-01-01

    The recombinant strains of the flavinogenic yeast Candida famata, which contain the DNA fragment consisting of the FMN1 gene (encoding the riboflavin kinase, enzyme that converts riboflavin to flavinmononucleotide) driven by the strong promoters (the regulated RIB1 or constitutive TEF1 promoter) were isolated. Riboflavin kinase activity in the isolated transformants was tested. The 6-8-fold increase of the riboflavin kinase activity was shown in the recombinant strains containing the integrated Debaryomyces hansenii FMN1 gene under the strong constitutive TEF1 promoter. The recombinant strains can be used for the following construction of flavinmononucleotide overproducers. PMID:17290783

  5. Significant enhancement of methionol production by co-expression of the aminotransferase gene ARO8 and the decarboxylase gene ARO10 in Saccharomyces cerevisiae.

    Science.gov (United States)

    Yin, Sheng; Lang, Tiandan; Xiao, Xiao; Liu, Li; Sun, Baoguo; Wang, Chengtao

    2015-03-01

    Methionol is an important volatile sulfur flavor compound, which can be produced via the Ehrlich pathway in Saccharomyces cerevisiae. Aminotransferase and decarboxylase are essential enzymes catalyzing methionol biosynthesis. In this work, two aminotransferase genes ARO8 and ARO9 and one decarboxylase gene ARO10 were introduced into S. cerevisiae S288c, respectively, via an expression vector. Over-expression of ARO8 resulted in higher aminotransferase activity than that of ARO9. And the cellular decarboxylase activity was remarkably increased by over-expression of ARO10. A co-expression vector carrying both ARO8 and ARO10 was further constructed to generate the recombinant strain S810. Shaking flask experiments showed that the methionol yield from S810 reached 1.27 g L(-1), which was increased by 51.8 and 68.8% compared to that from the wild-type strain and the control strain harboring the empty vector. The fed-batch fermentation by strain S810 produced 3.24 g L(-1) of methionol after 72 h of cultivation in a bioreactor. These results demonstrated that co-expression of ARO8 and ARO10 significantly boosted the methionol production. It is the first time that more than 3.0 g L(-1) of methionol produced by genetically engineered yeast strain was reported by co-expression of the aminotransferase and decarboxylase via the Ehrlich pathway. PMID:25743068

  6. Improvement of Ethanol Production in Saccharomyces cerevisiae by High-Efficient Disruption of the ADH2 Gene Using a Novel Recombinant TALEN Vector

    Science.gov (United States)

    Ye, Wei; Zhang, Weimin; Liu, Taomei; Tan, Guohui; Li, Haohua; Huang, Zilei

    2016-01-01

    Bioethanol is becoming increasingly important in energy supply and economic development. However, the low yield of bioethanol and the insufficiency of high-efficient genetic manipulation approaches limit its application. In this study, a novel transcription activator-like effector nuclease (TALEN) vector containing the left and right arms of TALEN was electroporated into Saccharomyces cerevisiae strain As2.4 to sequence the alcohol dehydrogenase gene ADH2 and the hygromycin-resistant gene hyg. Western blot analysis using anti-FLAG monoclonal antibody proved the successful expression of TALE proteins in As2.4 strains. qPCR and sequencing demonstrated the accurate knockout of the 17 bp target gene with 80% efficiency. The TALEN vector and ADH2 PCR product were electroporated into ΔADH2 to complement the ADH2 gene (ADH2+ As2.4). LC–MS and GC were employed to detect ethanol yields in the native As2.4, ΔADH2 As2.4, and ADH2+ As2.4 strains. Results showed that ethanol production was improved by 52.4 ± 5.3% through the disruption of ADH2 in As2.4. The bioethanol yield of ADH2+ As2.4 was nearly the same as that of native As2.4. This study is the first to report on the disruption of a target gene in S. cerevisiae by employing Fast TALEN technology to improve bioethanol yield. This work provides a novel approach for the disruption of a target gene in S. cerevisiae with high efficiency and specificity, thereby promoting the improvement of bioethanol production in S. cerevisiae by metabolic engineering. PMID:27462304

  7. Understanding the 3-hydroxypropionic acid tolerance mechanism in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kildegaard, Kanchana Rueksomtawin; Juncker, Agnieszka; Hallstrom, Bjorn; Jensen, Niels Bjerg; Maury, Jerome; Nielsen, Jen; Förster, Jochen; Borodina, Irina

    sustainable alternative for production of acrylic acid from renewable feedstocks. We are establishing Saccharomyces cerevisiae as an alternative host for 3HP production. However, 3HP also inhibits yeast grow th at level well below what is desired for commercial applications. Therefore, we are aiming to...... improve 3HP tolerance in S. cerevisiae by applying adaptive evolution approach. We have generated yeast strains with sign ificantly improved capacity for tolerating 3HP when compared to the wild-type. We will present physiolo gical characterization, genome re-sequencing, and transcriptome analysis of the...

  8. Accumulation of gold using Baker's yeast, Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Authors have reported preconcentration of 152Eu, a long-lived fission product, by yeast cells, Saccharomyces cerevisiae. Gold being a precious metal is used in electroplating, hydrogenation catalyst, etc. Heterogeneous composition of samples and low concentration offers renewed interest in its selective extraction of gold using various extractants. Gold can be recovered from different solutions using various chemical reagents like amines, organophosphorus compounds, and extractants containing sulphur as donor atom, etc. In the present work, two different strains of baker's yeast, Saccharomyces cerevisiae have been used to study the preconcentration of gold at various experimental conditions

  9. Establishment of a yeast platform strain for production of p-coumaric acid through metabolic engineering of aromatic amino acid biosynthesis.

    Science.gov (United States)

    Rodriguez, Angelica; Kildegaard, Kanchana R; Li, Mingji; Borodina, Irina; Nielsen, Jens

    2015-09-01

    Aromatic amino acids are precursors of numerous plant secondary metabolites with diverse biological functions. Many of these secondary metabolites are already being used as active pharmaceutical or nutraceutical ingredients, and there are numerous exploratory studies of other compounds with promising applications. p-Coumaric acid is derived from aromatic amino acids and, besides being a valuable chemical building block, it serves as precursor for biosynthesis of many secondary metabolites, such as polyphenols, flavonoids, and some polyketides. Here we developed a p-coumaric acid-overproducing Saccharomyces cerevisiae platform strain. First, we reduced by-product formation by knocking out phenylpyruvate decarboxylase ARO10 and pyruvate decarboxylase PDC5. Second, different versions of feedback-resistant DAHP synthase and chorismate mutase were overexpressed. Finally, we identified shikimate kinase as another important flux-controlling step in the aromatic amino acid pathway by overexpressing enzymes from Escherichia coli, homologous to the pentafunctional enzyme Aro1p and to the bifunctional chorismate synthase-flavin reductase Aro2p. The highest titer of p-coumaric acid of 1.93 ± 0.26 g L(-1) was obtained, when overexpressing tyrosine ammonia-lyase TAL from Flavobacterium johnsoniaeu, DAHP synthase ARO4(K229L), chorismate mutase ARO7(G141S) and E. coli shikimate kinase II (aroL) in Δpdc5Δaro10 strain background. To our knowledge this is the highest reported titer of an aromatic compound produced by yeast. The developed S. cerevisiae strain represents an attractive platform host for production of p-coumaric-acid derived secondary metabolites, such as flavonoids, polyphenols, and polyketides. PMID:26292030

  10. Nano-scale strain engineering of graphene and graphene-based devices

    OpenAIRE

    Yeh, N. -C.; Hsu, C.-C.; Teague, M. L.; Wang, J.-Q.; Boyd, D A; Chen, C.-C.

    2015-01-01

    Structural distortions in nano-materials can induce dramatic changes in their electronic properties. This situation is well manifested in graphene, a two-dimensional honeycomb structure of carbon atoms with only one atomic layer thickness. In particular, strained graphene can result in both charging effects and pseudo-magnetic fields, so that controlled strain on a perfect graphene lattice can be tailored to yield desirable electronic properties. Here we describe the theoretical foundation fo...

  11. Genetically Engineered Immunomodulatory Streptococcus thermophilus Strains Producing Antioxidant Enzymes Exhibit Enhanced Anti-Inflammatory Activities

    OpenAIRE

    del Carmen, Silvina; de Moreno de LeBlanc, Alejandra; Martin, Rebeca; Chain, Florian; Langella, Philippe; Bermúdez-Humarán, Luis G; LeBlanc, Jean Guy

    2014-01-01

    The aims of this study were to develop strains of lactic acid bacteria (LAB) having both immunomodulatory and antioxidant properties and to evaluate their anti-inflammatory effects both in vitro, in different cellular models, and in vivo, in a mouse model of colitis. Different Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains were cocultured with primary cultures of mononuclear cells. Analysis of the pro- and anti-inflammatory cytokines secreted by these cells...

  12. Strain-Engineering Anisotropic Electrical Conductance of Phosphorene and Few-Layer Black Phosphorus

    OpenAIRE

    Fei, Ruixiang; Yang, Li

    2014-01-01

    Newly fabricated monolayer phosphorene and its few-layer structures are expected to be promising for electronic and optical applications because of their finite direct band gaps and sizable but anisotropic electronic mobility. By first-principles simulations, we show that this unique anisotropic conductance can be controlled by using simple strain conditions. With the appropriate biaxial or uniaxial strain, we can rotate the preferred conducting direction by 90 degrees. This will be of useful...

  13. Interface Schottky barrier engineering via strain in metal-semiconductor composites.

    Science.gov (United States)

    Ma, Xiangchao; Dai, Ying; Yu, Lin; Huang, Baibiao

    2016-01-21

    The interfacial carrier transfer property, which is dominated by the interface Schottky barrier height (SBH), plays a crucial role in determining the performance of metal-semiconductor heterostructures in a variety of applications. Therefore, artificially controlling the interface SBH is of great importance for their industrial applications. As a model system, the Au/TiO2 (001) heterostructure is studied using first-principles calculations and the tight-binding method in the present study. Our investigation demonstrates that strain can be an effective way to decrease the interface SBH and that the n-type SBH can be more effectively decreased than the p-type SBH. Astonishingly, strain affects the interface SBH mainly by changing the intrinsic properties of Au and TiO2, whereas the interfacial potential alignment is almost independent of strain due to two opposite effects, which are induced by strain at the interfacial region. These observed trends can be understood on the basis of the general free-electron gas model of typical metals, the tight-binding theory and the crystal-field theory, which suggest that similar trends may be generalized for many other metal-semiconductor heterostructures. Given the commonness and tunability of strain in typical heterostructures, we anticipate that the tunability of the interface SBH with strain described here can provide an alternative effective way for realizing more efficient applications of relevant heterostructures. PMID:26511292

  14. Strain-engineering the anisotropic electrical conductance in ReS2 monolayer

    Science.gov (United States)

    Yu, Sheng; Zhu, Hao; Eshun, Kwesi; Shi, Chen; Zeng, Min; Li, Qiliang

    2016-05-01

    Rhenium disulfide (ReS2) is a semiconducting layered transition metal dichalcogenide that exhibits a stable distorted 1 T (Re in octahedral coordination) phase. The reduced symmetry in ReS2 leads to in-plane anisotropy in various material properties. In this work, we performed a comprehensive first-principle computational study of strain effect on the anisotropic mechanical and electronic properties of ReS2 monolayers. We found that the anisotropic ratio in electron mobility along two principle axes is 2.36 while the ratio in hole mobility reaches 7.76. The study of strain applied along different directions shows that the elastic modulus is largest for out-of-plane direction, and the strain along a-direction induces indirect bandgap while strain along b- or c-direction does not. In addition, the carrier mobility can be significantly improved by the c-direction tensile strain. This study indicates that the ReS2 monolayer has promising applications in nanoscale strain sensor and conductance-switch FETs.

  15. 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

    -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 ERG9 expression to glucose concentration its promoter was replaced by the HXT1 promoter. In a second approach, the HXT2 promoter was used to express an ERG9 antisense construct. Using the HXT1 promoter to control ERG9 expression, it was possible to divert the carbon flux from sterol synthesis...

  16. Interface Schottky barrier engineering via strain in metal-semiconductor composites

    Science.gov (United States)

    Ma, Xiangchao; Dai, Ying; Yu, Lin; Huang, Baibiao

    2016-01-01

    The interfacial carrier transfer property, which is dominated by the interface Schottky barrier height (SBH), plays a crucial role in determining the performance of metal-semiconductor heterostructures in a variety of applications. Therefore, artificially controlling the interface SBH is of great importance for their industrial applications. As a model system, the Au/TiO2 (001) heterostructure is studied using first-principles calculations and the tight-binding method in the present study. Our investigation demonstrates that strain can be an effective way to decrease the interface SBH and that the n-type SBH can be more effectively decreased than the p-type SBH. Astonishingly, strain affects the interface SBH mainly by changing the intrinsic properties of Au and TiO2, whereas the interfacial potential alignment is almost independent of strain due to two opposite effects, which are induced by strain at the interfacial region. These observed trends can be understood on the basis of the general free-electron gas model of typical metals, the tight-binding theory and the crystal-field theory, which suggest that similar trends may be generalized for many other metal-semiconductor heterostructures. Given the commonness and tunability of strain in typical heterostructures, we anticipate that the tunability of the interface SBH with strain described here can provide an alternative effective way for realizing more efficient applications of relevant heterostructures.The interfacial carrier transfer property, which is dominated by the interface Schottky barrier height (SBH), plays a crucial role in determining the performance of metal-semiconductor heterostructures in a variety of applications. Therefore, artificially controlling the interface SBH is of great importance for their industrial applications. As a model system, the Au/TiO2 (001) heterostructure is studied using first-principles calculations and the tight-binding method in the present study. Our investigation

  17. Interfacially engineered oxygen octahedral rotations and their impact on strain relief in coherently grown SrRu O3 films

    Science.gov (United States)

    Kan, Daisuke; Wakabayashi, Yusuke; Tajiri, Hiroo; Shimakawa, Yuichi

    2016-07-01

    We report synchrotron x-ray diffraction investigations of interfacially engineered oxygen octahedral rotations and their impact on strain relief in perovskite SrRu O3 films. We show that octahedral rotations with distinct patterns and magnitudes can be accommodated into coherently grown films. The SrRu O3 film grown directly on the GdSc O3 substrate has the Ru O6 octahedral rotation with the a-b+c- pattern in the Glazer notation and the rotation angles of αrot=6.6 ±0 .2∘ , βrot=5.5 ±0 .2∘ , and γrot=3.6 ±0 .2∘ . On the other hand, when a 1-nm-thick BaTi O3 layer without Ti O6 rotations is inserted between the SrRu O3 and GdSc O3 , the SrRu O3 has the Ru O6 rotation with a-b0c+ , and αrot=5.6 ±0 .8∘ and γrot=3.6 ±0 .8∘ . These results indicate that there are some degrees of freedom in the octahedral rotations accommodated in SrRu O3 depending on the interface structure and that the γrot rotations play the important roles in the film's structural properties when the rotation about the [010] pc axis is blocked. We also found that the strain relief in the film is influenced by the interfacially engineered octahedral rotations. The interfacial BaTi O3 layer results in the in-plane periodic lattice modulation in the t-SRO film, allowing for the anisotropic relief of the substrate-induced strain. The results highlight the importance of the interface structure as a factor, determining not only octahedral rotations in coherently grown SRO films but also the strain reliefs in them.

  18. Engineering control of bacterial cellulose production using a genetic toolkit and a new cellulose-producing strain

    Science.gov (United States)

    Florea, Michael; Hagemann, Henrik; Santosa, Gabriella; Micklem, Chris N.; Spencer-Milnes, Xenia; de Arroyo Garcia, Laura; Paschou, Despoina; Lazenbatt, Christopher; Kong, Deze; Chughtai, Haroon; Jensen, Kirsten; Freemont, Paul S.; Kitney, Richard; Reeve, Benjamin; Ellis, Tom

    2016-01-01

    Bacterial cellulose is a strong and ultrapure form of cellulose produced naturally by several species of the Acetobacteraceae. Its high strength, purity, and biocompatibility make it of great interest to materials science; however, precise control of its biosynthesis has remained a challenge for biotechnology. Here we isolate a strain of Komagataeibacter rhaeticus (K. rhaeticus iGEM) that can produce cellulose at high yields, grow in low-nitrogen conditions, and is highly resistant to toxic chemicals. We achieved external control over its bacterial cellulose production through development of a modular genetic toolkit that enables rational reprogramming of the cell. To further its use as an organism for biotechnology, we sequenced its genome and demonstrate genetic circuits that enable functionalization and patterning of heterologous gene expression within the cellulose matrix. This work lays the foundations for using genetic engineering to produce cellulose-based materials, with numerous applications in basic science, materials engineering, and biotechnology. PMID:27247386

  19. Engineering control of bacterial cellulose production using a genetic toolkit and a new cellulose-producing strain.

    Science.gov (United States)

    Florea, Michael; Hagemann, Henrik; Santosa, Gabriella; Abbott, James; Micklem, Chris N; Spencer-Milnes, Xenia; de Arroyo Garcia, Laura; Paschou, Despoina; Lazenbatt, Christopher; Kong, Deze; Chughtai, Haroon; Jensen, Kirsten; Freemont, Paul S; Kitney, Richard; Reeve, Benjamin; Ellis, Tom

    2016-06-14

    Bacterial cellulose is a strong and ultrapure form of cellulose produced naturally by several species of the Acetobacteraceae Its high strength, purity, and biocompatibility make it of great interest to materials science; however, precise control of its biosynthesis has remained a challenge for biotechnology. Here we isolate a strain of Komagataeibacter rhaeticus (K. rhaeticus iGEM) that can produce cellulose at high yields, grow in low-nitrogen conditions, and is highly resistant to toxic chemicals. We achieved external control over its bacterial cellulose production through development of a modular genetic toolkit that enables rational reprogramming of the cell. To further its use as an organism for biotechnology, we sequenced its genome and demonstrate genetic circuits that enable functionalization and patterning of heterologous gene expression within the cellulose matrix. This work lays the foundations for using genetic engineering to produce cellulose-based materials, with numerous applications in basic science, materials engineering, and biotechnology. PMID:27247386

  20. Metabolic Engineering of Probiotic Saccharomyces boulardii.

    Science.gov (United States)

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

    2016-04-15

    Saccharomyces boulardiiis a probiotic yeast that has been used for promoting gut health as well as preventing diarrheal diseases. This yeast not only exhibits beneficial phenotypes for gut health but also can stay longer in the gut thanSaccharomyces cerevisiae Therefore,S. boulardiiis an attractive host for metabolic engineering to produce biomolecules of interest in the gut. However, the lack of auxotrophic strains with defined genetic backgrounds has hampered the use of this strain for metabolic engineering. Here, we report the development of well-defined auxotrophic mutants (leu2,ura3,his3, andtrp1) through clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9-based genome editing. The resulting auxotrophic mutants can be used as a host for introducing various genetic perturbations, such as overexpression or deletion of a target gene, using existing genetic tools forS. cerevisiae We demonstrated the overexpression of a heterologous gene (lacZ), the correct localization of a target protein (red fluorescent protein) into mitochondria by using a protein localization signal, and the introduction of a heterologous metabolic pathway (xylose-assimilating pathway) in the genome ofS. boulardii We further demonstrated that human lysozyme, which is beneficial for human gut health, could be secreted byS. boulardii Our results suggest that more sophisticated genetic perturbations to improveS. boulardiican be performed without using a drug resistance marker, which is a prerequisite forin vivoapplications using engineeredS. boulardii. PMID:26850302

  1. Strain-Engineering the Gauge Potential of Dirac fermions in PECVD-grown Graphene

    Science.gov (United States)

    Hsu, Chen-Chih; Teague, Marcus; Wang, Jaiqing; Yeh, Nai-Chang

    Non-trivial strain can induce pseudo-magnetic fields in graphene so that the electronic properties of Dirac fermions can be tuned by controlling the strain on graphene. Here we employ nearly strain-free single-domain PECVD-graphene1 to induce controlled strain by placing graphene on nanostructured substrates. Strain-induced gauge potentials and pseudo-magnetic fields can be manifested by the local tunneling conductance peaks at quantized energies.2,3 Additionally, pseudo-magnetic field-induced local spontaneous time-reversal symmetry breaking can be revealed by spatially alternating presence and absence of the zero mode in the tunneling conductance spectra.2,3 We also employ molecular dynamics simulations to determine the spatial distribution of the pseudo-magnetic field for a given nanostructure. We find that a tetrahedron-like nanostructure can be an effective ``valley splitter'' to separate the trajectories of Dirac fermions of opposite pseudo-spins. Proper design and arrangement of several valley filters can function as a ``valley propagator'' to guide valley-polarized currents. We plan to verify the valley Hall effect associated with a valley splitter and to assess the feasibility of realistic valleytronic applications.

  2. Strain-Engineered Graphene Grown on Hexagonal Boron Nitride by Molecular Beam Epitaxy

    Science.gov (United States)

    Summerfield, Alex; Davies, Andrew; Cheng, Tin S.; Korolkov, Vladimir V.; Cho, YongJin; Mellor, Christopher J.; Foxon, C. Thomas; Khlobystov, Andrei N.; Watanabe, Kenji; Taniguchi, Takashi; Eaves, Laurence; Novikov, Sergei V.; Beton, Peter H.

    2016-01-01

    Graphene grown by high temperature molecular beam epitaxy on hexagonal boron nitride (hBN) forms continuous domains with dimensions of order 20 μm, and exhibits moiré patterns with large periodicities, up to ~30 nm, indicating that the layers are highly strained. Topological defects in the moiré patterns are observed and attributed to the relaxation of graphene islands which nucleate at different sites and subsequently coalesce. In addition, cracks are formed leading to strain relaxation, highly anisotropic strain fields, and abrupt boundaries between regions with different moiré periods. These cracks can also be formed by modification of the layers with a local probe resulting in the contraction and physical displacement of graphene layers. The Raman spectra of regions with a large moiré period reveal split and shifted G and 2D peaks confirming the presence of strain. Our work demonstrates a new approach to the growth of epitaxial graphene and a means of generating and modifying strain in graphene. PMID:26928710

  3. Directed Evolution towards Increased Isoprenoid Production in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Carlsen, Simon; Nielsen, Michael Lynge; Kielland-Brandt, Morten;

    Due to declining drug discovery rates from organic synthetic libraries, pharmaceutical companies are turning their attention towards secondary metabolites. Isoprenoids, also known as terpenoids, constitute the largest known group of secondary metabolites isolated to date, encompassing more than 55...... pyrophosphate and geranylgeranyl pyrophosphate for large-scale microbial production of terpenoids. Saccharomyces cerevisiae was chosen as production platform due to its widespread use in industrial production and the waste number of molecular biology tools which is available for its manipulation. The effort...... for discovering new genetic perturbations, which would results in and increased production of isoprenoids by S. cerevisiae has been very limited. This project is focus on creating diversity within a lycopene producing S. cerevisiae strain by construction of gDNA-, cDNA-, and transposon-libraries. The diversified...

  4. Lung stress, strain, and energy load: engineering concepts to understand the mechanism of ventilator-induced lung injury (VILI).

    Science.gov (United States)

    Nieman, Gary F; Satalin, Joshua; Andrews, Penny; Habashi, Nader M; Gatto, Louis A

    2016-12-01

    It was recently shown that acute respiratory distress syndrome (ARDS) mortality has not been reduced in over 15 years and remains ~40 %, even with protective low tidal volume (LVt) ventilation. Thus, there is a critical need to develop novel ventilation strategies that will protect the lung and reduce ARDS mortality. Protti et al. have begun to analyze the impact of mechanical ventilation on lung tissue using engineering methods in normal pigs ventilated for 54 h. They used these methods to assess the impact of a mechanical breath on dynamic and static global lung strain and energy load. Strain is the change in lung volume in response to an applied stress (i.e., Tidal Volume-Vt). This study has yielded a number of exciting new concepts including the following: (1) Individual mechanical breath parameters (e.g., Vt or Plateau Pressure) are not directly correlated with VILI but rather any combination of parameters that subject the lung to excessive dynamic strain and energy/power load will cause VILI; (2) all strain is not equal; dynamic strain resulting in a dynamic energy load (i.e., kinetic energy) is more damaging to lung tissue than static strain and energy load (i.e., potential energy); and (3) a critical consideration is not just the size of the Vt but the size of the lung that is being ventilated by this Vt. This key concept merits attention since our current protective ventilation strategies are fixated on the priority of keeping the Vt low. If the lung is fully inflated, a large Vt is not necessarily injurious. In conclusion, using engineering concepts to analyze the impact of the mechanical breath on the lung is a novel new approach to investigate VILI mechanisms and to help design the optimally protective breath. Data generated using these methods have challenged some of the current dogma surrounding the mechanisms of VILI and of the components in the mechanical breath necessary for lung protection. PMID:27316442

  5. Electrochemical Probing of in Vivo 5-Hydroxymethyl Furfural Reduction in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kostesha, Natalie; Almeida, J.R.M.; Heiskanen, Arto;

    2009-01-01

    In this work, mediated amperometry was used to evaluate whether differences in intracellular nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) level could be observed between a genetically modified Saccharomyces cerevisiae strain, engineered for NADPH dependent 5-hydroxymethyl-2-furaldehyde...... amperometric signal during real-time monitoring of the concentration dependent HMF reduction in living cells could be translated into the cellular enzyme kinetic parameters: K-M,cell(app), V-MAX, k(cat,cell), and k(cat,cell)/K-M,cell(app). The results indicated that the overexpression of the ADH6 gene gave a...... 68% decrease in K-M,cell(app), and 42% increase in V-MAX, resulting in a 4-fold increase in k(cat,cell)/K-M,cell(app). These results demonstrate that the mediated amperometric method is useful for monitoring the short-term dynamics of NAD(P)H variations and determining cellular enzyme kinetic...

  6. CNT based nanosensor for defect identification and structural strain monitoring of engineering components

    International Nuclear Information System (INIS)

    This work reports the possibility of using PVDF ICNT composite as strain sensors. The PVDF ICNT films were prepared by solvent casting method and were bonded directly onto specimens by nonconductive adhesive. 4 % CNT- PVDF polymer nanocomposite film is characterised using various techniques like SEM, AFM, XRD and Raman spectroscopy and are reported. Microstructures and morphology were obtained using SEM and AFM. Lattice parameters were determined using XRD technique and reported. The shift in the Raman Spectrum due to the strain induced in the material is also discussed. The two electrodes were connected on to PVDF ICNT films with silver conducting epoxy to reduce contact resistance. Conventional Strain gauge is also attached to other end ofthe specimen for comparison. Zirconium Alloy, which is used in Pressurised Heavy Water Reactor (PHWR) as structural material is used for the present study. The results are discussed in detail in the paper

  7. Strain engineering the magnetic states of vacancy-doped monolayer MoSe2

    International Nuclear Information System (INIS)

    Highlights: • The effects of vacancy defects and biaxial strain on the magnetic property of MoSe2 were investigated. • VSe, VSe2 and VMoSe3 vacancy doped MoSe2 monolayer are nonmagnetic, while VMoSe6 vacancy geometry shows magnetic moment. • The combined vacancy doping and tensile strain can effectively tune the magnetism in MoSe2 monolayer. - Abstract: The effect of vacancy defect and strain on the magnetic property of monolayer MoSe2 was investigated by the first-principles calculation. Firstly, the most stable structure of monolayer MoSe2 and four kinds of vacancy, VSe, VSe2, VMoSe3 and VMoSe6 geometries were systematically studied. Our calculations show that 1H-MoSe2, as well as VSe, VSe2 and VMoSe3 vacancy doped MoSe2 monolayer are nonmagnetic, while VMoSe6 vacancy geometry shows magnetism and the magnetic moment mainly comes from the six Mo atoms around the vacancy although it is most difficult to form. Meanwhile, our calculations indicate that MoSe2 monolayer under tensile strain ranging from 0% to 10% does not show any magnetism. However, it is found that the combined tensile strain and VSe, or VSe2 and VMoSe3 vacancy indeed induce magnetism in MoSe2 monolayer by breaking the Mo–Mo metallic bonds. As for VMoSe6 vacancy geometry, the vacancy induced magnetic moment decrease under the imposed tensile strain

  8. Applied systems biology - vanillin production in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Strucko, Tomas; Eriksen, Carsten; Nielsen, J.;

    2012-01-01

    Vanillin is the most important aroma compound based on market value, and natural vanillin is extracted from the cured seed pods of the Vanilla orchid. Most of the world’s vanillin, however, is obtained by chemical synthesis from petrochemicals or wood pulp lignins. As an alternative, de novo...... biosynthesis of vanillin in baker’s yeast Saccharomyces cerevisiae was recently demonstrated by successfully introducing the metabolic pathway for vanillin production in yeast. Nevertheless, the amount of vanillin produced in this S. cerevisiae strain is insufficient for commercial production and improvements...... need to be done. We have introduced the genes necessary for vanillin production in an identical manner in two different yeast strains S288c and CEN.PK,where comprehensive – omics datasets are available, hence, allowing vanillin production in the two strain backgrounds to be evaluated and compared in a...

  9. Germanium-on-Silicon Strain Engineered Materials for Improved Device Performance Grown by Chemical Vapor Deposition

    Science.gov (United States)

    Bharathan, Jayesh Moorkoth

    The primary goal of this research is to develop a chemical vapor deposition process for growing epitaxial films of germanium on silicon (001) substrates with two-dimensional (2-D) morphology, and a low density of threading dislocations. Growth was carried out in a reduced-pressure chemical vapor deposition (RPCVD) system by a two-step growth technique. An accurate knowledge of elastic constants of thin films is important in understanding the effect of strain on material properties. Residual thermal strain was used to measure the Poisson ratio of Ge films grown on Si(001) substrates, by the sin2Psi method and highresolution x-ray diffraction. The Poisson ratio of the Ge films was measured to be 0.25, compared to the bulk value of 0.27. The result was found to be independent of film thickness and defect density, which confirmed that the strain is associated with the elastic response of the film. The study showed that the use of Poisson ratio instead of bulk compliance values yields a more accurate description of the state of in-plane strain present in the film. The experimentally measured in-plane strain in Ge films was found to be lower than the theoretical calculations based on the differential thermal expansion coefficients of Si and Ge. The mechanism of thermal misfit strain relaxation in epitaxial Ge films grown on Si(001) substrates was investigated by x-ray diffraction, and transmission electron microscopy. Lattice misfit strain associated with Ge/(001)Si mismatched epitaxy is relieved by a network of Lomer edge misfit dislocations during the first step of the growth technique. However, thermal misfit strain energy during growth is relieved by interdiffusion mechanism at the heterointerface. Two SiGe compositions containing 0.5 and 6.0 atomic percent Si were detected that relieve the thermal mismatch strain associated with the two steps of the growth process. This study discusses the importance of interdiffusion mechanism in relieving small misfit strains

  10. Reconstruction of the carnitine biosynthesis pathway from Neurospora crassa in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Franken, Jaco; Burger, Anita; Swiegers, Jan H; Bauer, Florian F

    2015-08-01

    Industrial synthesis of L-carnitine is currently performed by whole-cell biotransformation of industrial waste products, mostly D-carnitine and cronobetaine, through specific bacterial species. No comparable system has been established using eukaryotic microorganisms, even though there is a significant and growing international demand for either the pure compound or carnitine-enriched consumables. In eukaryotes, including the fungus Neurospora crassa, L-carnitine is biosynthesized through a four-step metabolic conversion of trimethyllysine to L-carnitine. In contrast, the industrial yeast, Saccharomyces cerevisiae lacks the enzymes of the eukaryotic biosynthesis pathway and is unable to synthesize carnitine. This study describes the cloning of all four of the N. crassa carnitine biosynthesis genes and the reconstruction of the entire pathway in S. cerevisiae. The engineered yeast strains were able to catalyze the synthesis of L-carnitine, which was quantified using hydrophilic interaction liquid chromatography electrospray ionization mass spectrometry (HILIC-ESI-MS) analyses, from trimethyllysine. Furthermore, the yeast threonine aldolase Gly1p was shown to effectively catalyze the second step of the pathway, fulfilling the role of a serine hydroxymethyltransferase. The analyses also identified yeast enzymes that interact with the introduced pathway, including Can1p, which was identified as the yeast transporter for trimethyllysine, and the two yeast serine hydroxymethyltransferases, Shm1p and Shm2p. Together, this study opens the possibility of using an engineered, carnitine-producing yeast in various industrial applications while providing insight into possible future strategies aimed at tailoring the production capacity of such strains. PMID:25851717

  11. Measurements of Creep Internal Stress Based on Constant Strain Rate and Its Application to Engineering

    Institute of Scientific and Technical Information of China (English)

    TAO Wen-liang; WEI Tao

    2006-01-01

    This research is carried out on the basis of Constant Strain Rate(CSR) to measure creep internal stress. Measurements of creep internal stress are conducted on the material test machine by using the CSR method. A mathematical model of creep internal stress is also proposed and its application is presented in this paper.

  12. Response of Saccharomyces cerevisiae to cadmium stress

    Energy Technology Data Exchange (ETDEWEB)

    Moreira, Luciana Mara Costa; Ribeiro, Frederico Haddad; Neves, Maria Jose [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil). Lab. de Radiobiologia], e-mail: luamatu@uol.com.br; Porto, Barbara Abranches Araujo; Amaral, Angela M.; Menezes, Maria Angela B.C. [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Lab. de Ativacao Neutronica], e-mail: menezes@cdtn.br; Rosa, Carlos Augusto [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Microbiologia], e-mail: carlrosa@icb.ufmg

    2009-07-01

    The intensification of industrial activity has been greatly contributing with the increase of heavy metals in the environment. Among these heavy metals, cadmium becomes a serious pervasive environmental pollutant. The cadmium is a heavy metal with no biological function, very toxic and carcinogenic at low concentrations. The toxicity of cadmium and several other metals can be mainly attributed to the multiplicity of coordination complexes and clusters that they can form. Some aspects of the cellular response to cadmium were extensively investigated in the yeast Saccharomyces cerevisiae. The primary site of interaction between many toxic metals and microbial cells is the plasma membrane. Plasma-membrane permeabilisation has been reported in a variety of microorganisms following cadmium exposure, and is considered one mechanism of cadmium toxicity in the yeast. In this work, using the yeast strain S. cerevisiae W303-WT, we have investigated the relationships between Cd uptake and release of cellular metal ions (K{sup +} and Na{sup +}) using neutron activation technique. The neutron activation was an easy, rapid and suitable technique for doing these metal determinations on yeast cells; was observed the change in morphology of the strains during the process of Cd accumulation, these alterations were observed by Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM) during incorporation of cadmium. (author)

  13. Use of pantothenate as a metabolic switch increases the genetic stability of farnesene producing Saccharomyces cerevisiae.

    Science.gov (United States)

    Sandoval, Celeste M; Ayson, Marites; Moss, Nathan; Lieu, Bonny; Jackson, Peter; Gaucher, Sara P; Horning, Tizita; Dahl, Robert H; Denery, Judith R; Abbott, Derek A; Meadows, Adam L

    2014-09-01

    We observed that removing pantothenate (vitamin B5), a precursor to co-enzyme A, from the growth medium of Saccharomyces cerevisiae engineered to produce β-farnesene reduced the strain׳s farnesene flux by 70%, but increased its viability, growth rate and biomass yield. Conversely, the growth rate and biomass yield of wild-type yeast were reduced. Cultivation in media lacking pantothenate eliminates the growth advantage of low-producing mutants, leading to improved production upon scale-up to lab-scale bioreactor testing. An omics investigation revealed that when exogenous pantothenate levels are limited, acyl-CoA metabolites decrease, β-oxidation decreases from unexpectedly high levels in the farnesene producer, and sterol and fatty acid synthesis likely limits the growth rate of the wild-type strain. Thus pantothenate supplementation can be utilized as a "metabolic switch" for tuning the synthesis rates of molecules relying on CoA intermediates and aid the economic scale-up of strains producing acyl-CoA derived molecules to manufacturing facilities. PMID:25076380

  14. Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Franken, Jaco; Brandt, Bianca A; Tai, Siew L; Bauer, Florian F

    2013-01-01

    Levans are fructose polymers synthesized by a broad range of micro-organisms and a limited number of plant species as non-structural storage carbohydrates. In microbes, these polymers contribute to the formation of the extracellular polysaccharide (EPS) matrix and play a role in microbial biofilm formation. Levans belong to a larger group of commercially important polymers, referred to as fructans, which are used as a source of prebiotic fibre. For levan, specifically, this market remains untapped, since no viable production strategy has been established. Synthesis of levan is catalysed by a group of enzymes, referred to as levansucrases, using sucrose as substrate. Heterologous expression of levansucrases has been notoriously difficult to achieve in Saccharomyces cerevisiae. As a strategy, this study used an invertase (Δsuc2) null mutant and two separate, engineered, sucrose accumulating yeast strains as hosts for the expression of the levansucrase M1FT, previously cloned from Leuconostoc mesenteroides. Intracellular sucrose accumulation was achieved either by expression of a sucrose synthase (Susy) from potato or the spinach sucrose transporter (SUT). The data indicate that in both Δsuc2 and the sucrose accumulating strains, the M1FT was able to catalyse fructose polymerisation. In the absence of the predicted M1FT secretion signal, intracellular levan accumulation was significantly enhanced for both sucrose accumulation strains, when grown on minimal media. Interestingly, co-expression of M1FT and SUT resulted in hyper-production and extracellular build-up of levan when grown in rich medium containing sucrose. This study presents the first report of levan production in S. cerevisiae and opens potential avenues for the production of levan using this well established industrial microbe. Furthermore, the work provides interesting perspectives when considering the heterologous expression of sugar polymerizing enzymes in yeast. PMID:24147008

  15. Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Jaco Franken

    Full Text Available Levans are fructose polymers synthesized by a broad range of micro-organisms and a limited number of plant species as non-structural storage carbohydrates. In microbes, these polymers contribute to the formation of the extracellular polysaccharide (EPS matrix and play a role in microbial biofilm formation. Levans belong to a larger group of commercially important polymers, referred to as fructans, which are used as a source of prebiotic fibre. For levan, specifically, this market remains untapped, since no viable production strategy has been established. Synthesis of levan is catalysed by a group of enzymes, referred to as levansucrases, using sucrose as substrate. Heterologous expression of levansucrases has been notoriously difficult to achieve in Saccharomyces cerevisiae. As a strategy, this study used an invertase (Δsuc2 null mutant and two separate, engineered, sucrose accumulating yeast strains as hosts for the expression of the levansucrase M1FT, previously cloned from Leuconostoc mesenteroides. Intracellular sucrose accumulation was achieved either by expression of a sucrose synthase (Susy from potato or the spinach sucrose transporter (SUT. The data indicate that in both Δsuc2 and the sucrose accumulating strains, the M1FT was able to catalyse fructose polymerisation. In the absence of the predicted M1FT secretion signal, intracellular levan accumulation was significantly enhanced for both sucrose accumulation strains, when grown on minimal media. Interestingly, co-expression of M1FT and SUT resulted in hyper-production and extracellular build-up of levan when grown in rich medium containing sucrose. This study presents the first report of levan production in S. cerevisiae and opens potential avenues for the production of levan using this well established industrial microbe. Furthermore, the work provides interesting perspectives when considering the heterologous expression of sugar polymerizing enzymes in yeast.

  16. S. cerevisiae Sit4 Phosphatase Is Active Irrespective of the Nitrogen Source Provided and Gln3 Phosphorylation Levels Become Nitrogen Source-Responsive In a sit4 Deleted Strain

    OpenAIRE

    Tate, Jennifer J.; Feller, André; Dubois, Evelyne; Cooper, Terrance G.

    2006-01-01

    Tor1,2 control of type-2A-related phosphatase activities in S. cerevisiae has been reported to be responsible for the regulation of Gln3 phosphorylation and intracellular localization in response to the nature of the nitrogen source available. According to the model, excess nitrogen stimulates Tor1,2 to phosphorylate Tip41 and/or Tap42. Tap42 then complexes with and inactivates Sit4 phosphatase, thereby preventing it from dephosphorylating Gln3. Phosphorylated Gln3 complexes with Ure2 and is ...

  17. Progress toward analytical description of the creep strain-time behavior of engineering alloys

    International Nuclear Information System (INIS)

    Elevated-temperature design methods in the United States often require a comprehensive description of the properties of the construction materials. These descriptions include representations for creep strain-time behavior as a function of stress, temperature, and material variability. Work conducted at this laboratory in the past five years toward the development of analytical techniques to derive such representations is summarized. Results for several common elevated-temperature structural materials are presented to illustrate the techniques

  18. Creation of High-Yield Polyhydroxyalkanoates Engineered Strains by Low Energy Ion Implantation

    Science.gov (United States)

    Qian, Shiquan; Cheng, Ying; Zhu, Suwen; Cheng, Beijiu

    2008-12-01

    Polyhydroxyalkanoates (PHAs), as a candidate for biodegradable plastic materials, can be synthesized by numerous microorganisms. However, as its production cost is high in comparison with those of chemically synthesized plastics, a lot of research has been focused on the efficient production of PHAs using different methods. In the present study, the mutation effects of PHAs production in strain pCB4 were investigated with implantation of low energy ions. It was found that under the implantation conditions of 7.8 × 1014 N+/cm2 at 10 keV, a high-yield PHAs strain with high genetic stability was generated from many mutants. After optimizing its fermentation conditions, the biomass, PHAs concentration and PHAs content of pCBH4 reached 2.26 g/L, 1.81 g/L, and 80.08% respectively, whereas its wild type controls were about 1.24 g/L, 0.61 g/L, and 49.20%. Moreover, the main constituent of PHAs was identified as poly-3-hydroxybutyrates (PHB) in the mutant stain and the yield of this compound was increased up to 41.33% in contrast to that of 27.78% in the wild type strain.

  19. Creation of High-Yield Polyhydroxyalkanoates Engineered Strains by Low Energy Ion Implantation

    Institute of Scientific and Technical Information of China (English)

    QIAN Shiquan; CHENG Ying; ZHU Suwen; CHENG Beijiu

    2008-01-01

    Polyhydroxyalkanoates (PHAs), as a candidate for biodegradable plastic materials, can be synthesized by numerous microorganisms. However, as its production cost is high in comparison with those of chemically synthesized plastics, a lot of research has been focused on the efficient production of PHAs using different methods. In the present study, the mutation effects of PHAs production in strain pCB4 were investigated with implantation of low energy ions. It was found that under the implantation conditions of 7.8×1014 N+/cm2 at 10 keV, a high-yield PHAs strain with high genetic stability was generated from many mutants. After optimizing its fermentation conditions, the biomass, PHAs concentration and PHAs content of pCBH4reached 2.26 g/L, 1.81 g/L, and 80.08% respectively, whereas its wild type controls were about 1.24 g/L, 0.61 g/L, and 49.20%. Moreover, the main constituent of PHAs was identified as poly-3-hydroxybutyrates (PHB) in the mutant stain and the yield of this compound was increased up to 41.33% in contrast to that of 27.78% in the wild type strain.

  20. Characteristics of Saccharomyces cerevisiae yeasts exhibiting rough colonies and pseudohyphal morphology with respect to alcoholic fermentation

    OpenAIRE

    Vanda Renata Reis; Ana Paula Guarnieri Bassi; Jessica Carolina Gomes da Silva; Sandra Regina Ceccato-Antonini

    2014-01-01

    Among the native yeasts found in alcoholic fermentation, rough colonies associated with pseudohyphal morphology belonging to the species Saccharomyces cerevisiae are very common and undesirable during the process. The aim of this work was to perform morphological and physiological characterisations of S. cerevisiae strains that exhibited rough and smooth colonies in an attempt to identify alternatives that could contribute to the management of rough colony yeasts in alcoholic fermentation. Ch...