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Sample records for saccharomyces cerevisiae heterologously

  1. Adaption of Saccharomyces cerevisiae expressing a heterologous protein

    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...

  2. Heat shock response improves heterologous protein secretion in Saccharomyces cerevisiae

    Hou, Jin; Österlund, Tobias; Liu, Zihe

    2013-01-01

    The yeast Saccharomyces cerevisiae is a widely used platform for the production of heterologous proteins of medical or industrial interest. However, heterologous protein productivity is often low due to limitations of the host strain. Heat shock response (HSR) is an inducible, global, cellular...... stress response, which facilitates the cell recovery from many forms of stress, e.g., heat stress. In S. cerevisiae, HSR is regulated mainly by the transcription factor heat shock factor (Hsf1p) and many of its targets are genes coding for molecular chaperones that promote protein folding and prevent...... the accumulation of mis-folded or aggregated proteins. In this work, we over-expressed a mutant HSF1 gene HSF1-R206S which can constitutively activate HSR, so the heat shock response was induced at different levels, and we studied the impact of HSR on heterologous protein secretion. We found that moderate and high...

  3. Functional expression and evaluation of heterologous phosphoketolases in Saccharomyces cerevisiae

    Bergman, Alexandra; Siewers, Verena; Nielsen, Jens

    2016-01-01

    Phosphoketolases catalyze an energy-and redox-independent cleavage of certain sugar phosphates. Hereby, the two-carbon (C2) compound acetyl-phosphate is formed, which enzymatically can be converted into acetyl-CoA-a key precursor in central carbon metabolism. Saccharomyces cerevisiae does...... not demonstrate efficient phosphoketolase activity naturally. In this study, we aimed to compare and identify efficient heterologous phosphoketolase enzyme candidates that in yeast have the potential to reduce carbon loss compared to the native acetyl-CoA producing pathway by redirecting carbon flux directly from...... C5 and C6 sugars towards C2-synthesis. Nine phosphoketolase candidates were expressed in S. cerevisiae of which seven produced significant amounts of acetyl-phosphate after provision of sugar phosphate substrates in vitro. The candidates showed differing substrate specificities, and some...

  4. Functional expression of a heterologous nickel-dependent, ATP-independent urease in Saccharomyces cerevisiae

    Milne, N.; Luttik, M.A.H.; Cueto Rojas, H.F.; Wahl, A.; Van Maris, A.J.A.; Pronk, J.T.; Daran, J.G.

    2015-01-01

    In microbial processes for production of proteins, biomass and nitrogen-containing commodity chemicals, ATP requirements for nitrogen assimilation affect product yields on the energy producing substrate. In Saccharomyces cerevisiae, a current host for heterologous protein production and potential

  5. Comparison of heterologous xylose transporters in recombinant Saccharomyces cerevisiae

    Hahn-Hägerdal Bärbel

    2010-03-01

    Full Text Available Abstract Background Baker's yeast (Saccharomyces cerevisiae has been engineered for xylose utilization to enable production of fuel ethanol from lignocellulose raw material. One unresolved challenge is that S. cerevisiae lacks a dedicated transport system for pentose sugars, which means that xylose is transported by non-specific Hxt transporters with comparatively low transport rate and affinity for xylose. Results In this study, we compared three heterologous xylose transporters that have recently been shown to improve xylose uptake under different experimental conditions. The transporters Gxf1, Sut1 and At5g59250 from Candida intermedia, Pichia stipitis and Arabidopsis thaliana, respectively, were expressed in isogenic strains of S. cerevisiae and the transport kinetics and utilization of xylose was evaluated. Expression of the Gxf1 and Sut1 transporters led to significantly increased affinity and transport rates of xylose. In batch cultivation at 4 g/L xylose concentration, improved transport kinetics led to a corresponding increase in xylose utilization, whereas no correlation could be demonstrated at xylose concentrations greater than 15 g/L. The relative contribution of native sugar transporters to the overall xylose transport capacity was also estimated during growth on glucose and xylose. Conclusions Kinetic characterization and aerobic batch cultivation of strains expressing the Gxf1, Sut1 and At5g59250 transporters showed a direct relationship between transport kinetics and xylose growth. The Gxf1 transporter had the highest transport capacity and the highest xylose growth rate, followed by the Sut1 transporter. The range in which transport controlled the growth rate was determined to between 0 and 15 g/L xylose. The role of catabolite repression in regulation of native transporters was also confirmed by the observation that xylose transport by native S. cerevisiae transporters increased significantly during cultivation in xylose and

  6. Yield improvement of heterologous peptides expressed in yps1-disrupted Saccharomyces cerevisiae strains.

    Egel-Mitani; Andersen; Diers; Hach; Thim; Hastrup; Vad

    2000-06-01

    Heterologous protein expression levels in Saccharomyces cerevisiae fermentations are highly dependent on the susceptibility to endogenous yeast proteases. Small peptides, such as glucagon and glucagon-like-peptides (GLP-1 and GLP-2), featuring an open structure are particularly accessible for proteolytic degradation during fermentation. Therefore, homogeneous products cannot be obtained. The most sensitive residues are found at basic amino acid residues in the peptide sequence. These heterologous peptides are degraded mainly by the YPS1-encoded aspartic protease, yapsin1, when produced in the yeast. In this article, distinct degradation products were analyzed by HPLC and mass spectrometry, and high yield of the heterologous peptide production has been achieved by the disruption of the YPS1 gene (previously called YAP3). By this technique, high yield continuous fermentation of glucagon in S. cerevisiae is now possible.

  7. Functional expression of a heterologous nickel-dependent, ATP-independent urease in Saccharomyces cerevisiae.

    Milne, N; Luttik, M A H; Cueto Rojas, H F; Wahl, A; van Maris, A J A; Pronk, J T; Daran, J M

    2015-07-01

    In microbial processes for production of proteins, biomass and nitrogen-containing commodity chemicals, ATP requirements for nitrogen assimilation affect product yields on the energy producing substrate. In Saccharomyces cerevisiae, a current host for heterologous protein production and potential platform for production of nitrogen-containing chemicals, uptake and assimilation of ammonium requires 1 ATP per incorporated NH3. Urea assimilation by this yeast is more energy efficient but still requires 0.5 ATP per NH3 produced. To decrease ATP costs for nitrogen assimilation, the S. cerevisiae gene encoding ATP-dependent urease (DUR1,2) was replaced by a Schizosaccharomyces pombe gene encoding ATP-independent urease (ure2), along with its accessory genes ureD, ureF and ureG. Since S. pombe ure2 is a Ni(2+)-dependent enzyme and Saccharomyces cerevisiae does not express native Ni(2+)-dependent enzymes, the S. pombe high-affinity nickel-transporter gene (nic1) was also expressed. Expression of the S. pombe genes into dur1,2Δ S. cerevisiae yielded an in vitro ATP-independent urease activity of 0.44±0.01 µmol min(-1) mg protein(-1) and restored growth on urea as sole nitrogen source. Functional expression of the Nic1 transporter was essential for growth on urea at low Ni(2+) concentrations. The maximum specific growth rates of the engineered strain on urea and ammonium were lower than those of a DUR1,2 reference strain. In glucose-limited chemostat cultures with urea as nitrogen source, the engineered strain exhibited an increased release of ammonia and reduced nitrogen content of the biomass. Our results indicate a new strategy for improving yeast-based production of nitrogen-containing chemicals and demonstrate that Ni(2+)-dependent enzymes can be functionally expressed in S. cerevisiae. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  8. Mitochondrial targeting increases specific activity of a heterologous valine assimilation pathway in Saccharomyces cerevisiae

    Kevin V. Solomon

    2016-12-01

    Full Text Available Bio-based isobutantol is a sustainable ‘drop in’ substitute for petroleum-based fuels. However, well-studied production routes, such as the Ehrlich pathway, have yet to be commercialized despite more than a century of research. The more versatile bacterial valine catabolism may be a competitive alternate route producing not only an isobutanol precursor but several carboxylic acids with applications as biomonomers, and building blocks for other advanced biofuels. Here, we transfer the first two committed steps of the pathway from pathogenic Pseudomonas aeruginosa PAO1 to yeast to evaluate their activity in a safer model organism. Genes encoding the heteroligomeric branched chain keto-acid dehydrogenase (BCKAD; bkdA1, bkdA2, bkdB, lpdV, and the homooligomeric acyl-CoA dehydrogenase (ACD; acd1 were tagged with fluorescence epitopes and targeted for expression in either the mitochondria or cytoplasm of S. cerevisiae. We verified the localization of our constructs with confocal fluorescence microscopy before measuring the activity of tag-free constructs. Despite reduced heterologous expression of mitochondria-targeted enzymes, their specific activities were significantly improved with total enzyme activities up to 138% greater than those of enzymes expressed in the cytoplasm. In total, our results demonstrate that the choice of protein localization in yeast has significant impact on heterologous activity, and suggests a new path forward for isobutanol production. Keywords: Pseudomonas, Isobutanol, Dehydrogenase, Mitochondria, Saccharomyces cerevisiae, Metabolic engineering

  9. [High-level expression of heterologous protein based on increased copy number in Saccharomyces cerevisiae].

    Zhang, Xinjie; He, Peng; Tao, Yong; Yang, Yi

    2013-11-04

    High-level expression system of heterologous protein mediated by internal ribosome entry site (IRES) in Saccharomyces cerevisiae was constructed, which could be used for other applications of S. cerevisiae in metabolic engineering. We constructed co-expression cassette (promoter-mCherry-TIF4631 IRES-URA3) containing promoters Pilv5, Padh2 and Ptdh3 and recombined the co-expression cassette into the genome of W303-1B-A. The URA3+ transformants were selected. By comparing the difference in the mean florescence value of mCherry in transformants, the effect of three promoters was detected in the co-expression cassette. The copy numbers of the interested genes in the genome were determined by Real-Time PCR. We analyzed genetic stability by continuous subculturing transformants in the absence of selection pressure. To verify the application of co-expression cassette, the ORF of mCherry was replaced by beta-galactosidase (LACZ) and xylose reductase (XYL1). The enzyme activities and production of beta-galactosidase and xylose reductase were detected. mCherry has been expressed in the highest-level in transformants with co-expression cassette containing Pilv5 promoter. The highest copy number of DNA fragment integrating in the genome was 47 in transformants containing Pilv5. The engineering strains showed good genetic stability. Xylose reductase was successfully expressed in the co-expression cassette containing Pilv5 promoter and TIF4631 IRES. The highest enzyme activity was 0. 209 U/mg crude protein in the transformants WIX-10. Beta-galactosidase was also expressed successfully. The transformants that had the highest enzyme activity was WIL-1 and the enzyme activity was 12.58 U/mg crude protein. The system mediated by Pilv5 promoter and TIF4631 IRES could express heterologous protein efficiently in S. cerevisiae. This study offered a new strategy for expression of heterologous protein in S. cerevisiae and provided sufficient experimental evidence for metabolic engineering

  10. Synthetic Promoters and Transcription Factors for Heterologous Protein Expression in Saccharomyces cerevisiae

    Fabian Machens

    2017-10-01

    Full Text Available Orthogonal systems for heterologous protein expression as well as for the engineering of synthetic gene regulatory circuits in hosts like Saccharomyces cerevisiae depend on synthetic transcription factors (synTFs and corresponding cis-regulatory binding sites. We have constructed and characterized a set of synTFs based on either transcription activator-like effectors or CRISPR/Cas9, and corresponding small synthetic promoters (synPs with minimal sequence identity to the host’s endogenous promoters. The resulting collection of functional synTF/synP pairs confers very low background expression under uninduced conditions, while expression output upon induction of the various synTFs covers a wide range and reaches induction factors of up to 400. The broad spectrum of expression strengths that is achieved will be useful for various experimental setups, e.g., the transcriptional balancing of expression levels within heterologous pathways or the construction of artificial regulatory networks. Furthermore, our analyses reveal simple rules that enable the tuning of synTF expression output, thereby allowing easy modification of a given synTF/synP pair. This will make it easier for researchers to construct tailored transcriptional control systems.

  11. Saccharomyces cerevisiae

    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...

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

    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. Keywords: Yeast, Cell factory, Strain choice, Heterologous production, Vanillin-glucoside, Shikimate pathway

  13. Altered Phenotypes in Saccharomyces cerevisiae by Heterologous Expression of Basidiomycete Moniliophthora perniciosa SOD2 Gene

    Sônia C. Melo

    2015-06-01

    Full Text Available Heterologous expression of a putative manganese superoxide dismutase gene (SOD2 of the basidiomycete Moniliophthora perniciosa complemented the phenotypes of a Saccharomyces cerevisiae sod2Δ mutant. Sequence analysis of the cloned M. perniciosa cDNA revealed an open reading frame (ORF coding for a 176 amino acid polypeptide with the typical metal-binding motifs of a SOD2 gene, named MpSOD2. Phylogenetic comparison with known manganese superoxide dismutases (MnSODs located the protein of M. perniciosa (MpSod2p in a clade with the basidiomycete fungi Coprinopsis cinerea and Laccaria bicolor. Haploid wild-type yeast transformants containing a single copy of MpSOD2 showed increased resistance phenotypes against oxidative stress-inducing hydrogen peroxide and paraquat, but had unaltered phenotype against ultraviolet–C (UVC radiation. The same transformants exhibited high sensitivity against treatment with the pro-mutagen diethylnitrosamine (DEN that requires oxidation to become an active mutagen/carcinogen. Absence of MpSOD2 in the yeast sod2Δ mutant led to DEN hyper-resistance while introduction of a single copy of this gene restored the yeast wild-type phenotype. The haploid yeast wild-type transformant containing two SOD2 gene copies, one from M. perniciosa and one from its own, exhibited DEN super-sensitivity. This transformant also showed enhanced growth at 37 °C on the non-fermentable carbon source lactate, indicating functional expression of MpSod2p. The pro-mutagen dihydroethidium (DHE-based fluorescence assay monitored basal level of yeast cell oxidative stress. Compared to the wild type, the yeast sod2Δ mutant had a much higher level of intrinsic oxidative stress, which was reduced to wild type (WT level by introduction of one copy of the MpSOD2 gene. Taken together our data indicates functional expression of MpSod2 protein in the yeast S. cerevisiae.

  14. Heterologous expression of the Crassostrea gigas (Pacific oyster) alternative oxidase in the yeast Saccharomyces cerevisiae.

    Robertson, Aaron; Schaltz, Kyle; Neimanis, Karina; Staples, James F; McDonald, Allison E

    2016-10-01

    Alternative oxidase (AOX) is a terminal oxidase within the inner mitochondrial membrane (IMM) present in many organisms where it functions in the electron transport system (ETS). AOX directly accepts electrons from ubiquinol and is therefore capable of bypassing ETS Complexes III and IV. The human genome does not contain a gene coding for AOX, so AOX expression has been suggested as a gene therapy for a range of human mitochondrial diseases caused by genetic mutations that render Complex III and/or IV dysfunctional. An effective means of screening mutations amenable to AOX treatment remains to be devised. We have generated such a tool by heterologously expressing AOX from the Pacific oyster (Crassostrea gigas) in the yeast Saccharomyces cerevisiae under the control of a galactose promoter. Our results show that this animal AOX is monomeric and is correctly targeted to yeast mitochondria. Moreover, when expressed in yeast, Pacific oyster AOX is a functional quinol oxidase, conferring cyanide-resistant growth and myxothiazol-resistant oxygen consumption to yeast cells and isolated mitochondria. This system represents a high-throughput screening tool for determining which Complex III and IV genetic mutations in yeast will be amenable to AOX gene therapy. As many human genes are orthologous to those found in yeast, our invention represents an efficient and cost-effective way to evaluate viable research avenues. In addition, this system provides the opportunity to learn more about the localization, structure, and regulation of AOXs from animals that are not easily reared or manipulated in the lab.

  15. Heterologous expression of a tannic acid-inducible laccase3 of Cryphonectria parasitica in Saccharomyces cerevisiae

    Kim Dae-Hyuk

    2010-02-01

    Full Text Available Abstract Background A tannic acid-inducible and mycoviral-regulated laccase3 (lac3 from the chestnut blight fungus Cryphonectria parasitica has recently been identified, but further characterization was hampered because of the precipitation of protein products by tannic acid supplementation. The present study investigated the heterologous expression of the functional laccase3 using a yeast Saccharomyces cerevisiae. Results Laccase activity in the culture broth of transformants measured using a laccase-specific substrate suggested that the lac3 gene was successfully expressed and the corresponding protein product secreted into the culture media. In addition, activity staining and Western blot analysis of a native gel revealed that the enzyme activity co-existed with the protein product specific to anti-laccase3 antibody, confirming that the cloned lac3 gene is responsible for the laccase activity. When transformants were grown on plates containing tannic acid-supplemented media, brown coloration was observed around transformed cells, indicating the oxidation of tannic acid. However, the enzymatic activity was measurable only in the selective ura- media and was negligible in nonselective nutrient-rich culture conditions. This was in part because of the increased plasmid instability in the nonselective media. Moreover, the protein product of lac3 appears to be sensitive to the cultured nonselective nutrient-rich broth, because a rapid decline in enzymatic activity was observed when the cultured broth of ura- media was mixed with that of nonselective nutrient-rich broth. In addition, constitutive expression of the lac3 gene resulted in a reduced cell number of the lac3 transformants compared to that of vector-only transformed control. However, the presence of recombinant vector without lac3 induction did not affect the growth of transformants. Conclusions The results suggest that expression of the lac3 gene has an inhibitory effect on the growth of

  16. Heterologous Expression of Membrane and Soluble Proteins Derepresses GCN4 mRNA Translation in the Yeast Saccharomyces cerevisiae

    Steffensen, L.; Pedersen, P. A.

    2006-01-01

    -ATPase also induced GCN4 translation. Derepression of GCN4 translation required phosphorylation of eIF-2 , the tRNA binding domain of Gcn2p, and the ribosome-associated proteins Gcn1p and Gcn20p. The increase in Gcn4p density in response to heterologous expression did not induce transcription from the HIS4...... promoter, a traditional Gcn4p target.......This paper describes the first physiological response at the translational level towards heterologous protein production in Saccharomyces cerevisiae. In yeast, the phosphorylation of eukaryotic initiation factor 2 (eIF-2 ) by Gcn2p protein kinase mediates derepression of GCN4 mRNA translation. Gcn4...

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

    Liu, Zihe; Liu, Lifang; Osterlund, Tobias

    2014-01-01

    this modification alone, the amylase secretion could be improved by 35%. As a complement to the identification of genomic variants, transcriptome analysis was also performed in order to understand on a global level the transcriptional changes associated with the improved amylase production caused by UV mutagenesis.......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 widely used as a cell factory for recombinant protein production. However, many recombinant proteins are produced at only 1% (or less) of the theoretical capacity due to the complexity of the secretory pathway, which has not been fully exploited. In this study, we applied the concept of inverse metabolic...

  18. Heterologous expression of a rice metallothionein isoform (OsMTI-1b in Saccharomyces cerevisiae enhances cadmium, hydrogen peroxide and ethanol tolerance

    Zahra Ansarypour

    Full Text Available Abstract Metallothioneins are a superfamily of low-molecular-weight, cysteine (Cys-rich proteins that are believed to play important roles in protection against metal toxicity and oxidative stress. The main purpose of this study was to investigate the effect of heterologous expression of a rice metallothionein isoform (OsMTI-1b on the tolerance of Saccharomyces cerevisiae to Cd2+, H2O2 and ethanol stress. The gene encoding OsMTI-1b was cloned into p426GPD as a yeast expression vector. The new construct was transformed to competent cells of S. cerevisiae. After verification of heterologous expression of OsMTI-1b, the new strain and control were grown under stress conditions. In comparison to control strain, the transformed S. cerevisiae cells expressing OsMTI-1b showed more tolerance to Cd2+ and accumulated more Cd2+ ions when they were grown in the medium containing CdCl2. In addition, the heterologous expression of GST-OsMTI-1b conferred H2O2 and ethanol tolerance to S. cerevisiae cells. The results indicate that heterologous expression of plant MT isoforms can enhance the tolerance of S. cerevisiae to multiple stresses.

  19. Functional expression of rat VPAC1 receptor in Saccharomyces cerevisiae

    Hansen, M.K.; Tams, J.W.; Fahrenkrug, Jan

    1999-01-01

    G protein-coupled receptor; heterologous expression; membrane protein; Saccharomyces cerevisiae, vasoactive intestinal polypeptide; yeast mating factor-pre-pro *Ga-leader peptide......G protein-coupled receptor; heterologous expression; membrane protein; Saccharomyces cerevisiae, vasoactive intestinal polypeptide; yeast mating factor-pre-pro *Ga-leader peptide...

  20. Production of a heterologous proteinase A by Saccharomyces kluyveri

    Møller, Kasper; Tidemand, L.D.; Winther, J.R.

    2001-01-01

    In order to evaluate the potential of Saccharomyces kluyveri for heterologous protein production, S. kluyveri Y159 was transformed with a S. cerevisiae-based multi-copy plasmid containing the S. cerevisiae PEP4 gene, which encodes proteinase A, under the control of its native promoter. As a refer......In order to evaluate the potential of Saccharomyces kluyveri for heterologous protein production, S. kluyveri Y159 was transformed with a S. cerevisiae-based multi-copy plasmid containing the S. cerevisiae PEP4 gene, which encodes proteinase A, under the control of its native promoter...

  1. Diversion of flux toward sesquiterpene production in Saccharomyces cerevisiae by fusion of host and heterologous enzymes

    Albertsen, Line; Chen, Yun; Bach, Lars Stougaard

    2011-01-01

    be limited by the inability of the heterologous enzymes to collaborate with the native yeast enzymes. This may cause loss of intermediates by diffusion or degradation or due to conversion of the intermediate through competitive pathways. To bypass this problem, we have pursued a strategy in which key enzymes...... increased the production of patchoulol, the main sesquiterpene produced by PTS, up to 2-fold. Moreover, we have demonstrated that the fusion strategy can be used in combination with traditional metabolic engineering to further increase the production of patchoulol. This simple test case of synthetic biology...

  2. Improving heterologous protein secretion at aerobic conditions by activating hypoxia-induced genes in Saccharomyces cerevisiae

    Liu, Lifang; Zhang, Yiming; Liu, Zihe

    2015-01-01

    Oxygen is important for normal aerobic metabolism, as well as for protein production where it is needed for oxidative protein folding. However, several studies have reported that anaerobic conditions seem to be more favorable in terms of recombinant protein production. We were interested in incre......Oxygen is important for normal aerobic metabolism, as well as for protein production where it is needed for oxidative protein folding. However, several studies have reported that anaerobic conditions seem to be more favorable in terms of recombinant protein production. We were interested...... in increasing recombinant protein production under aerobic conditions so we focused on Rox1p regulation. Rox1p is a transcriptional regulator, which in oxidative conditions represses genes induced in hypoxia. We deleted ROX1 and studied the effects on the production of recombinant proteins in Saccharomyces...

  3. Improving L-arabinose utilization of pentose fermenting Saccharomyces cerevisiae cells by heterologous expression of L-arabinose transporting sugar transporters

    Boles Eckhard

    2011-10-01

    Full Text Available Abstract Background Hydrolysates of plant biomass used for the production of lignocellulosic biofuels typically contain sugar mixtures consisting mainly of D-glucose and D-xylose, and minor amounts of L-arabinose. The yeast Saccharomyces cerevisiae is the preferred microorganism for the fermentative production of ethanol but is not able to ferment pentose sugars. Although D-xylose and L-arabinose fermenting S. cerevisiae strains have been constructed recently, pentose uptake is still a limiting step in mixed sugar fermentations. Results Here we described the cloning and characterization of two sugar transporters, AraT from the yeast Scheffersomyces stipitis and Stp2 from the plant Arabidopsis thaliana, which mediate the uptake of L-arabinose but not of D-glucose into S. cerevisiae cells. A yeast strain lacking all of its endogenous hexose transporter genes and expressing a bacterial L-arabinose utilization pathway could no longer take up and grow with L-arabinose as the only carbon source. Expression of the heterologous transporters supported uptake and utilization of L-arabinose especially at low L-arabinose concentrations but did not, or only very weakly, support D-glucose uptake and utilization. In contrast, the S. cerevisiae D-galactose transporter, Gal2, mediated uptake of both L-arabinose and D-glucose, especially at high concentrations. Conclusions Using a newly developed screening system we have identified two heterologous sugar transporters from a yeast and a plant which can support uptake and utilization of L-arabinose in L-arabinose fermenting S. cerevisiae cells, especially at low L-arabinose concentrations.

  4. Heterologous expression of MlcE in Saccharomyces cerevisiae provides resistance to natural and semi-synthetic statins

    Ana Ley

    2015-12-01

    Full Text Available Statins are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the key enzyme in cholesterol biosynthesis. Their extensive use in treatment and prevention of cardiovascular diseases placed statins among the best selling drugs. Construction of Saccharomyces cerevisiae cell factory for the production of high concentrations of natural statins will require establishment of a non-destructive self-resistance mechanism to overcome the undesirable growth inhibition effects of statins. To establish active export of statins from yeast, and thereby detoxification, we integrated a putative efflux pump-encoding gene mlcE from the mevastatin-producing Penicillium citrinum into the S. cerevisiae genome. The resulting strain showed increased resistance to both natural statins (mevastatin and lovastatin and semi-synthetic statin (simvastatin when compared to the wild type strain. Expression of RFP-tagged mlcE showed that MlcE is localized to the yeast plasma and vacuolar membranes. We provide a possible engineering strategy for improvement of future yeast based production of natural and semi-synthetic statins. Keywords: Polyketide, Statins, Saccharomyces cerevisiae, Transport, Cell factory, Resistance

  5. Requirement of a Functional Flavin Mononucleotide Prenyltransferase for the Activity of a Bacterial Decarboxylase in a Heterologous Muconic Acid Pathway in Saccharomyces cerevisiae.

    Weber, Heike E; Gottardi, Manuela; Brückner, Christine; Oreb, Mislav; Boles, Eckhard; Tripp, Joanna

    2017-05-15

    Biotechnological production of cis , cis -muconic acid from renewable feedstocks is an environmentally sustainable alternative to conventional, petroleum-based methods. Even though a heterologous production pathway for cis , cis -muconic acid has already been established in the host organism Saccharomyces cerevisiae , the generation of industrially relevant amounts of cis , cis -muconic acid is hampered by the low activity of the bacterial protocatechuic acid (PCA) decarboxylase AroY isomeric subunit C iso (AroY-C iso ), leading to secretion of large amounts of the intermediate PCA into the medium. In the present study, we show that the activity of AroY-C iso in S. cerevisiae strongly depends on the strain background. We could demonstrate that the strain dependency is caused by the presence or absence of an intact genomic copy of PAD1 , which encodes a mitochondrial enzyme responsible for the biosynthesis of a prenylated form of the cofactor flavin mononucleotide (prFMN). The inactivity of AroY-C iso in strain CEN.PK2-1 could be overcome by plasmid-borne expression of Pad1 or its bacterial homologue AroY subunit B (AroY-B). Our data reveal that the two enzymes perform the same function in decarboxylation of PCA by AroY-C iso , although coexpression of Pad1 led to higher decarboxylase activity. Conversely, AroY-B can replace Pad1 in its function in decarboxylation of phenylacrylic acids by ferulic acid decarboxylase Fdc1. Targeting of the majority of AroY-B to mitochondria by fusion to a heterologous mitochondrial targeting signal did not improve decarboxylase activity of AroY-C iso , suggesting that mitochondrial localization has no major impact on cofactor biosynthesis. IMPORTANCE In Saccharomyces cerevisiae , the decarboxylation of protocatechuic acid (PCA) to catechol is the bottleneck reaction in the heterologous biosynthetic pathway for production of cis , cis -muconic acid, a valuable precursor for the production of bulk chemicals. In our work, we demonstrate

  6. Promiscuous activities of heterologous enzymes lead to unintended metabolic rerouting in Saccharomyces cerevisiae engineered to assimilate various sugars from renewable biomass.

    Yun, Eun Ju; Oh, Eun Joong; Liu, Jing-Jing; Yu, Sora; Kim, Dong Hyun; Kwak, Suryang; Kim, Kyoung Heon; Jin, Yong-Su

    2018-01-01

    Understanding the global metabolic network, significantly perturbed upon promiscuous activities of foreign enzymes and different carbon sources, is crucial for systematic optimization of metabolic engineering of yeast Saccharomyces cerevisiae . Here, we studied the effects of promiscuous activities of overexpressed enzymes encoded by foreign genes on rerouting of metabolic fluxes of an engineered yeast capable of assimilating sugars from renewable biomass by profiling intracellular and extracellular metabolites. Unbiased metabolite profiling of the engineered S. cerevisiae strain EJ4 revealed promiscuous enzymatic activities of xylose reductase and xylitol dehydrogenase on galactose and galactitol, respectively, resulting in accumulation of galactitol and tagatose during galactose fermentation. Moreover, during glucose fermentation, a trisaccharide consisting of glucose accumulated outside of the cells probably owing to the promiscuous and transglycosylation activity of β-glucosidase expressed for hydrolyzing cellobiose. Meanwhile, higher accumulation of fatty acids and secondary metabolites was observed during xylose and cellobiose fermentations, respectively. The heterologous enzymes functionally expressed in S. cerevisiae showed promiscuous activities that led to unintended metabolic rerouting in strain EJ4. Such metabolic rerouting could result in a low yield and productivity of a final product due to the formation of unexpected metabolites. Furthermore, the global metabolic network can be significantly regulated by carbon sources, thus yielding different patterns of metabolite production. This metabolomic study can provide useful information for yeast strain improvement and systematic optimization of yeast metabolism to manufacture bio-based products.

  7. Synthesis of FAEEs from glycerol in engineered Saccharomyces cerevisiae using endogenously produced ethanol by heterologous expression of an unspecific bacterial acyltransferase.

    Yu, Kyung Ok; Jung, Ju; Kim, Seung Wook; Park, Chul Hwan; Han, Sung Ok

    2012-01-01

    The high price of petroleum-based diesel fuel has led to the development of alternative fuels, such as ethanol. Saccharomyces cerevisiae was metabolically engineered to utilize glycerol as a substrate for ethanol production. For the synthesis of fatty acid ethyl esters (FAEEs) by engineered S. cerevisiae that utilize glycerol as substrate, heterologous expression of an unspecific acyltransferase from Acinetobacter baylyi with glycerol utilizing genes was established. As a result, the engineered YPH499 (pGcyaDak, pGupWs-DgaTCas) strain produced 0.24 g/L FAEEs using endogenous ethanol produced from glycerol. And this study also demonstrated the possibility of increasing FAEE production by enhancing ethanol production by minimizing the synthesis of glycerol. The overall FAEE production in strain YPH499 fps1Δ gpd2Δ (pGcyaDak, pGupWs-DgaTCas) was 2.1-fold more than in YPH499 (pGcyaDak, pGupWs-DgaTCas), with approximately 0.52 g/L FAEEs produced, while nearly 17 g/L of glycerol was consumed. These results clearly indicated that FAEEs were synthesized in engineered S. cerevisiae by esterifying exogenous fatty acids with endogenously produced ethanol from glycerol. This microbial system acts as a platform in applying metabolic engineering that allows the production of FAEEs from cheap and abundant substrates specifically glycerol through the use of endogenous bioethanol. Copyright © 2011 Wiley Periodicals, Inc.

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

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

    2004-01-01

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

  9. [Saccharomyces cerevisiae infections].

    Souza Goebel, Cristine; de Mattos Oliveira, Flávio; Severo, Luiz Carlos

    2013-01-01

    Saccharomyces cerevisiae is an ubiquitous yeast widely used in industry and it is also a common colonizer of the human mucosae. However, the incidence of invasive infection by these fungi has significantly increased in the last decades. To evaluate the infection by S. cerevisiae in a hospital in southern Brazil during a period of 10 years (2000-2010). Review of medical records of patients infected by this fungus. In this period, 6 patients were found to be infected by S. cerevisiae. The age range of the patients was from 10 years to 84. Urine, blood, ascitic fluid, peritoneal dialysis fluid, and esophageal biopsy samples were analyzed. The predisposing factors were cancer, transplant, surgical procedures, renal failure, use of venous catheters, mechanical ventilation, hospitalization in Intensive Care Unit, diabetes mellitus, chemotherapy, corticosteroid use, and parenteral nutrition. Amphotericin B and fluconazole were the treatments of choice. Three of the patients died and the other 3 were discharged from hospital. We must take special precautions in emerging infections, especially when there are predisposing conditions such as immunosuppression or patients with serious illnesses. The rapid and specific diagnosis of S. cerevisiae infections is important for therapeutic decision. Furthermore, epidemiological and efficacy studies of antifungal agents are necessary for a better therapeutic approach. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  10. MiYA, an efficient machine-learning workflow in conjunction with the YeastFab assembly strategy for combinatorial optimization of heterologous metabolic pathways in Saccharomyces cerevisiae.

    Zhou, Yikang; Li, Gang; Dong, Junkai; Xing, Xin-Hui; Dai, Junbiao; Zhang, Chong

    2018-05-01

    Facing boosting ability to construct combinatorial metabolic pathways, how to search the metabolic sweet spot has become the rate-limiting step. We here reported an efficient Machine-learning workflow in conjunction with YeastFab Assembly strategy (MiYA) for combinatorial optimizing the large biosynthetic genotypic space of heterologous metabolic pathways in Saccharomyces cerevisiae. Using β-carotene biosynthetic pathway as example, we first demonstrated that MiYA has the power to search only a small fraction (2-5%) of combinatorial space to precisely tune the expression level of each gene with a machine-learning algorithm of an artificial neural network (ANN) ensemble to avoid over-fitting problem when dealing with a small number of training samples. We then applied MiYA to improve the biosynthesis of violacein. Feed with initial data from a colorimetric plate-based, pre-screened pool of 24 strains producing violacein, MiYA successfully predicted, and verified experimentally, the existence of a strain that showed a 2.42-fold titer improvement in violacein production among 3125 possible designs. Furthermore, MiYA was able to largely avoid the branch pathway of violacein biosynthesis that makes deoxyviolacein, and produces very pure violacein. Together, MiYA combines the advantages of standardized building blocks and machine learning to accelerate the Design-Build-Test-Learn (DBTL) cycle for combinatorial optimization of metabolic pathways, which could significantly accelerate the development of microbial cell factories. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  11. Fungal genomics beyond Saccharomyces cerevisiae?

    Hofmann, Gerald; Mcintyre, Mhairi; Nielsen, Jens

    2003-01-01

    Fungi are used extensively in both fundamental research and industrial applications. Saccharomyces cerevisiae has been the model organism for fungal research for many years, particularly in functional genomics. However, considering the diversity within the fungal kingdom, it is obvious...

  12. Effects of fermentation by Saccharomyces cerevisiae and ...

    yassine

    2013-02-13

    Feb 13, 2013 ... Effect of Saccharomyces cerevisiae fermentation on the ... beetroot, fermentation, Saccharomyces cerevisiae, betalain compounds. ... by Saccharomyces cerevisiae strains (González et al., .... Both red and yellow pigments were influenced during S. .... in beverages such as white wine, grape fruit, and green.

  13. Engineering of aromatic amino acid metabolism in Saccharomyces cerevisiae

    Vuralhan, Z.

    2006-01-01

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

  14. Glucose repression in Saccharomyces cerevisiae

    Kayikci, Omur; Nielsen, Jens

    2015-01-01

    Glucose is the primary source of energy for the budding yeast Saccharomyces cerevisiae. Although yeast cells can utilize a wide range of carbon sources, presence of glucose suppresses molecular activities involved in the use of alternate carbon sources as well as it represses respiration and gluc......Glucose is the primary source of energy for the budding yeast Saccharomyces cerevisiae. Although yeast cells can utilize a wide range of carbon sources, presence of glucose suppresses molecular activities involved in the use of alternate carbon sources as well as it represses respiration...

  15. Induction of homologous recombination in Saccharomyces cerevisiae.

    Simon, J R; Moore, P D

    1988-09-01

    We have investigated the effects of UV irradiation of Saccharomyces cerevisiae in order to distinguish whether UV-induced recombination results from the induction of enzymes required for homologous recombination, or the production of substrate sites for recombination containing regions of DNA damage. We utilized split-dose experiments to investigate the induction of proteins required for survival, gene conversion, and mutation in a diploid strain of S. cerevisiae. We demonstrate that inducing doses of UV irradiation followed by a 6 h period of incubation render the cells resistant to challenge doses of UV irradiation. The effects of inducing and challenge doses of UV irradiation upon interchromosomal gene conversion and mutation are strictly additive. Using the yeast URA3 gene cloned in non-replicating single- and double-stranded plasmid vectors that integrate into chromosomal genes upon transformation, we show that UV irradiation of haploid yeast cells and homologous plasmid DNA sequences each stimulate homologous recombination approximately two-fold, and that these effects are additive. Non-specific DNA damage has little effect on the stimulation of homologous recombination, as shown by studies in which UV-irradiated heterologous DNA was included in transformation/recombination experiments. We further demonstrate that the effect of competing single- and double-stranded heterologous DNA sequences differs in UV-irradiated and unirradiated cells, suggesting an induction of recombinational machinery in UV-irradiated S. cerevisiae cells.

  16. levadura Saccharomyces Cerevisiae

    B. Aguilar Uscanga

    2005-01-01

    Full Text Available La pared celular de levaduras representa entre 20 a 30 % de la célula en peso seco. Está compuesta de polisacáridos complejos de β-glucanos, manoproteínas y quitina. Se estudió la composición de los polisacáridos contenidos en la pared celular de la Saccharomyces cerevisiae CEN.PK 113 y se observó el efecto de la variación de la fuente carbono (glucosa, sacarosa, galactosa, maltosa, manosa, etanol y pH (3, 4, 5, 6 en un medio mineral “cell factory”. Las células fueron recolectadas en fase exponencial y se extrajo la pared celular. Los extractos de pared se hidrolizaron con H2SO4 al 72% y las muestras fueron analizadas por cromatografía HPLC. Se realizó una prueba de resistencia al rompimiento celular con una β(1,3-glucanasa, y las células cultivadas a diferentes fuentes carbono y pH. Los resultados del análisis por HPLC, mostraron que la composición de los polisacáridos en la pared celular, varía considerablemente con las modificaciones del medio de cultivo. Se observó que las levaduras cultivadas en sacarosa tienen mayor porcentaje de pared celular (25% y mayor cantidad de glucanos (115µg/mg peso seco y mananos (131µg/mg peso seco, que aquellas levaduras cultivadas en etanol (13% en peso seco. Las levaduras cultivadas a pH 5 presentaron 19% de pared celular en peso seco, mientras que a pH 6 el porcentaje fue menor (14%. El análisis de resistencia al rompimiento celular, mostró que las células cultivadas en etanol y galactosa fueron resistentes al rompimiento enzimático. Se comparó este resultado con el contenido de polisacáridos en la pared celular y concluimos que la resistencia de la célula al rompimiento, no está ligada con la cantidad de β-glucanos contenidos en la pared celular, sino que va a depender del número de enlaces β(1,3 y β(1,6-glucanos, los cuales juegan un rol importante durante el ensamblaje de la pared

  17. Heterologous expression of Fusarium oxysporum tomatinase in Saccharomyces cerevisiae increases its resistance to saponins and improves ethanol production during the fermentation of Agave tequilana Weber var. azul and Agave salmiana must.

    Cira, Luis Alberto; González, Gloria Angélica; Torres, Juan Carlos; Pelayo, Carlos; Gutiérrez, Melesio; Ramírez, Jesús

    2008-03-01

    This paper describes the effect of the heterologous expression of tomatinase from Fusarium oxysporum f. sp lycopersici in Saccharomyces cerevisiae. The gene FoTom1 under the control of the S. cerevisiae phosphoglycerate kinase (PGK1) promoter was cloned into pYES2. S. cerevisiae strain Y45 was transformed with this vector and URA3 transformant strains were selected for resistance to alpha-tomatine. Two transformants were randomly selected for further study (designated Y45-1 and Y45-2). Control strain Y45 was inhibited at 50 muM alpha-tomatine, in contrast, transformants Y45-1 and Y45-2 did not show inhibition at 200 muM. Tomatinase activity was detected by HPLC monitoring tomatine disappearance and tomatidine appearance in the supernatants of culture medium. Maximum tomatinase activity was observed in the transformants after 6 h, remaining constant during the following 24 h. No tomatinase activity was detected in the parental strain. Moreover, the transformants were able to grow and produce ethanol in a mix of Agave tequilana Weber var. azul and Agave salmiana must, contrary to the Y45 strain which was unable to grow and ferment under these conditions.

  18. Molecular genetic diversity of the Saccharomyces yeasts in Taiwan: Saccharomyces arboricola, Saccharomyces cerevisiae and Saccharomyces kudriavzevii.

    Naumov, Gennadi I; Lee, Ching-Fu; Naumova, Elena S

    2013-01-01

    Genetic hybridization, sequence and karyotypic analyses of natural Saccharomyces yeasts isolated in different regions of Taiwan revealed three biological species: Saccharomyces arboricola, Saccharomyces cerevisiae and Saccharomyces kudriavzevii. Intraspecies variability of the D1/D2 and ITS1 rDNA sequences was detected among S. cerevisiae and S. kudriavzevii isolates. According to molecular and genetic analyses, the cosmopolitan species S. cerevisiae and S. kudriavzevii contain local divergent populations in Taiwan, Malaysia and Japan. Six of the seven known Saccharomyces species are documented in East Asia: S. arboricola, S. bayanus, S. cerevisiae, S. kudriavzevii, S. mikatae, and S. paradoxus.

  19. Kinetics of phosphomevalonate kinase from Saccharomyces cerevisiae.

    David E Garcia

    Full Text Available The mevalonate-based isoprenoid biosynthetic pathway is responsible for producing cholesterol in humans and is used commercially to produce drugs, chemicals, and fuels. Heterologous expression of this pathway in Escherichia coli has enabled high-level production of the antimalarial drug artemisinin and the proposed biofuel bisabolane. Understanding the kinetics of the enzymes in the biosynthetic pathway is critical to optimize the pathway for high flux. We have characterized the kinetic parameters of phosphomevalonate kinase (PMK, EC 2.7.4.2 from Saccharomyces cerevisiae, a previously unstudied enzyme. An E. coli codon-optimized version of the S. cerevisiae gene was cloned into pET-52b+, then the C-terminal 6X His-tagged protein was expressed in E. coli BL21(DE3 and purified on a Ni²⁺ column. The KM of the ATP binding site was determined to be 98.3 µM at 30°C, the optimal growth temperature for S. cerevisiae, and 74.3 µM at 37°C, the optimal growth temperature for E. coli. The K(M of the mevalonate-5-phosphate binding site was determined to be 885 µM at 30°C and 880 µM at 37°C. The V(max was determined to be 4.51 µmol/min/mg enzyme at 30°C and 5.33 µmol/min/mg enzyme at 37°C. PMK is Mg²⁺ dependent, with maximal activity achieved at concentrations of 10 mM or greater. Maximum activity was observed at pH = 7.2. PMK was not found to be substrate inhibited, nor feedback inhibited by FPP at concentrations up to 10 µM FPP.

  20. Acetylation dynamics and stoichiometry in Saccharomyces cerevisiae

    Weinert, Brian Tate; Iesmantavicius, Vytautas; Moustafa, Tarek

    2014-01-01

    Lysine acetylation is a frequently occurring posttranslational modification; however, little is known about the origin and regulation of most sites. Here we used quantitative mass spectrometry to analyze acetylation dynamics and stoichiometry in Saccharomyces cerevisiae. We found that acetylation...

  1. Glucose repression in Saccharomyces cerevisiae.

    Kayikci, Ömur; Nielsen, Jens

    2015-09-01

    Glucose is the primary source of energy for the budding yeast Saccharomyces cerevisiae. Although yeast cells can utilize a wide range of carbon sources, presence of glucose suppresses molecular activities involved in the use of alternate carbon sources as well as it represses respiration and gluconeogenesis. This dominant effect of glucose on yeast carbon metabolism is coordinated by several signaling and metabolic interactions that mainly regulate transcriptional activity but are also effective at post-transcriptional and post-translational levels. This review describes effects of glucose repression on yeast carbon metabolism with a focus on roles of the Snf3/Rgt2 glucose-sensing pathway and Snf1 signal transduction in establishment and relief of glucose repression. © FEMS 2015.

  2. Heterooligomeric phosphoribosyl diphosphate synthase of Saccharomyces cerevisiae

    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...

  3. Fatal Saccharomyces Cerevisiae Aortic Graft Infection

    Meyer, Michael (Technical Monitor); Smith, Davey; Metzgar, David; Wills, Christopher; Fierer, Joshua

    2002-01-01

    Saccharomyces cerevisiae is a yeast commonly used in baking and a frequent colonizer of human mucosal surfaces. It is considered relatively nonpathogenic in immunocompetent adults. We present a case of S. cerevisiae fungemia and aortic graft infection in an immunocompetent adult. This is the first reported case of S. cerevisiue fungemia where the identity of the pathogen was confirmed by rRNA sequencing.

  4. Effects of fermentation by Saccharomyces cerevisiae and ...

    yassine

    2013-02-13

    Feb 13, 2013 ... Full Length Research Paper. Effect of Saccharomyces cerevisiae fermentation on the ... 2003). Besides, several alcoholic beverages such as wine or liqueurs are obtained from fruit juices fermented by Saccharomyces ..... (2003). Kinetics of pigment release from hairy root cultures of Beta vulgaris under the ...

  5. Increasing NADH oxidation reduces overflow metabolism in Saccharomyces cerevisiae

    Vemuri, Goutham; Eiteman, M.A; McEwen, J.E

    2007-01-01

    effect is due to limited respiratory capacity or is caused by glucose-mediated repression of respiration. When respiration in S. cerevisiae was increased by introducing a heterologous alternative oxidase, we observed reduced aerobic ethanol formation. In contrast, increasing nonrespiratory NADH oxidation...... Crabtree effect.’’ The yeast Saccharomyces cerevisiae has served as an important model organism for studying the Crabtree effect. When subjected to increasing glycolytic fluxes under aerobic conditions, there is a threshold value of the glucose uptake rate at which the metabolism shifts from purely...... respiratory to mixed respiratory and fermentative. It is well known that glucose repression of respiratory pathways occurs at high glycolytic fluxes, resulting in a decrease in respiratory capacity. Despite many years of detailed studies on this subject, it is not known whether the onset of the Crabtree...

  6. Fatty acid metabolism in Saccharomyces cerevisiae

    van Roermund, C. W. T.; Waterham, H. R.; IJlst, L.; Wanders, R. J. A.

    2003-01-01

    Peroxisomes are essential subcellular organelles involved in a variety of metabolic processes. Their importance is underlined by the identification of a large group of inherited diseases in humans in which one or more of the peroxisomal functions are impaired. The yeast Saccharomyces cerevisiae has

  7. Ferrofluid modified Saccharomyces cerevisiae cells for biocatalysis

    Šafaříková, Miroslava; Maděrová, Zdeňka; Šafařík, Ivo

    2009-01-01

    Roč. 42, - (2009), s. 521-524 ISSN 0963-9969 R&D Projects: GA MPO 2A-1TP1/094; GA MŠk(CZ) OC 157 Institutional research plan: CEZ:AV0Z60870520 Keywords : Saccharomyces cerevisiae * magnetic fluid * hydrogen peroxide Subject RIV: EI - Biotechnology ; Bionics Impact factor: 2.414, year: 2009

  8. Excision repair and mutagenesis in Saccharomyces cerevisiae

    Kilbey, Brian

    1987-01-01

    This and succeeding letters discuss the James and Kilbey (1977 and 1978) model for the initiation of u.v. mutagenesis in Saccharomyces cerevisiae and its application to include a number of chemical mutagens. The Baranowska et al (1987) results indicating the role of DNA replication, the differing mechanisms in Escherichia coli, are all discussed. (UK)

  9. Nitrogen Catabolite Repression in Saccharomyces cerevisiae

    Hofman-Bang, H Jacob Peider

    1999-01-01

    In Saccharomyces cerevisiae the expression of all known nitrogen catabolite pathways are regulated by four regulators known as Gln3, Gat1, Da180, and Deh1. This is known as nitrogen catabolite repression (NCR). They bind to motifs in the promoter region to the consensus sequence S' GATAA 3'. Gln3...

  10. Characterisation of Saccharomyces cerevisiae hybrids selected for ...

    Wine yeasts (Saccharomyces cerevisiae) vary in their ability to develop the full aroma potential of Sauvignon blanc wine due to an inability to release volatile thiols. Subsequently, the use of 'thiolreleasing' wine yeasts (TRWY) has increased in popularity. However, anecdotal evidence suggests that some commercially ...

  11. Hybridization of Palm Wine Yeasts ( Saccharomyces Cerevisiae ...

    Haploid auxotrophic strains of Saccharomyces cerevisiae were selected from palm wine and propagated by protoplast fusion with Brewers yeast. Fusion resulted in an increase in both ethanol production and tolerance against exogenous ethanol. Mean fusion frequencies obtained for a mating types ranged between 8 x ...

  12. Substrate Channelling and Energetics of Saccharomyces cerevisiae ...

    Data collected during the high-cell-density cultivation of Saccharomyces cerevisiae DSM 2155 on glucose in a simulated five-phase feeding strategy of fed-batch process, executed on the Universal BIoprocess CONtrol (UBICON) system using 150L bioreactor over a period of 24h have been analysed. The consistency of the ...

  13. 21 CFR 866.5785 - Anti-Saccharomyces cerevisiae (S. cerevisiae) antibody (ASCA) test systems.

    2010-04-01

    ...) antibody (ASCA) test systems. 866.5785 Section 866.5785 Food and Drugs FOOD AND DRUG ADMINISTRATION... Immunological Test Systems § 866.5785 Anti-Saccharomyces cerevisiae (S. cerevisiae) antibody (ASCA) test systems. (a) Identification. The Anti-Saccharomyces cerevisiae (S. cerevisiae) antibody (ASCA) test system is...

  14. Saccharomyces cerevisiae metabolism in ecological context

    Jouhten, Paula; Ponomarova, Olga; González García, Ramón; Patil, Kiran R.

    2016-01-01

    The architecture and regulation of Saccharomyces cerevisiae metabolic network are among the best studied owing to its widespread use in both basic research and industry. Yet, several recent studies have revealed notable limitations in explaining genotype?metabolic phenotype relations in this yeast, especially when concerning multiple genetic/environmental perturbations. Apparently unexpected genotype?phenotype relations may originate in the evolutionarily shaped cellular operating principles ...

  15. [Urinary infection by Saccharomyces cerevisiae: Emerging yeast?].

    Elkhihal, B; Elhalimi, M; Ghfir, B; Mostachi, A; Lyagoubi, M; Aoufi, S

    2015-12-01

    Saccharomyces cerevisiae is a commensal yeast of the digestive, respiratory and genito-urinary tract. It is widely used as a probiotic for the treatment of post-antibiotic diarrhea. It most often occurs in immunocompromised patients frequently causing fungemia. We report the case of an adult diabetic patient who had a urinary tract infection due to S. cerevisiae. The disease started with urination associated with urinary frequency burns without fever. The diagnosis was established by the presence of yeasts on direct examination and positivity of culture on Sabouraud-chloramphenicol three times. The auxanogramme gallery (Auxacolor BioRad(®)) allowed the identification of S. cerevisiae. The patient was put on fluconazole with good outcome. This observation points out that this is an opportunistic yeast in immunocompromised patients. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  16. Investigation of autonomous cell cycle oscillation in Saccharomyces cerevisiae

    Hansen, Morten Skov

    2007-01-01

    Autonome Oscillationer i kontinuert kultivering af Saccharomyces cerevisiae Udgangspunktet for dette Ph.d. projekt var at søge at forstå, hvad der gør det muligt at opnå multiple statiske tilstande ved kontinuert kultivering af Saccharomyces cerevisiae med glukose som begrænsende substrat...

  17. Apoptosis - Triggering Effects: UVB-irradiation and Saccharomyces cerevisiae.

    Behzadi, Payam; Behzadi, Elham

    2012-12-01

    The pathogenic disturbance of Saccharomyces cerevisiae is known as a rare but invasive nosocomial fungal infection. This survey is focused on the evaluation of apoptosis-triggering effects of UVB-irradiation in Saccharomyces cerevisiae. The well-growth colonies of Saccharomyces cerevisiae on Sabouraud Dextrose Agar (SDA) were irradiated within an interval of 10 minutes by UVB-light (302 nm). Subsequently, the harvested DNA molecules of control and UV-exposed yeast colonies were run through the 1% agarose gel electrophoresis comprising the luminescent dye of ethidium bromide. No unusual patterns including DNA laddering bands or smears were detected. The applied procedure for UV exposure was not effective for inducing apoptosis in Saccharomyces cerevisiae. So, it needs another UV-radiation protocol for inducing apoptosis phenomenon in Saccharomyces cerevisiae.

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

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

    2010-01-01

    Background: Cost-effective fermentation of lignocellulosic hydrolysate to ethanol by Saccharomyces cerevisiae requires efficient mixed sugar utilization. Notably, the rate and yield of xylose and arabinose co-fermentation to ethanol must be enhanced. Results: Evolutionary engineering was used...... to improve the simultaneous conversion of xylose and arabinose to ethanol in a recombinant industrial Saccharomyces cerevisiae strain carrying the heterologous genes for xylose and arabinose utilization pathways integrated in the genome. The evolved strain TMB3130 displayed an increased consumption rate...... of our knowledge, this is the first report that characterizes the molecular mechanisms for improved mixed-pentose utilization obtained by evolutionary engineering of a recombinant S. cerevisiae strain. Increased transport of pentoses and increased activities of xylose converting enzymes contributed...

  19. 2μ plasmid in Saccharomyces species and in Saccharomyces cerevisiae.

    Strope, Pooja K; Kozmin, Stanislav G; Skelly, Daniel A; Magwene, Paul M; Dietrich, Fred S; McCusker, John H

    2015-12-01

    We determined that extrachromosomal 2μ plasmid was present in 67 of the Saccharomyces cerevisiae 100-genome strains; in addition to variation in the size and copy number of 2μ, we identified three distinct classes of 2μ. We identified 2μ presence/absence and class associations with populations, clinical origin and nuclear genotypes. We also screened genome sequences of S. paradoxus, S. kudriavzevii, S. uvarum, S. eubayanus, S. mikatae, S. arboricolus and S. bayanus strains for both integrated and extrachromosomal 2μ. Similar to S. cerevisiae, we found no integrated 2μ sequences in any S. paradoxus strains. However, we identified part of 2μ integrated into the genomes of some S. uvarum, S. kudriavzevii, S. mikatae and S. bayanus strains, which were distinct from each other and from all extrachromosomal 2μ. We identified extrachromosomal 2μ in one S. paradoxus, one S. eubayanus, two S. bayanus and 13 S. uvarum strains. The extrachromosomal 2μ in S. paradoxus, S. eubayanus and S. cerevisiae were distinct from each other. In contrast, the extrachromosomal 2μ in S. bayanus and S. uvarum strains were identical with each other and with one of the three classes of S. cerevisiae 2μ, consistent with interspecific transfer. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Sporulation in the Budding Yeast Saccharomyces cerevisiae

    Neiman, Aaron M.

    2011-01-01

    In response to nitrogen starvation in the presence of a poor carbon source, diploid cells of the yeast Saccharomyces cerevisiae undergo meiosis and package the haploid nuclei produced in meiosis into spores. The formation of spores requires an unusual cell division event in which daughter cells are formed within the cytoplasm of the mother cell. This process involves the de novo generation of two different cellular structures: novel membrane compartments within the cell cytoplasm that give rise to the spore plasma membrane and an extensive spore wall that protects the spore from environmental insults. This article summarizes what is known about the molecular mechanisms controlling spore assembly with particular attention to how constitutive cellular functions are modified to create novel behaviors during this developmental process. Key regulatory points on the sporulation pathway are also discussed as well as the possible role of sporulation in the natural ecology of S. cerevisiae. PMID:22084423

  1. Mobilomics in Saccharomyces cerevisiae strains.

    Menconi, Giulia; Battaglia, Giovanni; Grossi, Roberto; Pisanti, Nadia; Marangoni, Roberto

    2013-03-20

    Mobile Genetic Elements (MGEs) are selfish DNA integrated in the genomes. Their detection is mainly based on consensus-like searches by scanning the investigated genome against the sequence of an already identified MGE. Mobilomics aims at discovering all the MGEs in a genome and understanding their dynamic behavior: The data for this kind of investigation can be provided by comparative genomics of closely related organisms. The amount of data thus involved requires a strong computational effort, which should be alleviated. Our approach proposes to exploit the high similarity among homologous chromosomes of different strains of the same species, following a progressive comparative genomics philosophy. We introduce a software tool based on our new fast algorithm, called regender, which is able to identify the conserved regions between chromosomes. Our case study is represented by a unique recently available dataset of 39 different strains of S.cerevisiae, which regender is able to compare in few minutes. By exploring the non-conserved regions, where MGEs are mainly retrotransposons called Tys, and marking the candidate Tys based on their length, we are able to locate a priori and automatically all the already known Tys and map all the putative Tys in all the strains. The remaining putative mobile elements (PMEs) emerging from this intra-specific comparison are sharp markers of inter-specific evolution: indeed, many events of non-conservation among different yeast strains correspond to PMEs. A clustering based on the presence/absence of the candidate Tys in the strains suggests an evolutionary interconnection that is very similar to classic phylogenetic trees based on SNPs analysis, even though it is computed without using phylogenetic information. The case study indicates that the proposed methodology brings two major advantages: (a) it does not require any template sequence for the wanted MGEs and (b) it can be applied to infer MGEs also for low coverage genomes

  2. Mobilomics in Saccharomyces cerevisiae strains

    2013-01-01

    Background Mobile Genetic Elements (MGEs) are selfish DNA integrated in the genomes. Their detection is mainly based on consensus–like searches by scanning the investigated genome against the sequence of an already identified MGE. Mobilomics aims at discovering all the MGEs in a genome and understanding their dynamic behavior: The data for this kind of investigation can be provided by comparative genomics of closely related organisms. The amount of data thus involved requires a strong computational effort, which should be alleviated. Results Our approach proposes to exploit the high similarity among homologous chromosomes of different strains of the same species, following a progressive comparative genomics philosophy. We introduce a software tool based on our new fast algorithm, called regender, which is able to identify the conserved regions between chromosomes. Our case study is represented by a unique recently available dataset of 39 different strains of S.cerevisiae, which regender is able to compare in few minutes. By exploring the non–conserved regions, where MGEs are mainly retrotransposons called Tys, and marking the candidate Tys based on their length, we are able to locate a priori and automatically all the already known Tys and map all the putative Tys in all the strains. The remaining putative mobile elements (PMEs) emerging from this intra–specific comparison are sharp markers of inter–specific evolution: indeed, many events of non–conservation among different yeast strains correspond to PMEs. A clustering based on the presence/absence of the candidate Tys in the strains suggests an evolutionary interconnection that is very similar to classic phylogenetic trees based on SNPs analysis, even though it is computed without using phylogenetic information. Conclusions The case study indicates that the proposed methodology brings two major advantages: (a) it does not require any template sequence for the wanted MGEs and (b) it can be applied to

  3. Heterologous Expression of the Carrot Hsp17.7 gene Increased Growth, Cell Viability, and Protein Solubility in Transformed Yeast (Saccharomyces cerevisiae) under Heat, Cold, Acid, and Osmotic Stress Conditions.

    Ko, Eunhye; Kim, Minhye; Park, Yunho; Ahn, Yeh-Jin

    2017-08-01

    In industrial fermentation of yeast (Saccharomyces cerevisiae), culture conditions are often modified from the optimal growth conditions of the cells to maintain large-scale cultures and/or to increase recombinant protein production. However, altered growth conditions can be stressful to yeast cells resulting in reduced cell growth and viability. In this study, a small heat shock protein gene from carrot (Daucus carota L.), Hsp17.7, was inserted into the yeast genome via homologous recombination to increase tolerance to stress conditions that can occur during industrial culture. A DNA construct, Translational elongation factor gene promoter-carrot Hsp17.7 gene-Phosphoribosyl-anthranilate isomerase gene (an auxotrophic marker), was generated by a series of PCRs and introduced into the chromosome IV of the yeast genome. Immunoblot analysis showed that carrot Hsp17.7 accumulated in the transformed yeast cell lines. Growth rates and cell viability of these cell lines were higher than control cell lines under heat, cold, acid, and hyperosmotic stress conditions. Soluble protein levels were higher in the transgenic cell lines than control cell lines under heat and cold conditions, suggesting the molecular chaperone function of the recombinant Hsp17.7. This study showed that a recombinant DNA construct containing a HSP gene from carrot was successfully expressed in yeast by homologous recombination and increased tolerances to abiotic stress conditions.

  4. Enhanced isoprenoid production from xylose by engineered Saccharomyces cerevisiae.

    Kwak, Suryang; Kim, Soo Rin; Xu, Haiqing; Zhang, Guo-Chang; Lane, Stephan; Kim, Heejin; Jin, Yong-Su

    2017-11-01

    Saccharomyces cerevisiae has limited capabilities for producing fuels and chemicals derived from acetyl-CoA, such as isoprenoids, due to a rigid flux partition toward ethanol during glucose metabolism. Despite numerous efforts, xylose fermentation by engineered yeast harboring heterologous xylose metabolic pathways was not as efficient as glucose fermentation for producing ethanol. Therefore, we hypothesized that xylose metabolism by engineered yeast might be a better fit for producing non-ethanol metabolites. We indeed found that engineered S. cerevisiae on xylose showed higher expression levels of the enzymes involved in ethanol assimilation and cytosolic acetyl-CoA synthesis than on glucose. When genetic perturbations necessary for overproducing squalene and amorphadiene were introduced into engineered S. cerevisiae capable of fermenting xylose, we observed higher titers and yields of isoprenoids under xylose than glucose conditions. Specifically, co-overexpression of a truncated HMG1 (tHMG1) and ERG10 led to substantially higher squalene accumulation under xylose than glucose conditions. In contrast to glucose utilization producing massive amounts of ethanol regardless of aeration, xylose utilization allowed much less amounts of ethanol accumulation, indicating ethanol is simultaneously re-assimilated with xylose consumption and utilized for the biosynthesis of cytosolic acetyl-CoA. In addition, xylose utilization by engineered yeast with overexpression of tHMG1, ERG10, and ADS coding for amorphadiene synthase, and the down-regulation of ERG9 resulted in enhanced amorphadiene production as compared to glucose utilization. These results suggest that the problem of the rigid flux partition toward ethanol production in yeast during the production of isoprenoids and other acetyl-CoA derived chemicals can be bypassed by using xylose instead of glucose as a carbon source. Biotechnol. Bioeng. 2017;114: 2581-2591. © 2017 Wiley Periodicals, Inc. © 2017 Wiley

  5. Organization of Replication of Ribosomal DNA in Saccharomyces cerevisiae

    Linskens, Maarten H.K.; Huberman, Joel A.

    1988-01-01

    Using recently developed replicon mapping techniques, we have analyzed the replication of the ribosomal DNA in Saccharomyces cerevisiae. The results show that (i) the functional origin of replication colocalizes with an autonomously replicating sequence element previously mapped to the

  6. Genome-scale reconstruction of the Saccharomyces cerevisiae metabolic network

    Förster, Jochen; Famili, I.; Fu, P.

    2003-01-01

    The metabolic network in the yeast Saccharomyces cerevisiae was reconstructed using currently available genomic, biochemical, and physiological information. The metabolic reactions were compartmentalized between the cytosol and the mitochondria, and transport steps between the compartments...

  7. Effect of Saccharomyces cerevisiae fermentation on the colorants of ...

    Effect of Saccharomyces cerevisiae fermentation on the colorants of heated red beetroot extracts. Hayet Ben Haj Koubaier, Ismahen Essaidi, Ahmed Snoussi, Slim Zgoulli, Mohamed Moncef Chaabouni, Phillipe Thonart, Nabiha Bouzouita ...

  8. Molecular Basis for Saccharomyces cerevisiae Biofilm Development

    Andersen, Kaj Scherz

    In this study, I sought to identify genes regulating the global molecular program for development of sessile multicellular communities, also known as biofilm, of the eukaryotic microorganism, Saccharomyces cerevisiae (yeast). Yeast biofilm has a clinical interest, as biofilms can cause chronic...... infections in humans. Biofilm is also interesting from an evolutionary standpoint, as an example of primitive multicellularity. By using a genome-wide screen of yeast deletion mutants, I show that 71 genes are essential for biofilm formation. Two-thirds of these genes are required for transcription of FLO11......, but only a small subset is previously described as regulators of FLO11. These results reveal that the regulation of biofilm formation and FLO11 is even more complex than what has previously been described. I find that the molecular program for biofilm formation shares many essential components with two...

  9. Modification of mutation frequency in Saccharomyces Cerevisiae

    Vashishat, R.K.; Kakar, S.N.

    1976-01-01

    In a reverse mutation system, using haploid, histidine-requirinq strain of Saccharomyces cerevisiae, the frequency of uv-induced prototrophs increased if the post-irradiation minimal medium was supplemented with limited amounts of histidine. Addition of natural amino acids or RNA bases in the post-irradiation minimal medium, with or without histidine, also increased the uv-induced mutation frequency. Thus, post-irradiation conditions favouring protein and RNA synthesis, are effective in increasing uv-induced mutations in yeast. As compared to uv light, nitrous acid was more effective in inducing reversions in this strain and the frequency increased if the treated cells were plated on minimal medium supplemented with limited amounts of histidine. However, the addition of amino acids or RNA bases decreased the number of revertants. An additional inclusion of histidine reversed the suppressive effect of these metabolites. The mutation induction processes are thus different or differently modifiable in uv and nitrous acid. (author)

  10. Study on biosorption of uranium by alginate immobilized saccharomyces cerevisiae

    Wang Baoe; Xu Weichang; Xie Shuibo; Guo Yangbin

    2005-01-01

    Saccharomyces cerevisiae has great capability of biosorption of uranium. The maxium uptake is 172.4 mg/g according to this study. To adapt to the application of the biomass in the field, the biosorption of uranium by cross-linked and alginate calcium immobilized Saccharomyces cerevisiae is studied. Results indicate the maxium uptake is 185.2 mg/g by formaldehyde cross-linked biomass, and it is 769.2 mg/g by alginate calcium immobilized biomass. (authors)

  11. Saccharomyces cerevisiae var. boulardii fungemia following probiotic treatment

    Appel-da-Silva, Marcelo C.; Narvaez, Gabriel A.; Perez, Leandro R.R.; Drehmer, Laura; Lewgoy, Jairo

    2017-01-01

    Probiotics are commonly prescribed as an adjuvant in the treatment of antibiotic-associated diarrhea caused by Clostridium difficile. We report the case of an immunocompromised 73-year-old patient on chemotherapy who developed Saccharomyces cerevisiae var. boulardii fungemia in a central venous catheter during treatment of antibiotic-associated pseudomembranous colitis with the probiotic Saccharomyces cerevisiae var. boulardii. Fungemia was resolved after interruption of probiotic administrat...

  12. Response of Saccharomyces cerevisiae to cadmium stress

    Moreira, Luciana Mara Costa; Ribeiro, Frederico Haddad; Neves, Maria Jose; Porto, Barbara Abranches Araujo; Amaral, Angela M.; Menezes, Maria Angela B.C.; Rosa, Carlos Augusto

    2009-01-01

    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 + and Na + ) 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. Response of Saccharomyces cerevisiae to cadmium stress

    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)

  14. Stoichiometric network constraints on xylose metabolism by recombinant Saccharomyces cerevisiae

    Yong-Su Jin; Thomas W. Jeffries

    2004-01-01

    Metabolic pathway engineering is constrained by the thermodynamic and stoichiometric feasibility of enzymatic activities of introduced genes. Engineering of xylose metabolism in Saccharomyces cerevisiae has focused on introducing genes for the initial xylose assimilation steps from Pichia stipitis, a xylose-fermenting yeast, into S. cerevisiae, a yeast raditionally...

  15. Saccharomyces cerevisiae var. boulardii fungemia following probiotic treatment

    Marcelo C. Appel-da-Silva

    2017-12-01

    Full Text Available Probiotics are commonly prescribed as an adjuvant in the treatment of antibiotic-associated diarrhea caused by Clostridium difficile. We report the case of an immunocompromised 73-year-old patient on chemotherapy who developed Saccharomyces cerevisiae var. boulardii fungemia in a central venous catheter during treatment of antibiotic-associated pseudomembranous colitis with the probiotic Saccharomyces cerevisiae var. boulardii. Fungemia was resolved after interruption of probiotic administration without the need to replace the central venous line. Keywords: Saccharomyces, Probiotics, Fungemia, Critical illness, Clostridium difficile

  16. Pathways of ultraviolet mutability in Saccharomyces cerevisiae

    Lemontt, J.F.

    1977-01-01

    Non-allelic mutants of Saccharomyces cerevisiae with reduced capacity for ultraviolet light (UV)-induced forward mutation from CAN1 to can1 were assigned to seven distinct genetic loci, each with allele designations umr1-1, umr2-1, ..., umr7-1 to indicate UV mutation resistance. None conferred a great deal of UV sensitivity. When assayed on yeast extract-peptone-dextrose complex growth agar, umr1, umr3, and umr7 were the most UV-sensitive. When assayed on synthetic agar lacking arginine, however, umr3 was the most UV-sensitive. All strains carrying each of the seven umr genes exhibited varying degrees of defective UV mutability, compact with wild types. Normal UV revertibility of three different alleles was observed in strains carrying either umr4, umr5, umr6, or umr7. Five a/α homozygous umr diploids failed to sporulate. One of these, umr7, blocked normal secretion of alpha hormone in α segregants and could not conjugate with a strains. The phenotypes of umr mutants are consistent with the existence of branched UV mutation pathways of different specificity

  17. Sugar and Glycerol Transport in Saccharomyces cerevisiae.

    Bisson, Linda F; Fan, Qingwen; Walker, Gordon A

    2016-01-01

    In Saccharomyces cerevisiae the process of transport of sugar substrates into the cell comprises a complex network of transporters and interacting regulatory mechanisms. Members of the large family of hexose (HXT) transporters display uptake efficiencies consistent with their environmental expression and play physiological roles in addition to feeding the glycolytic pathway. Multiple glucose-inducing and glucose-independent mechanisms serve to regulate expression of the sugar transporters in yeast assuring that expression levels and transporter activity are coordinated with cellular metabolism and energy needs. The expression of sugar transport activity is modulated by other nutritional and environmental factors that may override glucose-generated signals. Transporter expression and activity is regulated transcriptionally, post-transcriptionally and post-translationally. Recent studies have expanded upon this suite of regulatory mechanisms to include transcriptional expression fine tuning mediated by antisense RNA and prion-based regulation of transcription. Much remains to be learned about cell biology from the continued analysis of this dynamic process of substrate acquisition.

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

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

    2005-01-01

    .6-16.8%) recorded for two isolates from blue veined cheeses. Merely 25% of the S. cerevisiae var. boulardii strains displayed good adhesive properties (16.2-28.0%). The expression of the proinflammatory cytokine IL-1α decreased strikingly in IPEC-J2 cells exposed to a Shiga-like toxin 2e producing Escherichia coli...... strain when the cells were pre- and coincubated with S. cerevisiae var. boulardii even though this yeast strain was low adhesive (5.4%), suggesting that adhesion is not a mandatory prerequisite for such a probiotic effect. A strain of S. cerevisiae isolated from West African sorghum beer exerted similar......The probiotic potential of IS Saccharomyces cerevisiae strains used for production of foods or bevel-ages or isolated from such, and eight strains of Saccharomyces cerevisiae var. boulardii, was investigated. All strains included were able to withstand pH 2.5 and 0.3% Ox-all. Adhesion...

  19. Effects of gene orientation and use of multiple promoters on the expression of XYL1 and XYL2 in Saccharomyces cerevisiae

    Ju Yun Bae; Jose Laplaza; Thomas W. Jeffries

    2008-01-01

    Orientation of adjacent genes has been reported to affect their expression in eukaryotic systems, and metabolic engineering also often makes repeated use of a few promoters to obtain high expression. To improve transcriptional control in heterologous expression, we examined how these factors affect gene expression and enzymatic activity in Saccharomyces cerevisiae. We...

  20. An apoptotic cell cycle mutant in Saccharomyces cerevisiae

    Villadsen, Ingrid

    1996-01-01

    The simple eukaryote Saccharomyces cerevisiae has proved to be a useful organism for elucidating the mechanisms that govern cell cycle progression in eukaryotic cells. The excellent in vivo system permits a cell cycle study using temperature sensitive mutants. In addition, it is possible to study...... many genes and gene products from higher eukaryotes in Saccharomyces cerevisiae because many genes and biological processes are homologous or similar in lower and in higher eukaryotes. The highly developed methods of genetics and molecular biology greatly facilitates studies of higher eukaryotic...... processes.Programmmed cell death with apoptosis plays a major role in development and homeostatis in most, if not all, animal cells. Apoptosis is a morphologically distinct form of death, that requires the activation of a highly regulated suicide program. Saccharomyces cerevisiae provides a new system...

  1. Accumulation of gold using Baker's yeast, Saccharomyces cerevisiae

    Roy, Kamalika; Lahiri, Susanta; Sinha, P.

    2006-01-01

    Authors have reported preconcentration of 152 Eu, 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

  2. Regulation of Cation Balance in Saccharomyces cerevisiae

    Cyert, Martha S.; Philpott, Caroline C.

    2013-01-01

    All living organisms require nutrient minerals for growth and have developed mechanisms to acquire, utilize, and store nutrient minerals effectively. In the aqueous cellular environment, these elements exist as charged ions that, together with protons and hydroxide ions, facilitate biochemical reactions and establish the electrochemical gradients across membranes that drive cellular processes such as transport and ATP synthesis. Metal ions serve as essential enzyme cofactors and perform both structural and signaling roles within cells. However, because these ions can also be toxic, cells have developed sophisticated homeostatic mechanisms to regulate their levels and avoid toxicity. Studies in Saccharomyces cerevisiae have characterized many of the gene products and processes responsible for acquiring, utilizing, storing, and regulating levels of these ions. Findings in this model organism have often allowed the corresponding machinery in humans to be identified and have provided insights into diseases that result from defects in ion homeostasis. This review summarizes our current understanding of how cation balance is achieved and modulated in baker’s yeast. Control of intracellular pH is discussed, as well as uptake, storage, and efflux mechanisms for the alkali metal cations, Na+ and K+, the divalent cations, Ca2+ and Mg2+, and the trace metal ions, Fe2+, Zn2+, Cu2+, and Mn2+. Signal transduction pathways that are regulated by pH and Ca2+ are reviewed, as well as the mechanisms that allow cells to maintain appropriate intracellular cation concentrations when challenged by extreme conditions, i.e., either limited availability or toxic levels in the environment. PMID:23463800

  3. Microbially induced separation of quartz from calcite using Saccharomyces cerevisiae.

    Padukone, S Usha; Natarajan, K A

    2011-11-01

    Cells of Saccharomyces cerevisiae and their metabolites were successfully utilized to achieve selective separation of quartz and calcite through microbially induced flotation and flocculation. S. cerevisiae was adapted to calcite and quartz minerals. Adsorption studies and electrokinetic investigations were carried out to understand the changes in the surface chemistry of yeast cells and the minerals after mutual interaction. Possible mechanisms in microbially induced flotation and flocculation are outlined. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Improvement of Lead Tolerance of Saccharomyces cerevisiae by Random Mutagenesis of Transcription Regulator SPT3.

    Zhu, Liying; Gao, Shan; Zhang, Hongman; Huang, He; Jiang, Ling

    2018-01-01

    Bioremediation of heavy metal pollution with biomaterials such as bacteria and fungi usually suffer from limitations because of microbial sensitivity to high concentration of heavy metals. Herein, we adopted a novel random mutagenesis technique called RAISE to manipulate the transcription regulator SPT3 of Saccharomyces cerevisiae to improve cell lead tolerance. The best strain Mutant VI was selected from the random mutagenesis libraries on account of the growth performance, with higher specific growth rate than the control strain (0.068 vs. 0.040 h -1 ) at lead concentration as high as 1.8 g/L. Combined with the transcriptome analysis of S. cerevisiae, expressing the SPT3 protein was performed to make better sense of the global regulatory effects of SPT3. The data analysis revealed that 57 of S. cerevisiae genes were induced and 113 genes were suppressed, ranging from those for trehalose synthesis, carbon metabolism, and nucleotide synthesis to lead resistance. Especially, the accumulation of intracellular trehalose in S. cerevisiae under certain conditions of stress is considered important to lead resistance. The above results represented that SPT3 was acted as global transcription regulator in the exponential phase of strain and accordingly improved heavy metal tolerance in the heterologous host S. cerevisiae. The present study provides a route to complex phenotypes that are not readily accessible by traditional methods.

  5. Rationally designed, heterologous S. cerevisiae transcripts expose novel expression determinants

    Ben-Yehezkel, Tuval; Atar, Shimshi; Zur, Hadas; Diament, Alon; Goz, Eli; Marx, Tzipy; Cohen, Rafael; Dana, Alexandra; Feldman, Anna; Shapiro, Ehud; Tuller, Tamir

    2015-01-01

    Deducing generic causal relations between RNA transcript features and protein expression profiles from endogenous gene expression data remains a major unsolved problem in biology. The analysis of gene expression from heterologous genes contributes significantly to solving this problem, but has been heavily biased toward the study of the effect of 5′ transcript regions and to prokaryotes. Here, we employ a synthetic biology driven approach that systematically differentiates the effect of different regions of the transcript on gene expression up to 240 nucleotides into the ORF. This enabled us to discover new causal effects between features in previously unexplored regions of transcripts, and gene expression in natural regimes. We rationally designed, constructed, and analyzed 383 gene variants of the viral HRSVgp04 gene ORF, with multiple synonymous mutations at key positions along the transcript in the eukaryote S. cerevisiae. Our results show that a few silent mutations at the 5′UTR can have a dramatic effect of up to 15 fold change on protein levels, and that even synonymous mutations in positions more than 120 nucleotides downstream from the ORF 5′end can modulate protein levels up to 160%–300%. We demonstrate that the correlation between protein levels and folding energy increases with the significance of the level of selection of the latter in endogenous genes, reinforcing the notion that selection for folding strength in different parts of the ORF is related to translation regulation. Our measured protein abundance correlates notably(correlation up to r = 0.62 (p=0.0013)) with mean relative codon decoding times, based on ribosomal densities (Ribo-Seq) in endogenous genes, supporting the conjecture that translation elongation and adaptation to the tRNA pool can modify protein levels in a causal/direct manner. This report provides an improved understanding of transcript evolution, design principles of gene expression regulation, and suggests simple

  6. Capturing of the monoterpene olefin limonene produced in Saccharomyces cerevisiae

    Jongedijk, E.J.; Cankar, K.; Ranzijn, J.; Krol, van der A.R.; Bouwmeester, H.J.; Beekwilder, M.J.

    2015-01-01

    Monoterpene olefins such as limonene are plant compounds with applications as flavouring and fragrance agents, as solvents and potentially also in polymer and fuel chemistry. We engineered baker's yeast Saccharomyces cerevisiae to express a (-)-limonene synthase from Perilla frutescens and a

  7. Reducing the genetic complexity of glycolysis in Saccharomyces cerevisiae

    Solis Escalante, D.

    2015-01-01

    Glycolysis, a biochemical pathway that oxidizes glucose to pyruvate, is at the core of sugar metabolism in Saccharomyces cerevisiae (bakers’ yeast). Glycolysis is not only a catabolic route involved in energy conservation, but also provides building blocks for anabolism. From an applied perspective,

  8. Silver Uptake and Reuse of Biomass by Saccharomyces cerevisiae ...

    Studies were carried out on the recovery of bound silver and reuse of Chlorella emersonii and Saccharomyces cerevisiae biomass for further silver uptake after they were placed in contact with 20mg/l silver for 30 minutes to allow for maximum binding. It was found that 0.16M nitric acid gave the best recovery rates of silver.

  9. Production of Saccharomyces cerevisiae biomass in papaya extract ...

    Extracts of papaya fruit were used as substrate for single cell protein (SCP) production using Saccharomyces cerevisiae. A 500 g of papaya fruit was extracted with different volumes of sterile distilled water. Extraction with 200 mL of sterile distilled water sustained highest cell growth. Biochemical analysis of dry biomass ...

  10. Novel feeding strategies for Saccharomyces cerevisiae DS2155 ...

    The dual behavior of Saccharomyces cerevisiae on glucose feed as function of the dilution rate near the critical specific growth rate (ì=0.25) is a bottleneck in industrial production, hence the need for more efficient feeding strategies. In this work novel feeding strategies have been generated and evaluated. For each feeding ...

  11. Adsorption and Interfacial Electron Transfer of Saccharomyces Cerevisiae

    Hansen, Allan Glargaard; Boisen, Anja; Nielsen, Jens Ulrik

    2003-01-01

    We have studied the adsorption and electron-transfer dynamics of Saccharomyces cerevisiae (yeast) iso-l-cytochrome c adsorbed on Au(lll) electrodes in aqueous phosphate buffer media. This cytochrome possesses a thiol group dos e to the protein surface (Cysl02) suitable for linking the protein...

  12. Genetic Basis for Saccharomyces cerevisiae Biofilm in Liquid Medium

    Andersen, Kaj Scherz; Bojsen, Rasmus Kenneth; Gro Rejkjær Sørensen, Laura

    2014-01-01

    than free-living cells. We investigated the genetic basis for yeast, Saccharomyces cerevisiae, biofilm on solid surfaces in liquid medium by screening a comprehensive deletion mutant collection in the S1278b background and found 71 genes that were essential for biofilm development. Quantitative...

  13. Evidence against a photoprotective component of photoreactivation in Saccharomyces cerevisiae

    MacQuillan, A.M.; Green, G.; Perry, W.G.

    1981-01-01

    Photoreactivation-deficient (phr - ) mutants of Saccharomyces cerevisiae were shown to lack in vitro DNA-photolyase activity. A phr - mutant was then compared with a phr + strain for near-UV induced photoprotection from far-UV irradiation. Neither strain exhibited a photoprotective effect. (author)

  14. Kinetics of formation of induced mutants of Saccharomyces cerevisiae

    Chepurnoj, A.I.; Levkovich, N.V.; Mikhova-Tsenova, N.; Mel'nikova, L.A.

    1990-01-01

    UV and γ-radiation mutagenic effect an various strains of Saccharomyces cerevisiae was studied by analyzing formation kinetics of induced mutants at the period of postirradiation incubation. Mechanisms of induced reverse formation was suggested. The presented analysis is considered to be differential taking account of more subtle aspects of induced mutagenesis. 8 refs.; 10 figs.; 3 tabs

  15. Reconstitution of an efficient thymidine salvage pathway in Saccharomyces cerevisiae

    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. Analysis of the RNA Content of the Yeast "Saccharomyces Cerevisiae"

    Deutch, Charles E.; Marshall, Pamela A.

    2008-01-01

    In this article, the authors describe an interconnected set of relatively simple laboratory experiments in which students determine the RNA content of yeast cells and use agarose gel electrophoresis to separate and analyze the major species of cellular RNA. This set of experiments focuses on RNAs from the yeast "Saccharomyces cerevisiae", a…

  17. Mead features fermented by Saccharomyces cerevisiae (lalvin k1 ...

    Eduardo Morales

    Full Length Research Paper. Mead features fermented by Saccharomyces cerevisiae. (lalvin k1-1116). Eduardo Marin MORALES1*, Valmir Eduardo ALCARDE2 and Dejanira de Franceschi de. ANGELIS1. 1Department of Biochemistry and Microbiology, Institute of Biosciences, UNESP - Univ Estadual Paulista, Av. 24-A,.

  18. Potential application of Saccharomyces cerevisiae strains for the ...

    This paper aimed at evaluating the fermentation behavior of selected Saccharomyces cerevisiae strains in banana pulp and they were compared with commercial yeast (baker's yeast) for subsequent production of distilled spirits. Five types of microorganisms were used: Four yeast strains obtained from accredited ...

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

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

    2012-01-01

    Industrial biotechnology aims to develop robust microbial cell factories, such as Saccharomyces cerevisiae, to produce an array of added value chemicals presently dominated by petrochemical processes. Xylose is the second most abundant monosaccharide after glucose and the most prevalent pentose s...

  20. Effects of dietary L-threonine and Saccharomyces cerevisiae on ...

    threonine (0, 2.5, 5 and 7.5 g/kg) with or without Saccharomyces cerevisiae (SC) on performance, carcass characteristics, intestinal morphology and immune system of broiler chickens. A total of 360 1-d-old male broiler chicks were randomly ...

  1. Switching the mode of sucrose utilization by Saccharomyces cerevisiae

    Badotti, Fernanda; Dário, Marcelo G; Alves, Sergio L; Cordioli, Maria Luiza A; Miletti, Luiz C; de Araujo, Pedro S; Stambuk, Boris U

    2008-01-01

    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 sucro...

  2. The Plasma Membrane of Saccharomyces cerevisiae : Structure, Function, and Biogenesis

    VANDERREST, ME; KAMMINGA, AH; NAKANO, A; ANRAKU, Y; POOLMAN, B; KONINGS, WN

    The composition of phospholipids, sphingolipids, and sterols in the plasma membrane has a strong influence on the activity of the proteins associated or embedded in the lipid bilayer. Since most lipid-synthesizing enzymes in Saccharomyces cerevisiae are located in intracellular organelles, an

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

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

    2005-05-01

    The probiotic potential of 18 Saccharomyces cerevisiae strains used for production of foods or beverages or isolated from such, and eight strains of Saccharomyces cerevisiae var. boulardii, was investigated. All strains included were able to withstand pH 2.5 and 0.3% Oxgall. Adhesion to the nontumorigenic porcine jejunal epithelial cell line (IPEC-J2) was investigated by incorporation of 3H-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 (13.6-16.8%) recorded for two isolates from blue veined cheeses. Merely 25% of the S. cerevisiae var. boulardii strains displayed good adhesive properties (16.2-28.0%). The expression of the proinflammatory cytokine IL-1alpha decreased strikingly in IPEC-J2 cells exposed to a Shiga-like toxin 2e producing Escherichia coli strain when the cells were pre- and coincubated with S. cerevisiae var. boulardii even though this yeast strain was low adhesive (5.4%), suggesting that adhesion is not a mandatory prerequisite for such a probiotic effect. A strain of S. cerevisiae isolated from West African sorghum beer exerted similar effects hence indicating that food-borne strains of S. cerevisiae may possess probiotic properties in spite of low adhesiveness.

  4. Lycopene overproduction in Saccharomyces cerevisiae through combining pathway engineering with host engineering.

    Chen, Yan; Xiao, Wenhai; Wang, Ying; Liu, Hong; Li, Xia; Yuan, Yingjin

    2016-06-21

    Microbial production of lycopene, a commercially and medically important compound, has received increasing concern in recent years. Saccharomyces cerevisiae is regarded as a safer host for lycopene production than Escherichia coli. However, to date, the lycopene yield (mg/g DCW) in S. cerevisiae was lower than that in E. coli and did not facilitate downstream extraction process, which might be attributed to the incompatibility between host cell and heterologous pathway. Therefore, to achieve lycopene overproduction in S. cerevisiae, both host cell and heterologous pathway should be delicately engineered. In this study, lycopene biosynthesis pathway was constructed by integration of CrtE, CrtB and CrtI in S. cerevisiae CEN.PK2. When YPL062W, a distant genetic locus, was deleted, little acetate was accumulated and approximately 100 % increase in cytosolic acetyl-CoA pool was achieved relative to that in parental strain. Through screening CrtE, CrtB and CrtI from diverse species, an optimal carotenogenic enzyme combination was obtained, and CrtI from Blakeslea trispora (BtCrtI) was found to have excellent performance on lycopene production as well as lycopene proportion in carotenoid. Then, the expression level of BtCrtI was fine-tuned and the effect of cell mating types was also evaluated. Finally, potential distant genetic targets (YJL064W, ROX1, and DOS2) were deleted and a stress-responsive transcription factor INO2 was also up-regulated. Through the above modifications between host cell and carotenogenic pathway, lycopene yield was increased by approximately 22-fold (from 2.43 to 54.63 mg/g DCW). Eventually, in fed-batch fermentation, lycopene production reached 55.56 mg/g DCW, which is the highest reported yield in yeasts. Saccharomyces cerevisiae was engineered to produce lycopene in this study. Through combining host engineering (distant genetic loci and cell mating types) with pathway engineering (enzyme screening and gene fine-tuning), lycopene yield was

  5. Signature pathway expression of xylose utilization in the genetically engineered industrial yeast Saccharomyces cerevisiae

    Background: The limited xylose utilizing ability of native Saccharomyces cerevisiae has been a major obstacle for efficient cellulosic ethanol production from lignocellulosic materials. Haploid laboratory strains of S. cerevisiae are commonly used for genetic engineering to enable its xylose utiliza...

  6. Purification of Arp2/3 complex from Saccharomyces cerevisiae

    Doolittle, Lynda K.; Rosen, Michael K.; Padrick, Shae B.

    2014-01-01

    Summary Much of cellular control over actin dynamics comes through regulation of actin filament initiation. At the molecular level, this is accomplished through a collection of cellular protein machines, called actin nucleation factors, which position actin monomers to initiate a new actin filament. The Arp2/3 complex is a principal actin nucleation factor used throughout the eukaryotic family tree. The budding yeast Saccharomyces cerevisiae has proven to be not only an excellent genetic platform for the study of the Arp2/3 complex, but also an excellent source for the purification of endogenous Arp2/3 complex. Here we describe a protocol for the preparation of endogenous Arp2/3 complex from wild type Saccharomyces cerevisiae. This protocol produces material suitable for biochemical study, and yields milligram quantities of purified Arp2/3 complex. PMID:23868593

  7. Saccharomyces cerevisiae: a sexy yeast with a prion problem.

    Kelly, Amy C; Wickner, Reed B

    2013-01-01

    Yeast prions are infectious proteins that spread exclusively by mating. The frequency of prions in the wild therefore largely reflects the rate of spread by mating counterbalanced by prion growth slowing effects in the host. We recently showed that the frequency of outcross mating is about 1% of mitotic doublings with 23-46% of total matings being outcrosses. These findings imply that even the mildest forms of the [PSI+], [URE3] and [PIN+] prions impart > 1% growth/survival detriment on their hosts. Our estimate of outcrossing suggests that Saccharomyces cerevisiae is far more sexual than previously thought and would therefore be more responsive to the adaptive effects of natural selection compared with a strictly asexual yeast. Further, given its large effective population size, a growth/survival detriment of > 1% for yeast prions should strongly select against prion-infected strains in wild populations of Saccharomyces cerevisiae.

  8. Efficient screening of environmental isolates for Saccharomyces cerevisiae strains that are suitable for brewing.

    Fujihara, Hidehiko; Hino, Mika; Takashita, Hideharu; Kajiwara, Yasuhiro; Okamoto, Keiko; Furukawa, Kensuke

    2014-01-01

    We developed an efficient screening method for Saccharomyces cerevisiae strains from environmental isolates. MultiPlex PCR was performed targeting four brewing S. cerevisiae genes (SSU1, AWA1, BIO6, and FLO1). At least three genes among the four were amplified from all S. cerevisiae strains. The use of this method allowed us to successfully obtain S. cerevisiae strains.

  9. Influence of organic acids and organochlorinated insecticides on metabolism of Saccharomyces cerevisiae

    Pejin Dušanka J.

    2005-01-01

    Full Text Available Saccharomyces cerevisiae is exposed to different stress factors during the production: osmotic, temperature, oxidative. The response to these stresses is the adaptive mechanism of cells. The raw materials Saccharomyces cerevisiae is produced from, contain metabolism products of present microorganisms and protective agents used during the growth of sugar beet for example the influence of acetic and butyric acid and organochlorinated insecticides, lindan and heptachlor, on the metabolism of Saccharomyces cerevisiae was investigated and presented in this work. The mentioned compounds affect negatively the specific growth rate, yield, content of proteins, phosphorus, total ribonucleic acids. These compounds influence the increase of trechalose and glycogen content in the Saccharomyces cerevisiae cells.

  10. Applied systems biology - vanillin production in Saccharomyces cerevisiae

    Strucko, Tomas; Eriksen, Jens Christian; Nielsen, J.; Mortensen, Uffe Hasbro

    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...

  11. Hydrogen peroxide removal with magnetically responsive Saccharomyces cerevisiae cells

    Šafařík, Ivo; Maděrová, Zdeňka; Šafaříková, Miroslava

    2008-01-01

    Roč. 56, - (2008), s. 7925-7928 ISSN 0021-8561 R&D Projects: GA MPO 2A-1TP1/094; GA MŠk OC 157 Institutional research plan: CEZ:AV0Z60870520 Keywords : magnetic alginate beads * catalase * magnetic separation * Saccharomyces cerevisiae cells * hydrogen peroxide Subject RIV: GM - Food Processing Impact factor: 2.562, year: 2008

  12. Intracellular metabolite profiling of Saccharomyces cerevisiae evolved under furfural

    Jung, Young Hoon; Kim, Sooah; Yang, Jungwoo; Seo, Jin?Ho; Kim, Kyoung Heon

    2016-01-01

    Summary Furfural, one of the most common inhibitors in pre?treatment hydrolysates, reduces the cell growth and ethanol production of yeast. Evolutionary engineering has been used as a selection scheme to obtain yeast strains that exhibit furfural tolerance. However, the response of Saccharomyces cerevisiae to furfural at the metabolite level during evolution remains unknown. In this study, evolutionary engineering and metabolomic analyses were applied to determine the effects of furfural on y...

  13. Oligoadenylate is present in the mitochondrial RNA of Saccharomyces cerevisiae

    Yuckenberg, P.D.; Phillips, S.L.

    1982-01-01

    The authors examined Saccharomyces cerevisiae mitochondrial RNA for polyadenylate. Using hybridization to [/sup 3/H]polyuridylate as the assay for adenylate sequences, they found adenylate-rich oligonucleotides approximately 8 residues long. Longer polyadenylate was not detected. Most of the adenylate-rich sequence is associated with the large mitochondrial rRNA. The remainder is associated with the 10-12S group of transcripts

  14. Dynamics of Storage Carbohydrates Metabolism in Saccharomyces cerevisiae

    Suarez-Mendez, C.A.

    2015-01-01

    Production of chemicals via biotechnological routes are becoming rapidly an alternative to oil-based processes. Several microorganisms including yeast, bacteria, fungi and algae can transform feedstocks into high-value molecules at industrial scale. Improvement of the bioprocess performance is a key factor for making this technology economically feasible. Despite the vast knowledge on microbial metabolism, some gaps still remain open. In Saccharomyces cerevisiae, metabolism of storage carbohy...

  15. Biosynthesis and engineering of kaempferol in Saccharomyces cerevisiae

    Duan, Lijin; Ding, Wentao; Liu, Xiaonan; Cheng, Xiaozhi; Cai, Jing; Hua, Erbing; Jiang, Huifeng

    2017-01-01

    Background Kaempferol is a flavonol with broad bioactivity of anti-oxidant, anti-cancer, anti-diabetic, anti-microbial, cardio-protective and anti-asthma. Microbial synthesis of kaempferol is a promising strategy because of the low content in primary plant source. Methods In this study, the biosynthesis pathway of kaempferol was constructed in the budding yeast Saccharomyces cerevisiae to produce kaempferol de novo, and several biological measures were taken for high production. Results First...

  16. [Saccharomyces cerevisiae invasive infection: The first reported case in Morocco].

    Maleb, A; Sebbar, E; Frikh, M; Boubker, S; Moussaoui, A; El Mekkaoui, A; Khannoussi, W; Kharrasse, G; Belefquih, B; Lemnouer, A; Ismaili, Z; Elouennass, M

    2017-06-01

    Saccharomyces cerevisiae is a cosmopolitan yeast, widely used in agro-alimentary and pharmaceutical industry. Its impact in human pathology is rare, but maybe still underestimated compared to the real situation. This yeast is currently considered as an emerging and opportunistic pathogen. Risk factors are immunosuppression and intravascular device carrying. Fungemias are the most frequent clinical forms. We report the first case of S. cerevisiae invasive infection described in Morocco, and to propose a review of the literature cases of S. cerevisiae infections described worldwide. A 77-year-old patient, with no notable medical history, who was hospitalized for a upper gastrointestinal stenosis secondary to impassable metastatic gastric tumor. Its history was marked by the onset of septic shock, with S. cerevisiae in his urine and in his blood, with arguments for confirmation of invasion: the presence of several risk factors in the patient, positive direct microbiological examination, abundant and exclusive culture of S. cerevisiae from clinical samples. Species identification was confirmed by the study of biochemical characteristics of the isolated yeast. Confirmation of S. cerevisiae infection requires a clinical suspicion in patients with risk factors, but also a correct microbiological diagnosis. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

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

    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.

  18. Metabolic engineering of Saccharomyces cerevisiae for overproduction of triacylglycerols

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

    2018-01-01

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

  19. Value-added biotransformation of cellulosic sugars by engineered Saccharomyces cerevisiae.

    Lane, Stephan; Dong, Jia; Jin, Yong-Su

    2018-07-01

    The substantial research efforts into lignocellulosic biofuels have generated an abundance of valuable knowledge and technologies for metabolic engineering. In particular, these investments have led to a vast growth in proficiency of engineering the yeast Saccharomyces cerevisiae for consuming lignocellulosic sugars, enabling the simultaneous assimilation of multiple carbon sources, and producing a large variety of value-added products by introduction of heterologous metabolic pathways. While microbial conversion of cellulosic sugars into large-volume low-value biofuels is not currently economically feasible, there may still be opportunities to produce other value-added chemicals as regulation of cellulosic sugar metabolism is quite different from glucose metabolism. This review summarizes these recent advances with an emphasis on employing engineered yeast for the bioconversion of lignocellulosic sugars into a variety of non-ethanol value-added products. Copyright © 2018 Elsevier Ltd. All rights reserved.

  20. Transcriptome-Based Characterization of Interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in Lactose-Grown Chemostat Cocultures

    Mendes, F.; Sieuwerts, S.; De Hulster, E.; Almering, M.J.; Luttik, M.A.; Pronk, J.T.; Smid, E.J.; Bron, P.A.; Daran-Lapujade, P.

    2013-01-01

    Mixed populations of Saccharomyces cerevisiae yeasts and lactic acid bacteria occur in many dairy, food, and beverage fermentations, but knowledge about their interactions is incomplete. In the present study, interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp.

  1. Transcriptome-based characterization of interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in lactose-grown chemostat cocultures

    Mendes, F.; Sieuwerts, S.; Hulster, de E.; Almering, M.J.; Luttik, M.A.H.; Pronk, J.T.; Smid, E.J.; Baron, P.A.; Daran-Lapujade, P.

    2013-01-01

    Mixed populations of Saccharomyces cerevisiae yeasts and lactic acid bacteria occur in many dairy, food, and beverage fermentations, but knowledge about their interactions is incomplete. In the present study, interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp.

  2. Glucose-based microbial production of the hormone melatonin in yeast Saccharomyces cerevisiae

    Germann, Susanne Manuela; Jacobsen, Simo Abdessamad; Schneider, Konstantin

    2016-01-01

    performed by complex chemical synthesis. In this study, we demonstrate microbial production of melatonin and related compounds, such as serotonin and N-acetylserotonin. We generated Saccharomyces cerevisiae strains that comprise heterologous genes encoding one or more variants of an L-tryptophan hydroxylase...... accomplished increased product titers by altering expression levels of selected pathway enzymes and boosting co-factor supply. The final yeast strain produced melatonin at a titer of 14.50 ± 0.57 mg L−1 in a 76h fermentation using simulated fed-batch medium with glucose as sole carbon source. Our study lays...

  3. Characterization of vacuolar amino acid transporter from Fusarium oxysporum in Saccharomyces cerevisiae.

    Lunprom, Siriporn; Pongcharoen, Pongsanat; Sekito, Takayuki; Kawano-Kawada, Miyuki; Kakinuma, Yoshimi; Akiyama, Koichi

    2015-01-01

    Fusarium oxysporum causes wilt disease in many plant families, and many genes are involved in its development or growth in host plants. A recent study revealed that vacuolar amino acid transporters play an important role in spore formation in Schizosaccharomyces pombe and Saccharomyces cerevisiae. To investigate the role of vacuolar amino acid transporters of this phytopathogenic fungus, the FOXG_11334 (FoAVT3) gene from F. oxysporum was isolated and its function was characterized. Transcription of FoAVT3 was upregulated after rapamycin treatment. A green fluorescent protein fusion of FoAvt3p was localized to vacuolar membranes in both S. cerevisiae and F. oxysporum. Analysis of the amino acid content of the vacuolar fraction and amino acid transport activities using vacuolar membrane vesicles from S. cerevisiae cells heterologously expressing FoAVT3 revealed that FoAvt3p functions as a vacuolar amino acid transporter, exporting neutral amino acids. We conclude that the FoAVT3 gene encodes a vacuolar neutral amino acid transporter.

  4. Identification and characterization of antifungal compounds using a Saccharomyces cerevisiae reporter bioassay.

    Brad Tebbets

    Full Text Available New antifungal drugs are urgently needed due to the currently limited selection, the emergence of drug resistance, and the toxicity of several commonly used drugs. To identify drug leads, we screened small molecules using a Saccharomyces cerevisiae reporter bioassay in which S. cerevisiae heterologously expresses Hik1, a group III hybrid histidine kinase (HHK from Magnaporthe grisea. Group III HHKs are integral in fungal cell physiology, and highly conserved throughout this kingdom; they are absent in mammals, making them an attractive drug target. Our screen identified compounds 13 and 33, which showed robust activity against numerous fungal genera including Candida spp., Cryptococcus spp. and molds such as Aspergillus fumigatus and Rhizopus oryzae. Drug-resistant Candida albicans from patients were also highly susceptible to compounds 13 and 33. While the compounds do not act directly on HHKs, microarray analysis showed that compound 13 induced transcripts associated with oxidative stress, and compound 33, transcripts linked with heavy metal stress. Both compounds were highly active against C. albicans biofilm, in vitro and in vivo, and exerted synergy with fluconazole, which was inactive alone. Thus, we identified potent, broad-spectrum antifungal drug leads from a small molecule screen using a high-throughput, S. cerevisiae reporter bioassay.

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

    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.

  6. Expression of an endoglucanase from Tribolium castaneum (TcEG1) in Saccharomyces cerevisiae.

    Shirley, Derek; Oppert, Cris; Reynolds, Todd B; Miracle, Bethany; Oppert, Brenda; Klingeman, William E; Jurat-Fuentes, Juan Luis

    2014-10-01

    Insects are a largely unexploited resource in prospecting for novel cellulolytic enzymes to improve the production of ethanol fuel from lignocellulosic biomass. The cost of lignocellulosic ethanol production is expected to decrease by the combination of cellulose degradation (saccharification) and fermentation of the resulting glucose to ethanol in a single process, catalyzed by the yeast Saccharomyces cerevisiae transformed to express efficient cellulases. While S. cerevisiae is an established heterologous expression system, there are no available data on the functional expression of insect cellulolytic enzymes for this species. To address this knowledge gap, S. cerevisiae was transformed to express the full-length cDNA encoding an endoglucanase from the red flour beetle, Tribolium castaneum (TcEG1), and evaluated the activity of the transgenic product (rTcEG1). Expression of the TcEG1 cDNA in S. cerevisiae was under control of the strong glyceraldehyde-3 phosphate dehydrogenase promoter. Cultured transformed yeast secreted rTcEG1 protein as a functional β-1,4-endoglucanase, which allowed transformants to survive on selective media containing cellulose as the only available carbon source. Evaluation of substrate specificity for secreted rTcEG1 demonstrated endoglucanase activity, although some activity was also detected against complex cellulose substrates. Potentially relevant to uses in biofuel production rTcEG1 activity increased with pH conditions, with the highest activity detected at pH 12. Our results demonstrate the potential for functional production of an insect cellulase in S. cerevisiae and confirm the stability of rTcEG1 activity in strong alkaline environments. © 2013 Institute of Zoology, Chinese Academy of Sciences.

  7. Habitat Predicts Levels of Genetic Admixture in Saccharomyces cerevisiae

    Viranga Tilakaratna

    2017-09-01

    Full Text Available Genetic admixture can provide material for populations to adapt to local environments, and this process has played a crucial role in the domestication of plants and animals. The model yeast, Saccharomyces cerevisiae, has been domesticated multiple times for the production of wine, sake, beer, and bread, but the high rate of admixture between yeast lineages has so far been treated as a complication for population genomic analysis. Here, we make use of the low recombination rate at centromeres to investigate admixture in yeast using a classic Bayesian approach and a locus-by-locus phylogenetic approach. Using both approaches, we find that S. cerevisiae from stable oak woodland habitats are less likely to show recent genetic admixture compared with those isolated from transient habitats such as fruits, wine, or human infections. When woodland yeast strains do show recent genetic admixture, the degree of admixture is lower than in strains from other habitats. Furthermore, S. cerevisiae populations from oak woodlands are genetically isolated from each other, with only occasional migration between woodlands and local fruit habitats. Application of the phylogenetic approach suggests that there is a previously undetected population in North Africa that is the closest outgroup to the European S. cerevisiae, including the domesticated Wine population. Careful testing for admixture in S. cerevisiae leads to a better understanding of the underlying population structure of the species and will be important for understanding the selective processes underlying domestication in this economically important species.

  8. Habitat Predicts Levels of Genetic Admixture in Saccharomyces cerevisiae.

    Tilakaratna, Viranga; Bensasson, Douda

    2017-09-07

    Genetic admixture can provide material for populations to adapt to local environments, and this process has played a crucial role in the domestication of plants and animals. The model yeast, Saccharomyces cerevisiae , has been domesticated multiple times for the production of wine, sake, beer, and bread, but the high rate of admixture between yeast lineages has so far been treated as a complication for population genomic analysis. Here, we make use of the low recombination rate at centromeres to investigate admixture in yeast using a classic Bayesian approach and a locus-by-locus phylogenetic approach. Using both approaches, we find that S. cerevisiae from stable oak woodland habitats are less likely to show recent genetic admixture compared with those isolated from transient habitats such as fruits, wine, or human infections. When woodland yeast strains do show recent genetic admixture, the degree of admixture is lower than in strains from other habitats. Furthermore, S. cerevisiae populations from oak woodlands are genetically isolated from each other, with only occasional migration between woodlands and local fruit habitats. Application of the phylogenetic approach suggests that there is a previously undetected population in North Africa that is the closest outgroup to the European S. cerevisiae , including the domesticated Wine population. Careful testing for admixture in S. cerevisiae leads to a better understanding of the underlying population structure of the species and will be important for understanding the selective processes underlying domestication in this economically important species. Copyright © 2017 Tilakaratna and Bensasson.

  9. Metabolic alterations during ascosporogenesis of Saccharomyces cerevisiae

    Carvalho, Sandra; Nadkarni, G.B.

    1977-01-01

    Sporulation of S. cerevisiae has been shown to alter the profiles of enzymes involved in gluconeogenesis and glycolysis. The enhancement in the levels of total cellular carbohydrates could be correlated with the enhancement in fructose 1,6-diphosphatase and trehalose-phosphate synthetase. The latter activity could account for the 15-fold increase in trehalose levels in sporulating cells. Glucose-6-phosphatase, pyruvate kinase and phosphofructokinase showed continuous decline during ascosporogenesis. The relative incorporation of radioactivity from possible precursors of gluconeogenesis indicated that acetate-2- 14 C alone could contribute to carbohydrate synthesis. (author)

  10. [Mitochondria inheritance in yeast saccharomyces cerevisiae].

    Fizikova, A Iu

    2011-01-01

    The review is devoted to the main mechanisms of mitochondria inheritance in yeast Saccharonmyces cerevisiae. The genetic mechanisms of functionally active mitochondria inheritance in eukaryotic cells is one of the most relevant in modem researches. A great number of genetic diseases are associated with mitochondria dysfunction. Plasticity of eukaryotic cell metabolism according to the environmental changes is ensured by adequate mitochondria functioning by means of ATP synthesis coordination, reactive oxygen species accumulation, apoptosis regulation and is an important factor of cell adaptation to stress. Mitochondria participation in important for cell vitality processes masters the presence of accurate mechanisms of mitochondria functions regulation according to environment fluctuations. The mechanisms of mitochondria division and distribution are highly conserved. Baker yeast S. cerevisiae is an ideal model object for mitochondria researches due to energetic metabolism lability, ability to switch over respiration to fermentation, and petite-positive phenotype. Correction of metabolism according to the environmental changes is necessary for cell vitality. The influence of respiratory, carbon, amino acid and phosphate metabolism on mitochondria functions was shown. As far as the mechanisms that stabilize functions of mitochondria and mtDNA are highly conserve, we can project yeast regularities on higher eukaryotes systems. This makes it possible to approximate understanding the etiology and pathogenesis of a great number of human diseases.

  11. Point mutation in calcium-binding domain of mouse polyomavirus VP1 protein does not prevent virus-like particle formation, but changes VP1 interactions with Saccharomyces cerevisiae cell structures

    Adamec, T.; Palková, Zdena; Velková, K.; Štokrová, Jitka; Forstová, J.

    2005-01-01

    Roč. 5, 4-5 (2005), s. 331-340 ISSN 1567-1356 R&D Projects: GA ČR GA204/03/0593 Institutional research plan: CEZ:AV0Z5052915 Keywords : polyomavirus VP1 * Saccharomyces cerevisiae * heterologous expression Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.477, year: 2005

  12. Construction of killer industrial yeast Saccharomyces cerevisiae HAU-1 and its fermentation performance

    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.

  13. On cycles in the transcription network of Saccharomyces cerevisiae

    Berman Piotr

    2008-01-01

    Full Text Available Abstract Background We investigate the cycles in the transcription network of Saccharomyces cerevisiae. Unlike a similar network of Escherichia coli, it contains many cycles. We characterize properties of these cycles and their place in the regulatory mechanism of the cell. Results Almost all cycles in the transcription network of Saccharomyces cerevisiae are contained in a single strongly connected component, which we call LSCC (L for "largest", except for a single cycle of two transcription factors. The fact that LSCC includes almost all cycles is well explained by the properties of a random graph with the same in- and out-degrees of the nodes. Among different physiological conditions, cell cycle has the most significant relationship with LSCC, as the set of 64 transcription interactions that are active in all phases of the cell cycle has overlap of 27 with the interactions of LSCC (of which there are 49. Conversely, if we remove the interactions that are active in all phases of the cell cycle (25% of interactions to transcription factors, the LSCC would have only three nodes and 5 edges, many fewer than expected. This subgraph of the transcription network consists mostly of interactions that are active only in the stress response subnetwork. We also characterize the role of LSCC in the topology of the network. We show that LSCC can be used to define a natural hierarchy in the network and that in every physiological subnetwork LSCC plays a pivotal role. Conclusion Apart from those well-defined conditions, the transcription network of Saccharomyces cerevisiae is devoid of cycles. It was observed that two conditions that were studied and that have no cycles of their own are exogenous: diauxic shift and DNA repair, while cell cycle and sporulation are endogenous. We claim that in a certain sense (slow recovery stress response is endogenous as well.

  14. Removal of Pyrimethanil and Fenhexamid from Saccharomyces cerevisiae Liquid Cultures

    Etjen Bizaj

    2011-01-01

    Full Text Available The capacity for the removal of pyrimethanil and fenhexamid, two fungicides commonly used for the control of Botrytis cinerea in vineyards, has been evaluated during an alcoholic fermentation process in batch system. Commercial and wild strains of Saccharomyces cerevisiae were used. Batch fermentations were carried out in yeast extract-malt extract medium (YM with 18.0 % (by mass glucose, and the fungicides were added separately at three concentrations: 0.1, 1.0 and 10.0 mg/L. The removal capacity of yeast strains was also examined in stationary phase cultures of Saccharomyces cerevisiae. Stationary assays were performed with yeast biomass harvested from the stationary phase of an anaerobic fermentation process, with separate additions of 0.1, 1.0 and 10.0 mg/L of both fungicides. Removal studies with stationary phase cells were performed with viable and non-viable cells inactivated with sodium azide. This study clearly shows that both Saccharomyces cerevisiae strains were able to remove fenhexamid and pyrimethanil in stationary and fermentative assays. The removal potential is shown to be strain dependent in stationary but not in fermentative assays. However, the removal potential is dependent on the type of fungicide in both stationary and fermentative assays. In stationary phase cultures no significant difference in fungicide removal potential between viable and non-viable cells was observed, indicating that both pesticides were not degraded by metabolically active cells. However, the presence of both pesticides influenced fermentation kinetics and only pyrimethanil at 10.0 mg/L increased the production of volatile acidity of both strains.

  15. Protein expression of saccharomyces cerevisiae in response to uranium exposure

    Sakamoto, Fuminori; Nankawa, Takuya; Kozai, Naofumi; Ohnuki, Toshihiko; Fujii, Tsutomu; Iefuji, Haruyuki; Francis, A.J.

    2007-01-01

    Protein expression of Saccharomyces cerevisiae grown in the medium containing 238 U (VI) and 233 U (VI) was examined by two-dimensional gel electrophoresis. Saccharomyces cerevisiae of BY4743 was grown in yeast nitrogen base medium containing glucose and glycerol 2-phosphate and 238 U of 0, 2.0, and 5.0 x 10 -4 M or 233 U of 2.5 x 10 -6 M (radioactivity was higher by 350 times than 2.0 x 10 -4 M 238 U) and 5.0 x 10 -6 M for 112 h at 30 degC. The growth of Saccharomyces cerevisiae was monitored by measuring OD 600 at 112 h after the inoculation. Uranium concentrations in the media also were measured by radiometry using a liquid scintillation counter. The growths of the yeast grown in the above media were in the following order: control>2.5 x 10 -6 M 233 U>2.0 x 10 -4 M 238 U>5.0 x 10 -6 M 233 U>5.0 x 10 -4 M 238 U. This result indicated that not only radiological but also chemical effect of U reduced the growth of the yeast. The concentrations of U in the medium containing 238 U or 233 U decreased, suggesting U accumulation by the yeast cells. The 2-D gel electrophoresis analysis showed the appearance of several spots after exposure to 238 U or to 233 U but not in the control containing no uranium. These results show that the yeast cells exposed to U express several specific proteins. (author)

  16. Division of labour in the yeast: Saccharomyces cerevisiae.

    Wloch-Salamon, Dominika M; Fisher, Roberta M; Regenberg, Birgitte

    2017-10-01

    Division of labour between different specialized cell types is a central part of how we describe complexity in multicellular organisms. However, it is increasingly being recognized that division of labour also plays an important role in the lives of predominantly unicellular organisms. Saccharomyces cerevisiae displays several phenotypes that could be considered a division of labour, including quiescence, apoptosis and biofilm formation, but they have not been explicitly treated as such. We discuss each of these examples, using a definition of division of labour that involves phenotypic variation between cells within a population, cooperation between cells performing different tasks and maximization of the inclusive fitness of all cells involved. We then propose future research directions and possible experimental tests using S. cerevisiae as a model organism for understanding the genetic mechanisms and selective pressures that can lead to the evolution of the very first stages of a division of labour. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  17. Energy-dependent effects of resveratrol in Saccharomyces cerevisiae.

    Madrigal-Perez, Luis Alberto; Canizal-Garcia, Melina; González-Hernández, Juan Carlos; Reynoso-Camacho, Rosalia; Nava, Gerardo M; Ramos-Gomez, Minerva

    2016-06-01

    The metabolic effects induced by resveratrol have been associated mainly with the consumption of high-calorie diets; however, its effects with standard or low-calorie diets remain unclear. To better understand the interactions between resveratrol and cellular energy levels, we used Saccharomyces cerevisiae as a model. Herein it is shown that resveratrol: (a) decreased cell viability in an energy-dependent manner; (b) lessening of cell viability occurred specifically when cells were under cellular respiration; and (c) inhibition of oxygen consumption in state 4 occurred at low and standard energy levels, whereas at high energy levels oxygen consumption was promoted. These findings indicate that the effects of resveratrol are dependent on the cellular energy status and linked to metabolic respiration. Importantly, our study also revealed that S. cerevisiae is a suitable and useful model to elucidate the molecular targets of resveratrol under different nutritional statuses. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  18. Properties of promoters cloned randomly from the Saccharomyces cerevisiae genome.

    Santangelo, G M; Tornow, J; McLaughlin, C S; Moldave, K

    1988-01-01

    Promoters were isolated at random from the genome of Saccharomyces cerevisiae by using a plasmid that contains a divergently arrayed pair of promoterless reporter genes. A comprehensive library was constructed by inserting random (DNase I-generated) fragments into the intergenic region upstream from the reporter genes. Simple in vivo assays for either reporter gene product (alcohol dehydrogenase or beta-galactosidase) allowed the rapid identification of promoters from among these random fragments. Poly(dA-dT) homopolymer tracts were present in three of five randomly cloned promoters. With two exceptions, each RNA start site detected was 40 to 100 base pairs downstream from a TATA element. All of the randomly cloned promoters were capable of activating reporter gene transcription bidirectionally. Interestingly, one of the promoter fragments originated in a region of the S. cerevisiae rDNA spacer; regulated divergent transcription (presumably by RNA polymerase II) initiated in the same region. Images PMID:2847031

  19. Gains and Losses of Transcription Factor Binding Sites in Saccharomyces cerevisiae and Saccharomyces paradoxus.

    Schaefke, Bernhard; Wang, Tzi-Yuan; Wang, Chuen-Yi; Li, Wen-Hsiung

    2015-07-27

    Gene expression evolution occurs through changes in cis- or trans-regulatory elements or both. Interactions between transcription factors (TFs) and their binding sites (TFBSs) constitute one of the most important points where these two regulatory components intersect. In this study, we investigated the evolution of TFBSs in the promoter regions of different Saccharomyces strains and species. We divided the promoter of a gene into the proximal region and the distal region, which are defined, respectively, as the 200-bp region upstream of the transcription starting site and as the 200-bp region upstream of the proximal region. We found that the predicted TFBSs in the proximal promoter regions tend to be evolutionarily more conserved than those in the distal promoter regions. Additionally, Saccharomyces cerevisiae strains used in the fermentation of alcoholic drinks have experienced more TFBS losses than gains compared with strains from other environments (wild strains, laboratory strains, and clinical strains). We also showed that differences in TFBSs correlate with the cis component of gene expression evolution between species (comparing S. cerevisiae and its sister species Saccharomyces paradoxus) and within species (comparing two closely related S. cerevisiae strains). © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  20. Gains and Losses of Transcription Factor Binding Sites in Saccharomyces cerevisiae and Saccharomyces paradoxus

    Schaefke, Bernhard; Wang, Tzi-Yuan; Wang, Chuen-Yi; Li, Wen-Hsiung

    2015-01-01

    Gene expression evolution occurs through changes in cis- or trans-regulatory elements or both. Interactions between transcription factors (TFs) and their binding sites (TFBSs) constitute one of the most important points where these two regulatory components intersect. In this study, we investigated the evolution of TFBSs in the promoter regions of different Saccharomyces strains and species. We divided the promoter of a gene into the proximal region and the distal region, which are defined, respectively, as the 200-bp region upstream of the transcription starting site and as the 200-bp region upstream of the proximal region. We found that the predicted TFBSs in the proximal promoter regions tend to be evolutionarily more conserved than those in the distal promoter regions. Additionally, Saccharomyces cerevisiae strains used in the fermentation of alcoholic drinks have experienced more TFBS losses than gains compared with strains from other environments (wild strains, laboratory strains, and clinical strains). We also showed that differences in TFBSs correlate with the cis component of gene expression evolution between species (comparing S. cerevisiae and its sister species Saccharomyces paradoxus) and within species (comparing two closely related S. cerevisiae strains). PMID:26220934

  1. Sucrose and Saccharomyces cerevisiae: a relationship most sweet.

    Marques, Wesley Leoricy; Raghavendran, Vijayendran; Stambuk, Boris Ugarte; Gombert, Andreas Karoly

    2016-02-01

    Sucrose is an abundant, readily available and inexpensive substrate for industrial biotechnology processes and its use is demonstrated with much success in the production of fuel ethanol in Brazil. Saccharomyces cerevisiae, which naturally evolved to efficiently consume sugars such as sucrose, is one of the most important cell factories due to its robustness, stress tolerance, genetic accessibility, simple nutrient requirements and long history as an industrial workhorse. This minireview is focused on sucrose metabolism in S. cerevisiae, a rather unexplored subject in the scientific literature. An analysis of sucrose availability in nature and yeast sugar metabolism was performed, in order to understand the molecular background that makes S. cerevisiae consume this sugar efficiently. A historical overview on the use of sucrose and S. cerevisiae by humans is also presented considering sugarcane and sugarbeet as the main sources of this carbohydrate. Physiological aspects of sucrose consumption are compared with those concerning other economically relevant sugars. Also, metabolic engineering efforts to alter sucrose catabolism are presented in a chronological manner. In spite of its extensive use in yeast-based industries, a lot of basic and applied research on sucrose metabolism is imperative, mainly in fields such as genetics, physiology and metabolic engineering. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Metabolic impact of redox cofactor perturbations in Saccharomyces cerevisiae

    Hou, Jin; Lages, Nuno; Oldiges, M.

    2009-01-01

    to induce widespread changes in metabolism. We present a detailed analysis of the impact of perturbations in redox cofactors in the cytosol or mitochondria on glucose and energy metabolism in Saccharomyces cerevisiae to aid metabolic engineering decisions that involve cofactor engineering. We enhanced NADH...... oxidation by introducing NADH oxidase or alternative oxidase, its ATP-mediated conversion to NADPH using NADH kinase as well as the interconversion of NADH and NADPH independent of ATP by the soluble, non-proton-translocating bacterial transhydrogenase. Decreasing cytosolic NADH level lowered glycerol...

  3. Studies of the Saccharomyces cerevisiae Cultivation under Oscillatory Mixing Conditions

    M?ris Rikmanis

    2005-12-01

    Full Text Available Saccharomyces cerevisiae was cultivated under non-aerated conditions in a 5 l laboratory bioreactor. Using the experimental data and the regression analysis method, some mathematical correlations for stirrer rotational speed oscillation frequency and the reaction of the yeast were established. It has been found that different growth parameters are influenced variously by stirrer rotational speed and stirrer rotational speed oscillation frequency. Stirring oscillations can be among the methods for stimulation of biotechnological processes. The obtained results can be used for designing bioreactors and optimizing working conditions.

  4. mRNA decapping enzyme from ribosomes of Saccharomyces cerevisiae

    Stevens, A.

    1980-01-01

    By use of [ 3 H]methyl-5'-capped [ 14 C]mRNA from yeast as a substrate, a decapping enzyme activity has been detected in enzyme fractions derived from a high salt wash of ribosomes of Saccharomyces cerevisiae. The product of the decapping reaction is [ 3 H]m 7 GDP. That the enzyme is not a non-specific pyrophosphatase is suggested by the finding that the diphosphate product, m 7 GpppA(G), and UDP-glucose are not hydrolyzed

  5. Magnetically altered ethanol fermentation capacity of Saccharomyces cerevisiae

    Galonja-Corghill Tamara

    2009-01-01

    Full Text Available We studied the effect of static magnetic fields on ethanol production by yeast Saccharomyces cerevisiae 424A (LNH-ST using sugar cane molasses during the fermentation in an enclosed bioreactor. Two static NdFeB magnets were attached to a cylindrical tube reactor with their opposite poles (north to south, creating 150 mT magnetic field inside the reactor. Comparable differences emerged between the results of these two experimental conditions. We found ethanol productivity to be 15% higher in the samples exposed to 150 mT magnetic field.

  6. How Saccharomyces cerevisiae copes with toxic metals and metalloids.

    Wysocki, Robert; Tamás, Markus J

    2010-11-01

    Toxic metals and metalloids are widespread in nature and can locally reach fairly high concentrations. To ensure cellular protection and survival in such environments, all organisms possess systems to evade toxicity and acquire tolerance. This review provides an overview of the molecular mechanisms that contribute to metal toxicity, detoxification and tolerance acquisition in budding yeast Saccharomyces cerevisiae. We mainly focus on the metals/metalloids arsenic, cadmium, antimony, mercury, chromium and selenium, and emphasize recent findings on sensing and signalling mechanisms and on the regulation of tolerance and detoxification systems that safeguard cellular and genetic integrity.

  7. Studies of Saccharomyces cerevisiae and Non-Saccharomyces Yeasts during Alcoholic Fermentation

    Kemsawasd, Varongsiri

    The early death of non-Saccharomyces yeasts during mixed culture spontaneous wine fermentation has traditionally been attributed to the lower capacity of these yeast species to withstand high levels of ethanol, low pH, and other media properties that are a part of progressing fermentation. However......, other yeast-yeast interactions, such as cell-cell contact mediated growth arrest and/or toxininduced death may also be a significant factor in the relative fragility of these non-Saccharomyces yeasts in mixed culture fermentation. In the present work we evaluate the combined roles of cell-cell contact...... and/or antimicrobial peptides on the early death of Lachancea thermotolerans during mixed culture fermentations with Saccharomyces cerevisiae. Using a specially designed double compartment fermentation system, we established that both cell-to-cell contact and antimicrobial peptides contribute...

  8. Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae.

    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.

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

    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. Expression of monellin in a food-grade delivery system in Saccharomyces cerevisiae.

    Liu, Jun; Yan, Da-zhong; Zhao, Sheng-jun

    2015-10-01

    Genetically modified (GM) foods have caused much controversy. Construction of a food-grade delivery system is a desirable technique with presumptive impact on industrial applications from the perspective of bio-safety. The aim of this study was to construct a food-grade delivery system for Saccharomyces cerevisiae and to study the expression of monellin from the berries of the West African forest plant Dioscoreophyllum cumminsii in this system. A food-grade system for S. cerevisiae was constructed based on ribosomal DNA (rDNA)-mediated homologous recombination to enable high-copy-number integration of the expression cassette inserted into the rDNA locus. A copper resistance gene (CUP1) was used as the selection marker for yeast transformation. Because variants of transformants containing different copy numbers at the CUP1 locus can be readily selected after growth in the presence of elevated copper levels, we suggest that this system would prove useful in the generation of tandemly iterated gene clusters. Using this food-grade system, a single-chain monellin gene was heterologously expressed. The yield of monellin reached a maximum of 675 mg L(-1) . This system harbors exclusively S. cerevisiae DNA with no antibiotic resistance genes, and it should therefore be appropriate for safe use in the food industry. Monellin was shown to be expressed in this food-grade delivery system. To our knowledge, this is the first report so far on expression of monellin in a food-grade expression system in S. cerevisiae. © 2014 Society of Chemical Industry.

  11. Growth of non-Saccharomyces yeasts affects nutrient availability for Saccharomyces cerevisiae during wine fermentation.

    Medina, Karina; Boido, Eduardo; Dellacassa, Eduardo; Carrau, Francisco

    2012-07-02

    Yeast produces numerous secondary metabolites during fermentation that impact final wine quality. Although it is widely recognized that growth of diverse non-Saccharomyces (NS) yeast can positively affect flavor complexity during Saccharomyces cerevisiae wine fermentation, the inability to control spontaneous or co-fermentation processes by NS yeast has restricted their use in winemaking. We selected two NS yeasts from our Uruguayan native collection to study NS-S. cerevisiae interactions during wine fermentation. The selected strains of Hanseniaspora vineae and Metschnikowia pulcherrima had different yeast assimilable nitrogen consumption profiles and had different effects on S. cerevisiae fermentation and growth kinetics. Studies in which we varied inoculum size and using either simultaneous or sequential inoculation of NS yeast and S. cerevisiae suggested that competition for nutrients had a significant effect on fermentation kinetics. Sluggish fermentations were more pronounced when S. cerevisiae was inoculated 24h after the initial stage of fermentation with a NS strain compared to co-inoculation. Monitoring strain populations using differential WL nutrient agar medium and fermentation kinetics of mixed cultures allowed for a better understanding of strain interactions and nutrient addition effects. Limitation of nutrient availability for S. cerevisiae was shown to result in stuck fermentations as well as to reduce sensory desirability of the resulting wine. Addition of diammonium phosphate (DAP) and a vitamin mix to a defined medium allowed for a comparison of nutrient competition between strains. Addition of DAP and the vitamin mix was most effective in preventing stuck fermentations. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Glucose-based microbial production of the hormone melatonin in yeast Saccharomyces cerevisiae.

    Germann, Susanne M; Baallal Jacobsen, Simo A; Schneider, Konstantin; Harrison, Scott J; Jensen, Niels B; Chen, Xiao; Stahlhut, Steen G; Borodina, Irina; Luo, Hao; Zhu, Jiangfeng; Maury, Jérôme; Forster, Jochen

    2016-05-01

    Melatonin is a natural mammalian hormone that plays an important role in regulating the circadian cycle in humans. It is a clinically effective drug exhibiting positive effects as a sleep aid and a powerful antioxidant used as a dietary supplement. Commercial melatonin production is predominantly performed by complex chemical synthesis. In this study, we demonstrate microbial production of melatonin and related compounds, such as serotonin and N-acetylserotonin. We generated Saccharomyces cerevisiae strains that comprise heterologous genes encoding one or more variants of an L-tryptophan hydroxylase, a 5-hydroxy-L-tryptophan decarboxylase, a serotonin acetyltransferase, an acetylserotonin O-methyltransferase, and means for providing the cofactor tetrahydrobiopterin via heterologous biosynthesis and recycling pathways. We thereby achieved de novo melatonin biosynthesis from glucose. We furthermore accomplished increased product titers by altering expression levels of selected pathway enzymes and boosting co-factor supply. The final yeast strain produced melatonin at a titer of 14.50 ± 0.57 mg L(-1) in a 76h fermentation using simulated fed-batch medium with glucose as sole carbon source. Our study lays the basis for further developing a yeast cell factory for biological production of melatonin. © 2015 The Authors. Biotechnology Journal published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    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

  14. De novo biosynthesis of vanillin in fission yeast (Schizosaccharomyces pombe) and baker's yeast (Saccharomyces cerevisiae).

    Hansen, Esben H; Møller, Birger Lindberg; Kock, Gertrud R; Bünner, Camilla M; Kristensen, Charlotte; Jensen, Ole R; Okkels, Finn T; Olsen, Carl E; Motawia, Mohammed S; Hansen, Jørgen

    2009-05-01

    Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast, Saccharomyces cerevisiae. Productivities were 65 and 45 mg/liter, after introduction of three and four heterologous genes, respectively. The engineered pathways involve incorporation of 3-dehydroshikimate dehydratase from the dung mold Podospora pauciseta, an aromatic carboxylic acid reductase (ACAR) from a bacterium of the Nocardia genus, and an O-methyltransferase from Homo sapiens. In S. cerevisiae, the ACAR enzyme required activation by phosphopantetheinylation, and this was achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol was achieved by knockout of the host alcohol dehydrogenase ADH6. In S. pombe, the biosynthesis was further improved by introduction of an Arabidopsis thaliana family 1 UDP-glycosyltransferase, converting vanillin into vanillin beta-D-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts. These de novo pathways represent the first examples of one-cell microbial generation of these valuable compounds from glucose. S. pombe yeast has not previously been metabolically engineered to produce any valuable, industrially scalable, white biotech commodity.

  15. Iterative optimization of xylose catabolism in Saccharomyces cerevisiae using combinatorial expression tuning.

    Latimer, Luke N; Dueber, John E

    2017-06-01

    A common challenge in metabolic engineering is rapidly identifying rate-controlling enzymes in heterologous pathways for subsequent production improvement. We demonstrate a workflow to address this challenge and apply it to improving xylose utilization in Saccharomyces cerevisiae. For eight reactions required for conversion of xylose to ethanol, we screened enzymes for functional expression in S. cerevisiae, followed by a combinatorial expression analysis to achieve pathway flux balancing and identification of limiting enzymatic activities. In the next round of strain engineering, we increased the copy number of these limiting enzymes and again tested the eight-enzyme combinatorial expression library in this new background. This workflow yielded a strain that has a ∼70% increase in biomass yield and ∼240% increase in xylose utilization. Finally, we chromosomally integrated the expression library. This library enriched for strains with multiple integrations of the pathway, which likely were the result of tandem integrations mediated by promoter homology. Biotechnol. Bioeng. 2017;114: 1301-1309. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  16. Recombinant production of human Aquaporin-1 to an exceptional high membrane density in Saccharomyces cerevisiae.

    Julie Bomholt

    Full Text Available In the present paper we explored the capacity of yeast Saccharomyces cerevisiae as host for heterologous expression of human Aquaporin-1. Aquaporin-1 cDNA was expressed from a galactose inducible promoter situated on a plasmid with an adjustable copy number. Human Aquaporin-1 was C-terminally tagged with yeast enhanced GFP for quantification of functional expression, determination of sub-cellular localization, estimation of in vivo folding efficiency and establishment of a purification protocol. Aquaporin-1 was found to constitute 8.5 percent of total membrane protein content after expression at 15°C in a yeast host over-producing the Gal4p transcriptional activator and growth in amino acid supplemented minimal medium. In-gel fluorescence combined with western blotting showed that low accumulation of correctly folded recombinant Aquaporin-1 at 30°C was due to in vivo mal-folding. Reduction of the expression temperature to 15°C almost completely prevented Aquaporin-1 mal-folding. Bioimaging of live yeast cells revealed that recombinant Aquaporin-1 accumulated in the yeast plasma membrane. A detergent screen for solubilization revealed that CYMAL-5 was superior in solubilizing recombinant Aquaporin-1 and generated a monodisperse protein preparation. A single Ni-affinity chromatography step was used to obtain almost pure Aquaporin-1. Recombinant Aquaporin-1 produced in S. cerevisiae was not N-glycosylated in contrast to the protein found in human erythrocytes.

  17. Saccharomyces cerevisiae as a model organism: a comparative study.

    Hiren Karathia

    Full Text Available BACKGROUND: Model organisms are used for research because they provide a framework on which to develop and optimize methods that facilitate and standardize analysis. Such organisms should be representative of the living beings for which they are to serve as proxy. However, in practice, a model organism is often selected ad hoc, and without considering its representativeness, because a systematic and rational method to include this consideration in the selection process is still lacking. METHODOLOGY/PRINCIPAL FINDINGS: In this work we propose such a method and apply it in a pilot study of strengths and limitations of Saccharomyces cerevisiae as a model organism. The method relies on the functional classification of proteins into different biological pathways and processes and on full proteome comparisons between the putative model organism and other organisms for which we would like to extrapolate results. Here we compare S. cerevisiae to 704 other organisms from various phyla. For each organism, our results identify the pathways and processes for which S. cerevisiae is predicted to be a good model to extrapolate from. We find that animals in general and Homo sapiens in particular are some of the non-fungal organisms for which S. cerevisiae is likely to be a good model in which to study a significant fraction of common biological processes. We validate our approach by correctly predicting which organisms are phenotypically more distant from S. cerevisiae with respect to several different biological processes. CONCLUSIONS/SIGNIFICANCE: The method we propose could be used to choose appropriate substitute model organisms for the study of biological processes in other species that are harder to study. For example, one could identify appropriate models to study either pathologies in humans or specific biological processes in species with a long development time, such as plants.

  18. Data on dynamic study of cytoophidia in Saccharomyces cerevisiae

    Hui Li

    2016-09-01

    Full Text Available The data in this paper are related to the research article entitled “Filamentation of metabolic enzymes in Saccharomyces cerevisiae” Q.J. Shen et al. (2016 [1]. Cytoophidia are filamentous structures discovered in fruit flies (doi:10.1016/S1673-8527(0960046-1 J.L. Liu (2010 [2], bacteria (doi:10.1038/ncb2087 M. Ingerson-Mahar et al. (2010 [3], yeast (doi:10.1083/jcb.201003001; doi:10.1242/bio.20149613 C. Noree et al. (2010 and J. Zhang, L. Hulme, J.L. Liu (2014 [4,5] and human cells (doi:10.1371/journal.pone.0029690; doi:10.1016/j.jgg.2011.08.004 K. Chen et al. (2011 and W.C. Carcamo et al. (2011 ( [6,7]. However, there is little research on the motility of the cytoophidia. Here we selected cytoophidia formed by 6 filament-forming proteins in the budding yeast S. cerevisiae, and performed living-cell imaging of cells expressing the proteins fused with GFP. The dynamic features of the six types of cytoophidia were analyzed. In the data, both raw movies and analysed results of the dynamics of cytoophidia are presented. Keywords: Saccharomyces cerevisiae, CTP synthase, Cytoophidium, Metabolism, Filamentation

  19. Omics analysis of acetic acid tolerance in Saccharomyces cerevisiae.

    Geng, Peng; Zhang, Liang; Shi, Gui Yang

    2017-05-01

    Acetic acid is an inhibitor in industrial processes such as wine making and bioethanol production from cellulosic hydrolysate. It causes energy depletion, inhibition of metabolic enzyme activity, growth arrest and ethanol productivity losses in Saccharomyces cerevisiae. Therefore, understanding the mechanisms of the yeast responses to acetic acid stress is essential for improving acetic acid tolerance and ethanol production. Although 329 genes associated with acetic acid tolerance have been identified in the Saccharomyces genome and included in the database ( http://www.yeastgenome.org/observable/resistance_to_acetic_acid/overview ), the cellular mechanistic responses to acetic acid remain unclear in this organism. Post-genomic approaches such as transcriptomics, proteomics, metabolomics and chemogenomics are being applied to yeast and are providing insight into the mechanisms and interactions of genes, proteins and other components that together determine complex quantitative phenotypic traits such as acetic acid tolerance. This review focuses on these omics approaches in the response to acetic acid in S. cerevisiae. Additionally, several novel strains with improved acetic acid tolerance have been engineered by modifying key genes, and the application of these strains and recently acquired knowledge to industrial processes is also discussed.

  20. Characterization of an MMS sensitive mutant of Saccharomyces cerevisiae

    Martin, P.S.

    1979-01-01

    We have characterized a methyl methanesulfonate sensitive mutant of the yeast Saccharomyces cerevisiae in order to learn more about DNA repair and mutagenesis in this organism. The mutation, designated mms3-1, also confers sensitivity to ultraviolet light and to ethyl methanesulfonate in both haploids and homozygous diploids. Its effect on γ-ray sensitivity, however, is a function of the ploidy of the cell and its effect on induced mutation is a function of both the ploidy of the cell and the nature of the inducing agent. Our major findings are discussed. Our data indicate that: (1) Saccharomyces cerevisiae has an error prone pathway for the repair of uv damage controlled by the MMS3 gene product operating in and only in, and possibly induced by conditions present only in, a/α diploids; (2) in diploids, at least, there exists at least one step in the error prone repair of uv induced damage which is different from a step in the error prone repair of EMS induced damage; (3) a/α mms3-1/mms3-1 diploids may be defective in a step common to the repair of mutagenic lesions following uv irradiation and lethal lesions following γ irradiation; and (4) there are steps in the repair of MMS induced lethal damage that are different from steps in the repair of EMS induced lethal damage

  1. A vaccine grade of yeast Saccharomyces cerevisiae expressing mammalian myostatin

    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.

  2. Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae.

    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.

  3. On the origins and industrial applications of Saccharomyces cerevisiae × Saccharomyces kudriavzevii hybrids.

    Peris, David; Pérez-Torrado, Roberto; Hittinger, Chris Todd; Barrio, Eladio; Querol, Amparo

    2018-01-01

    Companies based on alcoholic fermentation products, such as wine, beer and biofuels, use yeasts to make their products. Each industrial process utilizes different media conditions, which differ in sugar content, the presence of inhibitors and fermentation temperature. Saccharomyces cerevisiae has traditionally been the main yeast responsible for most fermentation processes. However, the market is changing due to consumer demand and external factors such as climate change. Some processes, such as biofuel production or winemaking, require new yeasts to solve specific challenges, especially those associated with sustainability, novel flavours and altered alcohol content. One of the proposed solutions is the application of yeast hybrids. The lager beer market has been dominated by S. cerevisiae × S. eubayanus hybrids. However, several less thoroughly studied hybrids have been isolated from other diverse industrial processes. Here we focus on S. cerevisiae × S. kudriavzevii hybrids, which have been isolated from diverse industrial conditions that include wine, ale beer, cider and dietary supplements. Emerging data suggest an extended and complex story of adaptation of these hybrids to traditional industrial conditions. S. cerevisiae × S. kudriavzevii hybrids are also being explored for new industrial applications, such as biofuels. This review describes the past, present and future of S. cerevisiae × S. kudriavzevii hybrids. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  4. Sucrose fermentation by Saccharomyces cerevisiae lacking hexose transport.

    Batista, Anderson S; Miletti, Luiz C; Stambuk, Boris U

    2004-01-01

    Sucrose is the major carbon source used by Saccharomyces cerevisiae during production of baker's yeast, fuel ethanol and several distilled beverages. It is generally accepted that sucrose fermentation proceeds through extracellular hydrolysis of the sugar, mediated by the periplasmic invertase, producing glucose and fructose that are transported into the cells and metabolized. In the present work we analyzed the contribution to sucrose fermentation of a poorly characterized pathway of sucrose utilization by S. cerevisiae cells, the active transport of the sugar through the plasma membrane and its intracellular hydrolysis. A yeast strain that lacks the major hexose transporters (hxt1-hxt7 and gal2) is incapable of growing on or fermenting glucose or fructose. Our results show that this hxt-null strain is still able to ferment sucrose due to direct uptake of the sugar into the cells. Deletion of the AGT1 gene, which encodes a high-affinity sucrose-H(+) symporter, rendered cells incapable of sucrose fermentation. Since sucrose is not an inducer of the permease, expression of the AGT1 must be constitutive in order to allow growth of the hxt-null strain on sucrose. The molecular characterization of active sucrose transport and fermentation by S. cerevisiae cells opens new opportunities to optimize yeasts for sugarcane-based industrial processes.

  5. Saccharomyces cerevisiae in the Production of Fermented Beverages

    Graeme M Walker

    2016-11-01

    Full Text Available Alcoholic beverages are produced following the fermentation of sugars by yeasts, mainly (but not exclusively strains of the species, Saccharomyces cerevisiae. The sugary starting materials may emanate from cereal starches (which require enzymatic pre-hydrolysis in the case of beers and whiskies, sucrose-rich plants (molasses or sugar juice from sugarcane in the case of rums, or from fruits (which do not require pre-hydrolysis in the case of wines and brandies. In the presence of sugars, together with other essential nutrients such as amino acids, minerals and vitamins, S. cerevisiae will conduct fermentative metabolism to ethanol and carbon dioxide (as the primary fermentation metabolites as the cells strive to make energy and regenerate the coenzyme NAD+ under anaerobic conditions. Yeasts will also produce numerous secondary metabolites which act as important beverage flavour congeners, including higher alcohols, esters, carbonyls and sulphur compounds. These are very important in dictating the final flavour and aroma characteristics of beverages such as beer and wine, but also in distilled beverages such as whisky, rum and brandy. Therefore, yeasts are of vital importance in providing the alcohol content and the sensory profiles of such beverages. This Introductory Chapter reviews, in general, the growth, physiology and metabolism of S. cerevisiae in alcoholic beverage fermentations.

  6. Functional relevance of water and glycerol channels in Saccharomyces cerevisiae.

    Sabir, Farzana; Loureiro-Dias, Maria C; Soveral, Graça; Prista, Catarina

    2017-05-01

    Our understanding of the functional relevance of orthodox aquaporins and aquaglyceroporins in Saccharomyces cerevisiae is essentially based on phenotypic variations obtained by expression/overexpression/deletion of these major intrinsic proteins in selected strains. These water/glycerol channels are considered crucial during various life-cycle phases, such as sporulation and mating and in some life processes such as rapid freeze-thaw tolerance, osmoregulation and phenomena associated with cell surface. Despite their putative functional roles not only as channels but also as sensors, their underlying mechanisms and their regulation are still poorly understood. In the present review, we summarize and discuss the physiological relevance of S. cerevisiae aquaporins (Aqy1 and Aqy2) and aquaglyceroporins (Fps1 and Yfl054c). In particular, the fact that most S. cerevisiae laboratory strains harbor genes coding for non-functional aquaporins, while wild and industrial strains possess at least one functional aquaporin, suggests that aquaporin activity is required for cell survival under more harsh conditions. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. Irradiation effects on the alcohol fermentation ability of saccharomyces cerevisiae

    Sadi, Suharni

    1987-01-01

    Irradiation effects on the alcohol fermentation ability of saccharomyces cerevisiae. S. cerevisiae suspensions of 1.5x10 8 clls/ml were exposed to single and fractionated doses of gamma irradiation, i.e. 0; 0.30; 0.60; 0.90; and 1.20 kGy in aerobic condition at dose rate of 1.63 kGy/hour. The fractionated doses were given with time interval of 15, 30 and 45 minutes. The fermentation was held at 30 0 C for 40 hours. It is seen that an increase of alcohol production was obtained when cells were irradiated at 0.60 kGy, although the result has no significant difference statistically with control. At the dose of 1.20 kGy the alcohol fermentation ability of S. cerevisiae decreased drastically as compared to control. Irradiation using single or fractionated doses with time interval of 15-45 minutes did not influence the alcohol production. Comparing the time interval of 45 minutes at 0.60 kGy and at 1.20 kGy, it appeared that the yield of alcohol was different. (author). 17 refs.; 4 figs

  8. Osmo-, thermo- and ethanol- tolerances of Saccharomyces cerevisiae S1

    Sandrasegarampillai Balakumar

    2012-03-01

    Full Text Available Saccharomyces cerevisiae S1, which is a locally isolated and improved strain showed viability at 40, 45 and 50ºC and produced ethanol at 40, 43 and 45ºC. When the cells were given heat shock at 45ºC for 30min and grown at 40ºC, 100% viability was observed for 60h, and addition of 200gl-1 ethanol has led to complete cell death at 30h. Heat shock given at 45ºC (for 30min has improved the tolerance to temperature induced ethanol shock leading to 37% viability at 30h. when the cells were subjected to ethanol (200gl-1 for 30 min and osmotic shock (sorbitol 300gl-1, trehalose contents in the cells were increased. The heat shocked cells showed better viability in presence of added ethanol. Soy flour supplementation has improved the viability of S. cerevisiae S1 to 80% in presence of 100gl-1 added ethanol and to 60% in presence of 300gl-1 sorbitol. In presence of sorbitol (200gl-1 and ethanol (50gl-1 at 40ºC, 46% viability was retained by S. cerevisiae S1 at 48h and it was improved to 80% by soy flour supplementation.

  9. Early manifestations of replicative aging in the yeast Saccharomyces cerevisiae

    Maksim I. Sorokin

    2014-01-01

    Full Text Available The yeast Saccharomyces cerevisiae is successfully used as a model organism to find genes responsible for lifespan control of higher organisms. As functional decline of higher eukaryotes can start as early as one quarter of the average lifespan, we asked whether S. cerevisiae can be used to model this manifestation of aging. While the average replicative lifespan of S. cerevisiae mother cells ranges between 15 and 30 division cycles, we found that resistances to certain stresses start to decrease much earlier. Looking into the mechanism, we found that knockouts of genes responsible for mitochondriato-nucleus (retrograde signaling, RTG1 or RTG3, significantly decrease the resistance of cells that generated more than four daughters, but not of the younger ones. We also found that even young mother cells frequently contain mitochondria with heterogeneous transmembrane potential and that the percentage of such cells correlates with replicative age. Together, these facts suggest that retrograde signaling starts to malfunction in relatively young cells, leading to accumulation of heterogeneous mitochondria within one cell. The latter may further contribute to a decline in stress resistances.

  10. Saccharomyces cerevisiae in the Production of Whisk(ey

    Graeme M. Walker

    2016-12-01

    Full Text Available Whisk(ey is a major global distilled spirit beverage. Whiskies are produced from cereal starches that are saccharified, fermented and distilled prior to spirit maturation. The strain of Saccharomyces cerevisiae employed in whisky fermentations is crucially important not only in terms of ethanol yields, but also for production of minor yeast metabolites which collectively contribute to development of spirit flavour and aroma characteristics. Distillers must therefore pay very careful attention to the strain of yeast exploited to ensure consistency of fermentation performance and spirit congener profiles. In the Scotch whisky industry, initiatives to address sustainability issues facing the industry (for example, reduced energy and water usage have resulted in a growing awareness regarding criteria for selecting new distilling yeasts with improved efficiency. For example, there is now a desire for Scotch whisky distilling yeasts to perform under more challenging conditions such as high gravity wort fermentations. This article highlights the important roles of S. cerevisiae strains in whisky production (with particular emphasis on Scotch and describes key fermentation performance attributes sought in distiller’s yeast, such as high alcohol yields, stress tolerance and desirable congener profiles. We hope that the information herein will be useful for whisky producers and yeast suppliers in selecting new distilling strains of S. cerevisiae, and for the scientific community to stimulate further research in this area.

  11. The adsorption of Sr(II) and Cs(I) ions by irradiated Saccharomyces cerevisiae

    Yiming Tan; Jundong Feng; Liang Qiu; Zhentian Zhao; Xiaohong Zhang; Haiqian Zhang

    2017-01-01

    Adsorption behavior and mechanism of Sr(II) and Cs(I) in single and binary solutions using irradiated Saccharomyces cerevisiae was investigated. The effects of several environmental factors on Sr(II) and Cs(I) adsorption to irradiated Saccharomyces cerevisiae was determined. The equilibrium experimental data were simulated by different kinetic models and isotherm models. The combined effect of Sr(II) and Cs(I) on Saccharomyces cerevisiae is generally antagonistic. SEM and EDS analyses indicate that crystals formed on the cell surface are precipitate of Sr(II) and Cs(I), respectively. (author)

  12. Uranium removal from acidic aqueous solutions by Saccharomyces cerevisiae, Debaryomyces hansenii, Kluyveromyces marxianus and Candida colliculosa

    Sarri, S.; Misaelides, P.; Papanikolaou, M.; Zamboulis, D.

    2009-01-01

    The sorption of uranium from acidic aqueous solutions (pH 4.5, C init = 10 to 1000 mg U/L) by Saccharomyces cerevisiae, Debaryomyces hansenii, Kluyveromyces marxianus and Candida colliculosa was investigated using a batch technique. The U-sorption onto Saccharomyces cerevisiae and Debaryomyces hansenii followed a Langmuir, while that onto Kluyveromyces marxianus and Candida colliculosa a Freundlich isotherm. The results demonstrated that all investigated biomasses could effectively remove uranium from acidic aqueous solutions. From all sorbents, Saccharomyces cerevisiae appeared to be the most effective with a maximum sorption capacity of 127.7 mg U/g dry biomass. (author)

  13. Intracellular Signal Triggered by Cholera Toxin in Saccharomyces boulardii and Saccharomyces cerevisiae

    Brandão, Rogelio L.; Castro, Ieso M.; Bambirra, Eduardo A.; Amaral, Sheila C.; Fietto, Luciano G.; Tropia, Maria José M.; Neves, Maria José; Dos Santos, Raquel G.; Gomes, Newton C. M.; Nicoli, Jacques R.

    1998-01-01

    As is the case for Saccharomyces boulardii, Saccharomyces cerevisiae W303 protects Fisher rats against cholera toxin (CT). The addition of glucose or dinitrophenol to cells of S. boulardii grown on a nonfermentable carbon source activated trehalase in a manner similar to that observed for S. cerevisiae. The addition of CT to the same cells also resulted in trehalase activation. Experiments performed separately on the A and B subunits of CT showed that both are necessary for activation. Similarly, the addition of CT but not of its separate subunits led to a cyclic AMP (cAMP) signal in both S. boulardii and S. cerevisiae. These data suggest that trehalase stimulation by CT probably occurred through the cAMP-mediated protein phosphorylation cascade. The requirement of CT subunit B for both the cAMP signal and trehalase activation indicates the presence of a specific receptor on the yeasts able to bind to the toxin, a situation similar to that observed for mammalian cells. This hypothesis was reinforced by experiments with 125I-labeled CT showing specific binding of the toxin to yeast cells. The adhesion of CT to a receptor on the yeast surface through the B subunit and internalization of the A subunit (necessary for the cAMP signal and trehalase activation) could be one more mechanism explaining protection against the toxin observed for rats treated with yeasts. PMID:9464394

  14. Probiotic Activity of Saccharomyces cerevisiae var. boulardii Against Human Pathogens

    Katarzyna Rajkowska

    2012-01-01

    Full Text Available Infectious diarrhoea is associated with a modification of the intestinal microflora and colonization of pathogenic bacteria. Tests were performed for seven probiotic yeast strains of Saccharomyces cerevisiae var. boulardii, designated for the prevention and treatment of diarrhoea. To check their possible effectiveness against diarrhoea of different etiologies, the activity against a variety of human pathogenic or opportunistic bacteria was investigated in vitro. In mixed cultures with S. cerevisiae var. boulardii, a statistically significant reduction was observed in the number of cells of Listeria monocytogenes, Pseudomonas aeruginosa and Staphylococcus aureus, by even 55.9 % in the case of L. monocytogenes compared with bacterial monocultures. The influence of yeasts was mostly associated with the shortening of the bacterial lag phase duration, more rapid achievement of the maximum growth rates, and a decrease by 4.4–57.1 % (L. monocytogenes, P. aeruginosa, or an increase by 1.4–70.6 % (Escherichia coli, Enterococcus faecalis, Salmonella Typhimurium in the exponential growth rates. Another issue included in the research was the ability of S. cerevisiae var. boulardii to bind pathogenic bacteria to its cell surface. Yeasts have shown binding capacity of E. coli, S. Typhimurium and additionally of S. aureus, Campylobacter jejuni and E. faecalis. However, no adhesion of L. monocytogenes and P. aeruginosa to the yeast cell wall was noted. The probiotic activity of S. cerevisiae var. boulardii against human pathogens is related to a decrease in the number of viable and active cells of bacteria and the binding capacity of yeasts. These processes may limit bacterial invasiveness and prevent bacterial adherence and translocation in the human intestines.

  15. Establishing very long-chain fatty alcohol and wax ester biosynthesis in Saccharomyces cerevisiae.

    Wenning, Leonie; Yu, Tao; David, Florian; Nielsen, Jens; Siewers, Verena

    2017-05-01

    Wax esters (WEs) are neutral lipids and can be used for a broad range of commercial applications, including personal care products, lubricants, or coatings. They are synthesized by enzymatic reactions catalyzed by a fatty acyl reductase (FAR) and a wax ester synthase (WS). At present, commercially used WEs are mainly isolated from Simmondsia chinensis (jojoba), but the high extraction costs and limited harvest areas constrain their use. The use of FARs in combination with different WSs to achieve a synthesis of jojoba-like WEs in bacteria and yeast has been reported previously, but the products were restricted to C28-C36 WEs. These rather short WEs make up only a very small percentage of the total WEs in natural jojoba oil. The synthesis of longer chain WEs (up to C44) in Saccharomyces cerevisiae has so far only been achieved after substrate feeding. Here we identified new routes for producing very long-chain fatty alcohols (VLCFOHs) up to a chain length of C22 by heterologous expression of a FAR derived from Apis mellifera (AmFAR1) or Marinobacter aquaeolei VT8 (Maqu_2220) in S. cerevisiae and achieved maximum yields of 3.22 ± 0.36 mg/g cell dry weight (CDW) and 7.84 ± 3.09 mg/g CDW, respectively, after 48 h. Moreover, we enabled the synthesis of jojoba-like WEs up to a chain length of C42, catalyzed by a combination of Maqu_2220 together with the WS from S. chinensis (SciWS) and the S. cerevisiae elongase Elo2p, with a maximum yield of 12.24 ± 3.35 mg/g CDW after 48 h. Biotechnol. Bioeng. 2017;114: 1025-1035. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Dominance of Saccharomyces cerevisiae in alcoholic fermentation processes

    Albergaria, Helena; Arneborg, Nils

    2016-01-01

    Winemaking, brewing and baking are some of the oldest biotechnological processes. In all of them, alcoholic fermentation is the main biotransformation and Saccharomyces cerevisiae the primary microorganism. Although a wide variety of microbial species may participate in alcoholic fermentation and...

  17. Evaluation of molecular typing techniques to assign genetic diversity among Saccharomyces cerevisiae strains

    Baleiras Couto, M.M.; Eijsma, B.; Hofstra, H.; Huis in 't Veld, J.H.J.; Vossen, J.M.B.M. van der

    1996-01-01

    Discrimination of strains within the species Saccharomyces cerevisiae was demonstrated by the use of four different techniques to type 15 strains isolated from spoiled wine and beer. Random amplified polymorphic DNA with specific oligonucleotides and PCR fingerprinting with the microsatellite

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

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

    2014-01-01

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

  19. Reconstruction of the biosynthetic pathway for the core fungal polyketide scaffold rubrofusarin in Saccharomyces cerevisiae

    Rugbjerg, Peter; Naesby, Michael; Mortensen, Uffe Hasbro

    2013-01-01

    production in easily fermentable and genetically engineerable organisms, such as Saccharomyces cerevisiae and Escherichia coli are desirable. Rubrofusarin is an orange polyketide pigment that is a common intermediate in many different fungal biosynthetic pathways. RESULTS: In this study, we established...

  20. Multiplexed CRISPR/Cas9 Genome Editing and Gene Regulation Using Csy4 in Saccharomyces cerevisiae

    Ferreira, Raphael; Skrekas, Christos; Nielsen, Jens

    2018-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) technology has greatly accelerated the field of strain engineering. However, insufficient efforts have been made toward developing robust multiplexing tools in Saccharomyces cerevisiae. Here, we exploit the RNA processing capacity...

  1. Tolerance of budding yeast Saccharomyces cerevisiae to ultra high pressure

    Shibata, M.; Torigoe, M.; Matsumoto, Y.; Yamamoto, M.; Takizawa, N.; Hada, Y.; Mori, Y.; Takarabe, K.; Ono, F.

    2014-05-01

    Our studies on the tolerance of plants and animals against very high pressure of several GPa have been extended to a smaller sized fungus, the budding yeast Saccharomyces cerevisiae. Several pieces of budding yeast (dry yeast) were sealed in a small teflon capsule with a liquid pressure medium fluorinate, and exposed to 7.5 GPa by using a cubic anvil press. The pressure was kept constant for various duration of time from 2 to 24 h. After the pressure was released, the specimens were brought out from the teflon capsule, and they were cultivated on a potato dextrose agar. It was found that the budding yeast exposed to 7.5 GPa for up to 6 h showed multiplication. However, those exposed to 7.5 GPa for longer than 12 h were found dead. The high pressure tolerance of budding yeast is a little weaker than that of tardigrades.

  2. Mechanisms and Regulation of Mitotic Recombination in Saccharomyces cerevisiae

    Symington, Lorraine S.; Rothstein, Rodney; Lisby, Michael

    2014-01-01

    Homology-dependent exchange of genetic information between DNA molecules has a profound impact on the maintenance of genome integrity by facilitating error-free DNA repair, replication, and chromosome segregation during cell division as well as programmed cell developmental events. This chapter will focus on homologous mitotic recombination in budding yeast Saccharomyces cerevisiae. However, there is an important link between mitotic and meiotic recombination (covered in the forthcoming chapter by Hunter et al. 2015) and many of the functions are evolutionarily conserved. Here we will discuss several models that have been proposed to explain the mechanism of mitotic recombination, the genes and proteins involved in various pathways, the genetic and physical assays used to discover and study these genes, and the roles of many of these proteins inside the cell. PMID:25381364

  3. Radioimmunoassay for yeast killer toxin from Saccharomyces cerevisiae

    Siddiqui, F.A.; Bussey, H.

    1981-01-01

    A radioimmunoassay was developed for the K1 killer toxin from strain T158C/S14a of Saccharomyces cerevisiae. Iodine 125-labelled toxin was made to a specific activity of 100 μCi/mg of protein. Antibody to purified toxin was prepared in rabbits using toxin cross-linked to itself. These antibodies, partially purified by 50 percent ammonium sulfate precipitation and Sepharose CL-6B column chromatography, produced one precipitation band with killer toxin and bound 125 I-labelled toxin in a radioimmunoassay. The antibody preparation also bound with the toxins from another K1 killer, A364A, and three chromosomal superkiller mutants derived from it. (auth)

  4. Higher-order structure of Saccharomyces cerevisiae chromatin

    Lowary, P.T.; Widom, J.

    1989-01-01

    We have developed a method for partially purifying chromatin from Saccharomyces cerevisiae (baker's yeast) to a level suitable for studies of its higher-order folding. This has required the use of yeast strains that are free of the ubiquitous yeast killer virus. Results from dynamic light scattering, electron microscopy, and x-ray diffraction show that the yeast chromatin undergoes a cation-dependent folding into 30-nm filaments that resemble those characteristic of higher-cell chromatin; moreover, the packing of nucleosomes within the yeast 30-nm filaments is similar to that of higher cells. These results imply that yeast has a protein or protein domain that serves the role of the histone H 1 found in higher cells; physical and genetic studies of the yeast activity could help elucidate the structure and function of H 1. Images of the yeast 30-nm filaments can be used to test crossed-linker models for 30-nm filament structure

  5. Interaction among Saccharomyces cerevisiae pheromone receptors during endocytosis

    Chien-I Chang

    2014-03-01

    Full Text Available This study investigates endocytosis of Saccharomyces cerevisiae α-factor receptor and the role that receptor oligomerization plays in this process. α-factor receptor contains signal sequences in the cytoplasmic C-terminal domain that are essential for ligand-mediated endocytosis. In an endocytosis complementation assay, we found that oligomeric complexes of the receptor undergo ligand-mediated endocytosis when the α-factor binding site and the endocytosis signal sequences are located in different receptors. Both in vitro and in vivo assays suggested that ligand-induced conformational changes in one Ste2 subunit do not affect neighboring subunits. Therefore, recognition of the endocytosis signal sequence and recognition of the ligand-induced conformational change are likely to be two independent events.

  6. ACTIVITY OF SUPEROXIDE DISMUTASE ENZYME IN YEAST SACCHAROMYCES CEREVISIAE

    Blažena Lavová

    2014-02-01

    Full Text Available Reactive oxygen species (ROS with reactive nitrogen species (RNS are known to play dual role in biological systems, they can be harmful or beneficial to living systems. ROS can be important mediators of damage to cell structures, including proteins, lipids and nucleic acids termed as oxidative stress. The antioxidant enzymes protect the organism against the oxidative damage caused by active oxygen forms. The role of superoxide dismutase (SOD is to accelerate the dismutation of the toxic superoxide radical, produced during oxidative energy processes, to hydrogen peroxide and molecular oxygen. In this study, SOD activity of three yeast strains Saccharomyces cerevisiae was determined. It was found that SOD activity was the highest (23.7 U.mg-1 protein in strain 612 after 28 hours of cultivation. The lowest SOD activity from all tested strains was found after 56 hours of cultivation of strain Gyöng (0.7 U.mg-1 protein.

  7. Global mapping of DNA conformational flexibility on Saccharomyces cerevisiae.

    Giulia Menconi

    2015-04-01

    Full Text Available In this study we provide the first comprehensive map of DNA conformational flexibility in Saccharomyces cerevisiae complete genome. Flexibility plays a key role in DNA supercoiling and DNA/protein binding, regulating DNA transcription, replication or repair. Specific interest in flexibility analysis concerns its relationship with human genome instability. Enrichment in flexible sequences has been detected in unstable regions of human genome defined fragile sites, where genes map and carry frequent deletions and rearrangements in cancer. Flexible sequences have been suggested to be the determinants of fragile gene proneness to breakage; however, their actual role and properties remain elusive. Our in silico analysis carried out genome-wide via the StabFlex algorithm, shows the conserved presence of highly flexible regions in budding yeast genome as well as in genomes of other Saccharomyces sensu stricto species. Flexibile peaks in S. cerevisiae identify 175 ORFs mapping on their 3'UTR, a region affecting mRNA translation, localization and stability. (TAn repeats of different extension shape the central structure of peaks and co-localize with polyadenylation efficiency element (EE signals. ORFs with flexible peaks share common features. Transcripts are characterized by decreased half-life: this is considered peculiar of genes involved in regulatory systems with high turnover; consistently, their function affects biological processes such as cell cycle regulation or stress response. Our findings support the functional importance of flexibility peaks, suggesting that the flexible sequence may be derived by an expansion of canonical TAYRTA polyadenylation efficiency element. The flexible (TAn repeat amplification could be the outcome of an evolutionary neofunctionalization leading to a differential 3'-end processing and expression regulation in genes with peculiar function. Our study provides a new support to the functional role of flexibility in

  8. Global mapping of DNA conformational flexibility on Saccharomyces cerevisiae.

    Menconi, Giulia; Bedini, Andrea; Barale, Roberto; Sbrana, Isabella

    2015-04-01

    In this study we provide the first comprehensive map of DNA conformational flexibility in Saccharomyces cerevisiae complete genome. Flexibility plays a key role in DNA supercoiling and DNA/protein binding, regulating DNA transcription, replication or repair. Specific interest in flexibility analysis concerns its relationship with human genome instability. Enrichment in flexible sequences has been detected in unstable regions of human genome defined fragile sites, where genes map and carry frequent deletions and rearrangements in cancer. Flexible sequences have been suggested to be the determinants of fragile gene proneness to breakage; however, their actual role and properties remain elusive. Our in silico analysis carried out genome-wide via the StabFlex algorithm, shows the conserved presence of highly flexible regions in budding yeast genome as well as in genomes of other Saccharomyces sensu stricto species. Flexibile peaks in S. cerevisiae identify 175 ORFs mapping on their 3'UTR, a region affecting mRNA translation, localization and stability. (TA)n repeats of different extension shape the central structure of peaks and co-localize with polyadenylation efficiency element (EE) signals. ORFs with flexible peaks share common features. Transcripts are characterized by decreased half-life: this is considered peculiar of genes involved in regulatory systems with high turnover; consistently, their function affects biological processes such as cell cycle regulation or stress response. Our findings support the functional importance of flexibility peaks, suggesting that the flexible sequence may be derived by an expansion of canonical TAYRTA polyadenylation efficiency element. The flexible (TA)n repeat amplification could be the outcome of an evolutionary neofunctionalization leading to a differential 3'-end processing and expression regulation in genes with peculiar function. Our study provides a new support to the functional role of flexibility in genomes and a

  9. Copper Tolerance and Biosorption of Saccharomyces cerevisiae during Alcoholic Fermentation

    Liu, Ling-ling; Jia, Bo; Zhao, Fang; Huang, Wei-dong; Zhan, Ji-cheng

    2015-01-01

    At high levels, copper in grape mash can inhibit yeast activity and cause stuck fermentations. Wine yeast has limited tolerance of copper and can reduce copper levels in wine during fermentation. This study aimed to understand copper tolerance of wine yeast and establish the mechanism by which yeast decreases copper in the must during fermentation. Three strains of Saccharomyces cerevisiae (lab selected strain BH8 and industrial strains AWRI R2 and Freddo) and a simple model fermentation system containing 0 to 1.50 mM Cu2+ were used. ICP-AES determined Cu ion concentration in the must decreasing differently by strains and initial copper levels during fermentation. Fermentation performance was heavily inhibited under copper stress, paralleled a decrease in viable cell numbers. Strain BH8 showed higher copper-tolerance than strain AWRI R2 and higher adsorption than Freddo. Yeast cell surface depression and intracellular structure deformation after copper treatment were observed by scanning electron microscopy and transmission electron microscopy; electronic differential system detected higher surface Cu and no intracellular Cu on 1.50 mM copper treated yeast cells. It is most probably that surface adsorption dominated the biosorption process of Cu2+ for strain BH8, with saturation being accomplished in 24 h. This study demonstrated that Saccharomyces cerevisiae strain BH8 has good tolerance and adsorption of Cu, and reduces Cu2+ concentrations during fermentation in simple model system mainly through surface adsorption. The results indicate that the strain selected from China’s stress-tolerant wine grape is copper tolerant and can reduce copper in must when fermenting in a copper rich simple model system, and provided information for studies on mechanisms of heavy metal stress. PMID:26030864

  10. Copper Tolerance and Biosorption of Saccharomyces cerevisiae during Alcoholic Fermentation.

    Xiang-Yu Sun

    Full Text Available At high levels, copper in grape mash can inhibit yeast activity and cause stuck fermentations. Wine yeast has limited tolerance of copper and can reduce copper levels in wine during fermentation. This study aimed to understand copper tolerance of wine yeast and establish the mechanism by which yeast decreases copper in the must during fermentation. Three strains of Saccharomyces cerevisiae (lab selected strain BH8 and industrial strains AWRI R2 and Freddo and a simple model fermentation system containing 0 to 1.50 mM Cu2+ were used. ICP-AES determined Cu ion concentration in the must decreasing differently by strains and initial copper levels during fermentation. Fermentation performance was heavily inhibited under copper stress, paralleled a decrease in viable cell numbers. Strain BH8 showed higher copper-tolerance than strain AWRI R2 and higher adsorption than Freddo. Yeast cell surface depression and intracellular structure deformation after copper treatment were observed by scanning electron microscopy and transmission electron microscopy; electronic differential system detected higher surface Cu and no intracellular Cu on 1.50 mM copper treated yeast cells. It is most probably that surface adsorption dominated the biosorption process of Cu2+ for strain BH8, with saturation being accomplished in 24 h. This study demonstrated that Saccharomyces cerevisiae strain BH8 has good tolerance and adsorption of Cu, and reduces Cu2+ concentrations during fermentation in simple model system mainly through surface adsorption. The results indicate that the strain selected from China's stress-tolerant wine grape is copper tolerant and can reduce copper in must when fermenting in a copper rich simple model system, and provided information for studies on mechanisms of heavy metal stress.

  11. Repair of UV-damaged incoming plasmid DNA in Saccharomyces cerevisiae

    Keszenman-Pereyra, David

    1990-01-01

    A whole-cell transformation assay was used for the repair of UV-damaged plasma DNA in highly-transformable haploid strains of Saccharomyces cerevisiae having different repair capabilities. The experiments described demonstrate that three epistasis groups (Friedberg 1988) are involved in the repair of UV-incoming DNA and that the repair processes act less efficiently on incoming DNA than they do on chromosomal DNA. The implications of these findings for UV repair in Saccharomyces cerevisiae are discussed. (author)

  12. Progress in terpene synthesis strategies through engineering of Saccharomyces cerevisiae.

    Paramasivan, Kalaivani; Mutturi, Sarma

    2017-12-01

    Terpenes are natural products with a remarkable diversity in their chemical structures and they hold a significant market share commercially owing to their distinct applications. These potential molecules are usually derived from terrestrial plants, marine and microbial sources. In vitro production of terpenes using plant tissue culture and plant metabolic engineering, although receiving some success, the complexity in downstream processing because of the interference of phenolics and product commercialization due to regulations that are significant concerns. Industrial workhorses' viz., Escherichia coli and Saccharomyces cerevisiae have become microorganisms to produce non-native terpenes in order to address critical issues such as demand-supply imbalance, sustainability and commercial viability. S. cerevisiae enjoys several advantages for synthesizing non-native terpenes with the most significant being the compatibility for expressing cytochrome P450 enzymes from plant origin. Moreover, achievement of high titers such as 40 g/l of amorphadiene, a sesquiterpene, boosts commercial interest and encourages the researchers to envisage both molecular and process strategies for developing yeast cell factories to produce these compounds. This review contains a brief consideration of existing strategies to engineer S. cerevisiae toward the synthesis of terpene molecules. Some of the common targets for synthesis of terpenes in S. cerevisiae are as follows: overexpression of tHMG1, ERG20, upc2-1 in case of all classes of terpenes; repression of ERG9 by replacement of the native promoter with a repressive methionine promoter in case of mono-, di- and sesquiterpenes; overexpression of BTS1 in case of di- and tetraterpenes. Site-directed mutagenesis such as Upc2p (G888A) in case of all classes of terpenes, ERG20p (K197G) in case of monoterpenes, HMG2p (K6R) in case of mono-, di- and sesquiterpenes could be some generic targets. Efforts are made to consolidate various studies

  13. Redox balancing in recombinant strains of Saccharomyces cerevisiae

    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

  14. PRODUCTION, PROPERTIES AND APPLICATION OF SACCHAROMYCES CEREVISIAE VGSH-2 INULINASE

    G. P. Shuvaeva

    2014-01-01

    Full Text Available Summary. Experimental data on an acid and thermal inactivation of a high refined inulinase (2,1-β-D- fructanfructanohydrolase, KF 3.2.17, produced by the race of Saccharomyces cerevisiae VGSh-2 yeast are presented. The strain of S. cerevisiae VGSh-2 was produced by the method of the induced mutagenesis and deposited to the collection of pure cultures of the chair of biochemistry and biotechnology of Voronezh state university of engineering technologies. The cells of source culture (S. cerevisiae XII were affected step-by-step by the ultra-violet radiation (UFR and UFR in a complex with a chemical mutagen (etilenimine. The culture was grown up by the method of liquid-phase deep cultivation on a constant nutrient medium. Refining conditions for inulinase are sorted out. Activity of enzyme dependence on physical and chemical factors (рН and temperature is obtained and numerical values of the main kinetic constants – Km and Vmax are determined. The structure of enzyme molecule is studied by an infrared-spectroscopy method: the type and relative quantity of elements of secondary structure of protein are defined. Substrate binding groups of the active center of an inulinase are found. The comparative analysis of the ability to hydrolysis of inulin in several enzyme preparations from Jerusalem artichoke and to the subsequent their fermentation by the VGSh-2 and XI S. cerevisiae yeasts is carried out. Optimum conditions of enzyme hydrolysis of inulin are selected. Research of the fermentation process of starchcontaining raw materials by yeasts of VGSh-2 and XI races is done. It is established that the using of VGSh-2 S. cerevisiae yeast for a grain wort and the Jerusalem artichoke fermentation, allows to increase an extraction of ethyl alcohol comparing to control race, to improve its quality characteristics, and also allows to predict the using of new race in the food industry for production ethanol from grain raw materials and a fermentation of

  15. Switching the mode of sucrose utilization by Saccharomyces cerevisiae

    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

  16. Switching the mode of sucrose utilization by Saccharomyces cerevisiae.

    Badotti, Fernanda; Dário, Marcelo G; Alves, Sergio L; Cordioli, Maria Luiza A; Miletti, Luiz C; de Araujo, Pedro S; Stambuk, Boris U

    2008-02-27

    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. 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. 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 effectively reducing sucrose uptake by the yeast cells

  17. The evolution of gene expression QTL in Saccharomyces cerevisiae.

    James Ronald

    2007-08-01

    Full Text Available Understanding the evolutionary forces that influence patterns of gene expression variation will provide insights into the mechanisms of evolutionary change and the molecular basis of phenotypic diversity. To date, studies of gene expression evolution have primarily been made by analyzing how gene expression levels vary within and between species. However, the fundamental unit of heritable variation in transcript abundance is the underlying regulatory allele, and as a result it is necessary to understand gene expression evolution at the level of DNA sequence variation. Here we describe the evolutionary forces shaping patterns of genetic variation for 1206 cis-regulatory QTL identified in a cross between two divergent strains of Saccharomyces cerevisiae. We demonstrate that purifying selection against mildly deleterious alleles is the dominant force governing cis-regulatory evolution in S. cerevisiae and estimate the strength of selection. We also find that essential genes and genes with larger codon bias are subject to slightly stronger cis-regulatory constraint and that positive selection has played a role in the evolution of major trans-acting QTL.

  18. Biosynthesis and engineering of kaempferol in Saccharomyces cerevisiae.

    Duan, Lijin; Ding, Wentao; Liu, Xiaonan; Cheng, Xiaozhi; Cai, Jing; Hua, Erbing; Jiang, Huifeng

    2017-09-26

    Kaempferol is a flavonol with broad bioactivity of anti-oxidant, anti-cancer, anti-diabetic, anti-microbial, cardio-protective and anti-asthma. Microbial synthesis of kaempferol is a promising strategy because of the low content in primary plant source. In this study, the biosynthesis pathway of kaempferol was constructed in the budding yeast Saccharomyces cerevisiae to produce kaempferol de novo, and several biological measures were taken for high production. Firstly, a high efficient flavonol synthases (FLS) from Populus deltoides was introduced into the biosynthetic pathway of kaempferol. Secondly, a S. cerevisiae recombinant was constructed for de novo synthesis of kaempferol, which generated about 6.97 mg/L kaempferol from glucose. To further promote kaempferol production, the acetyl-CoA biosynthetic pathway was overexpressed and p-coumarate was supplied as substrate, which improved kaempferol titer by about 23 and 120%, respectively. Finally, a fed-batch process was developed for better kaempferol fermentation performance, and the production reached 66.29 mg/L in 40 h. The titer of kaempferol in our engineered yeast is 2.5 times of the highest reported titer. Our study provides a possible strategy to produce kaempferol using microbial cell factory.

  19. Lactose fermentation by engineered Saccharomyces cerevisiae capable of fermenting cellobiose.

    Liu, Jing-Jing; Zhang, Guo-Chang; Oh, Eun Joong; Pathanibul, Panchalee; Turner, Timothy L; Jin, Yong-Su

    2016-09-20

    Lactose is an inevitable byproduct of the dairy industry. In addition to cheese manufacturing, the growing Greek yogurt industry generates excess acid whey, which contains lactose. Therefore, rapid and efficient conversion of lactose to fuels and chemicals would be useful for recycling the otherwise harmful acid whey. Saccharomyces cerevisiae, a popular metabolic engineering host, cannot natively utilize lactose. However, we discovered that an engineered S. cerevisiae strain (EJ2) capable of fermenting cellobiose can also ferment lactose. This finding suggests that a cellobiose transporter (CDT-1) can transport lactose and a β-glucosidase (GH1-1) can hydrolyze lactose by acting as a β-galactosidase. While the lactose fermentation by the EJ2 strain was much slower than the cellobiose fermentation, a faster lactose-fermenting strain (EJ2e8) was obtained through serial subcultures on lactose. The EJ2e8 strain fermented lactose with a consumption rate of 2.16g/Lh. The improved lactose fermentation by the EJ2e8 strain was due to the increased copy number of cdt-1 and gh1-1 genes. Looking ahead, the EJ2e8 strain could be exploited for the production of other non-ethanol fuels and chemicals from lactose through further metabolic engineering. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Intracellular metabolite profiling of Saccharomyces cerevisiae evolved under furfural.

    Jung, Young Hoon; Kim, Sooah; Yang, Jungwoo; Seo, Jin-Ho; Kim, Kyoung Heon

    2017-03-01

    Furfural, one of the most common inhibitors in pre-treatment hydrolysates, reduces the cell growth and ethanol production of yeast. Evolutionary engineering has been used as a selection scheme to obtain yeast strains that exhibit furfural tolerance. However, the response of Saccharomyces cerevisiae to furfural at the metabolite level during evolution remains unknown. In this study, evolutionary engineering and metabolomic analyses were applied to determine the effects of furfural on yeasts and their metabolic response to continuous exposure to furfural. After 50 serial transfers of cultures in the presence of furfural, the evolved strains acquired the ability to stably manage its physiological status under the furfural stress. A total of 98 metabolites were identified, and their abundance profiles implied that yeast metabolism was globally regulated. Under the furfural stress, stress-protective molecules and cofactor-related mechanisms were mainly induced in the parental strain. However, during evolution under the furfural stress, S. cerevisiae underwent global metabolic allocations to quickly overcome the stress, particularly by maintaining higher levels of metabolites related to energy generation, cofactor regeneration and recovery from cellular damage. Mapping the mechanisms of furfural tolerance conferred by evolutionary engineering in the present study will be led to rational design of metabolically engineered yeasts. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

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

    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.

  2. Genomic Evolution of Saccharomyces cerevisiae under Chinese Rice Wine Fermentation

    Li, Yudong; Zhang, Weiping; Zheng, Daoqiong; Zhou, Zhan; Yu, Wenwen; Zhang, Lei; Feng, Lifang; Liang, Xinle; Guan, Wenjun; Zhou, Jingwen; Chen, Jian; Lin, Zhenguo

    2014-01-01

    Rice wine fermentation represents a unique environment for the evolution of the budding yeast, Saccharomyces cerevisiae. To understand how the selection pressure shaped the yeast genome and gene regulation, we determined the genome sequence and transcriptome of a S. cerevisiae strain YHJ7 isolated from Chinese rice wine (Huangjiu), a popular traditional alcoholic beverage in China. By comparing the genome of YHJ7 to the lab strain S288c, a Japanese sake strain K7, and a Chinese industrial bioethanol strain YJSH1, we identified many genomic sequence and structural variations in YHJ7, which are mainly located in subtelomeric regions, suggesting that these regions play an important role in genomic evolution between strains. In addition, our comparative transcriptome analysis between YHJ7 and S288c revealed a set of differentially expressed genes, including those involved in glucose transport (e.g., HXT2, HXT7) and oxidoredutase activity (e.g., AAD10, ADH7). Interestingly, many of these genomic and transcriptional variations are directly or indirectly associated with the adaptation of YHJ7 strain to its specific niches. Our molecular evolution analysis suggested that Japanese sake strains (K7/UC5) were derived from Chinese rice wine strains (YHJ7) at least approximately 2,300 years ago, providing the first molecular evidence elucidating the origin of Japanese sake strains. Our results depict interesting insights regarding the evolution of yeast during rice wine fermentation, and provided a valuable resource for genetic engineering to improve industrial wine-making strains. PMID:25212861

  3. PRODUKSI ETANOL DARI TETES TEBU OLEH Saccharomyces cerevisiae PEMBENTUK FLOK (NRRL – Y 265 (Ethanol Production from Cane Molasses by Flocculant Saccharomyces cerevisiae (NRRL – Y 265

    Agustin Krisna Wardani

    2013-08-01

    Full Text Available The potential use of sugar cane molasses by flocculant Saccharomyces cerevisiae in ethanol production was investigated. In order to minimize the negative effect of calcium on yeast growth, pretreated sugar cane molasses with dilute acid was performed. The influence of process parameters such as sugar concentration and inoculum concentration were evaluated for enhancing bioethanol production. Result showed that maximum ethanol concentration of 8,792% (b/v was obtained at the best condition of inoculum concentration 10% (v/v and sugar concentration 15% (b/v. Based on the experimental data, maximum yield of ethanol production of 65% was obtained. This result demonstrated the potential of molasses as promising biomass resources for ethanol production. Keywords: Ethanol, preteated cane molasses, flocculant Saccharomyces cerevisiae, fermentation   ABSTRAK Efisiensi produksi bioetanol diperoleh melalui ketepatan pemilihan jenis mikroorganisme, bahan baku, dan kontrol proses fermentasi. Alternatif proses untuk meminimalisasi biaya produksi etanol adalah dengan mengeliminasi tahap pemisahan sentrifugasi sel dari produk karena memerlukan biaya instalasi dan biaya perawatan yang tinggi. Proses sentrifugasi merupakan tahapan penting untuk memisahkan sel mikroba dari medium fermentasi pada produksi bioetanol. Untuk meminimalisir biaya produksi akibat proses tersebut digunakan inokulum Saccharomyces cerevisiae pembentuk flok dan tetes tebu sebagai sumber gula. Penelitian ini bertujuan untuk mendapatkan konsentrasi penambahan inokulum Saccharomyces cerevisiae pembentuk flok dan konsentrasi sumber gula dalam tetes tebu yang tepat dalam produksi etanol yang maksimum. Saccharomyces cerevisiae sebanyak 5%, 10%, dan 15% (v/v diinokulasikan pada medium tetes tebu hasil pretreatment dengan kandungan gula 15%, 20%, dan 25% (b/v pada pH 5. Fermentasi dilakukan pada suhu 30°C dan agitasi 100 rpm selama 72 jam. Etanol tertinggi didapat pada kondisi konsentrasi inokulum

  4. Exploring the Saccharomyces cerevisiae Volatile Metabolome: Indigenous versus Commercial Strains

    Alves, Zélia; Melo, André; Figueiredo, Ana Raquel; Coimbra, Manuel A.; Gomes, Ana C.; Rocha, Sílvia M.

    2015-01-01

    Winemaking is a highly industrialized process and a number of commercial Saccharomyces cerevisiae strains are used around the world, neglecting the diversity of native yeast strains that are responsible for the production of wines peculiar flavours. The aim of this study was to in-depth establish the S. cerevisiae volatile metabolome and to assess inter-strains variability. To fulfill this objective, two indigenous strains (BT2652 and BT2453 isolated from spontaneous fermentation of grapes collected in Bairrada Appellation, Portugal) and two commercial strains (CSc1 and CSc2) S. cerevisiae were analysed using a methodology based on advanced multidimensional gas chromatography (HS-SPME/GC×GC-ToFMS) tandem with multivariate analysis. A total of 257 volatile metabolites were identified, distributed over the chemical families of acetals, acids, alcohols, aldehydes, ketones, terpenic compounds, esters, ethers, furan-type compounds, hydrocarbons, pyrans, pyrazines and S-compounds. Some of these families are related with metabolic pathways of amino acid, carbohydrate and fatty acid metabolism as well as mono and sesquiterpenic biosynthesis. Principal Component Analysis (PCA) was used with a dataset comprising all variables (257 volatile components), and a distinction was observed between commercial and indigenous strains, which suggests inter-strains variability. In a second step, a subset containing esters and terpenic compounds (C10 and C15), metabolites of particular relevance to wine aroma, was also analysed using PCA. The terpenic and ester profiles express the strains variability and their potential contribution to the wine aromas, specially the BT2453, which produced the higher terpenic content. This research contributes to understand the metabolic diversity of indigenous wine microflora versus commercial strains and achieved knowledge that may be further exploited to produce wines with peculiar aroma properties. PMID:26600152

  5. High level secretion of cellobiohydrolases by Saccharomyces cerevisiae

    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.

  6. Exploring the northern limit of the distribution of Saccharomyces cerevisiae and Saccharomyces paradoxus in North America.

    Charron, Guillaume; Leducq, Jean-Baptiste; Bertin, Chloé; Dubé, Alexandre K; Landry, Christian R

    2014-03-01

    We examined the northern limit of Saccharomyces cerevisiae and Saccharomyces paradoxus in northeast America. We collected 876 natural samples at 29 sites and applied enrichment methods for the isolation of mesophilic yeasts. We uncovered a large diversity of yeasts, in some cases, associated with specific substrates. Sequencing of the ITS1, 5.8S and ITS2 loci allowed to assign 226 yeast strains at the species level, including 41 S. paradoxus strains. Our intensive sampling suggests that if present, S. cerevisiae is rare at these northern latitudes. Our sampling efforts spread across several months of the year revealed that successful sampling increases throughout the summer and diminishes significantly at the beginning of the fall. The data obtained on the ecological context of yeasts corroborate what was previously reported on Pichiaceae, Saccharomycodaceae, Debaryomycetaceae and Phaffomycetaceae yeast families. We identified 24 yeast isolates that could not be assigned to any known species and that may be of taxonomic, medical, or biotechnological importance. Our study reports new data on the taxonomic diversity of yeasts and new resources for studying the evolution and ecology of S. paradoxus. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  7. Physiological impact and context dependency of transcriptional responses : A chemostat study in Saccharomyces cerevisiae

    Tai, S.L.

    2007-01-01

    This thesis is a compilation of a four-year PhD project on bakers' yeast (Saccharomyces cerevisiae). Since the entire S. cerevisiae genome sequence became available in 1996, DNA-microarray analysis has become a popular high-information-density tool for analyzing gene expression in this important

  8. Invertase SUC2 Is the Key Hydrolase for Inulin Degradation in Saccharomyces cerevisiae

    Wang, Shi-An; Li, Fu-Li

    2013-01-01

    Specific Saccharomyces cerevisiae strains were recently found to be capable of efficiently utilizing inulin, but genetic mechanisms of inulin hydrolysis in yeast remain unknown. Here we report functional characteristics of invertase SUC2 from strain JZ1C and demonstrate that SUC2 is the key enzyme responsible for inulin metabolism in S. cerevisiae.

  9. Creation of a synthetic xylose-inducible promoter for Saccharomyces cerevisiae

    Saccharomyces cerevisiae is currently used to produce ethanol from glucose, but it cannot utilize five-carbon sugars contained in the hemicellulose component of biomass feedstocks. S. cerevisiae strains engineered for xylose fermentation have been made using constitutive promoters to express the req...

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

    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 framework...

  11. Development of Efficient Xylose Fermentation in Saccharomyces cerevisiae : Xylose Isomerase as a Key Component

    Van Maris, A.J.A.; Winkler, A.A.; Kuyper, M.; De Laat, W.T.; Van Dijken, J.P.; Pronk, J.T.

    2007-01-01

    Metabolic engineering of Saccharomyces cerevisiae for ethanol production from d-xylose, an abundant sugar in plant biomass hydrolysates, has been pursued vigorously for the past 15 years. Whereas wild-type S. cerevisiae cannot ferment d-xylose, the ketoisomer d-xylulose can be metabolised slowly.

  12. ESCRT-independent budding of HIV-1 gag virus-like particles from Saccharomyces cerevisiae spheroplasts.

    Andrew P Norgan

    Full Text Available Heterologous expression of HIV-1 Gag in a variety of host cells results in its packaging into virus-like particles (VLPs that are subsequently released into the extracellular milieu. This phenomenon represents a useful tool for probing cellular factors required for viral budding and has contributed to the discovery of roles for ubiquitin ligases and the endosomal sorting complexes required for transport (ESCRTs in viral budding. These factors are highly conserved throughout eukaryotes and have been studied extensively in the yeast Saccharomyces cerevisiae, a model eukaryote previously utilized as a host for the production of VLPs. We used heterologous expression of HIV Gag in yeast spheroplasts to examine the role of ESCRTs and associated factors (Rsp5, a HECT ubiquitin ligase of the Nedd4 family; Bro1, a homolog of Alix; and Vps4, the AAA-ATPase required for ESCRT function in all contexts/organisms investigated in the generation of VLPs. Our data reveal: 1 characterized Gag-ESCRT interaction motifs (late domains are not required for VLP budding, 2 loss of function alleles of the essential HECT ubiquitin ligase Rsp5 do not display defects in VLP formation, and 3 ESCRT function is not required for VLP formation from spheroplasts. These results suggest that the egress of HIV Gag from yeast cells is distinct from the most commonly described mode of exit from mammalian cells, instead mimicking ESCRT-independent VLP formation observed in a subset of mammalian cells. As such, budding of Gag from yeast cells appears to represent ESCRT-independent budding relevant to viral replication in at least some situations. Thus the myriad of genetic and biochemical tools available in the yeast system may be of utility in the study of this aspect of viral budding.

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

    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. Keywords: CRISPR–Cas9, Genome editing, Industrial yeast, Biorefineries, Chemical production

  14. K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae.

    Orentaite, Irma; Poranen, Minna M; Oksanen, Hanna M; Daugelavicius, Rimantas; Bamford, Dennis H

    2016-03-01

    Saccharomyces cerevisiae cells produce killer toxins, such as K1, K2 and K28, that can modulate the growth of other yeasts giving advantage for the killer strains. Here we focused on the physiological changes induced by K2 toxin on a non-toxin-producing yeast strain as well as K1, K2 and K28 killer strains. Potentiometric measurements were adjusted to observe that K2 toxin immediately acts on the sensitive cells leading to membrane permeability. This correlated with reduced respiration activity, lowered intracellular ATP content and decrease in cell viability. However, we did not detect any significant ATP leakage from the cells treated by killer toxin K2. Strains producing heterologous toxins K1 and K28 were less sensitive to K2 than the non-toxin producing one suggesting partial cross-protection between the different killer systems. This phenomenon may be connected to the observed differences in respiratory activities of the killer strains and the non-toxin-producing strain at low pH. This might also have practical consequences in wine industry; both as beneficial ones in controlling contaminating yeasts and non-beneficial ones causing sluggish fermentation. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. Nonhomologous Synapsis and Reduced Crossing over in a Heterozygous Paracentric Inversion in Saccharomyces Cerevisiae

    Dresser, M. E.; Ewing, D. J.; Harwell, S. N.; Coody, D.; Conrad, M. N.

    1994-01-01

    Homologous chromosome synapsis (``homosynapsis'') and crossing over are well-conserved aspects of meiotic chromosome behavior. The long-standing assumption that these two processes are causally related has been challenged recently by observations in Saccharomyces cerevisiae of significant levels of crossing over (1) between small sequences at nonhomologous locations and (2) in mutants where synapsis is abnormal or absent. In order to avoid problems of local sequence effects and of mutation pleiotropy, we have perturbed synapsis by making a set of isogenic strains that are heterozygous and homozygous for a large chromosomal paracentric inversion covering a well marked genetic interval and then measured recombination. We find that reciprocal recombination in the marked interval in heterozygotes is reduced variably across the interval, on average to ~55% of that in the homozygotes, and that positive interference still modulates crossing over. Cytologically, stable synapsis across the interval is apparently heterologous rather than homologous, consistent with the interpretation that stable homosynapsis is required to initiate or consummate a large fraction of the crossing over observed in wild-type strains. When crossing over does occur in heterozygotes, dicentric and acentric chromosomes are formed and can be visualized and quantitated on blots though not demonstrated in viable spores. We find that there is no loss of dicentric chromosomes during the two meiotic divisions and that the acentric chromosome is recovered at only 1/3 to 1/2 of the expected level. PMID:7851761

  16. Fumaric acid production in Saccharomyces cerevisiae by simultaneous use of oxidative and reductive routes.

    Xu, Guoqiang; Chen, Xiulai; Liu, Liming; Jiang, Linghuo

    2013-11-01

    In this study, the simultaneous use of reductive and oxidative routes to produce fumaric acid was explored. The strain FMME003 (Saccharomyces cerevisiae CEN.PK2-1CΔTHI2) exhibited capability to accumulate pyruvate and was used for fumaric acid production. The fum1 mutant FMME004 could produce fumaric acid via oxidative route, but the introduction of reductive route derived from Rhizopus oryzae NRRL 1526 led to lower fumaric acid production. Analysis of the key factors associated with fumaric acid production revealed that pyruvate carboxylase had a low degree of control over the carbon flow to malic acid. The fumaric acid titer was improved dramatically when the heterologous gene RoPYC was overexpressed and 32 μg/L of biotin was added. Furthermore, under the optimal carbon/nitrogen ratio, the engineered strain FMME004-6 could produce up to 5.64 ± 0.16 g/L of fumaric acid. These results demonstrated that the proposed fermentative method is efficient for fumaric acid production. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Alleviating Redox Imbalance Enhances 7-Dehydrocholesterol Production in Engineered Saccharomyces cerevisiae.

    Wan Su

    Full Text Available Maintaining redox balance is critical for the production of heterologous secondary metabolites, whereas on various occasions the native cofactor balance does not match the needs in engineered microorganisms. In this study, 7-dehydrocholesterol (7-DHC, a crucial precursor of vitamin D3 biosynthesis pathway was constructed in Saccharomyces cerevisiae BY4742 with endogenous ergosterol synthesis pathway blocked by knocking out the erg5 gene (encoding C-22 desaturase. The deletion of erg5 led to redox imbalance with higher ratio of cytosolic free NADH/NAD+ and more glycerol and ethanol accumulation. To alleviate the redox imbalance, a water-forming NADH oxidase (NOX and an alternative oxidase (AOX1 were employed in our system based on cofactor regeneration strategy. Consequently, the production of 7-dehydrocholesterol was increased by 74.4% in shake flask culture. In the meanwhile, the ratio of free NADH/NAD+ and the concentration of glycerol and ethanol were reduced by 78.0%, 50.7% and 7.9% respectively. In a 5-L bioreactor, the optimal production of 7-DHC reached 44.49(±9.63 mg/L. This study provides a reference to increase the production of some desired compounds that are restricted by redox imbalance.

  18. De Novo Biosynthesis of Vanillin in Fission Yeast (Schizosaccharomyces pombe) and Baker's Yeast (Saccharomyces cerevisiae) ▿

    Hansen, Esben H.; Møller, Birger Lindberg; Kock, Gertrud R.; Bünner, Camilla M.; Kristensen, Charlotte; Jensen, Ole R.; Okkels, Finn T.; Olsen, Carl E.; Motawia, Mohammed S.; Hansen, Jørgen

    2009-01-01

    Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast, Saccharomyces cerevisiae. Productivities were 65 and 45 mg/liter, after introduction of three and four heterologous genes, respectively. The engineered pathways involve incorporation of 3-dehydroshikimate dehydratase from the dung mold Podospora pauciseta, an aromatic carboxylic acid reductase (ACAR) from a bacterium of the Nocardia genus, and an O-methyltransferase from Homo sapiens. In S. cerevisiae, the ACAR enzyme required activation by phosphopantetheinylation, and this was achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol was achieved by knockout of the host alcohol dehydrogenase ADH6. In S. pombe, the biosynthesis was further improved by introduction of an Arabidopsis thaliana family 1 UDP-glycosyltransferase, converting vanillin into vanillin β-d-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts. These de novo pathways represent the first examples of one-cell microbial generation of these valuable compounds from glucose. S. pombe yeast has not previously been metabolically engineered to produce any valuable, industrially scalable, white biotech commodity. PMID:19286778

  19. Directed evolution of xylose isomerase for improved xylose catabolism and fermentation in the yeast Saccharomyces cerevisiae.

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

    2012-08-01

    The heterologous expression of a highly functional xylose isomerase pathway in Saccharomyces cerevisiae would have significant advantages for ethanol yield, since the pathway bypasses cofactor requirements found in the traditionally used oxidoreductase pathways. However, nearly all reported xylose isomerase-based pathways in S. cerevisiae suffer from poor ethanol productivity, low xylose consumption rates, and poor cell growth compared with an oxidoreductase pathway and, additionally, often require adaptive strain evolution. Here, we report on the directed evolution of the Piromyces sp. xylose isomerase (encoded by xylA) for use in yeast. After three rounds of mutagenesis and growth-based screening, we isolated a variant containing six mutations (E15D, E114G, E129D, T142S, A177T, and V433I) that exhibited a 77% increase in enzymatic activity. When expressed in a minimally engineered yeast host containing a gre3 knockout and tal1 and XKS1 overexpression, the strain expressing this mutant enzyme improved its aerobic growth rate by 61-fold and both ethanol production and xylose consumption rates by nearly 8-fold. Moreover, the mutant enzyme enabled ethanol production by these yeasts under oxygen-limited fermentation conditions, unlike the wild-type enzyme. Under microaerobic conditions, the ethanol production rates of the strain expressing the mutant xylose isomerase were considerably higher than previously reported values for yeast harboring a xylose isomerase pathway and were also comparable to those of the strains harboring an oxidoreductase pathway. Consequently, this study shows the potential to evolve a xylose isomerase pathway for more efficient xylose utilization.

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

    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.

  1. Anaerobic α-Amylase Production and Secretion with Fumarate as the Final Electron Acceptor in Saccharomyces cerevisiae

    Liu, Zihe; Österlund, Tobias; Hou, Jin

    2013-01-01

    In this study, we focus on production of heterologous α-amylase in the yeast Saccharomyces cerevisiae under anaerobic conditions. We compare the metabolic fluxes and transcriptional regulation under aerobic and anaerobic conditions, with the objective of identifying the final electron acceptor...... reticulum are transferred to fumarate as the final electron acceptor. This model is supported by findings that the addition of fumarate under anaerobic (but not aerobic) conditions improves cell growth, specifically in the α-amylase-producing strain, in which it is not used as a carbon source. Our results...... provide a model for the molecular mechanism of anaerobic protein secretion using fumarate as the final electron acceptor, which may allow for further engineering of yeast for improved protein secretion under anaerobic growth conditions....

  2. Reconstruction of cytosolic fumaric acid biosynthetic pathways in Saccharomyces cerevisiae

    Xu Guoqiang

    2012-02-01

    Full Text Available Abstract Background Fumaric acid is a commercially important component of foodstuffs, pharmaceuticals and industrial materials, yet the current methods of production are unsustainable and ecologically destructive. Results In this study, the fumarate biosynthetic pathway involving reductive reactions of the tricarboxylic acid cycle was exogenously introduced in S. cerevisiae by a series of simple genetic modifications. First, the Rhizopus oryzae genes for malate dehydrogenase (RoMDH and fumarase (RoFUM1 were heterologously expressed. Then, expression of the endogenous pyruvate carboxylase (PYC2 was up-regulated. The resultant yeast strain, FMME-001 ↑PYC2 + ↑RoMDH, was capable of producing significantly higher yields of fumarate in the glucose medium (3.18 ± 0.15 g liter-1 than the control strain FMME-001 empty vector. Conclusions The results presented here provide a novel strategy for fumarate biosynthesis, which represents an important advancement in producing high yields of fumarate in a sustainable and ecologically-friendly manner.

  3. Genomic evolution of Saccharomyces cerevisiae under Chinese rice wine fermentation.

    Li, Yudong; Zhang, Weiping; Zheng, Daoqiong; Zhou, Zhan; Yu, Wenwen; Zhang, Lei; Feng, Lifang; Liang, Xinle; Guan, Wenjun; Zhou, Jingwen; Chen, Jian; Lin, Zhenguo

    2014-09-10

    Rice wine fermentation represents a unique environment for the evolution of the budding yeast, Saccharomyces cerevisiae. To understand how the selection pressure shaped the yeast genome and gene regulation, we determined the genome sequence and transcriptome of a S. cerevisiae strain YHJ7 isolated from Chinese rice wine (Huangjiu), a popular traditional alcoholic beverage in China. By comparing the genome of YHJ7 to the lab strain S288c, a Japanese sake strain K7, and a Chinese industrial bioethanol strain YJSH1, we identified many genomic sequence and structural variations in YHJ7, which are mainly located in subtelomeric regions, suggesting that these regions play an important role in genomic evolution between strains. In addition, our comparative transcriptome analysis between YHJ7 and S288c revealed a set of differentially expressed genes, including those involved in glucose transport (e.g., HXT2, HXT7) and oxidoredutase activity (e.g., AAD10, ADH7). Interestingly, many of these genomic and transcriptional variations are directly or indirectly associated with the adaptation of YHJ7 strain to its specific niches. Our molecular evolution analysis suggested that Japanese sake strains (K7/UC5) were derived from Chinese rice wine strains (YHJ7) at least approximately 2,300 years ago, providing the first molecular evidence elucidating the origin of Japanese sake strains. Our results depict interesting insights regarding the evolution of yeast during rice wine fermentation, and provided a valuable resource for genetic engineering to improve industrial wine-making strains. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  4. Direct conversion of starch to ethanol using recombınant Saccharomyces cerevisiae containing glucoamylase gene

    Purkan, P.; Baktir, A.; Puspaningsih, N. N. T.; Ni'mah, M.

    2017-09-01

    Saccharomyces cerevisiae is known for its high fermentative capacity, high ethanol yield and its high ethanol tolerance. The yeast is inability converting starch (relatively inexpensive substrate) into biofuel ethanol. Insertion of glucoamylase gene in yeast cell of Saccharomyces cerevisiae had been done to increase the yeast function in ethanol fermentation from starch. Transformation of yeast of S. cerevisiae with recombinant plasmid yEP-GLO1 carrying gene encoding glucoamylase (GLO1) produced the recombinant yeast which enable to degrade starch. Optimizing of bioconversion process of starch into ethanol by the yeast of recombinant Saccharomyces cerevisiae [yEP-GLO1] had been also done. Starch concentration which could be digested by recombinant yeast of S. cerevisiae [yEP-GLO1] was 10% (w/v). Bioconversion of starch having concentration 10% (b/v) using recombinant yeast of S. cerevisiae BY5207 [yEP-GLO1] could result ethanol as 20% (v/v) to alcoholmeter and 19,5% (v/v) to gas of chromatography. Otherwise, using recombinant yeast S. cerevisiae S. cerevisiae AS3324 [yEP-GLO1] resulted ethanol as 17% (v/v) to alcoholmeter and 17,5% (v/v) to gas of chromatography. The highest ethanol in starch bioconversion using both recombinant yeasts BY5207 and AS3324 could be resulted on 144 hours of fermentation time as well as in pH 5.

  5. A MultiSite GatewayTM vector set for the functional analysis of genes in the model Saccharomyces cerevisiae

    Nagels Durand Astrid

    2012-09-01

    Full Text Available Abstract Background Recombinatorial cloning using the GatewayTM technology has been the method of choice for high-throughput omics projects, resulting in the availability of entire ORFeomes in GatewayTM compatible vectors. The MultiSite GatewayTM system allows combining multiple genetic fragments such as promoter, ORF and epitope tag in one single reaction. To date, this technology has not been accessible in the yeast Saccharomyces cerevisiae, one of the most widely used experimental systems in molecular biology, due to the lack of appropriate destination vectors. Results Here, we present a set of three-fragment MultiSite GatewayTM destination vectors that have been developed for gene expression in S. cerevisiae and that allow the assembly of any promoter, open reading frame, epitope tag arrangement in combination with any of four auxotrophic markers and three distinct replication mechanisms. As an example of its applicability, we used yeast three-hybrid to provide evidence for the assembly of a ternary complex of plant proteins involved in jasmonate signalling and consisting of the JAZ, NINJA and TOPLESS proteins. Conclusion Our vectors make MultiSite GatewayTM cloning accessible in S. cerevisiae and implement a fast and versatile cloning method for the high-throughput functional analysis of (heterologous proteins in one of the most widely used model organisms for molecular biology research.

  6. Introducing a new breed of wine yeast: interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast and Saccharomyces mikatae.

    Bellon, Jennifer R; Schmid, Frank; Capone, Dimitra L; Dunn, Barbara L; Chambers, Paul J

    2013-01-01

    Interspecific hybrids are commonplace in agriculture and horticulture; bread wheat and grapefruit are but two examples. The benefits derived from interspecific hybridisation include the potential of generating advantageous transgressive phenotypes. This paper describes the generation of a new breed of wine yeast by interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast strain and Saccharomyces mikatae, a species hitherto not associated with industrial fermentation environs. While commercially available wine yeast strains provide consistent and reliable fermentations, wines produced using single inocula are thought to lack the sensory complexity and rounded palate structure obtained from spontaneous fermentations. In contrast, interspecific yeast hybrids have the potential to deliver increased complexity to wine sensory properties and alternative wine styles through the formation of novel, and wider ranging, yeast volatile fermentation metabolite profiles, whilst maintaining the robustness of the wine yeast parent. Screening of newly generated hybrids from a cross between a S. cerevisiae wine yeast and S. mikatae (closely-related but ecologically distant members of the Saccharomyces sensu stricto clade), has identified progeny with robust fermentation properties and winemaking potential. Chemical analysis showed that, relative to the S. cerevisiae wine yeast parent, hybrids produced wines with different concentrations of volatile metabolites that are known to contribute to wine flavour and aroma, including flavour compounds associated with non-Saccharomyces species. The new S. cerevisiae x S. mikatae hybrids have the potential to produce complex wines akin to products of spontaneous fermentation while giving winemakers the safeguard of an inoculated ferment.

  7. Introducing a new breed of wine yeast: interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast and Saccharomyces mikatae.

    Jennifer R Bellon

    Full Text Available Interspecific hybrids are commonplace in agriculture and horticulture; bread wheat and grapefruit are but two examples. The benefits derived from interspecific hybridisation include the potential of generating advantageous transgressive phenotypes. This paper describes the generation of a new breed of wine yeast by interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast strain and Saccharomyces mikatae, a species hitherto not associated with industrial fermentation environs. While commercially available wine yeast strains provide consistent and reliable fermentations, wines produced using single inocula are thought to lack the sensory complexity and rounded palate structure obtained from spontaneous fermentations. In contrast, interspecific yeast hybrids have the potential to deliver increased complexity to wine sensory properties and alternative wine styles through the formation of novel, and wider ranging, yeast volatile fermentation metabolite profiles, whilst maintaining the robustness of the wine yeast parent. Screening of newly generated hybrids from a cross between a S. cerevisiae wine yeast and S. mikatae (closely-related but ecologically distant members of the Saccharomyces sensu stricto clade, has identified progeny with robust fermentation properties and winemaking potential. Chemical analysis showed that, relative to the S. cerevisiae wine yeast parent, hybrids produced wines with different concentrations of volatile metabolites that are known to contribute to wine flavour and aroma, including flavour compounds associated with non-Saccharomyces species. The new S. cerevisiae x S. mikatae hybrids have the potential to produce complex wines akin to products of spontaneous fermentation while giving winemakers the safeguard of an inoculated ferment.

  8. Introducing a New Breed of Wine Yeast: Interspecific Hybridisation between a Commercial Saccharomyces cerevisiae Wine Yeast and Saccharomyces mikatae

    Bellon, Jennifer R.; Schmid, Frank; Capone, Dimitra L.; Dunn, Barbara L.; Chambers, Paul J.

    2013-01-01

    Interspecific hybrids are commonplace in agriculture and horticulture; bread wheat and grapefruit are but two examples. The benefits derived from interspecific hybridisation include the potential of generating advantageous transgressive phenotypes. This paper describes the generation of a new breed of wine yeast by interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast strain and Saccharomyces mikatae, a species hitherto not associated with industrial fermentation environs. While commercially available wine yeast strains provide consistent and reliable fermentations, wines produced using single inocula are thought to lack the sensory complexity and rounded palate structure obtained from spontaneous fermentations. In contrast, interspecific yeast hybrids have the potential to deliver increased complexity to wine sensory properties and alternative wine styles through the formation of novel, and wider ranging, yeast volatile fermentation metabolite profiles, whilst maintaining the robustness of the wine yeast parent. Screening of newly generated hybrids from a cross between a S. cerevisiae wine yeast and S. mikatae (closely-related but ecologically distant members of the Saccharomyces sensu stricto clade), has identified progeny with robust fermentation properties and winemaking potential. Chemical analysis showed that, relative to the S. cerevisiae wine yeast parent, hybrids produced wines with different concentrations of volatile metabolites that are known to contribute to wine flavour and aroma, including flavour compounds associated with non-Saccharomyces species. The new S. cerevisiae x S. mikatae hybrids have the potential to produce complex wines akin to products of spontaneous fermentation while giving winemakers the safeguard of an inoculated ferment. PMID:23614011

  9. Functional co-operation between the nuclei of Saccharomyces cerevisiae and mitochondria from other yeast species

    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....... italicus, S, oviformis, S. capensis and S. chevalieri) exhibited complete compatibility with S. cerevisiae nuclei. The closely related S. douglasii mitochondrial genome could also partially restore respiration-deficiency in rho (0) S. cerevisiae, whereas mitochondrial genomes from phylogenetically less...

  10. Compositions and methods for modeling Saccharomyces cerevisiae metabolism

    2012-01-01

    The invention provides an in silica model for determining a S. cerevisiae physiological function. The model includes a data structure relating a plurality of S. cerevisiae reactants to a plurality of S. cerevisiae reactions, a constraint set for the plurality of S. cerevisiae reactions, and comma...

  11. Efficient Sporulation of Saccharomyces cerevisiae in a 96 Multiwell Format.

    Paulissen, Scott M; Huang, Linda S

    2016-09-17

    During times of nutritional stress, Saccharomyces cerevisiae undergoes gametogenesis, known as sporulation. Diploid yeast cells that are starved for nitrogen and carbon will initiate the sporulation process. The process of sporulation includes meiosis followed by spore formation, where the haploid nuclei are packaged into environmentally resistant spores. We have developed methods for the efficient sporulation of budding yeast in 96 multiwell plates, to increase the throughput of screening yeast cells for sporulation phenotypes. These methods are compatible with screening with yeast containing plasmids requiring nutritional selection, when appropriate minimal media is used, or with screening yeast with genomic alterations, when a rich presporulation regimen is used. We find that for this method, aeration during sporulation is critical for spore formation, and have devised techniques to ensure sufficient aeration that are compatible with the 96 multiwell plate format. Although these methods do not achieve the typical ~80% level of sporulation that can be achieved in large-volume flask based experiments, these methods will reliably achieve about 50-60% level of sporulation in small-volume multiwell plates.

  12. Dynamics of the Saccharomyces cerevisiae Transcriptome during Bread Dough Fermentation

    Aslankoohi, Elham; Zhu, Bo; Rezaei, Mohammad Naser; Voordeckers, Karin; De Maeyer, Dries; Marchal, Kathleen; Dornez, Emmie

    2013-01-01

    The behavior of yeast cells during industrial processes such as the production of beer, wine, and bioethanol has been extensively studied. In contrast, our knowledge about yeast physiology during solid-state processes, such as bread dough, cheese, or cocoa fermentation, remains limited. We investigated changes in the transcriptomes of three genetically distinct Saccharomyces cerevisiae strains during bread dough fermentation. Our results show that regardless of the genetic background, all three strains exhibit similar changes in expression patterns. At the onset of fermentation, expression of glucose-regulated genes changes dramatically, and the osmotic stress response is activated. The middle fermentation phase is characterized by the induction of genes involved in amino acid metabolism. Finally, at the latest time point, cells suffer from nutrient depletion and activate pathways associated with starvation and stress responses. Further analysis shows that genes regulated by the high-osmolarity glycerol (HOG) pathway, the major pathway involved in the response to osmotic stress and glycerol homeostasis, are among the most differentially expressed genes at the onset of fermentation. More importantly, deletion of HOG1 and other genes of this pathway significantly reduces the fermentation capacity. Together, our results demonstrate that cells embedded in a solid matrix such as bread dough suffer severe osmotic stress and that a proper induction of the HOG pathway is critical for optimal fermentation. PMID:24056467

  13. Saccharomyces cerevisiae Fermentation Effects on Pollen: Archaeological Implications

    Crystal A. Dozier

    2016-03-01

    Full Text Available Pollen is the reproductive agent of flowering plants; palynology is utilized by archaeologists because sporopollenin, a major component in the exine of pollen grains, is resistant to decay and morphologically distinctive. Wine, beer, and mead have been identified in the archaeological record by palynological assessment due to indicator species or due to a pollen profile similar to that recovered from honey, a common source of sugar in a variety of fermented beverages. While most palynologists have assumed that pollen grains are resistant to alcoholic fermentation, a recent study in food science implies that pollen is a yeast nutrient because pollen-enriched meads produce more alcohol. The experiment presented here explores the potential distortion of the pollen record through fermentation by brewing a traditional, pollen-rich mead with Saccharomyces cerevisiae. In this experiment, the pollen grains did not undergo any discernible morphological changes nor were distorted in the pollen profile. Any nutrition that the yeast garners from the pollen therefore leaves sporopollenin intact. These results support palynological research on residues of alcoholic beverages and confirms that the fermentation process does not distort the pollen profile of the original substance. The paper concludes with the potential and limits of palynological study to assess fermentation within the archaeological record.

  14. Global gene response in Saccharomyces cerevisiae exposed to silver nanoparticles.

    Niazi, Javed H; Sang, Byoung-In; Kim, Yeon Seok; Gu, Man Bock

    2011-08-01

    Silver nanoparticles (AgNPs), exhibiting a broad size range and morphologies with highly reactive facets, which are widely applicable in real-life but not fully verified for biosafety and ecotoxicity, were subjected to report transcriptome profile in yeast Saccharomyces cerevisiae. A large number of genes accounted for ∼3% and ∼5% of the genome affected by AgNPs and Ag-ions, respectively. Principal component and cluster analysis suggest that the different physical forms of Ag were the major cause in differential expression profile. Among 90 genes affected by both AgNPs and Ag-ions, metalloprotein mediating high resistance to copper (CUP1-1 and CUP1-2) were strongly induced by AgNPs (∼45-folds) and Ag-ions (∼22-folds), respectively. A total of 17 genes, responsive to chemical stimuli, stress, and transport processes, were differentially induced by AgNPs. The differential expression was also seen with Ag-ions that affected 73 up- and 161 down-regulating genes, and most of these were involved in ion transport and homeostasis. This study provides new information on the knowledge for impact of nanoparticles on living microorganisms that can be extended to other nanoparticles.

  15. Bread, beer and wine: Saccharomyces cerevisiae diversity reflects human history.

    Legras, Jean-Luc; Merdinoglu, Didier; Cornuet, Jean-Marie; Karst, Francis

    2007-05-01

    Fermented beverages and foods have played a significant role in most societies worldwide for millennia. To better understand how the yeast species Saccharomyces cerevisiae, the main fermenting agent, evolved along this historical and expansion process, we analysed the genetic diversity among 651 strains from 56 different geographical origins, worldwide. Their genotyping at 12 microsatellite loci revealed 575 distinct genotypes organized in subgroups of yeast types, i.e. bread, beer, wine, sake. Some of these groups presented unexpected relatedness: Bread strains displayed a combination of alleles intermediate between beer and wine strains, and strains used for rice wine and sake were most closely related to beer and bread strains. However, up to 28% of genetic diversity between these technological groups was associated with geographical differences which suggests local domestications. Focusing on wine yeasts, a group of Lebanese strains were basal in an F(ST) tree, suggesting a Mesopotamia-based origin of most wine strains. In Europe, migration of wine strains occurred through the Danube Valley, and around the Mediterranean Sea. An approximate Bayesian computation approach suggested a postglacial divergence (most probable period 10,000-12,000 bp). As our results suggest intimate association between man and wine yeast across centuries, we hypothesize that yeast followed man and vine migrations as a commensal member of grapevine flora.

  16. Dynamics of the Saccharomyces cerevisiae transcriptome during bread dough fermentation.

    Aslankoohi, Elham; Zhu, Bo; Rezaei, Mohammad Naser; Voordeckers, Karin; De Maeyer, Dries; Marchal, Kathleen; Dornez, Emmie; Courtin, Christophe M; Verstrepen, Kevin J

    2013-12-01

    The behavior of yeast cells during industrial processes such as the production of beer, wine, and bioethanol has been extensively studied. In contrast, our knowledge about yeast physiology during solid-state processes, such as bread dough, cheese, or cocoa fermentation, remains limited. We investigated changes in the transcriptomes of three genetically distinct Saccharomyces cerevisiae strains during bread dough fermentation. Our results show that regardless of the genetic background, all three strains exhibit similar changes in expression patterns. At the onset of fermentation, expression of glucose-regulated genes changes dramatically, and the osmotic stress response is activated. The middle fermentation phase is characterized by the induction of genes involved in amino acid metabolism. Finally, at the latest time point, cells suffer from nutrient depletion and activate pathways associated with starvation and stress responses. Further analysis shows that genes regulated by the high-osmolarity glycerol (HOG) pathway, the major pathway involved in the response to osmotic stress and glycerol homeostasis, are among the most differentially expressed genes at the onset of fermentation. More importantly, deletion of HOG1 and other genes of this pathway significantly reduces the fermentation capacity. Together, our results demonstrate that cells embedded in a solid matrix such as bread dough suffer severe osmotic stress and that a proper induction of the HOG pathway is critical for optimal fermentation.

  17. Nuclear mitochondrial DNA activates replication in Saccharomyces cerevisiae.

    Laurent Chatre

    Full Text Available The nuclear genome of eukaryotes is colonized by DNA fragments of mitochondrial origin, called NUMTs. These insertions have been associated with a variety of germ-line diseases in humans. The significance of this uptake of potentially dangerous sequences into the nuclear genome is unclear. Here we provide functional evidence that sequences of mitochondrial origin promote nuclear DNA replication in Saccharomyces cerevisiae. We show that NUMTs are rich in key autonomously replicating sequence (ARS consensus motifs, whose mutation results in the reduction or loss of DNA replication activity. Furthermore, 2D-gel analysis of the mrc1 mutant exposed to hydroxyurea shows that several NUMTs function as late chromosomal origins. We also show that NUMTs located close to or within ARS provide key sequence elements for replication. Thus NUMTs can act as independent origins, when inserted in an appropriate genomic context or affect the efficiency of pre-existing origins. These findings show that migratory mitochondrial DNAs can impact on the replication of the nuclear region they are inserted in.

  18. De novo biosynthesis of anthocyanins in Saccharomyces cerevisiae.

    Eichenberger, Michael; Hansson, Anders; Fischer, David; Dürr, Lara; Naesby, Michael

    2018-06-01

    Anthocyanins (ACNs) are plant secondary metabolites responsible for most of the red, purple and blue colors of flowers, fruits and vegetables. They are increasingly used in the food and beverage industry as natural alternative to artificial colorants. Production of these compounds by fermentation of microorganisms would provide an attractive alternative. In this study, Saccharomyces cerevisiae was engineered for de novo production of the three basic anthocyanins, as well as the three main trans-flavan-3-ols. Enzymes from different plant sources were screened and efficient variants found for most steps of the biosynthetic pathway. However, the anthocyanidin synthase was identified as a major obstacle to efficient production. In yeast, this enzyme converts the majority of its natural substrates leucoanthocyanidins into the off-pathway flavonols. Nonetheless, de novo biosynthesis of ACNs was shown for the first time in yeast and for the first time in a single microorganism. It provides a framework for optimizing the activity of anthocyanidin synthase and represents an important step towards sustainable industrial production of these highly relevant molecules in yeast.

  19. Antimutators of mitochodrial and nuclear DNA in Saccharomyces cerevisiae

    Bianchi, L.; Foury, F.

    1982-01-01

    In Saccharomyces cerevisiae ten antimutator mutants have been isolated. The spontaneous occurrence of mitochondrial mutants resistant to erythromycin, oligomycin, and diuron is decreased 2-60-fold in these strains. The rate of forward and reverse spontaneous mutations of the nuclear genome is also reduced. The meiotic progenies arising from the crosses of seven mutants (LB 1 , LB 2 , LB 4 , LB 5 , LB 6 , LB 7 , LB 10 ) with an isogenic parental strain exhibit 2:2 segregations and therefore are the result of mutations in a single nuclear gene. The six mutants LB 1 , LB 2 , LB 4 , LB 6 , LB 7 , LB 10 are semidominant and determine six complementation groups. The mutant LB 5 is dominant and therefore cannot be assigned to any complementation group. The mutants. LB 1 , LB 4 and LB 1 0 are gamma-ray sensitive and, by tetrad analysis, it has been shown that gamma-ray sensitivity and spontaneous antimutability are the result of a single nuclear gene mutation. The other three mutants LB 3 , LB 8 and LB 9 exhibit complex tetrad segregations, typical of cytoplasmic inheritance and do not complement each other. However, although the mutations are semidominant, it has not been possible to detect any antimutator cytoductant among some 500 cytoductants carrying the karl 1-1 nucleus. (orig./AJ)

  20. Spontaneous and radiation induced gene conversion in Saccharomyces cerevisiae

    Rao, B.S.; Murthy, M.S.S.

    1977-01-01

    Spontaneous and radiation induced gene conversion to arginine independence was studied in a heteroallelic diploid strain of yeast Saccharomyces cerevisiae BZ 34. When stationary phase cells were incubated in phosphate buffer (pH 7 ) at 30 0 C under aerated condition for 48 hours, the conversion frequency increased by a factor of about 1000 times the background. This was found to be so even when the cells were incubated in saline (0.85%) or distilled water. Various conditions influencing this enhancement have been investigated. Conversion frequency enhancement was not significant under anoxic conditions and was absent at low temperatures and in log phase cells. Caffeine could inhibit this enhancement when present in the suspension medium. These results can be explained on the basis of the induction of meiosis in cells held in buffer. Microscopic examination confirmed this view. Under conditions not favourable for the onset of meiosis there is no significant enhancement in conversion frequency. In stationary phase cells exposed to series of gamma doses, the conversion frequency increases with dose. Post irradiation incubation in buffer further increases the conversion frequency. However, the increase expressed as the ratio of the conversion frequency on buffer holding to that on immediate plating decreased with increasing dose. This decrease in enhancement with increasing dose may be due to the dose dependent inhibition of meiosis. (author)

  1. Brazilian propolis protects Saccharomyces cerevisiae cells against oxidative stress

    Rafael A. de Sá

    2013-09-01

    Full Text Available Propolis is a natural product widely used for humans. Due to its complex composition, a number of applications (antimicrobial, antiinflammatory, anesthetic, cytostatic and antioxidant have been attributed to this substance. Using Saccharomyces cerevisiae as a eukaryotic model we investigated the mechanisms underlying the antioxidant effect of propolis from Guarapari against oxidative stress. Submitting a wild type (BY4741 and antioxidant deficient strains (ctt1∆, sod1∆, gsh1∆, gtt1∆ and gtt2∆ either to 15 mM menadione or to 2 mM hydrogen peroxide during 60 min, we observed that all strains, except the mutant sod1∆, acquired tolerance when previously treated with 25 µg/mL of alcoholic propolis extract. Such a treatment reduced the levels of ROS generation and of lipid peroxidation, after oxidative stress. The increase in Cu/Zn-Sod activity by propolis suggests that the protection might be acting synergistically with Cu/Zn-Sod.

  2. Water-Transfer Slows Aging in Saccharomyces cerevisiae.

    Aviv Cohen

    Full Text Available Transferring Saccharomyces cerevisiae cells to water is known to extend their lifespan. However, it is unclear whether this lifespan extension is due to slowing the aging process or merely keeping old yeast alive. Here we show that in water-transferred yeast, the toxicity of polyQ proteins is decreased and the aging biomarker 47Q aggregates at a reduced rate and to a lesser extent. These beneficial effects of water-transfer could not be reproduced by diluting the growth medium and depended on de novo protein synthesis and proteasomes levels. Interestingly, we found that upon water-transfer 27 proteins are downregulated, 4 proteins are upregulated and 81 proteins change their intracellular localization, hinting at an active genetic program enabling the lifespan extension. Furthermore, the aging-related deterioration of the heat shock response (HSR, the unfolded protein response (UPR and the endoplasmic reticulum-associated protein degradation (ERAD, was largely prevented in water-transferred yeast, as the activities of these proteostatic network pathways remained nearly as robust as in young yeast. The characteristics of young yeast that are actively maintained upon water-transfer indicate that the extended lifespan is the outcome of slowing the rate of the aging process.

  3. Water-Transfer Slows Aging in Saccharomyces cerevisiae.

    Cohen, Aviv; Weindling, Esther; Rabinovich, Efrat; Nachman, Iftach; Fuchs, Shai; Chuartzman, Silvia; Gal, Lihi; Schuldiner, Maya; Bar-Nun, Shoshana

    2016-01-01

    Transferring Saccharomyces cerevisiae cells to water is known to extend their lifespan. However, it is unclear whether this lifespan extension is due to slowing the aging process or merely keeping old yeast alive. Here we show that in water-transferred yeast, the toxicity of polyQ proteins is decreased and the aging biomarker 47Q aggregates at a reduced rate and to a lesser extent. These beneficial effects of water-transfer could not be reproduced by diluting the growth medium and depended on de novo protein synthesis and proteasomes levels. Interestingly, we found that upon water-transfer 27 proteins are downregulated, 4 proteins are upregulated and 81 proteins change their intracellular localization, hinting at an active genetic program enabling the lifespan extension. Furthermore, the aging-related deterioration of the heat shock response (HSR), the unfolded protein response (UPR) and the endoplasmic reticulum-associated protein degradation (ERAD), was largely prevented in water-transferred yeast, as the activities of these proteostatic network pathways remained nearly as robust as in young yeast. The characteristics of young yeast that are actively maintained upon water-transfer indicate that the extended lifespan is the outcome of slowing the rate of the aging process.

  4. Saccharomyces cerevisiae proteinase A excretion and wine making.

    Song, Lulu; Chen, Yefu; Du, Yongjing; Wang, Xibin; Guo, Xuewu; Dong, Jian; Xiao, Dongguang

    2017-11-09

    Proteinase A (PrA), the major protease in Saccharomyces cerevisiae, plays an essential role in zymogen activation, sporulation, and other physiological processes in vivo. The extracellular secretion of PrA often occurs during alcoholic fermentation, especially in the later stages when the yeast cells are under stress conditions, and affects the quality and safety of fermented products. Thus, the mechanism underlying PrA excretion must be explored to improve the quality and safety of fermented products. This paper briefly introduces the structure and physiological function of PrA. Two transport routes of PrA, namely, the Golgi-to-vacuole pathway and the constitutive Golgi-to-plasma membrane pathway, are also discussed. Moreover, the research history and developments on the mechanism of extracellular PrA secretion are described. In addition, it is briefly discussed that calcium homeostasis plays an important role in the secretory pathway of proteins, implying that the regulation of PrA delivery to the plasma membrane requires the involvement of calcium ion. Finally, this review focuses on the effects of PrA excretion on wine making (including Chinese rice wine, grape wine, and beer brewage) and presents strategies to control PrA excretion.

  5. Nutrient sensing and signaling in the yeast Saccharomyces cerevisiae

    Conrad, Michaela; Schothorst, Joep; Kankipati, Harish Nag; Van Zeebroeck, Griet; Rubio-Texeira, Marta; Thevelein, Johan M

    2014-01-01

    The yeast Saccharomyces cerevisiae has been a favorite organism for pioneering studies on nutrient-sensing and signaling mechanisms. Many specific nutrient responses have been elucidated in great detail. This has led to important new concepts and insight into nutrient-controlled cellular regulation. Major highlights include the central role of the Snf1 protein kinase in the glucose repression pathway, galactose induction, the discovery of a G-protein-coupled receptor system, and role of Ras in glucose-induced cAMP signaling, the role of the protein synthesis initiation machinery in general control of nitrogen metabolism, the cyclin-controlled protein kinase Pho85 in phosphate regulation, nitrogen catabolite repression and the nitrogen-sensing target of rapamycin pathway, and the discovery of transporter-like proteins acting as nutrient sensors. In addition, a number of cellular targets, like carbohydrate stores, stress tolerance, and ribosomal gene expression, are controlled by the presence of multiple nutrients. The protein kinase A signaling pathway plays a major role in this general nutrient response. It has led to the discovery of nutrient transceptors (transporter receptors) as nutrient sensors. Major shortcomings in our knowledge are the relationship between rapid and steady-state nutrient signaling, the role of metabolic intermediates in intracellular nutrient sensing, and the identity of the nutrient sensors controlling cellular growth. PMID:24483210

  6. Mating-Type Genes and MAT Switching in Saccharomyces cerevisiae

    Haber, James E.

    2012-01-01

    Mating type in Saccharomyces cerevisiae is determined by two nonhomologous alleles, MATa and MATα. These sequences encode regulators of the two different haploid mating types and of the diploids formed by their conjugation. Analysis of the MATa1, MATα1, and MATα2 alleles provided one of the earliest models of cell-type specification by transcriptional activators and repressors. Remarkably, homothallic yeast cells can switch their mating type as often as every generation by a highly choreographed, site-specific homologous recombination event that replaces one MAT allele with different DNA sequences encoding the opposite MAT allele. This replacement process involves the participation of two intact but unexpressed copies of mating-type information at the heterochromatic loci, HMLα and HMRa, which are located at opposite ends of the same chromosome-encoding MAT. The study of MAT switching has yielded important insights into the control of cell lineage, the silencing of gene expression, the formation of heterochromatin, and the regulation of accessibility of the donor sequences. Real-time analysis of MAT switching has provided the most detailed description of the molecular events that occur during the homologous recombinational repair of a programmed double-strand chromosome break. PMID:22555442

  7. MAP kinase pathways in the yeast Saccharomyces cerevisiae

    Gustin, M. C.; Albertyn, J.; Alexander, M.; Davenport, K.; McIntire, L. V. (Principal Investigator)

    1998-01-01

    A cascade of three protein kinases known as a mitogen-activated protein kinase (MAPK) cascade is commonly found as part of the signaling pathways in eukaryotic cells. Almost two decades of genetic and biochemical experimentation plus the recently completed DNA sequence of the Saccharomyces cerevisiae genome have revealed just five functionally distinct MAPK cascades in this yeast. Sexual conjugation, cell growth, and adaptation to stress, for example, all require MAPK-mediated cellular responses. A primary function of these cascades appears to be the regulation of gene expression in response to extracellular signals or as part of specific developmental processes. In addition, the MAPK cascades often appear to regulate the cell cycle and vice versa. Despite the success of the gene hunter era in revealing these pathways, there are still many significant gaps in our knowledge of the molecular mechanisms for activation of these cascades and how the cascades regulate cell function. For example, comparison of different yeast signaling pathways reveals a surprising variety of different types of upstream signaling proteins that function to activate a MAPK cascade, yet how the upstream proteins actually activate the cascade remains unclear. We also know that the yeast MAPK pathways regulate each other and interact with other signaling pathways to produce a coordinated pattern of gene expression, but the molecular mechanisms of this cross talk are poorly understood. This review is therefore an attempt to present the current knowledge of MAPK pathways in yeast and some directions for future research in this area.

  8. Biosynthesis of diphthamide in the yeast Saccharomyces cerevisiae

    Chen, J.Y.C.

    1985-01-01

    Inactivation of EF-2 by diphtheria toxin requires the presence of a posttranslationally synthesized amino acid residue, diphthamide. The present work was undertaken to study the biosynthetic mechanism of diphthamide synthesis in the yeast Saccharomyces cerevisiae in order to gain better understanding of the biological roles of this unique amino acid residue. Thirty-one haploid ADP-ribosylation-negative mutants, comprising 5 complementation groups, were obtained. One of these mutants contains a toxin-resistant form of EF-2 which can be converted to a toxin-sensitive form through the methylation reaction catalyzed by a S-AdoMet:EF-2 methyltransferase enzyme which is present in other yeast strains. The [ 3 He]methylated residue in the EF-2 modified by the methyltransferase in the presence of S-Ado-L-[ 3 H-methyl]-Met has been analyzed chromatographically following both acid and enzymatic hydrolysis. At the conclusion of the reaction, all of the radiolabel was recovered as diphthine (the unamidated form of diphthamide). The authors conclude that the S-AdoMet:EF-2-methyltransferase is specific for the addition of at least the last two of the three methyl groups present in diphthine

  9. Identification of genes affecting vacuole membrane fragmentation in Saccharomyces cerevisiae.

    Lydie Michaillat

    Full Text Available The equilibrium of membrane fusion and fission influences the volume and copy number of organelles. Fusion of yeast vacuoles has been well characterized but their fission and the mechanisms determining vacuole size and abundance remain poorly understood. We therefore attempted to systematically characterize factors necessary for vacuole fission. Here, we present results of an in vivo screening for deficiencies in vacuolar fragmentation activity of an ordered collection deletion mutants, representing 4881 non-essential genes of the yeast Saccharomyces cerevisiae. The screen identified 133 mutants with strong defects in vacuole fragmentation. These comprise numerous known fragmentation factors, such as the Fab1p complex, Tor1p, Sit4p and the V-ATPase, thus validating the approach. The screen identified many novel factors promoting vacuole fragmentation. Among those are 22 open reading frames of unknown function and three conspicuous clusters of proteins with known function. The clusters concern the ESCRT machinery, adaptins, and lipases, which influence the production of diacylglycerol and phosphatidic acid. A common feature of these factors of known function is their capacity to change membrane curvature, suggesting that they might promote vacuole fragmentation via this property.

  10. Proteasome dynamics between proliferation and quiescence stages of Saccharomyces cerevisiae.

    Yedidi, Ravikiran S; Fatehi, Amatullah K; Enenkel, Cordula

    The ubiquitin-proteasome system (UPS) plays a critical role in cellular protein homeostasis and is required for the turnover of short-lived and unwanted proteins, which are targeted by poly-ubiquitination for degradation. Proteasome is the key protease of UPS and consists of multiple subunits, which are organized into a catalytic core particle (CP) and a regulatory particle (RP). In Saccharomyces cerevisiae, proteasome holo-enzymes are engaged in degrading poly-ubiquitinated substrates and are mostly localized in the nucleus during cell proliferation. While in quiescence, the RP and CP are sequestered into motile and reversible storage granules in the cytoplasm, called proteasome storage granules (PSGs). The reversible nature of PSGs allows the proteasomes to be transported back into the nucleus upon exit from quiescence. Nuclear import of RP and CP through nuclear pores occurs via the canonical pathway that includes the importin-αβ heterodimer and takes advantage of the Ran-GTP gradient across the nuclear membrane. Dependent on the growth stage, either inactive precursor complexes or mature holo-enzymes are imported into the nucleus. The present review discusses the dynamics of proteasomes including their assembly, nucleo-cytoplasmic transport during proliferation and the sequestration of proteasomes into PSGs during quiescence. [Formula: see text].

  11. Degradation signals for ubiquitin system proteolysis in Saccharomyces cerevisiae.

    Gilon, T; Chomsky, O; Kulka, R G

    1998-01-01

    Combinations of different ubiquitin-conjugating (Ubc) enzymes and other factors constitute subsidiary pathways of the ubiquitin system, each of which ubiquitinates a specific subset of proteins. There is evidence that certain sequence elements or structural motifs of target proteins are degradation signals which mark them for ubiquitination by a particular branch of the ubiquitin system and for subsequent degradation. Our aim was to devise a way of searching systematically for degradation signals and to determine to which ubiquitin system subpathways they direct the proteins. We have constructed two reporter gene libraries based on the lacZ or URA3 genes which, in Saccharomyces cerevisiae, express fusion proteins with a wide variety of C-terminal extensions. From these, we have isolated clones producing unstable fusion proteins which are stabilized in various ubc mutants. Among these are 10 clones whose products are stabilized in ubc6, ubc7 or ubc6ubc7 double mutants. The C-terminal extensions of these clones, which vary in length from 16 to 50 amino acid residues, are presumed to contain degradation signals channeling proteins for degradation via the UBC6 and/or UBC7 subpathways of the ubiquitin system. Some of these C-terminal tails share similar sequence motifs, and a feature common to almost all of these sequences is a highly hydrophobic region such as is usually located inside globular proteins or inserted into membranes. PMID:9582269

  12. Ubiquitin regulates TORC1 in yeast Saccharomyces cerevisiae.

    Hu, Kejin; Guo, Shuguang; Yan, Gonghong; Yuan, Wenjie; Zheng, Yin; Jiang, Yu

    2016-04-01

    In the yeast Saccharomyces cerevisiae the TOR complex 1 (TORC1) controls many growth-related cellular processes and is essential for cell growth and proliferation. Macrolide antibiotic rapamycin, in complex with a cytosol protein named FKBP12, specifically inhibits TORC1, causing growth arrest. The FKBP12-rapamycin complex interferes with TORC1 function by binding to the FRB domain of the TOR proteins. In an attempt to understand the role of the FRB domain in TOR function, we identified a single point mutation (Tor2(W2041R) ) in the FRB domain of Tor2 that renders yeast cells rapamycin resistant and temperature sensitive. At the permissive temperature, the Tor2 mutant protein is partially defective for binding with Kog1 and TORC1 is impaired for membrane association. At the restrictive temperature, Kog1 but not the Tor2 mutant protein, is rapidly degraded. Overexpression of ubiquitin stabilizes Kog1 and suppresses the growth defect associated with the tor2 mutant at the nonpremissive temperature. We find that ubiquitin binds non-covalently to Kog1, prevents Kog1 from degradation and stabilizes TORC1. Our data reveal a unique role for ubiquitin in regulation of TORC1 and suggest that Kog1 requires association with the Tor proteins for stabilization. © 2016 John Wiley & Sons Ltd.

  13. Phosphatidylcholine Supply to Peroxisomes of the Yeast Saccharomyces cerevisiae.

    Flis, Vid V; Fankl, Ariane; Ramprecht, Claudia; Zellnig, Günther; Leitner, Erich; Hermetter, Albin; Daum, Günther

    2015-01-01

    In the yeast Saccharomyces cerevisiae, phosphatidylcholine (PC), the major phospholipid (PL) of all organelle membranes, is synthesized via two different pathways. Methylation of phosphatidylethanolamine (PE) catalyzed by the methyl transferases Cho2p/Pem1p and Opi3p/Pem2p as well as incorporation of choline through the CDP (cytidine diphosphate)-choline branch of the Kennedy pathway lead to PC formation. To determine the contribution of these two pathways to the supply of PC to peroxisomes (PX), yeast mutants bearing defects in the two pathways were cultivated under peroxisome inducing conditions, i.e. in the presence of oleic acid, and subjected to biochemical and cell biological analyses. Phenotype studies revealed compromised growth of both the cho20Δopi3Δ (mutations in the methylation pathway) and the cki1Δdpl1Δeki1Δ (mutations in the CDP-choline pathway) mutant when grown on oleic acid. Analysis of peroxisomes from the two mutant strains showed that both pathways produce PC for the supply to peroxisomes, although the CDP-choline pathway seemed to contribute with higher efficiency than the methylation pathway. Changes in the peroxisomal lipid pattern of mutants caused by defects in the PC biosynthetic pathways resulted in changes of membrane properties as shown by anisotropy measurements with fluorescent probes. In summary, our data define the origin of peroxisomal PC and demonstrate the importance of PC for peroxisome membrane formation and integrity.

  14. Damage-induced ectopic recombination in the yeast Saccharomyces cerevisiae.

    Kupiec, M; Steinlauf, R

    1997-06-09

    Mitotic recombination in the yeast Saccharomyces cerevisiae is induced when cells are irradiated with UV or X-rays, reflecting the efficient repair of damage by recombinational repair mechanisms. We have used multiply marked haploid strains that allow the simultaneous detection of several types of ectopic recombination events. We show that inter-chromosomal ectopic conversion of lys2 heteroalleles and, to a lesser extent, direct repeat recombination (DRR) between non-tandem repeats, are increased by DNA-damaging agents; in contrast, ectopic recombination of the naturally occurring Ty element is not induced. We have tested several hypotheses that could explain the preferential lack of induction of Ty recombination by DNA-damaging agents. We have found that the lack of induction cannot be explained by a cell cycle control or by an effect of the mating-type genes. We also found no role for the flanking long terminal repeats (LTRs) of the Ty in preventing the induction. Ectopic conversion, DRR, and forward mutation of artificial repeats show different kinetics of induction at various positions of the cell cycle, reflecting different mechanisms of recombination. We discuss the mechanistic and evolutionary aspects of these results.

  15. Effects of low X-ray doses in Saccharomyces cerevisiae

    Jordan, A.; Laskowski, W.

    1987-01-01

    Three strains of Saccharomyces cerevisiae with different capacities for repair of radiation damage (RAD, rad18, and rad52) have been tested for their colony forming ability (CFA) and growth rates after application of small X-ray doses from 3.8 mGy to 40 Gy. There was no reproducible increase in CFA observable after application of doses between 3.8 mGy and 4.7 Gy.X-ray doses of 40 Gy causing an inactivation of CFA from 90% to 50%, depending on the repair capacity of the strains used, caused a reduced increase in optical density during 2 h buffer treatment in comparison to unirradiated cells. This reduction however, is reversible as soon as the cells are transferred into nutrient medium. One hour after transfer into growh medium the portions of cells with large buds (Gs and M phase) and cells with small buds (S phase) are drastically different in irradiated cells from those obtained in unirradiated cells. The time necessary for separation of mother and daughter cells is prolonged by X-ray irradiation and the formation of new buds is retarded. (orig.)

  16. Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

    Orlean, Peter

    2012-01-01

    The wall gives a Saccharomyces cerevisiae cell its osmotic integrity; defines cell shape during budding growth, mating, sporulation, and pseudohypha formation; and presents adhesive glycoproteins to other yeast cells. The wall consists of β1,3- and β1,6-glucans, a small amount of chitin, and many different proteins that may bear N- and O-linked glycans and a glycolipid anchor. These components become cross-linked in various ways to form higher-order complexes. Wall composition and degree of cross-linking vary during growth and development and change in response to cell wall stress. This article reviews wall biogenesis in vegetative cells, covering the structure of wall components and how they are cross-linked; the biosynthesis of N- and O-linked glycans, glycosylphosphatidylinositol membrane anchors, β1,3- and β1,6-linked glucans, and chitin; the reactions that cross-link wall components; and the possible functions of enzymatic and nonenzymatic cell wall proteins. PMID:23135325

  17. Xylose Fermentation by Saccharomyces cerevisiae: Challenges and Prospects

    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.

  18. Metabolic engineering of Saccharomyces cerevisiae for caffeine and theobromine production.

    Lu Jin

    Full Text Available Caffeine (1, 3, 7-trimethylxanthine and theobromine (3, 7-dimethylxanthine are the major purine alkaloids in plants, e.g., tea (Camellia sinensis and coffee (Coffea arabica. Caffeine is a major component of coffee and is used widely in food and beverage industries. Most of the enzymes involved in the caffeine biosynthetic pathway have been reported previously. Here, we demonstrated the biosynthesis of caffeine (0.38 mg/L by co-expression of Coffea arabica xanthosine methyltransferase (CaXMT and Camellia sinensis caffeine synthase (TCS in Saccharomyces cerevisiae. Furthermore, we endeavored to develop this production platform for making other purine-based alkaloids. To increase the catalytic activity of TCS in an effort to increase theobromine production, we identified four amino acid residues based on structural analyses of 3D-model of TCS. Two TCS1 mutants (Val317Met and Phe217Trp slightly increased in theobromine accumulation and simultaneously decreased in caffeine production. The application and further optimization of this biosynthetic platform are discussed.

  19. Metabolic engineering of Saccharomyces cerevisiae for caffeine and theobromine production.

    Jin, Lu; Bhuiya, Mohammad Wadud; Li, Mengmeng; Liu, XiangQi; Han, Jixiang; Deng, WeiWei; Wang, Min; Yu, Oliver; Zhang, Zhengzhu

    2014-01-01

    Caffeine (1, 3, 7-trimethylxanthine) and theobromine (3, 7-dimethylxanthine) are the major purine alkaloids in plants, e.g., tea (Camellia sinensis) and coffee (Coffea arabica). Caffeine is a major component of coffee and is used widely in food and beverage industries. Most of the enzymes involved in the caffeine biosynthetic pathway have been reported previously. Here, we demonstrated the biosynthesis of caffeine (0.38 mg/L) by co-expression of Coffea arabica xanthosine methyltransferase (CaXMT) and Camellia sinensis caffeine synthase (TCS) in Saccharomyces cerevisiae. Furthermore, we endeavored to develop this production platform for making other purine-based alkaloids. To increase the catalytic activity of TCS in an effort to increase theobromine production, we identified four amino acid residues based on structural analyses of 3D-model of TCS. Two TCS1 mutants (Val317Met and Phe217Trp) slightly increased in theobromine accumulation and simultaneously decreased in caffeine production. The application and further optimization of this biosynthetic platform are discussed.

  20. ORGANIC ACIDS CONCENTRATION IN WINE STOCKS AFTER Saccharomyces cerevisiae FERMENTATION

    V. N. Bayraktar

    2013-04-01

    Full Text Available The biochemical constituents in wine stocks that influence the flavor and quality of wine are investigated in the paper. The tested parameters consist of volume fraction of ethanol, residual sugar, phenolic compounds, tartaric, malic, citric, lactic, acetic acids, titratable acidity and volatile acids. The wine stocks that were received from white and red grape varieties Tairov`s selection were tested. There was a correlation between titratable acidity and volatile acids in the wine stocks from white and red grape varieties. High correlation was also found between lactic and acetic acids, between volatile acids, acetic acid and sugar. It was determined that wine stocks with a high concentration of ethanol originated from those yeast strains of Saccharomyces cerevisiae, in a fermented grape must of high speed of enzyme activity. The taste of wine stocks correlated with the ratio of tartaric to malic acid. Analysis showed significant differences between the varieties of white and red wine stocks in concentrations of organic acids, phenolic compounds, residual sugar, and volume fraction of ethanol. Positive correlation was indicated for both studied groups for volatile acids and acetic acid, tartaric, malic, lactic acids and total sugar. Prospective yeast cultures with high productivity of alcohol (ethanol were selected for winemaking biotechnology.

  1. Calcium dependence of eugenol tolerance and toxicity in Saccharomyces cerevisiae.

    Stephen K Roberts

    Full Text Available Eugenol is a plant-derived phenolic compound which has recognised therapeutical potential as an antifungal agent. However little is known of either its fungicidal activity or the mechanisms employed by fungi to tolerate eugenol toxicity. A better exploitation of eugenol as a therapeutic agent will therefore depend on addressing this knowledge gap. Eugenol initiates increases in cytosolic Ca2+ in Saccharomyces cerevisiae which is partly dependent on the plasma membrane calcium channel, Cch1p. However, it is unclear whether a toxic cytosolic Ca2+elevation mediates the fungicidal activity of eugenol. In the present study, no significant difference in yeast survival was observed following transient eugenol treatment in the presence or absence of extracellular Ca2+. Furthermore, using yeast expressing apoaequorin to report cytosolic Ca2+ and a range of eugenol derivatives, antifungal activity did not appear to be coupled to Ca2+ influx or cytosolic Ca2+ elevation. Taken together, these results suggest that eugenol toxicity is not dependent on a toxic influx of Ca2+. In contrast, careful control of extracellular Ca2+ (using EGTA or BAPTA revealed that tolerance of yeast to eugenol depended on Ca2+ influx via Cch1p. These findings expose significant differences between the antifungal activity of eugenol and that of azoles, amiodarone and carvacrol. This study highlights the potential to use eugenol in combination with other antifungal agents that exhibit differing modes of action as antifungal agents to combat drug resistant infections.

  2. Ethylene production in relation to nitrogen metabolism in Saccharomyces cerevisiae.

    Johansson, Nina; Persson, Karl O; Quehl, Paul; Norbeck, Joakim; Larsson, Christer

    2014-11-01

    We have previously shown that ethylene production in Saccharomyces cerevisiae expressing the ethylene-forming enzyme (EFE) from Pseudomonas syringae is strongly influenced by variations in the mode of cultivation as well as the choice of nitrogen source. Here, we have studied the influence of nitrogen metabolism on the production of ethylene further. Using ammonium, glutamate, glutamate/arginine, and arginine as nitrogen sources, it was found that glutamate (with or without arginine) correlates with a high ethylene production, most likely linked to an observed increase in 2-oxoglutarate levels. Arginine as a sole nitrogen source caused a reduced ethylene production. A reduction of arginine levels, accomplished using an arginine auxotrophic ARG4-deletion strain in the presence of limiting amounts of arginine or through CAR1 overexpression, did however not correlate with an increased ethylene production. As expected, arginine was necessary for ethylene production as ethylene production in the ARG4-deletion strain ceased at the time when arginine was depleted. In conclusion, our data suggest that high levels of 2-oxoglutarate and a limited amount of arginine are required for successful ethylene production in yeast. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  3. Therapeutic activity of a Saccharomyces cerevisiae-based probiotic and inactivated whole yeast on vaginal candidiasis.

    Pericolini, Eva; Gabrielli, Elena; Ballet, Nathalie; Sabbatini, Samuele; Roselletti, Elena; Cayzeele Decherf, Amélie; Pélerin, Fanny; Luciano, Eugenio; Perito, Stefano; Jüsten, Peter; Vecchiarelli, Anna

    2017-01-02

    Vulvovaginal candidiasis is the most prevalent vaginal infection worldwide and Candida albicans is its major agent. Vulvovaginal candidiasis is characterized by disruption of the vaginal microbiota composition, as happens following large spectrum antibiotic usage. Recent studies support the effectiveness of oral and local probiotic treatment for prevention of recurrent vulvovaginal candidiasis. Saccharomyces cerevisiae is a safe yeast used as, or for, the production of ingredients for human nutrition and health. Here, we demonstrate that vaginal administration of probiotic Saccharomyces cerevisiae live yeast (GI) and, in part, inactivated whole yeast Saccharomyces cerevisiae (IY), used as post-challenge therapeutics, was able to positively influence the course of vaginal candidiasis by accelerating the clearance of the fungus. This effect was likely due to multiple interactions of Saccharomyces cerevisiae with Candida albicans. Both live and inactivated yeasts induced coaggregation of Candida and consequently inhibited its adherence to epithelial cells. However, only the probiotic yeast was able to suppress some major virulence factors of Candida albicans such as the ability to switch from yeast to mycelial form and the capacity to express several aspartyl proteases. The effectiveness of live yeast was higher than that of inactivated whole yeast suggesting that the synergy between mechanical effects and biological effects were dominant over purely mechanical effects. The protection of epithelial cells to Candida-induced damage was also observed. Overall, our data show for the first time that Saccharomyces cerevisiae-based ingredients, particularly the living cells, can exert beneficial therapeutic effects on a widespread vaginal mucosal infection.

  4. Impact of oxygenation on the performance of three non-Saccharomyces yeasts in co-fermentation with Saccharomyces cerevisiae.

    Shekhawat, Kirti; Bauer, Florian F; Setati, Mathabatha E

    2017-03-01

    The sequential or co-inoculation of grape must with non-Saccharomyces yeast species and Saccharomyces cerevisiae wine yeast strains has recently become a common practice in winemaking. The procedure intends to enhance unique aroma and flavor profiles of wine. The extent of the impact of non-Saccharomyces strains depends on their ability to produce biomass and to remain metabolically active for a sufficiently long period. However, mixed-culture wine fermentations tend to become rapidly dominated by S. cerevisiae, reducing or eliminating the non-Saccharomyces yeast contribution. For an efficient application of these yeasts, it is therefore essential to understand the environmental factors that modulate the population dynamics of such ecosystems. Several environmental parameters have been shown to influence population dynamics, but their specific effect remains largely uncharacterized. In this study, the population dynamics in co-fermentations of S. cerevisiae and three non-Saccharomyces yeast species: Torulaspora delbrueckii, Lachancea thermotolerans, and Metschnikowia pulcherrima, was investigated as a function of oxygen availability. In all cases, oxygen availability strongly influenced population dynamics, but clear species-dependent differences were observed. Our data show that L. thermotolerans required the least oxygen, followed by T. delbrueckii and M. pulcherrima. Distinct species-specific chemical volatile profiles correlated in all cases with increased persistence of non-Saccharomyces yeasts, in particular increases in some higher alcohols and medium chain fatty acids. The results highlight the role of oxygen in regulating the succession of yeasts during wine fermentations and suggests that more stringent aeration strategies would be necessary to support the persistence of non-Saccharomyces yeasts in real must fermentations.

  5. Prevalence reduction of pathogens in poultry fed with Saccharomyces cerevisiae

    Fanelli, A.

    2015-01-01

    Full Text Available Description of the subject. The growth of new antibiotic-resistant strains of pathogens represents a huge problem in poultry rearing. There is evidence that dietary yeast could be effective in the protection against a variety of pathogens that can affect poultry health and cause foodborne diseases in humans. Since still few or contradictory information are available for this topic. Objectives. The objective of this study was to investigate the effects of live yeast supplementation in broiler chickens on Salmonella enteritidis and Campylobacter jejuni content in feces, cecum, and skin. Method. Supplemented yeast consisted of Saccharomyces cerevisiae (Levucell® SB20, type boulardii I-1079, Lallemand, France and was administered at a rate of 1 x 106 CFU·g-1 of feed. On day ten of life, birds were orally challenged with S. enteritidis (1 x 105 CFU/bird and C. jejuni (3 x 105 CFU/bird. Growth performance, and coliforms, yeasts and lactobacilli enumeration were evaluated on day 0, 10, 20 and 38. Ten and eighteen days post infection (PI, 10 animals per replicate were slaughtered and pooled ceca content were analyzed for yeast enumeration and Salmonella and Campylobacter frequency and enumeration. The presence and the enumeration of Salmonella and Campylobacter in neck and breast skin were performed on one subject per replicate. Results. Dietary S. cerevisiae increased yeast and lactobacilli (p = 0.01 count, while Salmonella enumeration and frequency significantly decreased in neck (p = 0.03 and tended to decrease in cecum (p = 0.06, feces (p = 0.06, and breast (p = 0.08. On 10d PI Campylobacter presence was decreased in cecum (p = 0.01, feces (p < 0.01, breast skin (p = 0.04 and neck skin (p < 0.01, while the enumeration was found to be lower in feces (p < 0.01 and neck skin (p = 0.05. At the end of the trial the frequency of this pathogen was decreased in feces (p < 0.01, and breast skin (p = 0.02, while the enumeration was diminished in cecum (p

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

    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

  7. Saccharomyces Boulardii

    Saccharomyces boulardii is a yeast, which is a type of fungus. Saccharomyces boulardii was previously identified as a unique species of ... be a strain of Saccharomyces cerevisiae (baker's yeast). Saccharomyces boulardii is used as medicine. Saccharomyces boulardii is most ...

  8. Saccharomyces cerevisiae and non-Saccharomyces yeasts in grape varieties of the São Francisco Valley

    Camila M.P.B.S. de Ponzzes-Gomes

    2014-06-01

    Full Text Available The aims of this work was to characterise indigenous Saccharomyces cerevisiae strains in the naturally fermented juice of grape varieties Cabernet Sauvignon, Grenache, Tempranillo, Sauvignon Blanc and Verdejo used in the São Francisco River Valley, northeastern Brazil. In this study, 155 S. cerevisiae and 60 non-Saccharomyces yeasts were isolated and identified using physiological tests and sequencing of the D1/D2 domains of the large subunit of the rRNA gene. Among the non-Saccharomyces species, Rhodotorula mucilaginosa was the most common species, followed by Pichia kudriavzevii, Candida parapsilosis, Meyerozyma guilliermondii, Wickerhamomyces anomalus, Kloeckera apis, P. manshurica, C. orthopsilosis and C. zemplinina. The population counts of these yeasts ranged among 1.0 to 19 x 10(5 cfu/mL. A total of 155 isolates of S. cerevisiae were compared by mitochondrial DNA restriction analysis, and five molecular mitochondrial DNA restriction profiles were detected. Indigenous strains of S. cerevisiae isolated from grapes of the São Francisco Valley can be further tested as potential starters for wine production.

  9. Genes regulation encoding ADP/ATP carrier in yeasts Saccharomyces cerevisiae and Candida parapsilosis

    Nebohacova, M.

    2000-01-01

    Genes encoding a mitochondrial ADP/ATP carrier (AAC) in yeast Saccharomyces cerevisiae and Candida parapsilosis were investigated. AAC2 is coding for the major AAC isoform in S. cerevisiae. We suggest that AAC2 is a member of a syn-expression group of genes encoding oxidative phosphorylation proteins. Within our previous studies on the regulation of the AAC2 transcription an UAS (-393/-268) was identified that is essential for the expression of this gene. Two functional regulatory cis-elements are located within this UAS -binding sites for an ABFl factor and for HAP2/3/4/5 heteromeric complex. We examined relative contributions and mutual interactions of the ABFl and HAP2/3/4/5 factors in the activation of transcription from the UAS of the AAC2 gene. The whole UAS was dissected into smaller sub-fragments and tested for (i) the ability to form DNA-protein complexes with cellular proteins in vitro, (ii) the ability to confer heterologous expression using AAC3 gene lacking its own promoter, and (iii) the expression of AAC3-lacZ fusion instead of intact AAC3 gene. The obtained results demonstrated that: a) The whole UAS as well as sub-fragment containing only ABF1-binding site are able to form DNA-protein complexes with cellular proteins in oxygen- and heme- dependent manner. The experiments with antibody against the ABF1 showed that the ABF1 factor is one of the proteins binding to AAC2 promoter. We have been unsuccessful to prove the binding of cellular proteins to the HAP2/3/4/5-binding site. However, the presence of HAP2/3/4/5-binding site is necessary to drive a binding of cellular proteins to the ABF1-binding site in carbon source-dependent manner. b) The presence of both ABF1- and HAP2/3/4/5-binding sites and original spacing between them is necessary to confer the growth of Aaac2 mutant strain on non- fermentable carbon source when put in front of AAC3 gene introduced on centromeric vector to Aaac2 mutant strain. c) For the activation of AAC3-lacZ expression on

  10. Prevalence and susceptibility of Saccharomyces cerevisiae causing vaginitis in Greek women.

    Papaemmanouil, V; Georgogiannis, N; Plega, M; Lalaki, J; Lydakis, D; Dimitriou, M; Papadimitriou, A

    2011-12-01

    Saccharomyces cerevisiae is an ascomycetous yeast, that is traditionally used in wine bread and beer production. Vaginitis caused by S. cerevisiae is rare. The aim of this study was to evaluate the frequency of S. cerevisiae isolation from the vagina in two groups of women and determined the in vitro susceptibility of this fungus. Vaginal samples were collected from a total of 262 (asymptomatic and symptomatic) women with vaginitis attending the centre of family planning of General hospital of Piraeus. All blastomycetes that isolated from the vaginal samples were examined for microscopic morphological tests and identified by conventional methods: By API 20 C AUX and ID 32 C (Biomerieux). Antifungal susceptibility testing for amphotericin B,fluconazole itraconazole,voriconazole, posaconazole and caspofungin was performed by E -test (Ab BIODIKS SWEDEN) against S. cerevisiae. A total of 16 isolates of S. cerevisiae derived from vaginal sample of the referred women, average 6.10%. Susceptibility of 16 isolates of S. cerevisiae to a variety of antimycotic agents were obtained. So all isolates of S. cerevisiae were resistant to fluconazole, posaconazole and intraconazole, but they were sensitive to voriconazole caspofungin and Amphotericin B which were found sensitive (except 1/16 strains). None of the 16 patients had a history of occupational domestic use of baker's yeast. Vaginitis caused by S. cerevisiae occur, is rising and cannot be ignored. Treatment of Saccharomyces vaginitis constitutes a major challenge and may require selected and often prolonged therapy. Copyright © 2011 Elsevier Ltd. All rights reserved.

  11. Incorporating Protein Biosynthesis into the Saccharomyces cerevisiae Genome-scale Metabolic Model

    Olivares Hernandez, Roberto

    Based on stoichiometric biochemical equations that occur into the cell, the genome-scale metabolic models can quantify the metabolic fluxes, which are regarded as the final representation of the physiological state of the cell. For Saccharomyces Cerevisiae the genome scale model has been construc......Based on stoichiometric biochemical equations that occur into the cell, the genome-scale metabolic models can quantify the metabolic fluxes, which are regarded as the final representation of the physiological state of the cell. For Saccharomyces Cerevisiae the genome scale model has been...

  12. Screening of Non- Saccharomyces cerevisiae Strains for Tolerance to Formic Acid in Bioethanol Fermentation.

    Oshoma, Cyprian E; Greetham, Darren; Louis, Edward J; Smart, Katherine A; Phister, Trevor G; Powell, Chris; Du, Chenyu

    2015-01-01

    Formic acid is one of the major inhibitory compounds present in hydrolysates derived from lignocellulosic materials, the presence of which can significantly hamper the efficiency of converting available sugars into bioethanol. This study investigated the potential for screening formic acid tolerance in non-Saccharomyces cerevisiae yeast strains, which could be used for the development of advanced generation bioethanol processes. Spot plate and phenotypic microarray methods were used to screen the formic acid tolerance of 7 non-Saccharomyces cerevisiae yeasts. S. kudriavzeii IFO1802 and S. arboricolus 2.3319 displayed a higher formic acid tolerance when compared to other strains in the study. Strain S. arboricolus 2.3319 was selected for further investigation due to its genetic variability among the Saccharomyces species as related to Saccharomyces cerevisiae and availability of two sibling strains: S. arboricolus 2.3317 and 2.3318 in the lab. The tolerance of S. arboricolus strains (2.3317, 2.3318 and 2.3319) to formic acid was further investigated by lab-scale fermentation analysis, and compared with S. cerevisiae NCYC2592. S. arboricolus 2.3319 demonstrated improved formic acid tolerance and a similar bioethanol synthesis capacity to S. cerevisiae NCYC2592, while S. arboricolus 2.3317 and 2.3318 exhibited an overall inferior performance. Metabolite analysis indicated that S. arboricolus strain 2.3319 accumulated comparatively high concentrations of glycerol and glycogen, which may have contributed to its ability to tolerate high levels of formic acid.

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

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

    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 that wine making strains have been domesticated for wine production. In this study, we demonstrate that humans can distinguish between wines produced using wine strains and wild strains of S. cerevisiae as well as its sibling species, Saccharomyces paradoxus. Wine strains produced wine with fruity and floral characteristics, whereas wild strains produced wine with earthy and sulfurous characteristics. The differences that we observe between wine and wild strains provides further evidence that wine strains have evolved phenotypes that are distinct from their wild ancestors and relevant to their use in wine production. PMID:22093681

  14. Phenotypic and metabolic traits of commercial Saccharomyces cerevisiae yeasts.

    Barbosa, Catarina; Lage, Patrícia; Vilela, Alice; Mendes-Faia, Arlete; Mendes-Ferreira, Ana

    2014-01-01

    Currently, pursuing yeast strains that display both a high potential fitness for alcoholic fermentation and a favorable impact on quality is a major goal in the alcoholic beverage industry. This considerable industrial interest has led to many studies characterizing the phenotypic and metabolic traits of commercial yeast populations. In this study, 20 Saccharomyces cerevisiae strains from different geographical origins exhibited high phenotypic diversity when their response to nine biotechnologically relevant conditions was examined. Next, the fermentation fitness and metabolic traits of eight selected strains with a unique phenotypic profile were evaluated in a high-sugar synthetic medium under two nitrogen regimes. Although the strains exhibited significant differences in nitrogen requirements and utilization rates, a direct relationship between nitrogen consumption, specific growth rate, cell biomass, cell viability, acetic acid and glycerol formation was only observed under high-nitrogen conditions. In contrast, the strains produced more succinic acid under the low-nitrogen regime, and a direct relationship with the final cell biomass was established. Glucose and fructose utilization patterns depended on both yeast strain and nitrogen availability. For low-nitrogen fermentation, three strains did not fully degrade the fructose. This study validates phenotypic and metabolic diversity among commercial wine yeasts and contributes new findings on the relationship between nitrogen availability, yeast cell growth and sugar utilization. We suggest that measuring nitrogen during the stationary growth phase is important because yeast cells fermentative activity is not exclusively related to population size, as previously assumed, but it is also related to the quantity of nitrogen consumed during this growth phase.

  15. The effect of acetaminophen on ubiquitin homeostasis in Saccharomyces cerevisiae.

    Angelina Huseinovic

    Full Text Available Acetaminophen (APAP, although considered a safe drug, is one of the major causes of acute liver failure by overdose, and therapeutic chronic use can cause serious health problems. Although the reactive APAP metabolite N-acetyl-p-benzoquinoneimine (NAPQI is clearly linked to liver toxicity, toxicity of APAP is also found without drug metabolism of APAP to NAPQI. To get more insight into mechanisms of APAP toxicity, a genome-wide screen in Saccharomyces cerevisiae for APAP-resistant deletion strains was performed. In this screen we identified genes related to the DNA damage response. Next, we investigated the link between genotype and APAP-induced toxicity or resistance by performing a more detailed screen with a library containing mutants of 1522 genes related to nuclear processes, like DNA repair and chromatin remodelling. We identified 233 strains that had an altered growth rate relative to wild type, of which 107 showed increased resistance to APAP and 126 showed increased sensitivity. Gene Ontology analysis identified ubiquitin homeostasis, regulation of transcription of RNA polymerase II genes, and the mitochondria-to-nucleus signalling pathway to be associated with APAP resistance, while histone exchange and modification, and vesicular transport were connected to APAP sensitivity. Indeed, we observed a link between ubiquitin levels and APAP resistance, whereby ubiquitin deficiency conferred resistance to APAP toxicity while ubiquitin overexpression resulted in sensitivity. The toxicity profile of various chemicals, APAP, and its positional isomer AMAP on a series of deletion strains with ubiquitin deficiency showed a unique resistance pattern for APAP. Furthermore, exposure to APAP increased the level of free ubiquitin and influenced the ubiquitination of proteins. Together, these results uncover a role for ubiquitin homeostasis in APAP-induced toxicity.

  16. Metabolism and Regulation of Glycerolipids in the Yeast Saccharomyces cerevisiae

    Henry, Susan A.; Kohlwein, Sepp D.; Carman, George M.

    2012-01-01

    Due to its genetic tractability and increasing wealth of accessible data, the yeast Saccharomyces cerevisiae is a model system of choice for the study of the genetics, biochemistry, and cell biology of eukaryotic lipid metabolism. Glycerolipids (e.g., phospholipids and triacylglycerol) and their precursors are synthesized and metabolized by enzymes associated with the cytosol and membranous organelles, including endoplasmic reticulum, mitochondria, and lipid droplets. Genetic and biochemical analyses have revealed that glycerolipids play important roles in cell signaling, membrane trafficking, and anchoring of membrane proteins in addition to membrane structure. The expression of glycerolipid enzymes is controlled by a variety of conditions including growth stage and nutrient availability. Much of this regulation occurs at the transcriptional level and involves the Ino2–Ino4 activation complex and the Opi1 repressor, which interacts with Ino2 to attenuate transcriptional activation of UASINO-containing glycerolipid biosynthetic genes. Cellular levels of phosphatidic acid, precursor to all membrane phospholipids and the storage lipid triacylglycerol, regulates transcription of UASINO-containing genes by tethering Opi1 to the nuclear/endoplasmic reticulum membrane and controlling its translocation into the nucleus, a mechanism largely controlled by inositol availability. The transcriptional activator Zap1 controls the expression of some phospholipid synthesis genes in response to zinc availability. Regulatory mechanisms also include control of catalytic activity of glycerolipid enzymes by water-soluble precursors, products and lipids, and covalent modification of phosphorylation, while in vivo function of some enzymes is governed by their subcellular location. Genome-wide genetic analysis indicates coordinate regulation between glycerolipid metabolism and a broad spectrum of metabolic pathways. PMID:22345606

  17. Molecular Mechanism of Terbinafine Resistance in Saccharomyces cerevisiae

    Leber, Regina; Fuchsbichler, Sandra; Klobučníková, Vlasta; Schweighofer, Natascha; Pitters, Eva; Wohlfarter, Kathrin; Lederer, Mojca; Landl, Karina; Ruckenstuhl, Christoph; Hapala, Ivan; Turnowsky, Friederike

    2003-01-01

    Ten mutants of the yeast Saccharomyces cerevisiae resistant to the antimycotic terbinafine were isolated after chemical or UV mutagenesis. Molecular analysis of these mutants revealed single base pair exchanges in the ERG1 gene coding for squalene epoxidase, the target of terbinafine. The mutants did not show cross-resistance to any of the substrates of various pleiotropic drug resistance efflux pumps tested. The ERG1 mRNA levels in the mutants did not differ from those in the wild-type parent strains. Terbinafine resistance was transmitted with the mutated alleles in gene replacement experiments, proving that single amino acid substitutions in the Erg1 protein were sufficient to confer the resistance phenotype. The amino acid changes caused by the point mutations were clustered in two regions of the Erg1 protein. Seven mutants carried the amino acid substitutions F402L (one mutant), F420L (one mutant), and P430S (five mutants) in the C-terminal part of the protein; and three mutants carried an L251F exchange in the central part of the protein. Interestingly, all exchanges identified involved amino acids which are conserved in the squalene epoxidases of yeasts and mammals. Two mutations that were generated by PCR mutagenesis of the ERG1 gene and that conferred terbinafine resistance mapped in the same regions of the Erg1 protein, with one resulting in an L251F exchange and the other resulting in an F433S exchange. The results strongly indicate that these regions are responsible for the interaction of yeast squalene epoxidase with terbinafine. PMID:14638499

  18. Ecological and Genetic Barriers Differentiate Natural Populations of Saccharomyces cerevisiae.

    Clowers, Katie J; Heilberger, Justin; Piotrowski, Jeff S; Will, Jessica L; Gasch, Audrey P

    2015-09-01

    How populations that inhabit the same geographical area become genetically differentiated is not clear. To investigate this, we characterized phenotypic and genetic differences between two populations of Saccharomyces cerevisiae that in some cases inhabit the same environment but show relatively little gene flow. We profiled stress sensitivity in a group of vineyard isolates and a group of oak-soil strains and found several niche-related phenotypes that distinguish the populations. We performed bulk-segregant mapping on two of the distinguishing traits: The vineyard-specific ability to grow in grape juice and oak-specific tolerance to the cell wall damaging drug Congo red. To implicate causal genes, we also performed a chemical genomic screen in the lab-strain deletion collection and identified many important genes that fell under quantitative trait loci peaks. One gene important for growth in grape juice and identified by both the mapping and the screen was SSU1, a sulfite-nitrite pump implicated in wine fermentations. The beneficial allele is generated by a known translocation that we reasoned may also serve as a genetic barrier. We found that the translocation is prevalent in vineyard strains, but absent in oak strains, and presents a postzygotic barrier to spore viability. Furthermore, the translocation was associated with a fitness cost to the rapid growth rate seen in oak-soil strains. Our results reveal the translocation as a dual-function locus that enforces ecological differentiation while producing a genetic barrier to gene flow in these sympatric populations. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  19. Capturing of the monoterpene olefin limonene produced in Saccharomyces cerevisiae.

    Jongedijk, Esmer; Cankar, Katarina; Ranzijn, Jorn; van der Krol, Sander; Bouwmeester, Harro; Beekwilder, Jules

    2015-01-01

    Monoterpene olefins such as limonene are plant compounds with applications as flavouring and fragrance agents, as solvents and potentially also in polymer and fuel chemistry. We engineered baker's yeast Saccharomyces cerevisiae to express a (-)-limonene synthase from Perilla frutescens and a (+)-limonene synthase from Citrus limon. Both proteins were expressed either with their native plastid targeting signal or in a truncated form in which the plastidial sorting signal was removed. The yeast host strain for expression was AE9 K197G, which expresses a mutant Erg20 enzyme. This enzyme catalyses the formation of geranyl diphosphate, which is the precursor for monoterpenes. Several methods were tested to capture limonene produced by the yeast. Extraction from the culture medium by pentane, or by the addition of CaCl2 followed by solid-phase micro-extraction, did not lead to detectable limonene, indicating that limonene is rapidly lost from the culture medium. Volatile terpenes such as limonene may also be trapped in a dodecane phase added to the medium during fermentation. This method resulted in recovery of 0.028 mg/l (+)-limonene and 0.060 mg/l (-)-limonene in strains using the truncated Citrus and Perilla synthases, respectively. Trapping the headspace during culture of the limonene synthase-expressing strains resulted in higher titres, at 0.12 mg/l (+)-limonene and 0.49 mg/l (-)-limonene. These results show that the volatile properties of the olefins produced require specific methods for efficient recovery of these molecules from biotechnological production systems. Copyright © 2014 John Wiley & Sons, Ltd.

  20. Studies of anaerobic and aerobic glycolysis in Saccharomyces cerevisiae

    den Hollander, J.A.; Ugurbil, K.; Brown, T.R.; Bednar, M.; Redfield, C.; Shulman, R.G.

    1986-01-01

    Glucose metabolism was followed in suspensions of Saccharomyces cerevisiae by using 13C NMR and 14C radioactive labeling techniques and by Warburg manometer experiments. These experiments were performed for cells grown with various carbon sources in the growth medium, so as to evaluate the effect of catabolite repression. The rate of glucose utilization was most conveniently determined by the 13C NMR experiments, which measured the concentration of [1-13C]glucose, whereas the distribution of end products was determined from the 13C and the 14C experiments. By combining these measurements the flows into the various pathways that contribute to glucose catabolism were estimated, and the effect of oxygen upon glucose catabolism was evaluated. From these measurements, the Pasteur quotient (PQ) for glucose catabolism was calculated to be 2.95 for acetate-grown cells and 1.89 for cells grown on glucose into saturation. The Warburg experiments provided an independent estimate of glucose catabolism. The PQ estimated from Warburg experiments was 2.9 for acetate-grown cells in excellent agreement with the labeled carbon experiments and 4.6 for cells grown into saturation, which did not agree. Possible explanations of these differences are discussed. From these data an estimate is obtained of the net flow through the Embden-Meyerhof-Parnas pathway. The backward flow through fructose-1,6-bisphosphatase (Fru-1,6-P2-ase) was calculated from the scrambling of the 13C label of [1-13C]glucose into the C1 and C6 positions of trehalose. Combining these data allowed us to calculate the net flux through phosphofructokinase (PFK). For acetate-grown cells we found that the relative flow through PFK is a factor of 1.7 faster anaerobically than aerobically

  1. Immobilization of Saccharomyces Cerevisiae in Rice Hulls for Ethanol Production

    Edita Martini

    2011-05-01

    Full Text Available The whole cell immobilization in ethanol fermentation can be done by using natural carriers or through synthetic carriers. All of these methods have the same purpose of retaining high cell concentrations within a certain defined region of space which leads to higher ethanol productivity. Lignocellulosic plant substance represents one of highly potential sources in ethanol production. Some studies have found that cellulosic substances substances can also be used as a natural carrier in cell immobilization by re-circulating pre-culture medium into a reactor. In this experiment, rice hulls without any treatment were used to immobilize Saccharomyces cerevisiae through semi solid state incubation combined with re-circulating pre-culture medium. The scanning electron microscopy (SEM pictures of the carrier show that the yeast cells are absorbed and embedded to the rice hull pore. In liquid batch fermentation system with an initial sugar concentration of 50 g/L, nearly 100% total sugar was consumed after 48 hours. This resulted in an ethanol yield of 0.32 g ethanol/g glucose, which is 62.7% of the theoretical value. Ethanol productivity of 0.59 g/(L.h is 2.3 fold higher than that of free cells which is 0.26 g/(L.h. An effort to reuse the immobilized cells in liquid fermentation system showed poor results due to cell desorption in the first batch which led to high sugar concentration inhibitory effect in the second batch fermentation. This might be solved by using semi solid fermentation process in the future work.

  2. Predicting functional upstream open reading frames in Saccharomyces cerevisiae

    Kristiansson Erik

    2009-12-01

    Full Text Available Abstract Background Some upstream open reading frames (uORFs regulate gene expression (i.e., they are functional and can play key roles in keeping organisms healthy. However, how uORFs are involved in gene regulation is not yet fully understood. In order to get a complete view of how uORFs are involved in gene regulation, it is expected that a large number of experimentally verified functional uORFs are needed. Unfortunately, wet-experiments to verify that uORFs are functional are expensive. Results In this paper, a new computational approach to predicting functional uORFs in the yeast Saccharomyces cerevisiae is presented. Our approach is based on inductive logic programming and makes use of a novel combination of knowledge about biological conservation, Gene Ontology annotations and genes' responses to different conditions. Our method results in a set of simple and informative hypotheses with an estimated sensitivity of 76%. The hypotheses predict 301 further genes to have 398 novel functional uORFs. Three (RPC11, TPK1, and FOL1 of these 301 genes have been hypothesised, following wet-experiments, by a related study to have functional uORFs. A comparison with another related study suggests that eleven of the predicted functional uORFs from genes LDB17, HEM3, CIN8, BCK2, PMC1, FAS1, APP1, ACC1, CKA2, SUR1, and ATH1 are strong candidates for wet-lab experimental studies. Conclusions Learning based prediction of functional uORFs can be done with a high sensitivity. The predictions made in this study can serve as a list of candidates for subsequent wet-lab verification and might help to elucidate the regulatory roles of uORFs.

  3. Industrial Systems Biology of Saccharomyces cerevisiae Enables Novel Succinic Acid Cell Factory

    Otero, José Manuel; Cimini, Donatella; Patil, Kiran Raosaheb

    2013-01-01

    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......-direction of carbon fluxes in S. cerevisiae, and hence show proof of concept that this is a potentially attractive cell factory for over-producing different platform chemicals....

  4. Large-scale functional genomic analysis of sporulation and meiosis in Saccharomyces cerevisiae.

    Enyenihi, Akon H; Saunders, William S

    2003-01-01

    We have used a single-gene deletion mutant bank to identify the genes required for meiosis and sporulation among 4323 nonessential Saccharomyces cerevisiae annotated open reading frames (ORFs). Three hundred thirty-four sporulation-essential genes were identified, including 78 novel ORFs and 115 known genes without previously described sporulation defects in the comprehensive Saccharomyces Genome (SGD) or Yeast Proteome (YPD) phenotype databases. We have further divided the uncharacterized sp...

  5. Co-cultivation of non-conventional yeast with Saccharomyces cerevisiae to increase the aroma complexity of fermented beverages

    Rijswijck, van Irma M.H.

    2017-01-01

    Yeast are used as workhorses to convert hopped wort into beer. Conventionally, such yeasts belong to the genus Saccharomyces and most research on fermentation of wort for the production of beer has focussed on the species Saccharomyces cerevisiae and Saccharomyces

  6. Reconstruction of the High-Osmolarity Glycerol (HOG) Signaling Pathway from the Halophilic Fungus Wallemia ichthyophaga in Saccharomyces cerevisiae.

    Konte, Tilen; Terpitz, Ulrich; Plemenitaš, Ana

    2016-01-01

    The basidiomycetous fungus Wallemia ichthyophaga grows between 1.7 and 5.1 M NaCl and is the most halophilic eukaryote described to date. Like other fungi, W. ichthyophaga detects changes in environmental salinity mainly by the evolutionarily conserved high-osmolarity glycerol (HOG) signaling pathway. In Saccharomyces cerevisiae, the HOG pathway has been extensively studied in connection to osmotic regulation, with a valuable knock-out strain collection established. In the present study, we reconstructed the architecture of the HOG pathway of W. ichthyophaga in suitable S. cerevisiae knock-out strains, through heterologous expression of the W. ichthyophaga HOG pathway proteins. Compared to S. cerevisiae, where the Pbs2 (ScPbs2) kinase of the HOG pathway is activated via the SHO1 and SLN1 branches, the interactions between the W. ichthyophaga Pbs2 (WiPbs2) kinase and the W. ichthyophaga SHO1 branch orthologs are not conserved: as well as evidence of poor interactions between the WiSho1 Src-homology 3 (SH3) domain and the WiPbs2 proline-rich motif, the absence of a considerable part of the osmosensing apparatus in the genome of W. ichthyophaga suggests that the SHO1 branch components are not involved in HOG signaling in this halophilic fungus. In contrast, the conserved activation of WiPbs2 by the S. cerevisiae ScSsk2/ScSsk22 kinase and the sensitivity of W. ichthyophaga cells to fludioxonil, emphasize the significance of two-component (SLN1-like) signaling via Group III histidine kinase. Combined with protein modeling data, our study reveals conserved and non-conserved protein interactions in the HOG signaling pathway of W. ichthyophaga and therefore significantly improves the knowledge of hyperosmotic signal processing in this halophilic fungus.

  7. Functional expression and characterization of five wax ester synthases in Saccharomyces cerevisiae and their utility for biodiesel production

    Shi Shuobo

    2012-02-01

    Full Text Available Abstract Background Wax ester synthases (WSs can synthesize wax esters from alcohols and fatty acyl coenzyme A thioesters. The knowledge of the preferred substrates for each WS allows the use of yeast cells for the production of wax esters that are high-value materials and can be used in a variety of industrial applications. The products of WSs include fatty acid ethyl esters, which can be directly used as biodiesel. Results Here, heterologous WSs derived from five different organisms were successfully expressed and evaluated for their substrate preference in Saccharomyces cerevisiae. We investigated the potential of the different WSs for biodiesel (that is, fatty acid ethyl esters production in S. cerevisiae. All investigated WSs, from Acinetobacter baylyi ADP1, Marinobacter hydrocarbonoclasticus DSM 8798, Rhodococcus opacus PD630, Mus musculus C57BL/6 and Psychrobacter arcticus 273-4, have different substrate specificities, but they can all lead to the formation of biodiesel. The best biodiesel producing strain was found to be the one expressing WS from M. hydrocarbonoclasticus DSM 8798 that resulted in a biodiesel titer of 6.3 mg/L. To further enhance biodiesel production, acetyl coenzyme A carboxylase was up-regulated, which resulted in a 30% increase in biodiesel production. Conclusions Five WSs from different species were functionally expressed and their substrate preference characterized in S. cerevisiae, thus constructing cell factories for the production of specific kinds of wax ester. WS from M. hydrocarbonoclasticus showed the highest preference for ethanol compared to the other WSs, and could permit the engineered S. cerevisiae to produce biodiesel.

  8. Transcriptomic analysis of Saccharomyces cerevisiae x Saccharomyces kudriavzevii hybrids during low temperature winemaking [version 3; referees: 2 approved

    Jordi Tronchoni

    2017-09-01

    Full Text Available Background: Although Saccharomyces cerevisiae is the most frequently isolated species in wine fermentation, and the most studied species, other species and interspecific hybrids have greatly attracted the interest of researchers in this field in the last few years, given their potential to solve new winemaking industry challenges. S. cerevisiae x S. kudriavzevii hybrids exhibit good fermentative capabilities at low temperatures, and produce wines with smaller alcohol quantities and larger glycerol quantities, which can be very useful to solve challenges in the winemaking industry such as the necessity to enhance the aroma profile. Methods: In this study, we performed a transcriptomic study of S. cerevisiae x S. kudriavzevii hybrids in low temperature winemaking conditions. Results: The results revealed that the hybrids have acquired both fermentative abilities and cold adaptation abilities, attributed to S. cerevisiae and S. kudriavzevii parental species, respectively, showcasing their industrially relevant characteristics. For several key genes, we also studied the contribution to gene expression of each of the alleles of S. cerevisiae and S. kudriavzevii in the S. cerevisiae x S. kudriavzevii hybrids. From the results, it is not clear how important the differential expression of the specific parental alleles is to the phenotype of the hybrids. Conclusions: This study shows that the fermentative abilities of S. cerevisiae x S. kudriavzevii hybrids at low temperatures do not seem to result from differential expression of specific parental alleles of the key genes involved in this phenotype.

  9. An insight into the complex prion-prion interaction network in the budding yeast Saccharomyces cerevisiae.

    Du, Zhiqiang; Valtierra, Stephanie; Li, Liming

    2014-01-01

    The budding yeast Saccharomyces cerevisiae is a valuable model system for studying prion-prion interactions as it contains multiple prion proteins. A recent study from our laboratory showed that the existence of Swi1 prion ([SWI(+)]) and overproduction of Swi1 can have strong impacts on the formation of 2 other extensively studied yeast prions, [PSI(+)] and [PIN(+)] ([RNQ(+)]) (Genetics, Vol. 197, 685-700). We showed that a single yeast cell is capable of harboring at least 3 heterologous prion elements and these prions can influence each other's appearance positively and/or negatively. We also showed that during the de novo [PSI(+)] formation process upon Sup35 overproduction, the aggregation patterns of a preexisting inducer ([RNQ(+)] or [SWI(+)]) can undergo significant remodeling from stably transmitted dot-shaped aggregates to aggregates that co-localize with the newly formed Sup35 aggregates that are ring/ribbon/rod- shaped. Such co-localization disappears once the newly formed [PSI(+)] prion stabilizes. Our finding provides strong evidence supporting the "cross-seeding" model for prion-prion interactions and confirms earlier reports that the interactions among different prions and their prion proteins mostly occur at the initiation stages of prionogenesis. Our results also highlight a complex prion interaction network in yeast. We believe that elucidating the mechanism underlying the yeast prion-prion interaction network will not only provide insight into the process of prion de novo generation and propagation in yeast but also shed light on the mechanisms that govern protein misfolding, aggregation, and amyloidogenesis in higher eukaryotes.

  10. Peran Direct Fed Microbials (DFM Saccharomyces cerevisiae dan Aspergillus oryzae terhadap Produktivitas Ternak Ruminansia : Review

    H. Suryani

    2015-02-01

    Full Text Available Mikroorganisme yang biasa digunakan dalam pakan ternak ruminansia biasanya berupa probiotik. Probiotik memiliki makna yang bersepadanan dengan Direct Fed Microbials (DFM. Penambahan DFM jenis Saccharomyces cerevisiae dan Aspergillus oryzae pada pakan ternak ruminansia mampu memanipulasi rumen dengan meningkatkan populasi bakteri pemecah serat sehingga dapat meningkatkan kecernaan dan meningkatkan bobot badan. Mekanisme kerja S. cerevisiae dan A. oryzae yang masuk kedalam tubuh ternak dan mempengaruhi pencernaan atau penyerapan, ada yang sudah diketahui secara jelas tetapi ada juga yang masih berupa hipotesa. Pemanfaatan DFM jenis S. cerevisiae dan A. oryzae secara tunggal maupun kombinasi sebagian telah diamati dan memberikan respon positif.

  11. iTRAQ-based proteome profiling of Saccharomyces cerevisiae and cryotolerant species Saccharomyces uvarum and Saccharomyces kudriavzevii during low-temperature wine fermentation.

    García-Ríos, Estéfani; Querol, Amparo; Guillamón, José Manuel

    2016-09-02

    Temperature is one of the most important parameters to affect the duration and rate of alcoholic fermentation and final wine quality. Some species of the Saccharomyces genus have shown better adaptation at low temperature than Saccharomyces cerevisiae, which was the case of cryotolerant yeasts Saccharomyces uvarum and Saccharomyces kudriavzevii. In an attempt to detect inter-specific metabolic differences, we characterized the proteomic landscape of these cryotolerant species grown at 12°C and 28°C, which we compared with the proteome of S. cerevisiae (poorly adapted at low temperature). Our results showed that the main differences among the proteomic profiling of the three Saccharomyces strains grown at 12°C and 28°C lay in translation, glycolysis and amino acid metabolism. Our data corroborate previous transcriptomic results, which suggest that S. kudriavzevii is better adapted to grow at low temperature as a result of enhanced more efficient translation. Fitter amino acid biosynthetic pathways can also be mechanisms that better explain biomass yield in cryotolerant strains. Yet even at low temperature, S. cerevisiae is the most fermentative competitive species. A higher concentration of glycolytic and alcoholic fermentation enzymes in the S. cerevisiae strain might explain such greater fermentation activity. Temperature is one of the main relevant environmental variables that microorganisms have to cope with and it is also a key factor in some industrial processes that involve microorganisms. However, we are still far from understanding the molecular and physiological mechanisms of adaptation at low temperatures. The results obtained in this study provided a global atlas of the proteome changes triggered by temperature in three different species of the genus Saccharomyces with different degree of cryotolerance. These results would facilitate a better understanding of mechanisms for how yeast could adapt at the low temperature of growth. Copyright © 2016

  12. Saccharomyces kudriavzevii and Saccharomyces uvarum differ from Saccharomyces cerevisiae during the production of aroma-active higher alcohols and acetate esters using their amino acidic precursors.

    Stribny, Jiri; Gamero, Amparo; Pérez-Torrado, Roberto; Querol, Amparo

    2015-07-16

    Higher alcohols and acetate esters are important flavour and aroma components in the food industry. In alcoholic beverages these compounds are produced by yeast during fermentation. Although Saccharomyces cerevisiae is one of the most extensively used species, other species of the Saccharomyces genus have become common in fermentation processes. This study analyses and compares the production of higher alcohols and acetate esters from their amino acidic precursors in three Saccharomyces species: Saccharomyces kudriavzevii, Saccharomyces uvarum and S. cerevisiae. The global volatile compound analysis revealed that S. kudriavzevii produced large amounts of higher alcohols, whereas S. uvarum excelled in the production of acetate esters. Particularly from phenylalanine, S. uvarum produced the largest amounts of 2-phenylethyl acetate, while S. kudriavzevii obtained the greatest 2-phenylethanol formation from this precursor. The present data indicate differences in the amino acid metabolism and subsequent production of flavour-active higher alcohols and acetate esters among the closely related Saccharomyces species. This knowledge will prove useful for developing new enhanced processes in fragrance, flavour, and food industries. Copyright © 2015. Published by Elsevier B.V.

  13. Genome-wide transcription survey on flavour production in Saccharomyces cerevisiae

    Schoondermark-Stolk, Sung A.; Jansen, Michael; Verkleij, Arie J.; Verrips, C. Theo; Euverink, Gert-Jan W.; Dijkhuizen, Lubbert; Boonstra, Johannes

    2006-01-01

    The yeast Saccharomyces cerevisiae is widely used as aroma producer in the preparation of fermented foods and beverages. During food fermentations, secondary metabolites like 3-methyl-1-butanol, 4-methyl-2-oxopentanoate, 3-methyl-2-oxobutanoate and 3-methylbutyrate emerge. These four compounds have

  14. Saccharomyces cerevisiae Mixed Culture of Blackberry (Rubus ulmifolius L.) Juice: Synergism in the Aroma Compounds Production

    Ragazzo-Sánchez, Juan Arturo; Ortiz-Basurto, Rosa Isela; Luna-Solano, Guadalupe; Calderón-Santoyo, Montserrat

    2014-01-01

    Blackberry (Rubus sp.) juice was fermented using four different strains of Saccharomyces cerevisiae (Vitilevure-CM4457, Enoferm-T306, ICV-K1, and Greroche Rhona-L3574) recognized because of their use in the wine industry. A medium alcoholic graduation spirit (component analysis (PCA), and factorial discriminant analysis (DFA) permit to demonstrate the synergism between the strains. PMID:25506606

  15. Function of trehalose and glycogen in cell cycle progression and cell viability in Saccharomyces cerevisiae

    Silljé, H H; Paalman, J W; ter Schure, E G; Olsthoorn, S Q; Verkleij, A J; Boonstra, Johannes; Verrips, C T

    Trehalose and glycogen accumulate in Saccharomyces cerevisiae when growth conditions deteriorate. It has been suggested that aside from functioning as storage factors and stress protectants, these carbohydrates may be required for cell cycle progression at low growth rates under carbon limitation.

  16. Identification and regulation of genes involved in anaerobic growth of Saccharomyces cerevisiae

    Snoek, Isidora Sophia Ishtar

    2007-01-01

    Saccharomyces cerevisiae is one of the few yeast species that can grow equally well without molecular oxygen (anaerobic) as with this compound present (aerobic). This property has made it one of the most abundantly used yeasts in industry, since anaerobic incubation plays a major part in alcohol and

  17. Pathways for Holliday Junction Processing during Homologous Recombination in Saccharomyces cerevisiae

    Ashton, Thomas M; Mankouri, Hocine W; Heidenblut, Anna

    2011-01-01

    The Saccharomyces cerevisiae Rmi1 protein is a component of the highly conserved Sgs1-Top3-Rmi1 complex. Deletion of SGS1, TOP3, or RMI1 is synthetically lethal when combined with the loss of the Mus81-Mms4 or Slx1-Slx4 endonucleases, which have been implicated in Holliday junction (HJ) resolutio...

  18. Lipid Raft-Based Membrane Compartmentation of a Plant Transport Protein Expressed in Saccharomyces cerevisiae

    Grossmann, Q.; Opekarová, Miroslava; Nováková, L.; Stolz, J.; Tanner, W.

    2006-01-01

    Roč. 5, č. 6 (2006), s. 945-953 ISSN 1535-9778 R&D Projects: GA MŠk LC545 Institutional research plan: CEZ:AV0Z50200510 Keywords : saccharomyces cerevisiae * plant transport protein * hup1 Subject RIV: EE - Microbiology, Virology Impact factor: 3.707, year: 2006

  19. Saccharomyces cerevisiae of palm wine-enhanced ethanol production by using mutagens

    Uma, V.; Polasa, H.

    1990-01-01

    The newly isolated Saccharomyces cerevisiae of palm wine produced enhanced amounts of ethanol when cells were UV-irradiated and treated with N-methyl-N-nitro-N-nitrosoguanidine. A further increase of ethanol was observed in yeast extract, peptone, dextrose medium fortified with yeast extract, skimmed milk and soya flour. (author). 9 refs

  20. Aged mother cells of Saccharomyces cerevisiae show markers of oxidative stress and apoptosis

    Laun, P.; Pichová, Alena; Madeo, F.; Fuchs, J.; Ellinger, A.; Kohlwein, S.; Dawes, I.; Fröhlich, K. U.; Breitenbach, M.

    2001-01-01

    Roč. 39, č. 5 (2001), s. 1166-1173 ISSN 0950-382X R&D Projects: GA ČR GA204/97/0541 Institutional research plan: CEZ:AV0Z5020903 Keywords : Saccharomyces cerevisiae * genetic changes Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.398, year: 2001

  1. Transposon mutagenesis to improve the growth of recombinant Saccharomyces cerevisiae on D-xylose

    Haiying Ni; Jose M. Laplaza; Thomas W. Jeffries

    2007-01-01

    Saccharomyces cerevisiae L2612 transformed with genes for xylose reductase and xylitol dehydrogenase (XYL1 and XYL2) grows well on glucose but very poorly on D-xylose. When a gene for D-xylulokinase (XYL3 or XKS1) is overexpressed, growth on glucose is unaffected, but growth on xylose is blocked. Spontaneous or chemically induced mutants of this engineered yeast that...

  2. Effects of low-frequency magnetic fields on the viability of yeast Saccharomyces cerevisiae

    Novák, Jan; Strašák, Luděk; Fojt, Lukáš; Slaninová, I.; Vetterl, Vladimír

    2007-01-01

    Roč. 70, č. 1 (2007), s. 115-121 ISSN 1567-5394 R&D Projects: GA AV ČR(CZ) IAA4004404; GA AV ČR(CZ) IBS5004107 Institutional research plan: CEZ:AV0Z50040702 Keywords : low-frequency electromagnetic field * yeast * Saccharomyces cerevisiae Subject RIV: BO - Biophysics Impact factor: 2.992, year: 2007

  3. Hyper- and hyporesponsive mutant forms of the Saccharomyces cerevisiae Ssy1 amino acid sensor

    Poulsen, Peter; Gaber, Richard F.; Kielland-Brandt, Morten

    2008-01-01

    The Saccharomyces cerevisiae integral membrane protein Ssy1p functions with Ssy5p and Ptr3p to sense extracellular amino acids. Signal transduction leads to processing and nuclear localization of Stp1p and Stp2p, transcriptional activators of many amino acid transporter genes. Ssy1p is structural...

  4. Rad52 multimerization is important for its nuclear localization in Saccharomyces cerevisiae

    Plate, Iben; Albertsen, Line; Lisby, Michael

    2008-01-01

    Rad52 is essential for all homologous recombination and DNA double strand break repair events in Saccharomyces cerevisiae. This protein is multifunctional and contains several domains that allow it to interact with DNA as well as with different repair proteins. However, it has been unclear how Rad...

  5. Phenotypic characterization of glucose repression mutants of Saccharomyce cerevisiae usinge experiments with C-13-labelled glucose

    Vijayendran, Raghevendran; Gombert, A.K.; Christensen, B.

    2004-01-01

    techniques, which do not provide information about the integrated response a specific genetic modification has on the cellular function. In this study we have performed phenotypic characterization of several mutants of the yeast Saccharomyces cerevisiae through the use of experiments with C-13-labelled...

  6. Anaerobic and aerobic batch cultivations of Saccharomyces cerevisiae mutants impaired in glycerol synthesis

    Nissen, Torben Lauesgaard; Hamann, Claus Wendelboe; Kielland-Brandt, M. C.

    2000-01-01

    Glycerol is formed as a by-product in production of ethanol and baker's yeast during fermentation of Saccharomyces cerevisiae under anaerobic and aerobic growth conditions, respectively. One physiological role of glycerol formation by yeast is to reoxidize NADH, formed in synthesis of biomass...

  7. Intracellular pH distribution as a cell health indicator in Saccharomyces cerevisiae

    Aabo, Thomas; Glückstad, Jesper; Siegumfeldt, Henrik

    2011-01-01

    .d.(pHint)) to describe the internal pH distributions. The cellular pH distributional response to external stress such as heat has not previously been determined. In this study, the intracellular pH (pHi) and the s.d.(pHint) of Saccharomyces cerevisiae cells exposed to supralethal temperatures were measured using...

  8. In vivo dynamics of galactose metabolism in Saccharomyces cerevisiae: Metabolic fluxes and metabolite levels

    Østergaard, Simon; Olsson, Lisbeth; Nielsen, Jens

    2001-01-01

    The dynamics of galactose metabolism in Saccharomyces cerevisiae was studied by analyzing the metabolic response of the CEN.PK 113-7D wild-type strain when exposed to a galactose pulse during aerobic growth in a galactose-limited steady-state cultivation at a dilution rate of 0.097 h(-1). A fast...

  9. Optimization of ordered plasmid assembly by gap repair in Saccharomyces cerevisiae

    Eckert-Boulet, Nadine Valerie; Pedersen, Mette Louise; Krogh, Berit Olsen

    2012-01-01

    Combinatorial genetic libraries are powerful tools for diversifying and optimizing biomolecules. The process of library assembly is a major limiting factor for library complexity and quality. Gap repair by homologous recombination in Saccharomyces cerevisiae can facilitate in vivo assembly of DNA...

  10. Modulation of the acute phase response in feedlot steers supplemented with Saccharomyces cerevisiae

    This study was designed to determine the effect of supplementing feedlot steers with Saccharomyces cerevisiae CNCM I-1079 (SC) on the acute phase response to a lipopolysaccharide (LPS) challenge. Steers (n = 18; 266 ± 4 kilograms body weight) were separated into three treatment groups (n = 6/treatm...

  11. Benchmark data for identifying N6-methyladenosine sites in the Saccharomyces cerevisiae genome

    Wei Chen

    2015-12-01

    Full Text Available This data article contains the benchmark dataset for training and testing iRNA-Methyl, a web-server predictor for identifying N6-methyladenosine sites in RNA (Chen et al., 2015 [15]. It can also be used to develop other predictors for identifying N6-methyladenosine sites in the Saccharomyces cerevisiae genome.

  12. Increased xylose affinity of Hxt2 through gene shuffling of hexose transporters in Saccharomyces cerevisiae

    Nijland, Jeroen G; Shin, Hyun Yong; de Waal, Paul P; Klaassen, Paul; Driessen, Arnold J M

    AIMS: Optimizing D-xylose transport in Saccharomyces cerevisiae is essential for efficient bioethanol production from cellulosic materials. We have used a gene shuffling approach of hexose (Hxt) transporters in order to increase the affinity for D-xylose. METHODS AND RESULTS: Various libraries were

  13. Engineering of Saccharomyces cerevisiae for Efficient Anaerobic Alcoholic Fermentation of L-Arabinose

    Wisselink, H.W.; Toirkens, M.J.; Del Rosario Franco Berriel, M.; Winkler, A.A.; Van Dijken, J.P.; Pronk, J.T.; Van Maris, A.J.A.

    2007-01-01

    For cost-effective and efficient ethanol production from lignocellulosic fractions of plant biomass, the conversion of not only major constituents, such as glucose and xylose, but also less predominant sugars, such as L-arabinose, is required. Wild-type strains of Saccharomyces cerevisiae, the

  14. Saccharomyces cerevisiae Mixed Culture of Blackberry (Rubus ulmifolius L.) Juice: Synergism in the Aroma Compounds Production

    Bautista-Rosales, Pedro Ulises; Ragazzo-Sánchez, Juan Arturo; Ruiz-Montañez, Gabriela; Ortiz-Basurto, Rosa Isela; Luna-Solano, Guadalupe; Calderón-Santoyo, Montserrat

    2014-01-01

    Blackberry (Rubus sp.) juice was fermented using four different strains of Saccharomyces cerevisiae (Vitilevure-CM4457, Enoferm-T306, ICV-K1, and Greroche Rhona-L3574) recognized because of their use in the wine industry. A medium alcoholic graduation spirit (

  15. Saccharomyces cerevisiae strains tor second-generation ethanol production : from academie exploration to industrial implementation

    Jansen, Mickel L.A.; Bracher, J.M.; Papapetridis, I.; Verhoeven, M.D.; de Bruijn, J.A.; de Waal, P.; van Maris, A.J.A.; Klaassen, P; Pronk, J.T.

    2017-01-01

    The recent start-up of several full-scale ‘second generation’ ethanol plants marks a major milestone in the development of Saccharomyces cerevisiae strains for fermentation of lignocellulosic hydrolysates of agricultural residues and energy crops. After a discussion of the challenges that these

  16. Genomic Sequence of Saccharomyces cerevisiae BAW-6, a Yeast Strain Optimal for Brewing Barley Shochu.

    Kajiwara, Yasuhiro; Mori, Kazuki; Tashiro, Kosuke; Higuchi, Yujiro; Takegawa, Kaoru; Takashita, Hideharu

    2018-04-05

    Here, we report the draft genome sequence of Saccharomyces cerevisiae strain BAW-6, which is used for the production of barley shochu, a traditional Japanese spirit. This genomic information can be used to elucidate the genetic basis underlying the high alcohol production capacity and citric acid tolerance of shochu yeast. Copyright © 2018 Kajiwara et al.

  17. An in vitro assay for (1-->6)-beta-D-glucan synthesis in Saccharomyces cerevisiae.

    Vink, E.; Rodriguez-Suarez, R.J.; Gerard-Vincent, M.; Ribas, J.C.; de Nobel, J.G.; van den Ende, H.; Duran, A.; Klis, F.M.; Bussey, H.

    2004-01-01

    (1 --> 6)-beta-D-glucan is a key cell wall component of Saccharomyces cerevisiae and Candida albicans. Many genes are known to affect the levels or structure of this glucan, but their roles and a molecular description of the synthesis of (1 --> 6)-beta-D-glucan remain to be established and a method

  18. Prokaryotic diversity of the Saccharomyces cerevisiae Atx1p-mediated copper pathway.

    Bakel, H. van; Huynen, M.A.; Wijmenga, C.

    2004-01-01

    MOTIVATION: Several genes involved in the cellular import of copper and its subsequent incorporation into the high-affinity iron transport complex in Saccharomyces cerevisiae are known to be conserved between eukaryotes and prokaryotes. However, the degree to which these genes share their functional

  19. Growth rate-regulated expression of the hexose transporter HXT5 in Saccharomyces cerevisiae

    Verwaal, René

    2003-01-01

    Glucose, which is the most preferred carbon source for the yeast Saccharomyces cerevisiae, is transported across the plasma membrane into cells by hexose transporter (Hxt) proteins. The Hxt proteins are encoded by a multigene family consisting of 20 members. It was shown previously that HXT1-4 and

  20. The aspartic proteinase from Saccharomyces cerevisiae folds its own inhibitor into a helix

    Li, M; Phylip, L H; Lees, W E

    2000-01-01

    Aspartic proteinase A from yeast is specifically and potently inhibited by a small protein called IA3 from Saccharomyces cerevisiae. Although this inhibitor consists of 68 residues, we show that the inhibitory activity resides within the N-terminal half of the molecule. Structures solved at 2...

  1. Screening and identification of respiration deficiency mutants of yeasts (Saccharomyces Cerevisiae) induced by heavy ion irradiation

    Mao Shuhong; Chinese Academy of Sciences, Beijing; Jin Genming; Wei Zengquan; Xie Hongmei; Zhang Hong

    2006-01-01

    A screen of respiration deficiency mutants of Saccharomyces Cerevisiae induced by 5.19 MeV/u 22 Ne 5- ion irradiation is studied. Some respiration deficiency mutants, which are white colony phenotype in the selective culture of TTC medium, are obtained. The mutants are effectively identified by means of a new and simplified restriction analysis method. (authors)

  2. Mitochondrial genomic dysfunction causes dephosphorylation of Sch9 in the yeast Saccharomyces cerevisiae.

    Kawai, Shigeyuki; Urban, Jörg; Piccolis, Manuele; Panchaud, Nicolas; De Virgilio, Claudio; Loewith, Robbie

    2011-10-01

    TORC1-dependent phosphorylation of Saccharomyces cerevisiae Sch9 was dramatically reduced upon exposure to a protonophore or in respiration-incompetent ρ(0) cells but not in respiration-incompetent pet mutants, providing important insight into the molecular mechanisms governing interorganellar signaling in general and retrograde signaling in particular.

  3. Growth-rate regulated genes have profound impact on interpretation of transcriptome profiling in Saccharomyces cerevisiae

    Regenberg, Birgitte; Grotkjær, Thomas; Winther, Ole

    2006-01-01

    Growth rate is central to the development of cells in all organisms. However, little is known about the impact of changing growth rates. We used continuous cultures to control growth rate and studied the transcriptional program of the model eukaryote Saccharomyces cerevisiae, with generation time...

  4. Two NAD-linked redox shuttles maintain the peroxisomal redox balance in Saccharomyces cerevisiae

    Al-Saryi, Nadal A.; Al-Hejjaj, Murtakab Y.; van Roermund, Carlo W. T.; Hulmes, Georgia E.; Ekal, Lakhan; Payton, Chantell; Wanders, Ronald J. A.; Hettema, Ewald H.

    2017-01-01

    In Saccharomyces cerevisiae, peroxisomes are the sole site of fatty acid beta-oxidation. During this process, NAD(+) is reduced to NADH. When cells are grown on oleate medium, peroxisomal NADH is reoxidised to NAD(+) by malate dehydrogenase (Mdh3p) and reduction equivalents are transferred to the

  5. pH-Dependent Uptake of Fumaric Acid in Saccharomyces cerevisiae under Anaerobic Conditions

    Jamalzadeh, E.; Verheijen, P.J.; Heijnen, J.J.; Van Gulik, W.M.

    2011-01-01

    Microbial production of C4 dicarboxylic acids from renewable resources has gained renewed interest. The yeast Saccharomyces cerevisiae is known as a robust microorganism and is able to grow at low pH, which makes it a suitable candidate for biological production of organic acids. However, a

  6. Ethanol-independent biofilm formation by a flor wine yeast strain of Saccharomyces cerevisiae.

    Zara, Severino; Gross, Michael K; Zara, Giacomo; Budroni, Marilena; Bakalinsky, Alan T

    2010-06-01

    Flor strains of Saccharomyces cerevisiae form a biofilm on the surface of wine at the end of fermentation, when sugar is depleted and growth on ethanol becomes dependent on oxygen. Here, we report greater biofilm formation on glycerol and ethyl acetate and inconsistent formation on succinic, lactic, and acetic acids.

  7. Antioxidant properties and global metabolite screening of the probiotic yeast Saccharomyces cerevisiae var. boulardii.

    Datta, Suprama; Timson, David J; Annapure, Uday S

    2017-07-01

    Saccharomyces cerevisiae var. boulardii is the only yeast species with probiotic properties. It is considered to have therapeutic significance in gastrointestinal disorders. In the present study, a comparative physiological study between this yeast and Saccharomyces cerevisiae (BY4742) was performed by evaluating two prominent traits of probiotic species, responses to different stress conditions and antioxidant capacity. A global metabolite profile was also developed aiming to identify which therapeutically important secondary metabolites are produced. Saccharomyces cerevisiae var. boulardii showed no significant difference in growth patterns but greater stress tolerance compared to S. cerevisiae. It also demonstrated a six- to 10-fold greater antioxidant potential (judged by the 1,1-diphenyl-2-picrylhydrazyl assay), with a 70-fold higher total phenolic content and a 20-fold higher total flavonoid content in the extracellular fraction. These features were clearly differentiated by principal component analysis and further indicated by metabolite profiling. The extracellular fraction of the S. cerevisiae var. boulardii cultures was found to be rich in polyphenolic metabolites: vanillic acid, cinnamic acid, phenyl ethyl alcohol (rose oil), erythromycin, amphetamine and vitamin B 6 , which results in the antioxidant capacity of this strain. The present study presents a new perspective for differentiating the two genetically related strains of yeast, S. cerevisiae and S. cerevisiae var. boulardii by assessing their metabolome fingerprints. In addition to the correlation of the phenotypic properties with the secretory metabolites of these two yeasts, the present study also emphasizes the potential to exploit S. cerevisiae var. boulardii in the industrial production of these metabolites. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  8. "Ant" and "grasshopper" life-history strategies in Saccharomyces cerevisiae.

    Spor, Aymé; Wang, Shaoxiao; Dillmann, Christine; de Vienne, Dominique; Sicard, Delphine

    2008-02-13

    From the evolutionary and ecological points of view, it is essential to distinguish between the genetic and environmental components of the variability of life-history traits and of their trade-offs. Among the factors affecting this variability, the resource uptake rate deserves particular attention, because it depends on both the environment and the genetic background of the individuals. In order to unravel the bases of the life-history strategies in yeast, we grew a collection of twelve strains of Saccharomyces cerevisiae from different industrial and geographical origins in three culture media differing for their glucose content. Using a population dynamics model to fit the change of population size over time, we estimated the intrinsic growth rate (r), the carrying capacity (K), the mean cell size and the glucose consumption rate per cell. The life-history traits, as well as the glucose consumption rate, displayed large genetic and plastic variability and genetic-by-environment interactions. Within each medium, growth rate and carrying capacity were not correlated, but a marked trade-off between these traits was observed over the media, with high K and low r in the glucose rich medium and low K and high r in the other media. The cell size was tightly negatively correlated to carrying capacity in all conditions. The resource consumption rate appeared to be a clear-cut determinant of both the carrying capacity and the cell size in all media, since it accounted for 37% to 84% of the variation of those traits. In a given medium, the strains that consume glucose at high rate have large cell size and low carrying capacity, while the strains that consume glucose at low rate have small cell size but high carrying capacity. These two contrasted behaviors may be metaphorically defined as "ant" and "grasshopper" strategies of resource utilization. Interestingly, a strain may be "ant" in one medium and "grasshopper" in another. These life-history strategies are discussed

  9. Low doses effects of ionizing radiation on Saccharomyces cerevisiae

    Durand, J.; Broock, M. van; Gillette, V.H.

    2000-01-01

    The exposure of living cells to low doses of ionizing radiation induce in response the activation of cellular protection mechanisms against subsequent larger doses of radiation. This cellular adaptive response may vary depending on radiation intensity and time of exposure, and also on the testing probes used whether they were mammalian cells, yeast, bacteria and other organisms or cell types. The mechanisms involved are the genome activation, followed by DNA repair enzymes synthesis. Due to the prompt cell response, the cell cycle can be delayed, and the secondary detoxification of free radicals and/or activation of membrane bound receptors may proceed. All these phenomena are submitted to intense scientific research nowadays, and their elucidation will depend on the complexity of the organism under study. In the present work, the effects of low doses of ionizing radiation (gamma rays) over a suspension of the yeast Saccharomyces cerevisiae (Baker's yeast) was studied, mainly in respect to survival rate and radio-adaptive response. At first, the yeast surviving curve was assessed towards increasing doses, and an estimation of Lethal Dose 50 (LD50) was made. The irradiation tests were performed at LINAC (electrons Linear Accelerator) where electron energy reached approximately 2.65 MeV, and gamma-radiation was produced for bremsstrahlung process over an aluminium screen target. A series of experiments of conditioning doses was performed and an increment surviving fraction was observed when the dose was 2.3 Gy and a interval time between this and a higher dose (challenging dose) of 27 Gy was 90 minutes. A value of 58 ± 4 Gy was estimated for LD50, at a dose rate of 0.44 ± 0.03 Gy/min These quantities must be optimized. Besides data obtained over yeast survival, an unusual increasing amount of tiny yeast colonies appeared on the agar plates after incubation, and this number increased as increasing the time exposure. Preliminary results indicate these colonies as

  10. Network hubs buffer environmental variation in Saccharomyces cerevisiae.

    Sasha F Levy

    2008-11-01

    Full Text Available Regulatory and developmental systems produce phenotypes that are robust to environmental and genetic variation. A gene product that normally contributes to this robustness is termed a phenotypic capacitor. When a phenotypic capacitor fails, for example when challenged by a harsh environment or mutation, the system becomes less robust and thus produces greater phenotypic variation. A functional phenotypic capacitor provides a mechanism by which hidden polymorphism can accumulate, whereas its failure provides a mechanism by which evolutionary change might be promoted. The primary example to date of a phenotypic capacitor is Hsp90, a molecular chaperone that targets a large set of signal transduction proteins. In both Drosophila and Arabidopsis, compromised Hsp90 function results in pleiotropic phenotypic effects dependent on the underlying genotype. For some traits, Hsp90 also appears to buffer stochastic variation, yet the relationship between environmental and genetic buffering remains an important unresolved question. We previously used simulations of knockout mutations in transcriptional networks to predict that many gene products would act as phenotypic capacitors. To test this prediction, we use high-throughput morphological phenotyping of individual yeast cells from single-gene deletion strains to identify gene products that buffer environmental variation in Saccharomyces cerevisiae. We find more than 300 gene products that, when absent, increase morphological variation. Overrepresented among these capacitors are gene products that control chromosome organization and DNA integrity, RNA elongation, protein modification, cell cycle, and response to stimuli such as stress. Capacitors have a high number of synthetic-lethal interactions but knockouts of these genes do not tend to cause severe decreases in growth rate. Each capacitor can be classified based on whether or not it is encoded by a gene with a paralog in the genome. Capacitors with a

  11. Comparative proteomics of mitosis and meiosis in Saccharomyces cerevisiae.

    Kumar, Ravinder; Dhali, Snigdha; Srikanth, Rapole; Ghosh, Santanu Kumar; Srivastava, Sanjeeva

    2014-09-23

    Precise and timely segregation of genetic material and conservation of ploidy are the two foremost requirements for survival of a eukaryotic organism. Two highly regulated cell division processes, namely mitosis and meiosis are central to achieve this objective. The modes of chromosome segregation are distinct in these two processes that generate progeny cells of equal ploidy and half the ploidy in mitosis and meiosis, respectively. Additionally, the nutritional requirement and intracellular processing of biological cue also differ in these two processes. From this, it can be envisaged that proteome of mitotic and meiotic cells will differ significantly. Therefore, identification of proteins that differ in their level of expression between mitosis and meiosis would further reveal the mechanistic detail of these processes. In the present study, we have investigated the protein expression profile of mitosis and meiosis by comparing proteome of budding yeast cultures arrested at mitotic metaphase and metaphase-I of meiosis using proteomic approach. Approximately 1000 and 2000 protein spots were visualized on 2-DE and 2D-DIGE gels respectively, out of which 14 protein spots were significant in 2-DE and 22 in 2D-DIGE (pmitosis, an up-regulation of actin cytoskeleton and its negative regulator occurs in meiosis. Mitosis and meiosis are two different types of cell division cycles with entirely different outcomes with definite biological implication for almost all eukaryotic species. In this work, we investigated, for the first time, the differential proteomic profile of Saccharomyces cerevisiae culture arrested at mitotic metaphase (M) and metaphase-I (MI) of meiosis using 2-DE and 2D-DIGE. Our findings of up-regulation of actin and its negative regulator cofilin during meiosis suggest that the rate of actin cytoskeleton turnover is more in meiosis and actin cytoskeleton may play more crucial role during meiosis compared to mitosis. Present study also suggests that actin

  12. Acquisition of tolerance against oxidative damage in Saccharomyces cerevisiae

    Eleutherio Elis CA

    2001-07-01

    Full Text Available Abstract Background Living cells constantly sense and adapt to redox shifts by the induction of genes whose products act to maintain the cellular redox environment. In the eukaryote Saccharomyces cerevisiae, while stationary cells possess a degree of constitutive resistance towards oxidants, treatment of exponential phase cultures with sub-lethal stresses can lead to the transient induction of protection against subsequent lethal oxidant conditions. The sensors of oxidative stress and the corresponding transcription factors that activate gene expression under these conditions have not yet been completely identified. Results We report the role of SOD1, SOD2 and TPS1 genes (which encode the cytoplasmic Cu/Zn-superoxide dismutase, the mitochondrial Mn-isoform and trehalose-6-phosphate synthase, respectively in the development of resistance to oxidative stress. In all experimental conditions, the cultures were divided into two parts, one was immediately submitted to severe stress (namely: exposure to H2O2, heat shock or ethanol stress while the other was initially adapted to 40°C for 60 min. The deficiency in trehalose synthesis did not impair the acquisition of tolerance to H2O2, but this disaccharide played an essential role in tolerance against heat and ethanol stresses. We also verified that the presence of only one Sodp isoform was sufficient to improve cellular resistance to 5 mM H2O2. On the other hand, while the lack of Sod2p caused high cell sensitivity to ethanol and heat shock, the absence of Sod1p seemed to be beneficial to the process of acquisition of tolerance to these adverse conditions. The increase in oxidation-dependent fluorescence of crude extracts of sod1 mutant cells upon incubation at 40°C was approximately 2-fold higher than in sod2 and control strain extracts. Furthermore, in Western blots, we observed that sod mutants showed a different pattern of Hsp104p and Hsp26p expression also different from that in their control

  13. Micromechanical and surface adhesive properties of single saccharomyces cerevisiae cells

    Farzi, Bahman; Cetinkaya, Cetin

    2017-09-01

    The adhesion and mechanical properties of a biological cell (e.g. cell membrane elasticity and adhesiveness) are often strong indicators for the state of its health. Many existing techniques for determining mechanical properties of cells require direct physical contact with a single cell or a group of cells. Physical contact with the cell can trigger complex mechanotransduction mechanisms, leading to cellular responses, and consequently interfering with measurement accuracy. In the current work, based on ultrasonic excitation and interferometric (optical) motion detection, a non-contact method for characterizing the adhesion and mechanical properties of single cells is presented. It is experimentally demonstrated that the rocking (rigid body) motion and internal vibrational resonance frequencies of a single saccharomyces cerevisiae (SC) (baker’s yeast) cell can be acquired with the current approach, and the Young’s modulus and surface tension of the cell membrane as well as surface adhesion energy can be extracted from the values of these acquired resonance frequencies. The detected resonance frequency ranges for single SC cells include a rocking (rigid body) frequency of 330  ±  70 kHz and two breathing resonance frequencies of 1.53  ±  0.12 and 2.02  ±  0.31 MHz. Based on these values, the average work-of-adhesion of SC cells on a silicon substrate in aqueous medium is extracted, for the first time, as WASC-Si=16.2+/- 3.8 mJ {{m}-2} . Similarly, the surface tension and the Young’s modulus of the SC cell wall are predicted as {{σ }SC}=0.16+/- 0.02 N {{m}-1} and {{E}SC}= 9.20  ±  2.80 MPa, respectively. These results are compared to those reported in the literature by utilizing various methods, and good agreements are found. The current approach eliminates the measurement inaccuracies associated with the physical contact. Exciting and detecting cell dynamics at micro-second time-scales is significantly faster than the

  14. "Ant" and "grasshopper" life-history strategies in Saccharomyces cerevisiae.

    Aymé Spor

    Full Text Available From the evolutionary and ecological points of view, it is essential to distinguish between the genetic and environmental components of the variability of life-history traits and of their trade-offs. Among the factors affecting this variability, the resource uptake rate deserves particular attention, because it depends on both the environment and the genetic background of the individuals. In order to unravel the bases of the life-history strategies in yeast, we grew a collection of twelve strains of Saccharomyces cerevisiae from different industrial and geographical origins in three culture media differing for their glucose content. Using a population dynamics model to fit the change of population size over time, we estimated the intrinsic growth rate (r, the carrying capacity (K, the mean cell size and the glucose consumption rate per cell. The life-history traits, as well as the glucose consumption rate, displayed large genetic and plastic variability and genetic-by-environment interactions. Within each medium, growth rate and carrying capacity were not correlated, but a marked trade-off between these traits was observed over the media, with high K and low r in the glucose rich medium and low K and high r in the other media. The cell size was tightly negatively correlated to carrying capacity in all conditions. The resource consumption rate appeared to be a clear-cut determinant of both the carrying capacity and the cell size in all media, since it accounted for 37% to 84% of the variation of those traits. In a given medium, the strains that consume glucose at high rate have large cell size and low carrying capacity, while the strains that consume glucose at low rate have small cell size but high carrying capacity. These two contrasted behaviors may be metaphorically defined as "ant" and "grasshopper" strategies of resource utilization. Interestingly, a strain may be "ant" in one medium and "grasshopper" in another. These life

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

    Borodina, Irina; Nielsen, Jens

    2014-01-01

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

  16. Effects of Saccharomyces cerevisiae or boulardii yeasts on acute stress induced intestinal dysmotility.

    West, Christine; Stanisz, Andrew M; Wong, Annette; Kunze, Wolfgang A

    2016-12-28

    To investigate the capacity of Saccharomyces cerevisiae ( S. cerevisiae ) and Saccharomyces boulardii ( S. boulardii ) yeasts to reverse or to treat acute stress-related intestinal dysmotility. Adult Swiss Webster mice were stressed for 1 h in a wire-mesh restraint to induce symptoms of intestinal dysmotility and were subsequently killed by cervical dislocation. Jejunal and colon tissue were excised and placed within a tissue perfusion bath in which S. cerevisiae , S. boulardii , or their supernatants were administered into the lumen. Video recordings of contractility and gut diameter changes were converted to spatiotemporal maps and the velocity, frequency, and amplitude of propagating contractile clusters (PCC) were measured. Motility pre- and post-treatment was compared between stressed animals and unstressed controls. S. boulardii and S. cerevisiae helped to mediate the effects of stress on the small and large intestine. Restraint stress reduced jejunal transit velocity (mm/s) from 2.635 ± 0.316 to 1.644 ± 0.238, P boulardii helped to restore jejunal and colonic velocity towards the unstressed controls; 1.833 ± 0.688 to 2.627 ± 0.664, P boulardii or S. cerevisiae supernatants also helped to restore motility to unstressed values in similar capacity. There is a potential therapeutic role for S. cerevisiae and S. boulardii yeasts and their supernatants in the treatment of acute stress-related gut dysmotility.

  17. Outlining a future for non-Saccharomyces yeasts: selection of putative spoilage wine strains to be used in association with Saccharomyces cerevisiae for grape juice fermentation.

    Domizio, Paola; Romani, Cristina; Lencioni, Livio; Comitini, Francesca; Gobbi, Mirko; Mannazzu, Ilaria; Ciani, Maurizio

    2011-06-30

    The use of non-Saccharomyces yeasts that are generally considered as spoilage yeasts, in association with Saccharomyces cerevisiae for grape must fermentation was here evaluated. Analysis of the main oenological characteristics of pure cultures of 55 yeasts belonging to the genera Hanseniaspora, Pichia, Saccharomycodes and Zygosaccharomyces revealed wide biodiversity within each genus. Moreover, many of these non-Saccharomyces strains had interesting oenological properties in terms of fermentation purity, and ethanol and secondary metabolite production. The use of four non-Saccharomyces yeasts (one per genus) in mixed cultures with a commercial S. cerevisiae strain at different S. cerevisiae/non-Saccharomyces inoculum ratios was investigated. This revealed that most of the compounds normally produced at high concentrations by pure cultures of non-Saccharomyces, and which are considered detrimental to wine quality, do not reach threshold taste levels in these mixed fermentations. On the other hand, the analytical profiles of the wines produced by these mixed cultures indicated that depending on the yeast species and the S. cerevisiae/non-Saccharomyces inoculum ratio, these non-Saccharomyces yeasts can be used to increase production of polysaccharides and to modulate the final concentrations of acetic acid and volatile compounds, such as ethyl acetate, phenyl-ethyl acetate, 2-phenyl ethanol, and 2-methyl 1-butanol. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. Saccharomyces cerevisiae variety diastaticus friend or foe?-spoilage potential and brewing ability of different Saccharomyces cerevisiae variety diastaticus yeast isolates by genetic, phenotypic and physiological characterization.

    Meier-Dörnberg, Tim; Kory, Oliver Ingo; Jacob, Fritz; Michel, Maximilian; Hutzler, Mathias

    2018-06-01

    Saccharomyces cerevisiae variety diastaticus is generally considered to be an obligatory spoilage microorganism and spoilage yeast in beer and beer-mixed beverages. Their super-attenuating ability causes increased carbon dioxide concentrations, beer gushing and potential bottle explosion along with changes in flavor, sedimentation and increased turbidity. This research shows clear differences in the super-attenuating properties of S. cerevisiae var. diastaticus yeast strains and their potential for industrial brewing applications. Nineteen unknown spoilage yeast cultures were obtained as isolates and characterized using a broad spectrum of genetic and phenotypic methods. Results indicated that all isolates represent genetically different S. cerevisiae var. diastaticus strains except for strain TUM PI BA 124. Yeast strains were screened for their super-attenuating ability and sporulation. Even if the STA1 gene responsible for super-attenuation by encoding for the enzyme glucoamylase could be verified by real-time polymerase chain reaction, no correlation to the spoilage potential could be demonstrated. Seven strains were further characterized focusing on brewing and sensory properties according to the yeast characterization platform developed by Meier-Dörnberg. Yeast strain TUM 3-H-2 cannot metabolize dextrin and soluble starch and showed no spoilage potential or super-attenuating ability even when the strain belongs to the species S. cerevisiae var. diastaticus. Overall, the beer produced with S. cerevisiae var. diastaticus has a dry and winey body with noticeable phenolic off-flavors desirable in German wheat beers.

  19. A dynamic flux balance model and bottleneck identification of glucose, xylose, xylulose co-fermentation in Saccharomyces cerevisiae

    Economically viable production of lignocellulosic ethanol requires efficient conversion of feedstock sugars to ethanol. Saccharomyces cerevisiae cannot ferment xylose, the main five-carbon sugars in biomass, but can ferment xylulose, an enzymatically derived isomer. Xylulose fermentation is slow rel...

  20. Removal of Strontium Ions by Immobilized Saccharomyces Cerevisiae in Magnetic Chitosan Microspheres

    Yanan Yin

    2017-02-01

    Full Text Available A novel biosorbent, immobilized Saccharomyces cerevisiae in magnetic chitosan microspheres was prepared, characterized, and used for the removal of Sr2+ from aqueous solution. The structure and morphology of immobilized S. cerevisiae before and after Sr2+adsorption were observed using scanning electron microscopy with energy dispersive X-ray spectroscopy. The experimental results showed that the Langmuir and Freundlich isotherm models could be used to describe the Sr2+ adsorption onto immobilized S. cerevisiae microspheres. The maximal adsorption capacity (qm was calculated to be 81.96 mg/g by the Langmuir model. Immobilized S. cerevisiae was an effective adsorbent for the Sr2+ removal from aqueous solution.

  1. Microbial cells as biosorbents for heavy metals: accumulation of Uranium by Saccharomyces cerevisiae and Pseudomonas aeruginosa

    Strandberg, G.W.; Shumate, S.E. II; Parrott, J.R. Jr.

    1981-01-01

    Uranium accumulated extracellularly on the surfaces of Saccharomyces cerevisiae cells. The rate and extent of accumulation were subject to environmental parameters, such as pH, temperature, and interference by certain anions and cations. Uranium accumulation by Pseudomonas aeruginosa occurred intracellularly and was extremely rapid (<10 s), and no response to environmental parameters could be detected. Metabolism was not required for metal uptake by either organism. Cell-bound uranium reached a concentration of 10 to 15% of the dry cell weight, but only 32% of the S. cerevisiae cells and 44% of the P. aeruginosa cells within a given population possessed visible uranium deposits when examined by electron microscopy. Rates of uranium uptake by S. cerevisiae were increased by chemical pretreatment of the cells. Uranium could be removed chemically from S. cerevisiae cells, and the cells could then be reused as a biosorbent

  2. Evaluation of Lactobacillus plantarum and Saccharomyces cerevisiae in the Presence of Bifenthrin.

    Đorđević, Tijana M; Đurović-Pejčev, Rada D

    2016-06-01

    This work describes the effect of insecticide bifenthrin on Lactobacillus plantarum and Saccharomyces cerevisiae. Growths of used microorganisms in growth media supplemented with pesticide were studied. Determination of bacterial and yeast fermentation efficiency in wheat supplemented with bifenthrin was conducted. Additionally, investigation of bifenthrin dissipation during microbiological activity was performed. Experiments applying bifenthrin in different concentrations highlighted a negligible impact of the pesticide on the growth of L. plantarum and S. cerevisiae. This insecticide overall negatively affected the yeast fermentation of wheat, while its presence in wheat had a slight negative impact on lactic acid fermentation. The results of bifenthrin dissipation during lactic acid and yeast fermentations of wheat showed that activities of L. plantarum and S. cerevisiae caused lower pesticide reductions. Average bifenthrin residue reduction within samples fermented with L. plantarum was 5.4 % (maximum ~16 %), while within samples fermented with S. cerevisiae, it was 11.6 % (maximum ~17 %).

  3. Enhancing Fatty Acid Production of Saccharomyces cerevisiae as an Animal Feed Supplement.

    You, Seung Kyou; Joo, Young-Chul; Kang, Dae Hee; Shin, Sang Kyu; Hyeon, Jeong Eun; Woo, Han Min; Um, Youngsoon; Park, Chulhwan; Han, Sung Ok

    2017-12-20

    Saccharomyces cerevisiae is used for edible purposes, such as human food or as an animal feed supplement. Fatty acids are also beneficial as feed supplements, but S. cerevisiae produces small amounts of fatty acids. In this study, we enhanced fatty acid production of S. cerevisiae by overexpressing acetyl-CoA carboxylase, thioesterase, and malic enzyme associated with fatty acid metabolism. The enhanced strain pAMT showed 2.4-fold higher fatty acids than the wild-type strain. To further increase the fatty acids, various nitrogen sources were analyzed and calcium nitrate was selected as an optimal nitrogen source for fatty acid production. By concentration optimization, 672 mg/L of fatty acids was produced, which was 4.7-fold higher than wild-type strain. These results complement the low level fatty acid production and make it possible to obtain the benefits of fatty acids as an animal feed supplement while, simultaneously, maintaining the advantages of S. cerevisiae.

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

    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.

  5. Removal of strontium ions by immobilized saccharomyces cerevisiae in magnetic chitosan microspheres

    Yin, Yanan; Wang, Jian Long; Yang, Xiao Yong; Li, Weihua [Collaborative Innovation Center for Advanced Nuclear Energy Technology, Institute of Nuclear and New Energy Technology, Tsinghua University, Beijing (China)

    2017-02-15

    A novel biosorbent, immobilized Saccharomyces cerevisiae in magnetic chitosan microspheres was prepared, characterized, and used for the removal of Sr{sup 2+} from aqueous solution. The structure and morphology of immobilized S. cerevisiae before and after Sr{sup 2+}adsorption were observed using scanning electron microscopy with energy dispersive X-ray spectroscopy. The experimental results showed that the Langmuir and Freundlich isotherm models could be used to describe the Sr{sup 2+} adsorption onto immobilized S. cerevisiae microspheres. The maximal adsorption capacity (q{sub m}) was calculated to be 81.96 mg/g by the Langmuir model. Immobilized S. cerevisiae was an effective adsorbent for the Sr{sup 2+} removal from aqueous solution.

  6. Performance evaluation of Pichia kluyveri, Kluyveromyces marxianus and Saccharomyces cerevisiae in industrial tequila fermentation.

    Amaya-Delgado, L; Herrera-López, E J; Arrizon, Javier; Arellano-Plaza, M; Gschaedler, A

    2013-05-01

    Traditionally, industrial tequila production has used spontaneous fermentation or Saccharomyces cerevisiae yeast strains. Despite the potential of non-Saccharomyces strains for alcoholic fermentation, few studies have been performed at industrial level with these yeasts. Therefore, in this work, Agave tequilana juice was fermented at an industrial level using two non-Saccharomyces yeasts (Pichia kluyveri and Kluyveromyces marxianus) with fermentation efficiency higher than 85 %. Pichia kluyveri (GRO3) was more efficient for alcohol and ethyl lactate production than S. cerevisiae (AR5), while Kluyveromyces marxianus (GRO6) produced more isobutanol and ethyl-acetate than S. cerevisiae (AR5). The level of volatile compounds at the end of fermentation was compared with the tequila standard regulation. All volatile compounds were within the allowed range except for methanol, which was higher for S. cerevisiae (AR5) and K. marxianus (GRO6). The variations in methanol may have been caused by the Agave tequilana used for the tests, since this compound is not synthesized by these yeasts.

  7. Interactions between Lactobacillus kefiranofaciens and Saccharomyces cerevisiae in mixed culture for kefiran production.

    Cheirsilp, Benjamas; Shoji, Hirofumi; Shimizu, Hiroshi; Shioya, Suteaki

    2003-01-01

    Since a positive effect on the growth and kefiran production of Lactobacillus kefiranofaciens was observed in a mixed culture with Saccharomyces cerevisiae, the elucidation of the interactions between L. kefiranofaciens and S. cerevisiae may lead to higher productivity. Hence, the microbial interaction of each strain was investigated. Apart from the positive effect of a reduction in the amount of lactic acid by S. cerevisiae, a positive effect of S. cerevisiae on the growth and kefiran production of L. kefiranofaciens in a mixed culture was observed. Various experiments were carried out to study this effect. In this study, the observed increase in capsular kefiran in a mixed culture with inactivated S. cerevisiae correlated well to that in an anaerobic mixed culture. Differences in capsular kefiran production were observed for different initial S. cerevisiae concentrations under anaerobic conditions. From these fermentation results, it was concluded that the physical contact with S. cerevisiae mainly enhanced the capsular kefiran production of L. kefiranofaciens in a mixed culture. Therefore, in an anaerobic mixed culture, this direct contact resulted in higher capsular kefiran production than that in pure culture.

  8. Secretory Overexpression of Bacillus thermocatenulatus Lipase in Saccharomyces cerevisiae Using Combinatorial Library Strategy.

    Kajiwara, Shota; Yamada, Ryosuke; Ogino, Hiroyasu

    2018-04-10

    Simple and cost-effective lipase expression host microorganisms are highly desirable. A combinatorial library strategy is used to improve the secretory expression of lipase from Bacillus thermocatenulatus (BTL2) in the culture supernatant of Saccharomyces cerevisiae. A plasmid library including expression cassettes composed of sequences encoding one of each 15 promoters, 15 secretion signals, and 15 terminators derived from yeast species, S. cerevisiae, Pichia pastoris, and Hansenula polymorpha, is constructed. The S. cerevisiae transformant YPH499/D4, comprising H. polymorpha GAP promoter, S. cerevisiae SAG1 secretion signal, and P. pastoris AOX1 terminator, is selected by high-throughput screening. This transformant expresses BTL2 extra-cellularly with a 130-fold higher than the control strain, comprising S. cerevisiae PGK1 promoter, S. cerevisiae α-factor secretion signal, and S. cerevisiae PGK1 terminator, after cultivation for 72 h. This combinatorial library strategy holds promising potential for application in the optimization of the secretory expression of proteins in yeast. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    Guoqiang Xu

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

  10. Local isolate of Saccharomyces cerevisiae as biocompetitive agent of Aspergillus flavus

    Eni Kusumaningtyas

    2006-12-01

    Full Text Available Aspergillus flavus is a toxigenic fungus that contaminates feed and influences the animal health. Saccharomyces cerevisiae can be used as a biocompetitive agent to control the contamination. The ability of local isolate of S. cerevisiae as a biocompetitive agent for A. flavus was evaluated. A. flavus (30ml was swept on Sabouraud dextrose agar (SDA, while S. cerevisiae was swept on its left and right. Plates were incubated at 28oC for nine days. Lytic activity of S. cerevisiae was detected by pouring its suspension on the centre of the cross streaks of A. flavus. Plates were incubated at 28oC for five days. Growth inhibition of A. flavus by S. cerevisiae was determined by mixing the two fungi on Potato dextrose broth and incubated at 28oC for 24 hours. Total colony of A. flavus were then observed at incubation time of 2, 4, 6 and 24 hours by pour plates method on the SDA plates and incubated on 28oC for two days. Growth of hyphae of A. flavus sweep were inhibited with the swept of S. cerevisiae. The width of A. flavus colony treated with S. cerevisiae is narrower (3,02 cm than that of control ( 4,60 cm. The growth of A. flavus was also inhibited on the centre of cross streak where the S. cerevisiae poured. S. cerevisiae gradually reduced the colony number of A. flavus in the mixed culture of broth fungi ie. 14 x 103 CFU/ml while colony number of control is 80 x 103 CFU/ml. Results showed that S. cerevisiae could be used as biocompetitive agent of A. flavus.

  11. Estudio de nuevas levaduras Killer "Saccharomyces cerevisiae" y "Torulaspora delbrueckii" para elaborar vinos tranquilos y espumosos

    Velázquez Molinero, Rocío

    2016-01-01

    Se analizan dos nuevos tipos de levaduras vínicas killer de amplio espectro antifúngico: Sacharomyces cerevisiae Klus y Torulaspora delbrueckii Kbarr. Ambas matan a todos los tipos de levaduras S. cerevisiae conocidos, killer y sensibles, además de muchas otras especies de levaduras no-Saccharomyces. El receptor de la pared celular de las levaduras sensibles a ambas toxinas parece ser el beta-glucano. El fenotipo killer de estas levaduras está codificado en virus de dsRNA de tamaño mediano, M...

  12. Improved ethanol production from whey Saccharomyces cerevisiae using permeabilized cells of Kluyveromyces marxianus

    Rosenberg, M [Slovak Technical Univ., Bratislava (Slovakia). Dept. of Biochemical Technology; Tomaska, M [Slovak Technical Univ., Bratislava (Slovakia). Dept. of Biochemical Technology; Kanuch, J [Slovak Technical Univ., Bratislava (Slovakia). Dept. of Biochemical Technology; Sturdik, E [Slovak Technical Univ., Bratislava (Slovakia). Dept. of Biochemical Technology

    1996-12-31

    Permeabilized cells of Kluyveromyces marxianus CCY eSY2 were tested as the source of lactase in the ethanol fermentation of concentrated deproteinized whey (65-70 g/l lactose) by Saccharomyces cerevisiae CCY 10-13-14. Rapid lactose hydrolysis by small amounts of permeabilized cells following the fermentation of released glucose and galactose by S. cerevisiae resulted in a twofold enhancement of the overall volumetric productivity (1.03 g/lxh), compared to the fermentation in which the lactose was directly fermented by K. marxianus. (orig.)

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

    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...

  14. Saccharomyces cerevisiae UE-ME3 is a good strain for isoproturon biorremediation?

    Candeias, M; Alves-Pereira, I; Ferreira, R

    2010-01-01

    Isoproturon, an herbicide of pre- and pos-emergence of Autumn-Winter crops, persists occasionally in soil, groundwater and biological systems at levels above those established by European Directives. Saccharomyces cerevisiae UE-ME3 exposed in stationary phase to 50 and 100 mM isoproturon exhibit growth rates higher than control or exposed cells to 5 and 25 mM of this phenylurea. However, in S.cerevisiae UE-ME3 grown in the presence of 5 mM isoproturon, were observed a decrease of ...

  15. Expression of an Aspergillus niger Phytase Gene (phyA) in Saccharomyces cerevisiae

    Han, Yanming; Wilson, David B.; Lei, Xin gen

    1999-01-01

    Phytase improves the bioavailability of phytate phosphorus in plant foods to humans and animals and reduces phosphorus pollution of animal waste. Our objectives were to express an Aspergillus niger phytase gene (phyA) in Saccharomyces cerevisiae and to determine the effects of glycosylation on the phytase’s activity and thermostability. A 1.4-kb DNA fragment containing the coding region of the phyA gene was inserted into the expression vector pYES2 and was expressed in S. cerevisiae as an act...

  16. Growth-rate dependency of de novo resveratrol production in chemostat cultures of an engineered Saccharomyces cerevisiae strain

    Vos, T.; De la Torre Cortes, P.; Van Gulik, W.M.; Pronk, J.T.; Daran-Lapujade, P.A.S.

    2015-01-01

    Introduction: Saccharomyces cerevisiae has become a popular host for production of non-native compounds. The metabolic pathways involved generally require a net input of energy. To maximize the ATP yield on sugar in S. cerevisiae, industrial cultivation is typically performed in aerobic,

  17. Excessive by-product formation : A key contributor to low isobutanol yields of engineered Saccharomyces cerevisiae strains

    Milne, N.S.W.; Wahl, S.A.; Van Maris, A.J.A.; Pronk, J.T.; Daran, J.M.

    2016-01-01

    It is theoretically possible to engineer Saccharomyces cerevisiae strains in which isobutanol is the predominant catabolic product and high-yielding isobutanol-producing strains are already reported by industry. Conversely, isobutanol yields of engineered S. cerevisiae strains reported in the

  18. Optimizing anaerobic growth rate and fermentation kinetics in Saccharomyces cerevisiae strains expressing Calvin-cycle enzymes for improved ethanol yield

    Papapetridis, I.; Goudriaan, M.; De Keijzer, Nikita A.; van den Broek, M.A.; van Maris, A.J.A.; Pronk, J.T.

    2018-01-01

    Background: Reduction or elimination of by-product formation is of immediate economic relevance in fermentation processes for industrial bioethanol production with the yeast Saccharomyces cerevisiae. Anaerobic cultures of wild-type S. cerevisiae require formation of glycerol to maintain the

  19. An internal deletion in MTH1 enables growth on glucose of pyruvate-decarboxylase negative, non-fermentative Saccharomyces cerevisiae

    Oud, B.; Flores, C.L.; Gancedo, C.; Zhang, X.; Trueheart, J.; Daran, J.M.; Pronk, J.T.; Van Maris, A.J.A.

    2012-01-01

    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

  20. Integrated phospholipidomics and transcriptomics analysis of Saccharomyces cerevisiae with enhanced tolerance to a mixture of acetic acid, furfural, and phenol

    A mixture of acetic acid, furfural and phenol (AFP), three representative lignocellulose derived inhibitors, significantly inhibited the growth and bioethanol production of Saccharomyces cerevisiae. In order to uncover mechanisms behind the enhanced tolerance of an inhibitor-tolerant S.cerevisiae s...

  1. Directed Evolution towards Increased Isoprenoid Production in Saccharomyces cerevisiae

    Carlsen, Simon; Nielsen, Michael Lynge; Kielland-Brandt, Morten

    production can easily be scaled to meet current demands and it is also an environmental benign production method compared to organic synthesis. Thus it would be attractive to engineer a microorganism to produce high amounts of IPP and other immediate prenyl precursors such as geranyl pyrophosphate, farnesyl...... 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...... coloration which is the result of higher amount of lycopene is being produced and hence high amount of isoprenoid precursor being available. This will elucidate novel genetic targets for increasing isoprenoid production in S. cerevisiae...

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

    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:

  3. Co-cultivation of non-conventional yeast with Saccharomyces cerevisiae to increase the aroma complexity of fermented beverages

    Rijswijck, van, Irma M.H.

    2017-01-01

    Yeast are used as workhorses to convert hopped wort into beer. Conventionally, such yeasts belong to the genus Saccharomyces and most research on fermentation of wort for the production of beer has focussed on the species Saccharomyces cerevisiae and Saccharomyces pastorianus. Recently, there is an increasing interest in unravelling features of non-conventional yeast species for beer innovation. In this thesis, features of yeast isolates belonging to the species: Cyberlindnera fabianii, Pichi...

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

    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.

  5. Amperometric Biosensor for Monitoring Respiration Activity of Saccharomyces cerevisiae in the Presence of Cobalt and Zinc

    Miroslav Mikšaj

    2002-01-01

    Full Text Available For efficient control of heavy metal concentrations electrochemical methods, such as polarography and related techniques, are applied. Their advantages are simplicity, short analysis time and small quantities of samples needed. The presence of some heavy metals, such as zinc and cobalt, accelerates the growth of yeast. For the measurements of concentration changes, amperometric biosensor containing yeast Saccharomyces cerevisiae was used. The influence of zinc and cobalt on respiratory activity of the yeast Saccharomyces cerevisiae was estimated by measuring oxygen in the solution that was earlier enriched with cobalt or zinc. Measurements were performed using modified Clark’s oxygen electrode and the investigated concentrations of cobalt and zinc were up to 100 mg/L.

  6. ISOTERMAS DE ADSORÇÃO DE CÁDMIO POR Saccharomyces cerevisiae ISOTHERMS OF CADMIUM ADSORPTION BY Saccharomyces cerevisae

    Silvana ALBERTINI

    2001-08-01

    Full Text Available Com o objetivo de determinar as isotermas de adsorção de cádmio por Saccharomyces cerevisiae, foram utilizados os sais cloreto e nitrato de cádmio nas concentrações de 5, 10, 20, 40, 60, 80 e 100mg L-1. A biomassa foi produzida a partir de uma cultura "starter"de Saccharomyces cerevisiae IZ 1904. Após o contato de 16h entre o microrganismo e as soluções em estudo, a biomassa foi separada por centrifugação e o teor de cádmio residual foi determinado no sobrenadante por espectrofotometria de absorção atômica. Para os dois sais empregados foi observado um acúmulo crescente de cádmio nas concentrações de 5, 10, 20 e 40mg L-1. Nas concentrações de 60, 80 e 100mg L-1 foi observado que a levedura acumulou teores menores do metal, evidenciando danos na parede celular, nem sempre acompanhados de iguais danos da membrana citoplasmática, tais alterações da parede visualizadas por microscopia eletrônica de varredura.With the objective of determining the isotherms of cadmium the adsorption by Saccharomyces cerevisiae, the chloride and nitrate salts were used in the concentrations of 5, 10, 20, 40, 60, 80, and 100mg L-1. The biomass was produced from a starter culture of Saccharomyces cerevisiae IZ 1904. After a 16h contact between the microrganism and solutions of study the biomass was separated by a centrifuge and the cadmium residue content was determined at the supernatant by atomic adsorption spectrophotometry. For the two salts used a growing accumulation of cadmium was observed at concentrations of 5, 10, 20, and 40mg L-1. In the concentrations of 60, 80 and 100mg L-1 a decreasing of the accumulation of the metal was observed, evidencing damages of the cellular wall, which they're not accompanied always by damages of the citoplasmatic membrane, visualized by scanning electron microscopy.

  7. Rekayasa Glukosa Dari Tandan Kosong Kelapa Sawit Melalui Proses Fermentasi Dengan Saccharomyces cerevisiae Menjadi Bioetanol

    Nasruddin Nasruddin

    2013-06-01

    Full Text Available This research aims to study the performance of Saccharomyces cerevisiae in glucose engineering into bioethanol. Glucose comes from palm oil empty fruit bunches that had been pretreated by delignification and fermentation. Glucose solution result from hydrolysis for each treatment of 500 ml was fermented with Saccharomyces cerevisiae (2, 4, 6 and 8 g, fermentation time (4, 6, 8 and 10 days. Result of fermentation was distilled at 75°C ± 5°C for 60 minutes. Bioethanol produced were tested including: specific gravity by using picnometer and acidity was tested by volumetric methods. The analysis showed that the best bioethanol produced in this experiment, followed by laboratory tests obtained from the interaction between treatments for time of hydrolysis by Aspergillus niger for 6 days, with 4 grams of Saccharomyces cerevisiae fermentation for 6 days. Based on the test results of bioethanol obtained density 0.9873 g/cm3, percentage of bioethanol 9.2889% (v/v and acid number value 1.820 mg/L.ABSTRAKPenelitian ini bertujuan untuk mempelajarai kinerja Saccharomyces cerevisiae  merekayasa glukosa menjadi bioetanol. Glukosa berasal dari tandan kosong kelapa sawit yang telah dilakukan pretreatment dengan cara delignifikasi dan fermentasi. Larutan glukosa hasil hidrolisis untuk masing-masing perlakuan sebanyak 500 mL difermentasi dengan S. cerevisiae (2; 4; 6 dan 8 g, waktu fermentasi (4; 6; 8 dan 10 hari. Hasil fermentasi didestilasi pada suhu 75oC ± 5oC selama 60 menit. Bioetanol yang dihasilkan diuji yang meliputi : berat jenis dengan mengunakan piknometer dan keasaman diuji dengan metode volumetri. Hasil analisis menunjukkan bioetanol yang terbaik berdasarkan hasil percobaan yang dilanjutkan dengan uji laboratorium didapatkan dari interaksi antar perlakuan untuk waktu hidrolisis dengan Aspergilus niger selama 6 hari, fermentasi dengan 4 gram Saccharomyces cerevisiae selama 6 hari. Berdasarkan hasil uji bioetanol untuk berat jenis 0,9873 g/cm3

  8. Investigation of the effect of water exposed to nonequilibrium contact plasma onto saccharomyces cerevisiae yeast

    S. Mykolenko

    2015-05-01

    Full Text Available Introduction. Additional treatment of water by nonequilibrium contact plasma allows improving consumer characteristics of bakery goods considerably. Determination of the effect of plasma-chemically activated water on morphological, cultural and physiological properties of Saccharomyces cerevisiae yeast is important from the technological point of view. Materials and Methods. Experimental investigations were carried out in the conditions of bacteriological laboratory by seeding the culture of yeasts of ТМ “Lvivski” and “Kryvorizki” on Sabouraud dense liquid nutrient media. The quantity of viable cells of microorganisms was determined by the method of Gould sector seeds. Morphology of the yeast was investigated by phase-contrast microscopy. Biotechnological properties of yeasts were determined on Giss media. Results. The paper establishes the effect of water exposed to nonequilibrium contact plasma on the sensitivity of Saccharomyces cerevisiae and shows absence of suppressive action of treated water with regard to cultural properties of microorganisms. The experiments prove that with the use of plasma-chemically activated water morphological characteristics and biochemical properties of bakery yeasts produced by Lviv and Kryvyi Rig yeast plants are preserved. Culturing of Saccharomyces cerevisiae yeast on the nutrient media prepared with the use of water exposed to nonequilibrium contact plasm resulted in 6,5–15 times’ increase in quantity of viable microorganisms compared with the control on the mains drinking water. Conclusions. Physiological properties of Saccharomyces cerevisiae yeast improved owing to use water exposed to nonequilibrium contact plasma. Results of investigations are recommended for using in yeast production and bread making.

  9. The Impact of Single Amino Acids on Growth and Volatile Aroma Production by Saccharomyces cerevisiae Strains

    Samantha Fairbairn; Alexander McKinnon; Hannibal T. Musarurwa; António C. Ferreira; António C. Ferreira; Florian F. Bauer

    2017-01-01

    Nitrogen availability and utilization by Saccharomyces cerevisiae significantly influence fermentation kinetics and the production of volatile compounds important for wine aroma. Amino acids are the most important nitrogen source and have been classified based on how well they support growth. This study evaluated the effect of single amino acids on growth kinetics and major volatile production of two phenotypically different commercial wine yeast strains in synthetic grape must. Four growth p...

  10. Efectes de les concentracions de glucosa i etanol inicials en el creixement de Saccharomyces cerevisiae

    Cañadillas Castells, Jordi

    2014-01-01

    Saccharomyces cerevisiae Is one of the most used microorganisms to the alimentary industry. To optimise the process of fermentation is necessary to know the parameters that impact in his growth in order to modulate the specific speed in function of the biotechnological needs. They have described different parameters that impact in the growth of the yeast how are the concentration of sugars, of ethanol, pH, and temperature between others. The experimental work has for main aim study the kineti...

  11. Effects of furfural on the respiratory metabolism of Saccharomyces cerevisiae in glucose-limited chemostats,

    Sarvari Horvath, I; Franzén, C J; Taherzadeh, M J; Niklasson, C; Lidén, Gunnar

    2003-01-01

    Effects of furfural on the aerobic metabolism of the yeast Saccharomyces cerevisiae were studied by performing chemostat experiments, and the kinetics of furfural conversion was analyzed by performing dynamic experiments. Furfural, an important inhibitor present in lignocellulosic hydrolysates, was shown to have an inhibitory effect on yeast cells growing respiratively which was much greater than the inhibitory effect previously observed for anaerobically growing yeast cells. The residual fur...

  12. Saccharomyces cerevisiae has distinct adaptive responses to both hydrogen peroxide and menadione.

    Jamieson, D J

    1992-01-01

    Treatment of Saccharomyces cerevisiae cells with low concentrations of either hydrogen peroxide or menadione (a superoxide-generating agent) induces adaptive responses which protect cells from the lethal effects of subsequent challenge with higher concentrations of these oxidants. Pretreatment with menadione is protective against cell killing by hydrogen peroxide; however, pretreatment with hydrogen peroxide is unable to protect cells from subsequent challenge with menadione. This suggests th...

  13. Isolation of glutathione-deficient mutants of the yeast Saccharomyces cerevisiae

    Kistler, M.; Eckardt, F.; Summer, K.-H.

    1986-01-01

    Glutathione-deficient (gsh - ) mutants of the yeast Saccharomyces cerevisiae were isolated after UV treatment using MNNG as selective agent. For genetic and biochemical characterization 5 mutant strains were chosen which exhibited considerably decreased residual GSH contents varying from 2 to 6% of the wild-type levels. All 5 isolates showed a 2:2 segregation of the gsh - :GSH + phenotypes alluding to a monogenic recessive mutation. Complementation analysis indicates that all gsh - mutants belong to one complementation group. (Auth.)

  14. A special cell morphology of saccharomyces cerevisiae induced by low-temperature plasma

    Ling Dajun; Cao Jinxiang

    2003-01-01

    A special cell morphology, cavity-like cells, was found in posterities of Saccharomyces cerevisiae treated by low-temperature air plasma with different powers. The feature of the special morphology indicates that the cavity-like cells may be formed by cellular mutation effect induced by the plasma, instead of direct cellular damage by the plasma. The results suggest that the cellular mutation effect of the low-temperature plasma is a complex process

  15. Photoreactivity in Saccharomyces cerevisiae cells after irradiation with 25 MeV electrons

    Tsyb, T.S.; Seleva, N.G.; Myasnik, M.N.; Kabakova, N.M.

    1986-01-01

    Significant photoreactivation was noted in radio- and UV-sensitive rad-mutants of Saccharomyces cerevisiae cells exposed to 25 MeV electrons. In order to make the photoreactivable damage be manifest anoxic conditions of irradiation should be chosen as optimal ones. It was shown that the low oxygen effect was partially associated with the photoreactivable damage involved in the lethal effect of ionizing radiation

  16. The yeast Saccharomyces cerevisiae DNA polymerase IV: possible involvement in double strand break DNA repair.

    Leem, S H; Ropp, P A; Sugino, A

    1994-01-01

    We identified and purified a new DNA polymerase (DNA polymerase IV), which is similar to mammalian DNA polymerase beta, from Saccharomyces cerevisiae and suggested that it is encoded by YCR14C (POLX) on chromosome III. Here, we provided a direct evidence that the purified DNA polymerase IV is indeed encoded by POLX. Strains harboring a pol4 deletion mutation exhibit neither mitotic growth defect nor a meiosis defect, suggesting that DNA polymerase IV participates in nonessential functions in ...

  17. Lack of cortical endoplasmic reticulum protein Ist2 alters sodium accumulation in Saccharomyces cerevisiae cells

    Papoušková, Klára; Andršová, Markéta; Sychrová, Hana

    2017-01-01

    Roč. 17, č. 2 (2017), č. článku fox011. ISSN 1567-1356 R&D Projects: GA MŠk(CZ) LH14297 Institutional support: RVO:67985823 Keywords : Saccharomyces cerevisiae * Ist2 * alkali-metal- cation homeostasis * sodium tolerance * sodium uptake * alkali-metal- cation transporters Subject RIV: EE - Microbiology, Virology OBOR OECD: Mycology Impact factor: 3.299, year: 2016

  18. Specific distribution of the Saccharomyces cerevisiae linker histone homolog HHO1p in the chromatin

    Freidkin, Ilya; Katcoff, Don J.

    2001-01-01

    In virtually all eukaryotic organisms, linker DNA between nucleosomes is associated with a histone termed linker histone or histone H1. In Saccharomyces cerevisiae, HHO1 encodes a putative linker histone with very significant homology to histone H1. The encoded protein is expressed in the nucleus, but has not been shown to affect global chromatin structure, nor has its deletion shown any detectable phenotype. In vitro chromatin assembly experiments with recombinant HHO1p have shown that it is...

  19. Genetic diversity and molecular characterization of Saccharomyces cerevisiae strains from winemaking environments

    Schuller, Dorit Elisabeth

    2004-01-01

    Tese de doutoramento em Ciências The principal aim of the present work is to assess the genetic diversity of fermenting Saccharomyces cerevisiae strains found in vineyards belonging to the Vinho Verde Region in order to create a strain collection representing the region’s biodiversity wealth as a basis for future strain selection and improvement programs. Validation of molecular techniques for accurate genotyping is an indispensable prerequisite for biogeographical surveys. Molecular ty...

  20. Levels of acid-soluble polyphosphate in growing cultures of Saccharomyces cerevisiae.

    Solimene, R; Guerrini, A M; Donini, P

    1980-01-01

    Short-chain acid-soluble polyphosphates were extracted from growing cultures of Saccharomyces cerevisiae, and the changes in the levels of these compounds were determined. The production of acid-soluble polyphosphates correlated with the mitochondrial activities since it occurred in two bursts in respiration-competent yeast cells and in only one burst in respiration-deficient yeast cells. The possible role of these compounds is discussed.

  1. Effect of carbon source on the accumulation of cytochrome P-450 in the yeast Saccharomyces cerevisiae.

    Kärenlampi, S O; Marin, E; Hänninen, O O

    1981-01-01

    The appearance of cytochrome P-450 in the yeast Saccharomyces cerevisiae depended on the substrate supporting growth. Cytochrome P-450 was apparent in yeast cells grown on a strongly fermentable sugar such as D-glucose, D-fructose or sucrose. When yeast was grown on D-galactose, D-mannose or maltose, where fermentation and respiration occurred concomitantly, cytochrome P-450 was also formed. The cytochrome P-450 concentration was maximal at the beginning of the stationary phase of the culture...

  2. Effect of aeration on the fermentative activity of Saccharomyces cerevisiae cultured in apple juice

    Estela-Escalante, W.; Rychtera, M.; Melzoch, K.; Hatta-Sakoda, B.

    2012-01-01

    The influence of aeration on the fermentative activity of Saccharomyces cerevisiaeRTVE V 15-1-416 was studied in order to evaluate the synthesis of fermentation by-products. To achieve this, the strain was cultured in Erlenmeyer flasks and bioreactor containing sterilized and aroma removed apple juice. The chemical compounds produced during fermentations in shaken (200 min-¹) and static (without agitation) flasks and bioreactor, all in batch mode, were determined by GC and HPLC. The results s...

  3. Expansion and contraction of the DUP240 multigene family in Saccharomyces cerevisiae populations.

    Leh-Louis, Véronique; Wirth, Bénédicte; Potier, Serge; Souciet, Jean-Luc; Despons, Laurence

    2004-01-01

    The influence of duplicated sequences on chromosomal stability is poorly understood. To characterize chromosomal rearrangements involving duplicated sequences, we compared the organization of tandem repeats of the DUP240 gene family in 15 Saccharomyces cerevisiae strains of various origins. The DUP240 gene family consists of 10 members of unknown function in the reference strain S288C. Five DUP240 paralogs on chromosome I and two on chromosome VII are arranged as tandem repeats that are highl...

  4. Effects of an unusual poison identify a lifespan role for Topoisomerase 2 in Saccharomyces cerevisiae

    Tombline, Gregory; Millen, Jonathan I.; Polevoda, Bogdan; Rapaport, Matan; Baxter, Bonnie; Van Meter, Michael; Gilbertson, Matthew; Madrey, Joe; Piazza, Gary A.; Rasmussen, Lynn; Wennerberg, Krister; White, E. Lucile; Nitiss, John L.; Goldfarb, David S.

    2017-01-01

    A progressive loss of genome maintenance has been implicated as both a cause and consequence of aging. Here we present evidence supporting the hypothesis that an age-associated decay in genome maintenance promotes aging in Saccharomyces cerevisiae (yeast) due to an inability to sense or repair DNA damage by topoisomerase 2 (yTop2). We describe the characterization of LS1, identified in a high throughput screen for small molecules that shorten the replicative lifespan of yeast. LS1 accelerates...

  5. Relationship between solute permeability and osmotic remediability in a galactose-negative strain of Saccharomyces cerevisiae.

    Bassel, J; Douglas, H C

    1970-11-01

    An osmotic remedial allele, gal 7-1, in the galactose pathway of Saccharomyces cerevisiae responds to either penetrating (ethylene glycol and diethylene glycol) or nonpenetrating (KCl, NaCl, and sorbitol) solutes in the growth medium. Extracts from cells grown under restrictive conditions gave no increase in enzyme activity (gal-1-phosphate, uridylyl transferase) when exposed to the penetrating solutes; thus protein synthesis or possibly polymer assembly is proposed as the critical step remedied by the addition of the solutes.

  6. Purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae

    Elbing, Karin; McCartney, Rhonda R.; Schmidt, Martin C.

    2006-01-01

    Members of the Snf1/AMPK family of protein kinases are activated by distinct upstream kinases that phosphorylate a conserved threonine residue in the Snf1/AMPK activation loop. Recently, the identities of the Snf1- and AMPK-activating kinases have been determined. Here we describe the purification and characterization of the three Snf1-activating kinases of Saccharomyces cerevisiae. The identities of proteins associated with the Snf1-activating kinases were determined by peptide mass fingerpr...

  7. SORPTION OF Au(III BY Saccharomyces cerevisiae BIOMASS

    Amaria Amaria

    2010-07-01

    Full Text Available Au(III sorption by S. cerevisiae biomass extracted from beer waste industry was investigated. Experimentally, the sorption was conducted in batch method. This research involved five steps: 1 identification the functional groups present in the S. cerevisiae biomass by infrared spectroscopic technique, 2 determination of optimum pH, 3 determination of the sorption capacity and energy, 4 determination of the sorption type by conducting desorption of sorbed Au(III using specific eluents having different desorption capacity such as H2O (van der Waals, KNO3 (ion exchange, HNO3 (hydrogen bond, and tiourea (coordination bond, 5 determination of effective eluents in Au(III desorption by partial desorption of sorbed Au(III using thiourea, NaCN and KI. The remaining Au(III concentrations in filtrate were analyzed using Atomic Absorption Spectrophotometer. The results showed that: 1 Functional groups of S. cerevisiae biomass that involved in the sorption processes were hydroxyl (-OH, carboxylate (-COO- and amine (-NH2, 2 maximum sorption was occurred at pH 4, equal to 98.19% of total sorption, 3 The sorption capacity of biomass was 133.33 mg/g (6.7682E-04 mol/g and was involved sorption energy 23.03 kJ mol-1, 4 Sorption type was dominated by coordination bond, 5 NaCN was effective eluent to strip Au(III close to 100%.   Keywords: sorption, desorption, S. cerevisiae biomass, Au(III

  8. Recycling carbon dioxide during xylose fermentation by engineered Saccharomyces cerevisiae

    In this study, we introduced the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and phosphoribulokinase (PRK) into an engineered S. cerevisiae (SR8) harboring the XR/XDH pathway and up-regulated PPP 10, to enable CO2 recycling through a synthetic rPPP during xylose fermentation (Fig. 1). ...

  9. Dual N- and C-terminal helices are required for endoplasmic reticulum and lipid droplet association of alcohol acetyltransferases in Saccharomyces cerevisiae.

    Jyun-Liang Lin

    Full Text Available In the yeast Saccharomyces cerevisiae two alcohol acetyltransferases (AATases, Atf1 and Atf2, condense short chain alcohols with acetyl-CoA to produce volatile acetate esters. Such esters are, in large part, responsible for the distinctive flavors and aromas of fermented beverages including beer, wine, and sake. Atf1 and Atf2 localize to the endoplasmic reticulum (ER and Atf1 is known to localize to lipid droplets (LDs. The mechanism and function of these localizations are unknown. Here, we investigate potential mechanisms of Atf1 and Atf2 membrane association. Segments of the N- and C-terminal domains of Atf1 (residues 24-41 and 508-525, respectively are predicted to be amphipathic helices. Truncations of these helices revealed that the terminal domains are essential for ER and LD association. Moreover, mutations of the basic or hydrophobic residues in the N-terminal helix and hydrophobic residues in the C-terminal helix disrupted ER association and subsequent sorting from the ER to LDs. Similar amphipathic helices are found at both ends of Atf2, enabling ER and LD association. As was the case with Atf1, mutations to the N- and C-terminal helices of Atf2 prevented membrane association. Sequence comparison of the AATases from Saccharomyces, non-Saccharomyces yeast (K. lactis and P. anomala and fruits species (C. melo and S. lycopersicum showed that only AATases from Saccharomyces evolved terminal amphipathic helices. Heterologous expression of these orthologs in S. cerevisiae revealed that the absence of terminal amphipathic helices eliminates LD association. Combined, the results of this study suggest a common mechanism of membrane association for AATases via dual N- and C-terminal amphipathic helices.

  10. Novel feeding strategies for Saccharomyces cerevisiae DS2155 ...

    Administrator

    2007-05-02

    May 2, 2007 ... processes. The software also ensured the updating of the feed flow rate every 5 min for 24 h. The ... But, the exact location and amplitude ..... glucose effect in the Yeast Saccharomyces uvarum: involvement of short, and long ...

  11. Saccharomyces cerevisiae boulardii transient fungemia after intravenous self-inoculation

    Cohen, Lola; Ranque, Stéphane; Raoult, Didier

    2013-01-01

    We report the case of a young psychotic intravenous drug user injecting herself with Saccharomyces cervisiae (boulardii). She experienced a 24 h fever, resolving spontaneously confirming, quasi experimentally, the inocuity of this yeast in a non-immunocompromised host.

  12. Anti-Saccharomyces cerevisiae autoantibodies in autoimmune diseases: from bread baking to autoimmunity.

    Rinaldi, Maurizio; Perricone, Roberto; Blank, Miri; Perricone, Carlo; Shoenfeld, Yehuda

    2013-10-01

    Saccharomyces cerevisiae is best known as the baker's and brewer's yeast, but its residual traces are also frequent excipients in some vaccines. Although anti-S. cerevisiae autoantibodies (ASCAs) are considered specific for Crohn's disease, a growing number of studies have detected high levels of ASCAs in patients affected with autoimmune diseases as compared with healthy controls, including antiphospholipid syndrome, systemic lupus erythematosus, type 1 diabetes mellitus, and rheumatoid arthritis. Commensal microorganisms such as Saccharomyces are required for nutrition, proper development of Peyer's aggregated lymphoid tissue, and tissue healing. However, even the commensal nonclassically pathogenic microbiota can trigger autoimmunity when fine regulation of immune tolerance does not work properly. For our purposes, the protein database of the National Center for Biotechnology Information (NCBI) was consulted, comparing Saccharomyces mannan to several molecules with a pathogenetic role in autoimmune diseases. Thanks to the NCBI bioinformation technology tool, several overlaps in molecular structures (50-100 %) were identified when yeast mannan, and the most common autoantigens were compared. The autoantigen U2 snRNP B″ was found to conserve a superfamily protein domain that shares 83 % of the S. cerevisiae mannan sequence. Furthermore, ASCAs may be present years before the diagnosis of some associated autoimmune diseases as they were retrospectively found in the preserved blood samples of soldiers who became affected by Crohn's disease years later. Our results strongly suggest that ASCAs' role in clinical practice should be better addressed in order to evaluate their predictive or prognostic relevance.

  13. Crystallization and preliminary X-ray diffraction analysis of motif N from Saccharomyces cerevisiae Dbf4

    Matthews, Lindsay A.; Duong, Andrew; Prasad, Ajai A.; Duncker, Bernard P.; Guarné, Alba

    2009-01-01

    To understand the role of the Cdc7–Dbf4 complex in checkpoint responses, a fragment of Saccharomyces cerevisiae Dbf4 encompassing motif N was isolated, overproduced and crystallized. The Cdc7–Dbf4 complex plays an instrumental role in the initiation of DNA replication and is a target of replication-checkpoint responses in Saccharomyces cerevisiae. Cdc7 is a conserved serine/threonine kinase whose activity depends on association with its regulatory subunit, Dbf4. A conserved sequence near the N-terminus of Dbf4 (motif N) is necessary for the interaction of Cdc7–Dbf4 with the checkpoint kinase Rad53. To understand the role of the Cdc7–Dbf4 complex in checkpoint responses, a fragment of Saccharomyces cerevisiae Dbf4 encompassing motif N was isolated, overproduced and crystallized. A complete native data set was collected at 100 K from crystals that diffracted X-rays to 2.75 Å resolution and structure determination is currently under way

  14. Removal of lead, mercury and nickel using the yeast Saccharomyces cerevisiae

    Cherlys Infante J.

    2014-06-01

    Full Text Available Objective. In this study the biomass of the yeast Saccharomyces cerevisiae was used to remove lead, mercury and nickel in the form of ions dissolved in water. Materials and methods. Synthetic solutions were prepared containing the three heavy metals, which were put in contact with viable microorganisms at different conditions of pH, temperature, aeration and agitation. Results. Both individual variables and the interaction effects influenced the biosorption process. Throughout the experimental framework it was observed that the biomass of Saccharomyces cerevisiae removed a higher percentage of lead (86.4% as compared to mercury and nickel (69.7 and 47.8% respectively. When the pH was set at a value of 5 the effect was positive for all three metals. Conclusions. pH was the variable that had a greater influence on the biosorption of lead on the biomass of Saccharomyces cerevisiae. The affinity of the heavy metals for the biomass followed the order Pb>Hg>Ni.

  15. Anti-oxidant effects of pomegranate juice on Saccharomyces cerevisiae cell growth.

    Aslan, Abdullah; Can, Muhammed İsmail; Boydak, Didem

    2014-01-01

    Pomegranate juice has a number of positive effects on both human and animal subjects. Four groups were used in this study. i: Control group, ii: H2O2 group, iii: Pomegranate juice (PJ) group and iv: PJ + H2O2 group. Following the sterilization method for pomegranate juice (10%) and H2O2 (6% v/v), Saccharomyces cerevisiae cultures were added and the cultivation incubated at 35°C for 72 hours. Fatty acids and vitamin concentrations were measured using HPLC and GC and the total protein bands profile were determined by SDS-PAGE. According to our results statistically significant differences have been determined among the study groups in terms of fatty acids and vitamin (pPomegranate juice increased vitamins, fatty acids and total protein expression in Saccharomyces cerevisiae in comparison with the control. Pomegranate juice has a positive effect on fatty acid, vitamin and protein synthesis by Saccharomyces cerevisiae. Accordingly, we believe that it has significantly decreased oxidative damage thereby making a positive impact on yeast development.

  16. Parameter Optimization for Enhancement of Ethanol Yield by Atmospheric Pressure DBD-Treated Saccharomyces cerevisiae

    Dong Xiaoyu; Yuan Yulian; Tang Qian; Dou Shaohua; Di Lanbo; Zhang Xiuling

    2014-01-01

    In this study, Saccharomyces cerevisiae (S. cerevisiae) was exposed to dielectric barrier discharge plasma (DBD) to improve its ethanol production capacity during fermentation. Response surface methodology (RSM) was used to optimize the discharge-associated parameters of DBD for the purpose of maximizing the ethanol yield achieved by DBD-treated S. cerevisiae. According to single factor experiments, a mathematical model was established using Box-Behnken central composite experiment design, with plasma exposure time, power supply voltage, and exposed-sample volume as impact factors and ethanol yield as the response. This was followed by response surface analysis. Optimal experimental parameters for plasma discharge-induced enhancement in ethanol yield were plasma exposure time of 1 min, power voltage of 26 V, and an exposed sample volume of 9 mL. Under these conditions, the resulting yield of ethanol was 0.48 g/g, representing an increase of 33% over control. (plasma technology)

  17. Systems Biology of Saccharomyces cerevisiae Physiology and its DNA Damage Response

    Fazio, Alessandro

    The yeast Saccharomyces cerevisiae is a model organism in biology, being widely used in fundamental research, the first eukaryotic organism to be fully sequenced and the platform for the development of many genomics techniques. Therefore, it is not surprising that S. cerevisiae has also been widely...... used in the field of systems biology during the last decade. This thesis investigates S. cerevisiae growth physiology and DNA damage response by using a systems biology approach. Elucidation of the relationship between growth rate and gene expression is important to understand the mechanisms regulating...... set of growth dependent genes by using a multi-factorial experimental design. Moreover, new insights into the metabolic response and transcriptional regulation of these genes have been provided by using systems biology tools (Chapter 3). One of the prerequisite of systems biology should...

  18. Intracellular Ca2+ Regulation in Calcium Sensitive Phenotype of Saccharomyces cerevisiae

    HERMANSYAH

    2010-03-01

    Full Text Available Intracellular cytosolic Ca2+ concentration accumulation plays an essential information in Saccharomyces cerevisiae i.e. to explain cellular mechanism of Ca2+ sensitive phenotype. Disruption both S. cerevisiae PPase PTP2 and MSG5 genes showed an inhibited growth in the presence of Ca2+. On the other hand, by using Luminocounter with apoaequorin system, a method based upon luminescent photoprotein aequorin, intracellular Ca2+ concentration was accumulated as a consequence of calcium sensitive phenotype of S. cerevisiae. This fact indicated that PPase ptp2Δ and msg5Δ were involved in intracellular Ca2+ transport in addition their already known pathways i.e Mitogen Activated Protein Kinase cell wall integrity pathway, high osmolarity glycerol (HOG pathway, and pheromone response FUS3 pathway.

  19. The expression of glycerol facilitators from various yeast species improves growth on glycerol of Saccharomyces cerevisiae

    Mathias Klein

    2016-12-01

    Full Text Available Glycerol is an abundant by-product during biodiesel production and additionally has several assets compared to sugars when used as a carbon source for growing microorganisms in the context of biotechnological applications. However, most strains of the platform production organism Saccharomyces cerevisiae grow poorly in synthetic glycerol medium. It has been hypothesized that the uptake of glycerol could be a major bottleneck for the utilization of glycerol in S. cerevisiae. This species exclusively relies on an active transport system for glycerol uptake. This work demonstrates that the expression of predicted glycerol facilitators (Fps1 homologues from superior glycerol-utilizing yeast species such as Pachysolen tannophilus, Komagataella pastoris, Yarrowia lipolytica and Cyberlindnera jadinii significantly improves the growth performance on glycerol of the previously selected glycerol-consuming S. cerevisiae wild-type strain (CBS 6412-13A. The maximum specific growth rate increased from 0.13 up to 0.18 h−1 and a biomass yield coefficient of 0.56 gDW/gglycerol was observed. These results pave the way for exploiting the assets of glycerol in the production of fuels, chemicals and pharmaceuticals based on baker's yeast. Keywords: Yeast, Saccharomyces cerevisiae, Glycerol, Transport, Glycerol facilitator, Fps1, Stl1

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

    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. Engineering of a novel Saccharomyces cerevisiae wine strain with a respiratory phenotype at high external glucose concentrations.

    Henricsson, C; de Jesus Ferreira, M C; Hedfalk, K; Elbing, K; Larsson, C; Bill, R M; Norbeck, J; Hohmann, S; Gustafsson, L

    2005-10-01

    The recently described respiratory strain Saccharomyces cerevisiae KOY.TM6*P is, to our knowledge, the only reported strain of S. cerevisiae which completely redirects the flux of glucose from ethanol fermentation to respiration, even at high external glucose concentrations (27). In the KOY.TM6*P strain, portions of the genes encoding the predominant hexose transporter proteins, Hxt1 and Hxt7, were fused within the regions encoding transmembrane (TM) domain 6. The resulting chimeric gene, TM6*, encoded a chimera composed of the amino-terminal half of Hxt1 and the carboxy-terminal half of Hxt7. It was subsequently integrated into the genome of an hxt null strain. In this study, we have demonstrated the transferability of this respiratory phenotype to the V5 hxt1-7Delta strain, a derivative of a strain used in enology. We also show by using this mutant that it is not necessary to transform a complete hxt null strain with the TM6* construct to obtain a non-ethanol-producing phenotype. The resulting V5.TM6*P strain, obtained by transformation of the V5 hxt1-7Delta strain with the TM6* chimeric gene, produced only minor amounts of ethanol when cultured on external glucose concentrations as high as 5%. Despite the fact that glucose flux was reduced to 30% in the V5.TM6*P strain compared with that of its parental strain, the V5.TM6*P strain produced biomass at a specific rate as high as 85% that of the V5 wild-type strain. Even more relevant for the potential use of such a strain for the production of heterologous proteins and also of low-alcohol beverages is the observation that the biomass yield increased 50% with the mutant compared to its parental strain.

  2. Impact of mixed Torulaspora delbrueckii-Saccharomyces cerevisiae culture on high-sugar fermentation.

    Bely, Marina; Stoeckle, Philippe; Masneuf-Pomarède, Isabelle; Dubourdieu, Denis

    2008-03-20

    Conventional wine yeasts produce high concentrations of volatile acidity, mainly acetic acid, during high-sugar fermentation. This alcoholic fermentation by-product is highly detrimental to wine quality and, in some cases, levels may even exceed legal limits. In this study, a non-conventional species, Torulaspora delbrueckii, was used, in pure cultures and mixed with Saccharomyces cerevisiae yeast, to ferment botrytized musts. Fermentation rate, biomass growth, and the formation of volatile acidity, acetaldehyde, and glycerol were considered. This study demonstrated that T. delbrueckii, often described as a low acetic producer under standard conditions, retained this quality even in a high-sugar medium. Unlike S. cerevisiae, this species did not respond to the hyper-osmotic medium by increasing acetic production as soon as it is inoculated into the must. Nevertheless, this yeast produced low ethanol and biomass yields, and the fermentation was sluggish. As a result, T. delbrueckii fermentations do not reach the required ethanol content (14%vol.), although this species can survive at this concentration. A mixed culture of T. delbrueckii and S. cerevisiae was the best combination for improving the analytical profile of sweet wine, particularly volatile acidity and acetaldehyde production. A mixed T. delbrueckii/S. cerevisiae culture at a 20:1 ratio produced 53% less in volatile acidity and 60% less acetaldehyde than a pure culture of S. cerevisiae. Inoculating S. cerevisiae after 5 days' fermentation by T. delbrueckii had less effect on volatile acidity and acetaldehyde production and resulted in stuck fermentation. These results contribute to a better understanding of the behaviour of non-Saccharomyces and their potential application in wine industry.

  3. Bioethanol production by a flocculent hybrid, CHFY0321 obtained by protoplast fusion between Saccharomyces cerevisiae and Saccharomyces bayanus

    Choi, Gi-Wook; Kang, Hyun-Woo; Kim, Yule [Changhae Institute of Cassava and Ethanol Research, Changhae Ethanol Co., LTD, Palbok-Dong 829, Dukjin-Gu, Jeonju 561-203 (Korea); Um, Hyun-Ju; Kim, Mina; Kim, Yang-Hoon [Department of Microbiology, Chungbuk National University, 410 Sungbong-Ro, Heungduk-Gu, Cheongju 361-763 (Korea)

    2010-08-15

    Fusion hybrid yeast, CHFY0321, was obtained by protoplast fusion between non-flocculent-high ethanol fermentative Saccharomyces cerevisiae CHY1011 and flocculent-low ethanol fermentative Saccharomyces bayanus KCCM12633. The hybrid yeast was used together with the parental strains to examine ethanol production in batch fermentation. Under the conditions tested, the fusion hybrid CHFY0321 flocculated to the highest degree and had the capacity to ferment well at pH 4.5 and 32 C. Simultaneous saccharification and fermentation for ethanol production was carried out using a cassava (Manihot esculenta) powder hydrolysate medium containing 19.5% (w v{sup -1}) total sugar in a 5 l lab scale jar fermenter at 32 C for 65 h with an agitation speed of 2 Hz. Under these conditions, CHFY0321 showed the highest flocculating ability and the best fermentation efficiency for ethanol production compared with those of the wild-type parent strains. CHFY0321 gave a final ethanol concentration of 89.8 {+-} 0.13 g l{sup -1}, a volumetric ethanol productivity of 1.38 {+-} 0.13 g l{sup -1} h{sup -1}, and a theoretical yield of 94.2 {+-} 1.58%. These results suggest that CHFY0321 exhibited the fermentation characteristics of S. cerevisiae CHY1011 and the flocculent ability of S. bayanus KCCM12633. Therefore, the strong highly flocculent ethanol fermentative CHFY0321 has potential for improving biotechnological ethanol fermentation processes. (author)

  4. Bioethanol production by a flocculent hybrid, CHFY0321 obtained by protoplast fusion between Saccharomyces cerevisiae and Saccharomyces bayanus

    Choi, Gi-Wook; Um, Hyun-Ju; Kang, Hyun-Woo; Kim, Yule; Kim, Mina; Kim, Yang-Hoon

    2010-01-01

    Fusion hybrid yeast, CHFY0321, was obtained by protoplast fusion between non-flocculent-high ethanol fermentative Saccharomyces cerevisiae CHY1011 and flocculent-low ethanol fermentative Saccharomyces bayanus KCCM12633. The hybrid yeast was used together with the parental strains to examine ethanol production in batch fermentation. Under the conditions tested, the fusion hybrid CHFY0321 flocculated to the highest degree and had the capacity to ferment well at pH 4.5 and 32 o C. Simultaneous saccharification and fermentation for ethanol production was carried out using a cassava (Manihot esculenta) powder hydrolysate medium containing 19.5% (w v -1 ) total sugar in a 5 l lab scale jar fermenter at 32 o C for 65 h with an agitation speed of 2 Hz. Under these conditions, CHFY0321 showed the highest flocculating ability and the best fermentation efficiency for ethanol production compared with those of the wild-type parent strains. CHFY0321 gave a final ethanol concentration of 89.8 ± 0.13 g l -1 , a volumetric ethanol productivity of 1.38 ± 0.13 g l -1 h -1 , and a theoretical yield of 94.2 ± 1.58%. These results suggest that CHFY0321 exhibited the fermentation characteristics of S. cerevisiae CHY1011 and the flocculent ability of S. bayanus KCCM12633. Therefore, the strong highly flocculent ethanol fermentative CHFY0321 has potential for improving biotechnological ethanol fermentation processes.

  5. Activation of waste brewer's yeast Saccharomyces cerevisiae for bread production

    Popov Stevan D.; Dodić Siniša N.; Mastilović Jasna S.; Dodić Jelena M.; Popov-Raljić Jovanka V.

    2005-01-01

    The waste brewer's yeast S. cerevisiae (activated and non-activated) was compared with the commercial baker's yeast regarding the volume of developed gas in dough, volume and freshness stability of produced bread. The activation of waste brewer's yeast resulted in the increased volume of developed gas in dough by 100% compared to non-activated brewer's yeast, and the obtained bread is of more stable freshness compared to bread produced with baker's yeast. The activation of BY affects positive...

  6. Biosorption of the strontium ion by irradiated Saccharomyces cerevisiae under culture conditions.

    Qiu, Liang; Feng, Jundong; Dai, Yaodong; Chang, Shuquan

    2017-06-01

    As a new-emerging method for strontium disposal, biosorption has shown advantages such as high sorption capacity; low cost. In this study, we investigated the potential of Saccharomyces cerevisiae (S. cerevisiae) in strontium disposal under culture conditions and the effects of irradiation on their biosorption capabilities. We found that S. cerevisiae can survive irradiation and grow. Pre-exposure to irradiation rendered S. cerevisiae resistant to further irradiation. Surprisingly, the pre-exposure to irradiation can increase the biosorption capability of S. cerevisiae. We further investigated the factors that influenced the biosorption efficiency, which were (strongest to weakest): pH > strontium concentration > time > temperature. In our orthogonal experiment, the optimal conditions for strontium biosorption by irradiated S. cerevisiae were: pH 7, 150 mg L -1 strontium at the temperature of 32 °C with 30 h. The equilibrium of strontium biosorption was analyzed by Langmuir and Freundlich models, from which the formal model is found to provide a better fit for the experimental results. The kinetics of strontium biosorption by living irradiated S. cerevisiae was found to be comprised of three phases: dramatically increased during 0-9 h, decreased during 12-24 h, and increased during 30-50 h. These results provide a systematic understanding of the biosorption capabilities of irradiated S. cerevisiae, which can contribute to the development of remediating nuclear waste water. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Genomics and Biochemistry of Saccharomyces cerevisiae Wine Yeast Strains.

    Eldarov, M A; Kishkovskaia, S A; Tanaschuk, T N; Mardanov, A V

    2016-12-01

    Saccharomyces yeasts have been used for millennia for the production of beer, wine, bread, and other fermented products. Long-term "unconscious" selection and domestication led to the selection of hundreds of strains with desired production traits having significant phenotypic and genetic differences from their wild ancestors. This review summarizes the results of recent research in deciphering the genomes of wine Saccharomyces strains, the use of comparative genomics methods to study the mechanisms of yeast genome evolution under conditions of artificial selection, and the use of genomic and postgenomic approaches to identify the molecular nature of the important characteristics of commercial wine strains of Saccharomyces. Succinctly, data concerning metagenomics of microbial communities of grapes and wine and the dynamics of yeast and bacterial flora in the course of winemaking is provided. A separate section is devoted to an overview of the physiological, genetic, and biochemical features of sherry yeast strains used to produce biologically aged wines. The goal of the review is to convince the reader of the efficacy of new genomic and postgenomic technologies as tools for developing strategies for targeted selection and creation of new strains using "classical" and modern techniques for improving winemaking technology.

  8. Saccharomyces cerevisiae KTR4, KTR5 and KTR7 encode mannosyltransferases differentially involved in the N- and O-linked glycosylation pathways.

    Hernández, Nahúm V; López-Ramírez, Luz A; Díaz-Jiménez, Diana F; Mellado-Mojica, Erika; Martínez-Duncker, Iván; López, Mercedes G; Mora-Montes, Héctor M

    2017-10-01

    Saccharomyces cerevisiae is a model to understand basic aspects of protein glycosylation pathways. Although these metabolic routes have been thoroughly studied, there are still knowledge gaps; among them, the role of the MNT1/KRE2 gene family. This family is composed of nine members, with only six functionally characterized. The enzymes Ktr1, Ktr3, and Mnt1/Kre2 have overlapping activities in both O-linked and N-linked glycan synthesis; while Ktr2 and Yur1 participate exclusively in the elongation of the N-linked glycan outer chain. KTR6 encodes for a phosphomannosyltransferase that synthesizes the cell wall phosphomannan. Here, we aimed to establish the functional role of KTR4, KTR5 and KTR7 in the protein glycosylation pathways, by using heterologous complementation in Candida albicans null mutants lacking members of the MNT1/KRE2 gene family. The three S. cerevisiae genes restored defects in the C. albicans N-linked glycosylation pathway. KTR5 and KTR7 partially complemented a C. albicans null mutant with defects in the synthesis of O-linked glycans, and only KTR4 fully elongated the O-linked glycans like wild-type cells. Therefore, our results suggest that the three genes have a redundant activity in the S. cerevisiae N-linked glycosylation pathway, but KTR4 plays a major role in O-linked glycan synthesis. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  9. Loss of lager specific genes and subtelomeric regions define two different Saccharomyces cerevisiae lineages for Saccharomyces pastorianus Group I and II strains.

    Monerawela, Chandre; James, Tharappel C; Wolfe, Kenneth H; Bond, Ursula

    2015-03-01

    Lager yeasts, Saccharomyces pastorianus, are interspecies hybrids between S. cerevisiae and S. eubayanus and are classified into Group I and Group II clades. The genome of the Group II strain, Weihenstephan 34/70, contains eight so-called 'lager-specific' genes that are located in subtelomeric regions. We evaluated the origins of these genes through bioinformatic and PCR analyses of Saccharomyces genomes. We determined that four are of cerevisiae origin while four originate from S. eubayanus. The Group I yeasts contain all four S. eubayanus genes but individual strains contain only a subset of the cerevisiae genes. We identified S. cerevisiae strains that contain all four cerevisiae 'lager-specific' genes, and distinct patterns of loss of these genes in other strains. Analysis of the subtelomeric regions uncovered patterns of loss in different S. cerevisiae strains. We identify two classes of S. cerevisiae strains: ale yeasts (Foster O) and stout yeasts with patterns of 'lager-specific' genes and subtelomeric regions identical to Group I and II S. pastorianus yeasts, respectively. These findings lead us to propose that Group I and II S. pastorianus strains originate from separate hybridization events involving different S. cerevisiae lineages. Using the combined bioinformatic and PCR data, we describe a potential classification map for industrial yeasts. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  10. Comportamento celular e resposta antioxidante diferenciados de Saccharomyces cerevisiae e de Saccharomyces chevalieri ao metavanadato de amónio Different cellular behaviour and antioxidant response of Saccharomyces cerevisiae and Saccharomyces chevalieri growing in presence of ammonium metavanadate

    R. Ferreira

    2007-01-01

    Full Text Available A fermentação do vinho é um processo microbiológico complexo que requere a presença de leveduras adaptadas a condições de stresse. No ambiente celular de organismos aeróbios ocorrem naturalmente espécies reactivas de oxigénio (ROS como subprodutos da respiração mitocondrial. A elevada reactividade destas espécies químicas pode gerar danos moleculares que, em alguns casos, levam à morte celular. Em condições fisiológicas normais ou como resposta ao stresse oxidativo, a célula pode desencadear respostas adaptativas que envolvem mecanismos antioxidantes como os enzimas glutationo redutase (GR; EC 1.6.4.2 e catalases T (CAT T; EC 1.11.1.6 e A (CAT A; EC 1.11.1.6. O vanádio, um metal pesado presente em alguns fitofármacos, pode também com portar-se como um gerador de ROS, alterando o estado redox intracelular e exercendo efeitos nocivos em leveduras expostas a quantidade excessiva deste elemento. O principal objectivo deste trabalho foi comparar o efeito do metavanadato de amónio (NH4VO3, um sal pentavalente de vanádio, na viabilidade celular e nas actividades enzimáticas GR, CAT T e CAT A das leveduras vínicas Saccharomyces cerevisiae UE-ME3 e Saccharomyces chevalieri UE-ME1. Os resultados obtidos mostram que S. chevalieri UE-ME1 revelou menor tolerância ao NH4VO3 do que S. cerevisiae UE-ME3, uma vez que culturas de S. chevalieri não sobreviveram para valores de concentração do sal de vanádio superiores a 7,5 mM enquanto que células de S. cerevisiae mantiveram-se viáveis em presença de metavanadato de amónio 75 mM. As actividades enzimáticas estudadas apresentaram em S. chevalieri valores muito inferiores aos que foram determinados em S. cerevisiae embora em ambas as espécies de levedura o NH4VO3 pareça comportarse como um indutor de stresse oxidativo ao provocar um decréscimo significativo da actividade GR (PThe fermentation of wine is a complex microbiological process which requires yeast adaptation to stress

  11. Pichia pastoris versus Saccharomyces cerevisiae: a case study on the recombinant production of human granulocyte-macrophage colony-stimulating factor.

    Tran, Anh-Minh; Nguyen, Thanh-Thao; Nguyen, Cong-Thuan; Huynh-Thi, Xuan-Mai; Nguyen, Cao-Tri; Trinh, Minh-Thuong; Tran, Linh-Thuoc; Cartwright, Stephanie P; Bill, Roslyn M; Tran-Van, Hieu

    2017-04-04

    Recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) is a glycoprotein that has been approved by the FDA for the treatment of neutropenia and leukemia in combination with chemotherapies. Recombinant hGM-CSF is produced industrially using the baker's yeast, Saccharomyces cerevisiae, by large-scale fermentation. The methylotrophic yeast, Pichia pastoris, has emerged as an alternative host cell system due to its shorter and less immunogenic glycosylation pattern together with higher cell density growth and higher secreted protein yield than S. cerevisiae. In this study, we compared the pipeline from gene to recombinant protein in these two yeasts. Codon optimization in silico for both yeast species showed no difference in frequent codon usage. However, rhGM-CSF expressed from S. cerevisiae BY4742 showed a significant discrepancy in molecular weight from those of P. pastoris X33. Analysis showed purified rhGM-CSF species with molecular weights ranging from 30 to more than 60 kDa. Fed-batch fermentation over 72 h showed that rhGM-CSF was more highly secreted from P. pastoris than S. cerevisiae (285 and 64 mg total secreted protein/L, respectively). Ion exchange chromatography gave higher purity and recovery than hydrophobic interaction chromatography. Purified rhGM-CSF from P. pastoris was 327 times more potent than rhGM-CSF from S. cerevisiae in terms of proliferative stimulating capacity on the hGM-CSF-dependent cell line, TF-1. Our data support a view that the methylotrophic yeast P. pastoris is an effective recombinant host for heterologous rhGM-CSF production.

  12. Cellular responses of Saccharomyces cerevisiae to DNA damage

    Ciesla, Z.; Sledziewska-Gojska, E.; Nowicka, A.; Mieczkowski, P.; Fikus, M.U.; Koprowski, P.

    1998-01-01

    Full text. Several experimental strategies have been used to study responses of S. cerevisiae cells to DNA damage. One approach was based on the isolation of novel genes, the expression of which is induced by lesions in DNA. One of these genes, DIN7, was cloned and partially characterized previously. The product of DIN7 belongs to a large family of proteins involved in DNA repair and mutagenesis. This family includes Rad2, Rad27 and ExoI proteins of S. cerevisiae and their respective human homologues, all of which are endowed with DNA nuclease activity. To study cellular function of Din7 we constructed the pPK3 plasmid carrying DIN7 fused to the GAL1 promoter. Effects of DIN7 overproduction on the phenotypes of wild-type cells and of rad27 and exoI mutants were examined. Overproduction of Din7 does not seem to affect the proficiency of wild-type S. cerevisiae cells in recombination and mutagenesis. Also, overexpression of DIN7 does not suppress the deficiency of the EXOI gene product, the closest homologue of Din7, both in recombination and in controlling the fidelity of DNA replication. Unexpectedly, we found that elevated levels of Din7 result in a very high frequency of mitochondrial rho - mutants. A high frequency of production of rho - mutants wa s also observed in strains defective in the functioning of the Dun1 protein kinase involved in signal transmission in cells exposed to DNA damaging agents. Interestingly, deficiency of Dun1 results also in a significant derepression of the DIN7 gene. Experiments are under way to distinguish whether a high cellular level of Din7 specifically decreases stability of mitochondrial DNA or affects stability of chromosomal DNA as well. Analysis of previously constructed S. cerevisiae strains carrying random geno mic fusions with reporter lacZ gene, allowed us to identify the reading frame YBR173c, on chromosome II as a novel damage inducible gene - DIN8. We have shown that DIN8-lacZ fusion is induced in yeast cells treated

  13. Integrative proteomics and biochemical analyses define Ptc6p as the Saccharomyces cerevisiae pyruvate dehydrogenase phosphatase.

    Guo, Xiao; Niemi, Natalie M; Coon, Joshua J; Pagliarini, David J

    2017-07-14

    The pyruvate dehydrogenase complex (PDC) is the primary metabolic checkpoint connecting glycolysis and mitochondrial oxidative phosphorylation and is important for maintaining cellular and organismal glucose homeostasis. Phosphorylation of the PDC E1 subunit was identified as a key inhibitory modification in bovine tissue ∼50 years ago, and this regulatory process is now known to be conserved throughout evolution. Although Saccharomyces cerevisiae is a pervasive model organism for investigating cellular metabolism and its regulation by signaling processes, the phosphatase(s) responsible for activating the PDC in S. cerevisiae has not been conclusively defined. Here, using comparative mitochondrial phosphoproteomics, analyses of protein-protein interactions by affinity enrichment-mass spectrometry, and in vitro biochemistry, we define Ptc6p as the primary PDC phosphatase in S. cerevisiae Our analyses further suggest additional substrates for related S. cerevisiae phosphatases and describe the overall phosphoproteomic changes that accompany mitochondrial respiratory dysfunction. In summary, our quantitative proteomics and biochemical analyses have identified Ptc6p as the primary-and likely sole- S. cerevisiae PDC phosphatase, closing a key knowledge gap about the regulation of yeast mitochondrial metabolism. Our findings highlight the power of integrative omics and biochemical analyses for annotating the functions of poorly characterized signaling proteins. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. Oral administration of myostatin-specific recombinant Saccharomyces cerevisiae vaccine in rabbit.

    Liu, Zhongtian; Zhou, Gang; Ren, Chonghua; Xu, Kun; Yan, Qiang; Li, Xinyi; Zhang, Tingting; Zhang, Zhiying

    2016-04-29

    Yeast is considered as a simple and cost-effective host for protein expression, and our previous studies have proved that Saccharomyces cerevisiae can deliver recombinant protein and DNA into mouse dendritic cells and can further induce immune responses as novel vaccines. In order to know whether similar immune responses can be induced in rabbit by oral administration of such recombinant S. cerevisiae vaccine, we orally fed the rabbits with heat-inactivated myostatin-recombinant S. cerevisiae for 5 weeks, and then myostatin-specific antibody in serum was detected successfully by western blotting and ELISA assay. The rabbits treated with myostatin-recombinant S. cerevisiae vaccine grew faster and their muscles were much heavier than that of the control group. As a common experimental animal and a meat livestock with great economic value, rabbit was proved to be the second animal species that have been successfully orally immunized by recombinant S. cerevisiae vaccine after mice. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Response of Saccharomyces cerevisiae to D-limonene-induced oxidative stress.

    Liu, Jidong; Zhu, Yibo; Du, Guocheng; Zhou, Jingwen; Chen, Jian

    2013-07-01

    In the present study, we investigated the mode of cell response induced by D-limonene in Saccharomyces cerevisiae. D-limonene treatment was found to be accompanied by intracellular accumulation of reactive oxygen species (ROS). Since ROS impair cell membranes, an engineered strain with enhanced membrane biosynthesis exhibited a higher tolerance to D-limonene. Subsequent addition of an ROS scavenger significantly reduced the ROS level and alleviated cell growth inhibition. Thus, D-limonene-induced ROS accumulation plays an important role in cell death in S. cerevisiae. In D-limonene-treated S. cerevisiae strains, higher levels of antioxidants, antioxidant enzymes, and nicotinamide adenine dinucleotide phosphate (NADPH) were synthesized. Quantitative real-time PCR results also verified that D-limonene treatment triggered upregulation of genes involved in the antioxidant system and the regeneration of NADPH at the transcription level in S. cerevisiae. These data indicate that D-limonene treatment results in intracellular ROS accumulation, an important factor in cell death, and several antioxidant mechanisms in S. cerevisiae were enhanced in response to D-limonene treatment.

  16. Zinc oxide and silver nanoparticles toxicity in the baker's yeast, Saccharomyces cerevisiae.

    Galván Márquez, Imelda; Ghiyasvand, Mergan; Massarsky, Andrey; Babu, Mohan; Samanfar, Bahram; Omidi, Katayoun; Moon, Thomas W; Smith, Myron L; Golshani, Ashkan

    2018-01-01

    Engineered nanomaterials (ENMs) are increasingly incorporated into a variety of commercial applications and consumer products; however, ENMs may possess cytotoxic properties due to their small size. This study assessed the effects of two commonly used ENMs, zinc oxide nanoparticles (ZnONPs) and silver nanoparticles (AgNPs), in the model eukaryote Saccharomyces cerevisiae. A collection of ≈4600 S. cerevisiae deletion mutant strains was used to deduce the genes, whose absence makes S. cerevisiae more prone to the cytotoxic effects of ZnONPs or AgNPs. We demonstrate that S. cerevisiae strains that lack genes involved in transmembrane and membrane transport, cellular ion homeostasis, and cell wall organization or biogenesis exhibited the highest sensitivity to ZnONPs. In contrast, strains that lack genes involved in transcription and RNA processing, cellular respiration, and endocytosis and vesicular transport exhibited the highest sensitivity to AgNPs. Secondary assays confirmed that ZnONPs affected cell wall function and integrity, whereas AgNPs exposure decreased transcription, reduced endocytosis, and led to a dysfunctional electron transport system. This study supports the use of S. cerevisiae Gene Deletion Array as an effective high-throughput technique to determine cellular targets of ENM toxicity.

  17. Production of ethanol from blackstrap molasses by saccharomyces cerevisiae

    Elahi, S.; Hashmi, Abu-S.; Akhtar, C.M.; Ilahi, A.; Rajoka, M.I.

    1991-01-01

    Blackstrap molasses was analyzed for its composition and its fermentation was brought about by the yeast S. cerevisiae at predetermined optimal environmental conditions such as pH, temperature, Sugar concentration, and incubation period. The results revealed that sugar concentration 17%, pH 4.5, temperature 30 C and incubation period of 72 hours were the optimal conditions for producing maximum (73 g/l) ethanol. Clearance of molasses by 20% single superphosphate enhanced ethanol production by only 0.2%. (author)

  18. Increased mannoprotein content in wines produced by Saccharomyces kudriavzevii×Saccharomyces cerevisiae hybrids.

    Pérez-Través, Laura; Querol, Amparo; Pérez-Torrado, Roberto

    2016-11-21

    Several wine quality aspects are influenced by yeast mannoproteins on account of aroma compounds retention, lactic-acid bacterial growth stimulation, protection against protein haze and astringency reduction. Thus selecting a yeast strain that produces high levels of mannoproteins is important for the winemaking industry. In this work, we observed increased levels of mannoproteins in S. cerevisiae×S. kudriavzevii hybrids, compared to the S. cerevisiae strain, in wine fermentations. Furthermore, the expression of a key gene related to mannoproteins biosynthesis, PMT1, increased in the S. cerevisiae×S. kudriavzevii hybrid. We showed that artificially constructed S. cerevisiae×S. kudriavzevii hybrids also increased the levels of mannoproteins. This work demonstrates that either natural or artificial S. cerevisiae×S. kudriavzevii hybrids present mannoprotein overproducing capacity under winemaking conditions, a desirable physiological feature for this industry. These results suggest that genome interaction in hybrids generates a physiological environment that enhances the release of mannoproteins. Copyright © 2016 Elsevier B.V. All rights reserved.

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

    Xiaoling Tang

    2015-12-01

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

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

    Kanchana R. Kildegaard

    2015-12-01

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

  1. Microencapsulation of Saccharomyces cerevisiae and its evaluation to protect in simulated gastric conditions.

    Ghorbani-Choboghlo, Hassan; Zahraei-Salehi, Taghi; Ashrafi-Helan, Javad; Yahyaraeyat, Ramak; Pourjafar, Hadi; Nikaein, Donya; Balal, Asad; Khosravi, Ali-Reza

    2015-12-01

    Probiotic yeasts are used in production of functional foods and pharmaceutical products. They play an important role in promoting and maintaining human health. Until now, little work has been published on improving the survival of Saccharomyces in stimulated gastrointestinal condition. In this study the exposure of the yeast in the capsulate and free forms to artificial gastrointestinal conditions was assessed and the number of viable Saccharomyces cerevisiae cells during 0 to 120 mines in these conditions was evaluated by a pour plate method using sabouraud dextrose agar. Results showed the shape of the beads was generally spherical, sometimes elliptical with a mean diameter of about 50-90 μm. Also count of viable probiotic cells obtained for all the microcapsules were above the recommended levels for a probiotic food. Also decrease of approximately 4 logs was noted in the number of free cells after 2 h of incubation at pH 2 and 8, when compared to decreases of about 2 logs in the all microencapsulated S. cerevisiae under similar conditions. It is concluded that microencapsulation process was significantly able to increase the survival rate of Saccharomyces in a simulated gastrointestinal condition (p<0.05)..

  2. Activation of waste brewer's yeast Saccharomyces cerevisiae for bread production

    Popov Stevan D.

    2005-01-01

    Full Text Available The waste brewer's yeast S. cerevisiae (activated and non-activated was compared with the commercial baker's yeast regarding the volume of developed gas in dough, volume and freshness stability of produced bread. The activation of waste brewer's yeast resulted in the increased volume of developed gas in dough by 100% compared to non-activated brewer's yeast, and the obtained bread is of more stable freshness compared to bread produced with baker's yeast. The activation of BY affects positively the quality of produced bread regarding bread volume. The volume of developed gas in dough prepared with the use of non-activated BY was not sufficient, therefore, it should not be used as fermentation agent, but only as an additive in bread production process for bread freshness preservation. Intense mixing of dough results in more compressible crumb 48 hrs after baking compared to high-speed mixing.

  3. Adaptive answer to low ionizing radiation doses in Saccharomyces cerevisiae; Respuesta adaptativa a bajas dosis de radiacion ionizante en Saccharomyces cerevisiae

    Durand, Jorge L. [Comision Nacional de Energia Atomica, San Carlos de Bariloche (Argentina). Inst. Balseiro; Frati, Diego Libkind; Broock, Maria Van [Universidad Nacional del Comahue, Bariloche (Argentina). Centro Regional Universitario Bariloche; Gillette, Victor [Comision Nacional de Energia Atomica, San Carlos de Bariloche (Argentina). Centro Atomico

    2001-07-01

    The aim of this work is to verify the existence of the adaptive response phenomenon induced by low doses of ionizing radiation in living cells. It is known that low doses of ionizing radiation, called conditioning doses, may induce resistance in exposed organisms to higher doses, called challenging doses, which are applied after a period of time. The involved mechanisms in this phenomenon, called Adaptive Response, are diverse and complex. Among them, the most important are the activation of DNA-repair enzymes and nuclear recombination process. As the 'target' sample, it was utilized a 'wild type' strain of Saccharomyces cerevisiae in aqueous suspension. Adaptive Response was verified in a wide range of challenging doses. Conditioning doses, inductors of radio-resistance, were (0.44{+-}0.03) Gy and the waiting time between them and challenging doses was 2 hours at room temperature.(author)

  4. Selected non-Saccharomyces wine yeasts in controlled multistarter fermentations with Saccharomyces cerevisiae on alcoholic fermentation behaviour and wine aroma of cherry wines.

    Sun, Shu Yang; Gong, Han Sheng; Jiang, Xiao Man; Zhao, Yu Ping

    2014-12-01

    This study examined the effect of mixed fermentation of non-Saccharomyces (Torulaspora delbrueckii ZYMAFLORE Alpha(TD n. Sacch) and Metschnikowia pulcherrima JS22) and Saccharomyces cerevisiae yeasts (D254 and EC1118) on the production of cherry wines, in comparison with commonly used mono-culture. Results obtained during AF demonstrated that negligible inhibitory effect was observed in S. cerevisiae/Alpha pair, whereas a strong antagonistic effect was detected between MJS22 and S. cerevisiae strain, resulting in an early death of MJS22. For volatile components determined, S. cerevisiae/MJS22 couple was found to significantly boost the production of most detected compounds, more particularly in higher alcohols, esters, acids and terpenes; while the characteristic of S. cerevisiae/Alpha pair is an increase in fruity esters, higher alcohols and decrease in acid production. Sensory evaluation revealed that S. cerevisiae/MJS22 pair reinforced sweet, green and fatty notes to the cherry wines, and S. cerevisiae/Alpha trial enhanced the fruity odour and reduced green note. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Altering the Rate of Mitosis by Introducing Low-Gigahertz Radiation to Saccharomyces cerevisiae Cells

    Garg, S.; Ashby, C.

    2017-12-01

    This experiment aims to assess the impact of low-frequency radiation (from common technological tools such as cell phones, scanners, and wifi) on the mitotic rates of cells. In particular, the focus of the study was on the growth and development of Saccharomyces cerevisiae cultures that were exposed to radio waves from a wifi router, which were then compared to a cohort of the same species without exposure. Though routers emit a low gigahertz frequency, they are categorized as Group 2B radiation (possibly carcinogenic) by the International Agency for Research on Cancer of the World Health Organization, signifying that constant exposure poses a potential risk to humans. Twelve agar dishes of active Saccharomyces cerevisiae solution were prepared, with six dishes acting as the control under no added radiation and six acting as the experimental group under 2.4 GHz of radiation due to their proximity to the router. Data on how many cultures proliferated in each dish was collected every three days, with the experiment running for a total of twelve days. All subjects experienced growth curves until day 9 when the experimental group's growth peaked with an average of 62 colonies/dish. Three of the six dishes in this group lost colonies in the following three days, leaving the experimental group with an average of 61 colonies/dish on day 12, while the control group was still increasing by day 12 with an average of 48 colonies/dish, with only one dish undergoing a loss of colonies. Exposing the Saccharomyces cerevisiae cells to low grade radiation resulted in accelerated mitosis, and though the experimental group faced colony death after nine days, the loss was likely due to overpopulation in the dish.

  6. Glucose-free fructose production from Jerusalem artichoke using a recombinant inulinase-secreting Saccharomyces cerevisiae strain.

    Yu, Jing; Jiang, Jiaxi; Ji, Wangming; Li, Yuyang; Liu, Jianping

    2011-01-01

    Using inulin (polyfructose) obtained from Jerusalen artichokes, we have produced fructose free of residual glucose using a recombinant inulinase-secreting strain of Saccharomyces cerevisiae in a one-step fermentation of Jerusalem artichoke tubers. For producing fructose from inulin, a recombinant inulinase-producing Saccharomyce cerevisiae strain was constructed with a deficiency in fructose uptake by disruption of two hexokinase genes hxk1 and hxk2. The inulinase gene introduced into S. cerevisiae was cloned from Kluyveromyces cicerisporus. Extracellular inulinase activity of the recombinant hxk-mutated S. cerevisiae strain reached 31 U ml(-1) after 96 h growth. When grown in a medium containing Jerusalem artichoke tubers as the sole component without any additives, the recombinant yeast accumulated fructose up to 9.2% (w/v) in the fermentation broth with only 0.1% (w/v) glucose left after 24 h.

  7. Functional conservation between Schizosaccharomyces pombe ste8 and Saccharomyces cerevisiae STE11 protein kinases in yeast signal transduction

    Styrkársdóttir, U; Egel, R; Nielsen, O

    1992-01-01

    in signal transduction in budding yeast. Expression of the S. cerevisiae STE11 gene in S. pombe ste8 mutants restores the ability to transcribe mat1-Pm in response to pheromone. Also, such cells become capable of conjugation and sporulation. When mat1-Pm is artifically expressed from a heterologous promoter...

  8. Trichoderma virens β-glucosidase I (BGLI) gene; expression in Saccharomyces cerevisiae including docking and molecular dynamics studies.

    Wickramasinghe, Gammadde Hewa Ishan Maduka; Rathnayake, Pilimathalawe Panditharathna Attanayake Mudiyanselage Samith Indika; Chandrasekharan, Naduviladath Vishvanath; Weerasinghe, Mahindagoda Siril Samantha; Wijesundera, Ravindra Lakshman Chundananda; Wijesundera, Wijepurage Sandhya Sulochana

    2017-06-21

    Cellulose, a linear polymer of β 1-4, linked glucose, is the most abundant renewable fraction of plant biomass (lignocellulose). It is synergistically converted to glucose by endoglucanase (EG) cellobiohydrolase (CBH) and β-glucosidase (BGL) of the cellulase complex. BGL plays a major role in the conversion of randomly cleaved cellooligosaccharides into glucose. As it is well known, Saccharomyces cerevisiae can efficiently convert glucose into ethanol under anaerobic conditions. Therefore, S.cerevisiae was genetically modified with the objective of heterologous extracellular expression of the BGLI gene of Trichoderma virens making it capable of utilizing cellobiose to produce ethanol. The cDNA and a genomic sequence of the BGLI gene of Trichoderma virens was cloned in the yeast expression vector pGAPZα and separately transformed to Saccharomyces cerevisiae. The size of the BGLI cDNA clone was 1363 bp and the genomic DNA clone contained an additional 76 bp single intron following the first exon. The gene was 90% similar to the DNA sequence and 99% similar to the deduced amino acid sequence of 1,4-β-D-glucosidase of T. atroviride (AC237343.1). The BGLI activity expressed by the recombinant genomic clone was 3.4 times greater (1.7 x 10 -3  IU ml -1 ) than that observed for the cDNA clone (5 x 10 -4  IU ml -1 ). Furthermore, the activity was similar to the activity of locally isolated Trichoderma virens (1.5 x 10 -3  IU ml -1 ). The estimated size of the protein was 52 kDA. In fermentation studies, the maximum ethanol production by the genomic and the cDNA clones were 0.36 g and 0.06 g /g of cellobiose respectively. Molecular docking results indicated that the bare protein and cellobiose-protein complex behave in a similar manner with considerable stability in aqueous medium. The deduced binding site and the binding affinity of the constructed homology model appeared to be reasonable. Moreover, it was identified that the five hydrogen bonds formed

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

    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......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 hydrolysate. A particular biological problem are the pentoses, which are not naturally metabolized by the main industrial ethanol producer Saccharomyces cerevisiae. Several recombinant, mutated, and evolved xylose fermenting S. cerevisiae strains have been developed recently. We compare here the fermentation...

  10. Major sulfonate transporter Soa1 in Saccharomyces cerevisiae and considerable substrate diversity in its fungal family

    Holt, Sylvester; Kankipati, Harish; De Graeve, Stijn

    2017-01-01

    Sulfate is a well-established sulfur source for fungi; however, in soils sulfonates and sulfate esters, especially choline sulfate, are often much more prominent. Here we show that Saccharomyces cerevisiae YIL166C(SOA1) encodes an inorganic sulfur (sulfate, sulfite and thiosulfate) transporter...... that also catalyses sulfonate and choline sulfate uptake. Phylogenetic analysis of fungal SOA1 orthologues and expression of 20 members in the sul1 Delta sul2 Delta soa1 Delta strain, which is deficient in inorganic and organic sulfur compound uptake, reveals that these transporters have diverse substrate...... preferences for sulfur compounds. We further show that SOA2, a S. cerevisiae SOA1 paralogue found in S. uvarum, S. eubayanus and S. arboricola is likely to be an evolutionary remnant of the uncharacterized open reading frames YOL163W and YOL162W. Our work highlights the importance of sulfonates and choline...

  11. Secretory production of cell wall components by Saccharomyces cerevisiae protoplasts in static liquid culture.

    Aoyagi, Hideki; Ishizaka, Mikiko; Tanaka, Hideo

    2012-04-01

    When protoplasts of Saccharomyces cerevisiae T7 and IFO 0309 are cultured in a static liquid culture at 2.5 × 10(6) protoplasts/ml, cell wall regeneration does not occur and cell wall components (CWC) are released into the culture broth. By using a specialized fluorometer, the concentrations of CWC could be measured on the basis of the fluorescence intensity of the CWC after staining with Fluostain I. The inoculum concentration, pH, and osmotic pressure of the medium were important factors for the production of CWC in culture. Under optimal culture conditions, S. cerevisiae T7 protoplasts produced 0.91 mg/ml CWC after 24 h. The CWC induced the tumor necrosis factor-α production about 1.3 times higher than that of the commercially available β-1,3/1,6-glucan from baker's yeast cells.

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

    Borodina, Irina; Nielsen, Jens

    2014-05-01

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

  13. The effect of Saccharomyces cerevisiae on the stability of the herbicide glyphosate during bread leavening.

    Low, F L; Shaw, I C; Gerrard, J A

    2005-01-01

    To investigate the ability of baker's yeast (Saccharomyces cerevisiae) to degrade the herbicide glyphosate during the fermentation cycle of the breadmaking process. Aqueous glyphosate was added to bread ingredients and kneaded by commercially available breadmaking equipment into dough cultures. Cultures were incubated in the breadmaker throughout the fermentation cycle. The recovery of glyphosate levels following fermentation was determined, thus allowing an estimation of glyphosate degradation by yeast. It was shown, for the first time, that S. cerevisiae plays a role in metabolizing glyphosate during the fermentation stages of breadmaking. Approximately 21% was degraded within 1 h. As a result of projected increases in the glyphosate use on wheat and the role of bread as a dietary staple, this may contribute to more informed decisions being made relating to the use of glyphosate on glyphosate-resistant wheat, from a public health/regulatory perspective.

  14. Engineering high-level production of fatty alcohols by Saccharomyces cerevisiae from lignocellulosic feedstocks

    d'Espaux, Leo; Ghosh, Amit; Runguphan, Weerawat

    2017-01-01

    to similar to 20% of the maximum theoretical yield from glucose, the highest titers and yields reported to date in S. cerevisiae. We further demonstrate high-level production from lignocellulosic feedstocks derived from ionic-liquid treated switchgrass and sorghum, reaching 0.7 g/L in shake flasks......Fatty alcohols in the C12-C18 range are used in personal care products, lubricants, and potentially biofuels. These compounds can be produced from the fatty acid pathway by a fatty acid reductase (FAR), yet yields from the preferred industrial host Saccharomyces cerevisiae remain under 2......% of the theoretical maximum from glucose. Here we improved titer and yield of fatty alcohols using an approach involving quantitative analysis of protein levels and metabolic flux, engineering enzyme level and localization, pull-push-block engineering of carbon flux, and cofactor balancing. We compared four...

  15. Saccharomyces cerevisiae biofilm tolerance towards systemic antifungals depends on growth phase

    Bojsen, Rasmus Kenneth; Regenberg, Birgitte; Folkesson, Sven Anders

    2014-01-01

    Background : Biofilm-forming Candida species cause infections that can be difficult to eradicate, possibly because of antifungal drug tolerance mechanisms specific to biofilms. In spite of decades of research, the connection between biofilm and drug tolerance is not fully understood. Results : We...... used Saccharomyces cerevisiae as a model for drug susceptibility of yeast biofilms. Confocal laser scanning microscopy showed that S. cerevisiae and C. glabrata form similarly structured biofilms and that the viable cell numbers were significantly reduced by treatment of mature biofilms...... with amphotericin B but not voriconazole, flucytosine, or caspofungin. We showed that metabolic activity in yeast biofilm cells decreased with time, as visualized by FUN-1 staining, and mature, 48-hour biofilms contained cells with slow metabolism and limited growth. Time-kill studies showed that in exponentially...

  16. Isolation of beta-glucan from the cell wall of Saccharomyces cerevisiae.

    Shokri, Hojjatollah; Asadi, Farzad; Khosravi, Ali Reza

    2008-03-20

    Beta-glucan, one of the major cell wall components of Saccharomyces cerevisiae (S. cerevisiae), has been found to enhance immune functions. At present study, we developed an optimal procedure to extract and purify beta-glucan. At first, yeast cells were grown in sabouraud dextrose agar and then cultured in yeast extract-peptone-glucose (YPG) broth. After incubation, cells were harvested, washed and disrupted by means of sonication method. The obtained cell walls were used to prepare alkali-soluble beta-glucan (glucan-S1). In this regard, 2% sodium hydroxide (NaOH) and 3% acetic acid were used in alkaline-acid extraction, respectively. This preparation contained 2.4% protein. In the next step, DEAE sephacel chromatography was used to remove remaining proteins (glucan-S2). Subsequently this preparation was applied into concanavalin-A sepharose column to remove manann. Finally, beta-glucan free of mannoprotein complexes was prepared (glucan-S3).

  17. Genetic Approaches to Study Meiosis and Meiosis-Specific Gene Expression in Saccharomyces cerevisiae.

    Kassir, Yona; Stuart, David T

    2017-01-01

    The budding yeast Saccharomyces cerevisiae has a long history as a model organism for studies of meiosis and the cell cycle. The popularity of this yeast as a model is in large part due to the variety of genetic and cytological approaches that can be effectively performed with the cells. Cultures of the cells can be induced to synchronously progress through meiosis and sporulation allowing large-scale gene expression and biochemical studies to be performed. Additionally, the spore tetrads resulting from meiosis make it possible to characterize the haploid products of meiosis allowing investigation of meiotic recombination and chromosome segregation. Here we describe genetic methods for analysis progression of S. cerevisiae through meiosis and sporulation with an emphasis on strategies for the genetic analysis of regulators of meiosis-specific genes.

  18. Recombination-stable multimeric green fluorescent protein for characterization of weak promoter outputs in Saccharomyces cerevisiae

    Rugbjerg, Peter; Knuf, Christoph; Förster, Jochen

    2015-01-01

    a less leaky Cu2+-inducible promoter based on CUP1. The basal expression level of the new promoter was approx. 61% below the wild-type CUP1 promoter, thus expanding the absolute range of Cu2+-based gene control. The stability of 3vGFP towards direct-repeat recombination was assayed in S. cerevisiae......Green fluorescent proteins (GFPs) are widely used for visualization of proteins to track localization and expression dynamics. However, phenotypically important processes can operate at too low expression levels for routine detection, i.e. be overshadowed by autofluorescence noise. While GFP...... functions well in translational fusions, the use of tandem GFPs to amplify fluorescence signals is currently avoided in Saccharomyces cerevisiae and many other microorganisms due to the risk of loop-out by direct-repeat recombination. We increased GFP fluorescence by translationally fusing three different...

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

    Hong, Kuk-Ki; Nielsen, Jens

    2012-08-01

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

  20. Effects of aeration on formation and localization of the acetyl coenzyme A synthetases of Saccharomyces cerevisiae

    Klein, H. P.; Jahnke, L.

    1979-01-01

    Previous studies on the yeast Saccharomyces cerevisiae have shown that two different forms of the enzyme acetyl coenzyme A synthetase (ACS) are present, depending on the conditions under which the cells are grown. The paper evaluates the usefulness of a method designed to assay both synthetases simultaneously in yeast homogenates. The data presented confirm the possibility of simultaneous detection and estimation of the amount of both ACSs of S. cerevisiae in crude homogenates of this strain, making possible the study of physiological factors involved in the formation of these isoenzymes. One important factor for specifying which of the two enzymes is found in these yeast cells is the presence or absence of oxygen in their environment. Aeration not only affects the ratio of the two ACSs but also appears to affect the cellular distribution of these enzymes. Most of the data presented suggest the possibility that the nonaerobic ACS may serve as a precursor to the aerobic form.

  1. Bioaccumulation of uranium from waste water using different strains of Saccharomyces cerevisiae

    Tykva, R.; Novak, J.; Podracka, E.; Popa, K.

    2009-01-01

    Five different strains of Saccharomyces cerevisiae were tested for their abilities to accumulate uranium from waste water containing competitive ions. Samples of water passing out from a previous uranium mill were used. The strains tested possess different abilities to accumulate uranium. The kinetics of bioaccumulation, the leaching degree, the influence of cell density and their origin were investigated. Under the applied experimental conditions, more than a half of the total activity (uranium and the decay products) could be accumulated after 60 min contact time of 1 mL (S. cerevisiae) suspension and 5 mL of water. The other cations present in solution effectively competed for the uranium accumulation. 226 Ra and its decay products were completely retained using all tested strains. (authors)

  2. Budding yeast for budding geneticists: a primer on the Saccharomyces cerevisiae model system.

    Duina, Andrea A; Miller, Mary E; Keeney, Jill B

    2014-05-01

    The budding yeast Saccharomyces cerevisiae is a powerful model organism for studying fundamental aspects of eukaryotic cell biology. This Primer article presents a brief historical perspective on the emergence of this organism as a premier experimental system over the course of the past century. An overview of the central features of the S. cerevisiae genome, including the nature of its genetic elements and general organization, is also provided. Some of the most common experimental tools and resources available to yeast geneticists are presented in a way designed to engage and challenge undergraduate and graduate students eager to learn more about the experimental amenability of budding yeast. Finally, a discussion of several major discoveries derived from yeast studies highlights the far-reaching impact that the yeast system has had and will continue to have on our understanding of a variety of cellular processes relevant to all eukaryotes, including humans.

  3. Biocatalytic production of adipic acid from glucose using engineered Saccharomyces cerevisiae

    Kaushik Raj

    2018-06-01

    Full Text Available Adipic acid is an important industrial chemical used in the synthesis of nylon-6,6. The commercial synthesis of adipic acid uses petroleum-derived benzene and releases significant quantities of greenhouse gases. Biocatalytic production of adipic acid from renewable feedstocks could potentially reduce the environmental damage and eliminate the need for fossil fuel precursors. Recently, we have demonstrated the first enzymatic hydrogenation of muconic acid to adipic acid using microbial enoate reductases (ERs - complex iron-sulfur and flavin containing enzymes. In this work, we successfully expressed the Bacillus coagulans ER in a Saccharomyces cerevisiae strain producing muconic acid and developed a three-stage fermentation process enabling the synthesis of adipic acid from glucose. The ability to express active ERs and significant acid tolerance of S. cerevisiae highlight the applicability of the developed yeast strain for the biocatalytic production of adipic acid from renewable feedstocks. Keywords: Biosynthesis, Renewable resources, Yeast, Adipic acid, Synthetic biology

  4. Metabolic engineering of Saccharomyces cerevisiae ethanol strains PE-2 and CAT-1 for efficient lignocellulosic fermentation.

    Romaní, Aloia; Pereira, Filipa; Johansson, Björn; Domingues, Lucília

    2015-03-01

    In this work, Saccharomyces cerevisiae strains PE-2 and CAT-1, commonly used in the Brazilian fuel ethanol industry, were engineered for xylose fermentation, where the first fermented xylose faster than the latter, but also produced considerable amounts of xylitol. An engineered PE-2 strain (MEC1121) efficiently consumed xylose in presence of inhibitors both in synthetic and corn-cob hydrolysates. Interestingly, the S. cerevisiae MEC1121 consumed xylose and glucose simultaneously, while a CEN.PK based strain consumed glucose and xylose sequentially. Deletion of the aldose reductase GRE3 lowered xylitol production to undetectable levels and increased xylose consumption rate which led to higher final ethanol concentrations. Fermentation of corn-cob hydrolysate using this strain, MEC1133, resulted in an ethanol yield of 0.47 g/g of total sugars which is 92% of the theoretical yield. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. Saccharomyces cerevisiae GTPase complex: Gtr1p-Gtr2p regulates cell-proliferation through Saccharomyces cerevisiae Ran-binding protein, Yrb2p

    Wang Yonggang; Nakashima, Nobutaka; Sekiguchi, Takeshi; Nishimoto, Takeharu

    2005-01-01

    A Gtr1p GTPase, the GDP mutant of which suppresses both temperature-sensitive mutants of Saccharomyces cerevisiae RanGEF/Prp20p and RanGAP/Rna1p, was presently found to interact with Yrb2p, the S. cerevisiae homologue of mammalian Ran-binding protein 3. Gtr1p bound the Ran-binding domain of Yrb2p. In contrast, Gtr2p, a partner of Gtr1p, did not bind Yrb2p, although it bound Gtr1p. A triple mutant: yrb2Δ gtr1Δ gtr2Δ was lethal, while a double mutant: gtr1Δ gtr2Δ survived well, indicating that Yrb2p protected cells from the killing effect of gtr1Δ gtr2Δ. Recombinant Gtr1p and Gtr2p were purified as a complex from Escherichia coli. The resulting Gtr1p-Gtr2p complex was comprised of an equal amount of Gtr1p and Gtr2p, which inhibited the Rna1p/Yrb2 dependent RanGAP activity. Thus, the Gtr1p-Gtr2p cycle was suggested to regulate the Ran cycle through Yrb2p

  6. Genomic diversity of Saccharomyces cerevisiae yeasts associated with alcoholic fermentation of bacanora produced by artisanal methods.

    Álvarez-Ainza, M L; Zamora-Quiñonez, K A; Moreno-Ibarra, G M; Acedo-Félix, E

    2015-03-01

    Bacanora is a spirituous beverage elaborated with Agave angustifolia Haw in an artisanal process. Natural fermentation is mostly performed with native yeasts and bacteria. In this study, 228 strains of yeast like Saccharomyces were isolated from the natural alcoholic fermentation on the production of bacanora. Restriction analysis of the amplified region ITS1-5.8S-ITS2 of the ribosomal DNA genes (RFLPr) were used to confirm the genus, and 182 strains were identified as Saccharomyces cerevisiae. These strains displayed high genomic variability in their chromosomes profiles by karyotyping. Electrophoretic profiles of the strains evaluated showed a large number of chromosomes the size of which ranged between 225 and 2200 kpb approximately.

  7. Engineering and Evolution of Saccharomyces cerevisiae to Produce Biofuels and Chemicals.

    Turner, Timothy L; Kim, Heejin; Kong, In Iok; Liu, Jing-Jing; Zhang, Guo-Chang; Jin, Yong-Su

    To mitigate global climate change caused partly by the use of fossil fuels, the production of fuels and chemicals from renewable biomass has been attempted. The conversion of various sugars from renewable biomass into biofuels by engineered baker's yeast (Saccharomyces cerevisiae) is one major direction which has grown dramatically in recent years. As well as shifting away from fossil fuels, the production of commodity chemicals by engineered S. cerevisiae has also increased significantly. The traditional approaches of biochemical and metabolic engineering to develop economic bioconversion processes in laboratory and industrial settings have been accelerated by rapid advancements in the areas of yeast genomics, synthetic biology, and systems biology. Together, these innovations have resulted in rapid and efficient manipulation of S. cerevisiae to expand fermentable substrates and diversify value-added products. Here, we discuss recent and major advances in rational (relying on prior experimentally-derived knowledge) and combinatorial (relying on high-throughput screening and genomics) approaches to engineer S. cerevisiae for producing ethanol, butanol, 2,3-butanediol, fatty acid ethyl esters, isoprenoids, organic acids, rare sugars, antioxidants, and sugar alcohols from glucose, xylose, cellobiose, galactose, acetate, alginate, mannitol, arabinose, and lactose.

  8. Genomic reconstruction to improve bioethanol and ergosterol production of industrial yeast Saccharomyces cerevisiae.

    Zhang, Ke; Tong, Mengmeng; Gao, Kehui; Di, Yanan; Wang, Pinmei; Zhang, Chunfang; Wu, Xuechang; Zheng, Daoqiong

    2015-02-01

    Baker's yeast (Saccharomyces cerevisiae) is the common yeast used in the fields of bread making, brewing, and bioethanol production. Growth rate, stress tolerance, ethanol titer, and byproducts yields are some of the most important agronomic traits of S. cerevisiae for industrial applications. Here, we developed a novel method of constructing S. cerevisiae strains for co-producing bioethanol and ergosterol. The genome of an industrial S. cerevisiae strain, ZTW1, was first reconstructed through treatment with an antimitotic drug followed by sporulation and hybridization. A total of 140 mutants were selected for ethanol fermentation testing, and a significant positive correlation between ergosterol content and ethanol production was observed. The highest performing mutant, ZG27, produced 7.9 % more ethanol and 43.2 % more ergosterol than ZTW1 at the end of fermentation. Chromosomal karyotyping and proteome analysis of ZG27 and ZTW1 suggested that this breeding strategy caused large-scale genome structural variations and global gene expression diversities in the mutants. Genetic manipulation further demonstrated that the altered expression activity of some genes (such as ERG1, ERG9, and ERG11) involved in ergosterol synthesis partly explained the trait improvement in ZG27.

  9. Improved bread-baking process using Saccharomyces cerevisiae displayed with engineered cyclodextrin glucanotransferase.

    Shim, Jae-Hoon; Seo, Nam-Seok; Roh, Sun-Ah; Kim, Jung-Wan; Cha, Hyunju; Park, Kwan-Hwa

    2007-06-13

    A bread-baking process was developed using a potential novel enzyme, cyclodextrin glucanotransferase[3-18] (CGTase[3-18]), that had previously been engineered to have enhanced hydrolyzing activity with little cyclodextrin (CD) formation activity toward starch. CGTase[3-18] was primarily manipulated to be displayed on the cell surface of Saccharomyces cerevisiae. S. cerevisiae carrying pdeltaCGT integrated into the chromosome exhibited starch-hydrolyzing activity at the same optimal pH and temperature as the free enzyme. Volumes of the bread loaves and rice cakes prepared using S. cerevisiae/pdeltaCGT increased by 20% and 45%, respectively, with no detectable CD. Retrogradation rates of the bread and rice cakes decreased significantly during storage. In comparison to the wild type, S. cerevisiae/pdeltaCGT showed improved viability during four freeze-thaw cycles. The results indicated that CGTase[3-18] displayed on the surface of yeast hydrolyzed starch to glucose and maltose that can be used more efficiently for yeast fermentation. Therefore, display of an antistaling enzyme on the cell surface of yeast has potential for enhancing the baking process.

  10. Energetic and metabolic transient response of Saccharomyces cerevisiae to benzoic acid.

    Kresnowati, M T A P; van Winden, W A; van Gulik, W M; Heijnen, J J

    2008-11-01

    Saccharomyces cerevisiae is known to be able to adapt to the presence of the commonly used food preservative benzoic acid with a large energy expenditure. Some mechanisms for the adaptation process have been suggested, but its quantitative energetic and metabolic aspects have rarely been discussed. This study discusses use of the stimulus response approach to quantitatively study the energetic and metabolic aspects of the transient adaptation of S. cerevisiae to a shift in benzoic acid concentration, from 0 to 0.8 mM. The information obtained also serves as the basis for further utilization of benzoic acid as a tool for targeted perturbation of the energy system, which is important in studying the kinetics and regulation of central carbon metabolism in S. cerevisiae. Using this experimental set-up, we found significant fast-transient (< 3000 s) increases in O(2) consumption and CO(2) production rates, of approximately 50%, which reflect a high energy requirement for the adaptation process. We also found that with a longer exposure time to benzoic acid, S. cerevisiae decreases the cell membrane permeability for this weak acid by a factor of 10 and decreases the cell size to approximately 80% of the initial value. The intracellular metabolite profile in the new steady-state indicates increases in the glycolytic and tricarboxylic acid cycle fluxes, which are in agreement with the observed increases in specific glucose and O(2) uptake rates.

  11. Expression of a Dianthus flavonoid glucosyltransferase in Saccharomyces cerevisiae for whole-cell biocatalysis.

    Werner, Sean R; Morgan, John A

    2009-07-15

    Glycosyltransferases are promising biocatalysts for the synthesis of small molecule glycosides. In this study, Saccharomyces cerevisiae expressing a flavonoid glucosyltransferase (GT) from Dianthus caryophyllus (carnation) was investigated as a whole-cell biocatalyst. Two yeast expression systems were compared using the flavonoid naringenin as a model substrate. Under in vitro conditions, naringenin-7-O-glucoside was formed and a higher specific glucosyl transfer activity was found using a galactose inducible expression system compared to a constitutive expression system. However, S. cerevisiae expressing the GT constitutively was significantly more productive than the galactose inducible system under in vivo conditions. Interestingly, the glycosides were recovered directly from the culture broth and did not accumulate intracellularly. A previously uncharacterized naringenin glycoside formed using the D. caryophyllus GT was identified as naringenin-4'-O-glucoside. It was found that S. cerevisiae cells hydrolyze naringenin-7-O-glucoside during whole-cell biocatalysis, resulting in a low final glycoside titer. When phloretin was added as a substrate to the yeast strain expressing the GT constitutively, the natural product phlorizin was formed. This study demonstrates S. cerevisiae is a promising whole-cell biocatalyst host for the production of valuable glycosides.

  12. [Molecular evolution of the sulphite efflux gene SSU1 in Saccharomyces cerevisiae].

    Peng, Li-Xin; Sun, Fei-Fei; Huang, Yan-Yan; Li, Zhen-Chong

    2013-11-01

    The SSU1 gene encoding a membrane sulfite pump is a main facilitator invovled in sulfite efflux. In Saccharomyce cerevisiae, various range of resistance to sulfite was observed among strains. To explore the evolution traits of SSU1 gene, the population data of S. cerevisiae were collected and analyzed. The phylogenetic analysis indicated that S. cerevisiae population can be classified into three sub-populations, and the positive selection was detected in population by McDonald-Kreitman test. The anaylsis of Ka/Ks ratios further showed that S. cerevisiae sub-population was undergoing positive selection. This finding was also supported by PAML branch model. Nine potential positive selection sites were predicted by branch-site model, and four sites exclusively belong to the sub-population under positive seletion. The data from ssulp protein structure demonstrated that three sites are substitutions between polar and hydrophobic amino acids, and only one site of substitutaion from basic amino acid to basic amino acid (345R/K). Because amino acid pKa values are crucial for sulfite pump to maintain their routine function, positive selection of these amino acid substitutions might affect sulfite efflux efficient.

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

    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. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Genomic structural variation contributes to phenotypic change of industrial bioethanol yeast Saccharomyces cerevisiae.

    Zhang, Ke; Zhang, Li-Jie; Fang, Ya-Hong; Jin, Xin-Na; Qi, Lei; Wu, Xue-Chang; Zheng, Dao-Qiong

    2016-03-01

    Genomic structural variation (GSV) is a ubiquitous phenomenon observed in the genomes of Saccharomyces cerevisiae strains with different genetic backgrounds; however, the physiological and phenotypic effects of GSV are not well understood. Here, we first revealed the genetic characteristics of a widely used industrial S. cerevisiae strain, ZTW1, by whole genome sequencing. ZTW1 was identified as an aneuploidy strain and a large-scale GSV was observed in the ZTW1 genome compared with the genome of a diploid strain YJS329. These GSV events led to copy number variations (CNVs) in many chromosomal segments as well as one whole chromosome in the ZTW1 genome. Changes in the DNA dosage of certain functional genes directly affected their expression levels and the resultant ZTW1 phenotypes. Moreover, CNVs of large chromosomal regions triggered an aneuploidy stress in ZTW1. This stress decreased the proliferation ability and tolerance of ZTW1 to various stresses, while aneuploidy response stress may also provide some benefits to the fermentation performance of the yeast, including increased fermentation rates and decreased byproduct generation. This work reveals genomic characters of the bioethanol S. cerevisiae strain ZTW1 and suggests that GSV is an important kind of mutation that changes the traits of industrial S. cerevisiae strains. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. Bioethanol strains of Saccharomyces cerevisiae characterised by microsatellite and stress resistance.

    Reis, Vanda Renata; Antonangelo, Ana Teresa Burlamaqui Faraco; Bassi, Ana Paula Guarnieri; Colombi, Débora; Ceccato-Antonini, Sandra Regina

    Strains of Saccharomyces cerevisiae may display characteristics that are typical of rough-type colonies, made up of cells clustered in pseudohyphal structures and comprised of daughter buds that do not separate from the mother cell post-mitosis. These strains are known to occur frequently in fermentation tanks with significant lower ethanol yield when compared to fermentations carried out by smooth strains of S. cerevisiae that are composed of dispersed cells. In an attempt to delineate genetic and phenotypic differences underlying the two phenotypes, this study analysed 10 microsatellite loci of 22 S. cerevisiae strains as well as stress resistance towards high concentrations of ethanol and glucose, low pH and cell sedimentation rates. The results obtained from the phenotypic tests by Principal-Component Analysis revealed that unlike the smooth colonies, the rough colonies of S. cerevisiae exhibit an enhanced resistance to stressful conditions resulting from the presence of excessive glucose and ethanol and high sedimentation rate. The microsatellite analysis was not successful to distinguish between the colony phenotypes as phenotypic assays. The relevant industrial strain PE-2 was observed in close genetic proximity to rough-colony although it does not display this colony morphology. A unique genetic pattern specific to a particular phenotype remains elusive. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  16. Bioethanol strains of Saccharomyces cerevisiae characterised by microsatellite and stress resistance

    Vanda Renata Reis

    Full Text Available Abstract Strains of Saccharomyces cerevisiae may display characteristics that are typical of rough-type colonies, made up of cells clustered in pseudohyphal structures and comprised of daughter buds that do not separate from the mother cell post-mitosis. These strains are known to occur frequently in fermentation tanks with significant lower ethanol yield when compared to fermentations carried out by smooth strains of S. cerevisiae that are composed of dispersed cells. In an attempt to delineate genetic and phenotypic differences underlying the two phenotypes, this study analysed 10 microsatellite loci of 22 S. cerevisiae strains as well as stress resistance towards high concentrations of ethanol and glucose, low pH and cell sedimentation rates. The results obtained from the phenotypic tests by Principal-Component Analysis revealed that unlike the smooth colonies, the rough colonies of S. cerevisiae exhibit an enhanced resistance to stressful conditions resulting from the presence of excessive glucose and ethanol and high sedimentation rate. The microsatellite analysis was not successful to distinguish between the colony phenotypes as phenotypic assays. The relevant industrial strain PE-2 was observed in close genetic proximity to rough-colony although it does not display this colony morphology. A unique genetic pattern specific to a particular phenotype remains elusive.

  17. Torulaspora delbrueckii contribution in mixed brewing fermentations with different Saccharomyces cerevisiae strains.

    Canonico, Laura; Comitini, Francesca; Ciani, Maurizio

    2017-10-16

    In recent years, there has been growing demand for distinctive high quality beer. Fermentation management has a fundamental role in beer quality and the levels of aroma compounds. Use of non-conventional yeast has been proposed to enhance beer bioflavor. In the present work we investigated mixed fermentations using three commercial Saccharomyces cerevisiae strains, without and with addition of a selected Torulaspora delbrueckii strain evaluating their interactions, as well as the aroma profiles. At the S. cerevisiae/T. delbrueckii co-inoculation ratio of 1:20, viable cell counts indicated that T. delbrueckii dominated all of the three combinations. In the mixed fermentations, T. delbrueckii provided higher levels of higher alcohols (excepting of β-phenyl ethanol), in contrast to data obtained in winemaking, where higher alcohols had lower levels. Moreover, mixed fermentations showed significantly higher ethyl acetate (from 5 to 16mg/L) and isoamyl acetate (from 0.019 to 0.128mg/L), and were generally lower in ethyl hexanoate and ethyl octanoate. Therefore, irrespective of S. cerevisiae strain, T. delbrueckii influenced on all mixed fermentations. On the other hand, the mixed fermentations were also affected by each of the three S. cerevisiae strains, which resulted in beers with distinctive flavors. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Ethanol production from xylose in engineered Saccharomyces cerevisiae strains. Current state and perspectives

    Matsushika, Akinori; Inoue, Hiroyuki; Sawayama, Shigeki [National Inst. of Advanced Industrial Science and Technology (AIST), Hiroshima (JP). Biomass Technology Research Center (BTRC); Kodaki, Tsutomu [Kyoto Univ. (Japan). Inst. of Advanced Energy

    2009-08-15

    Bioethanol production from xylose is important for utilization of lignocellulosic biomass as raw materials. The research on yeast conversion of xylose to ethanol has been intensively studied especially for genetically engineered Saccharomyces cerevisiae during the last 20 years. S. cerevisiae, which is a very safe microorganism that plays a traditional and major role in industrial bioethanol production, has several advantages due to its high ethanol productivity, as well as its high ethanol and inhibitor tolerance. However, this yeast cannot ferment xylose, which is the dominant pentose sugar in hydrolysates of lignocellulosic biomass. A number of different strategies have been applied to engineer yeasts capable of efficiently producing ethanol from xylose, including the introduction of initial xylose metabolism and xylose transport, changing the intracellular redox balance, and overexpression of xylulokinase and pentose phosphate pathways. In this review, recent progress with regard to these studies is discussed, focusing particularly on xylose-fermenting strains of S. cerevisiae. Recent studies using several promising approaches such as host strain selection and adaptation to obtain further improved xylose-utilizing S. cerevisiae are also addressed. (orig.)

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

    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.

  20. Behavior of Lactobacillus plantarum and Saccharomyces cerevisiae in fresh and thermally processed orange juice.

    Alwazeer, Duried; Cachon, Remy; Divies, Charles

    2002-10-01

    Lactobacillus plantarum and Saccharomyces cerevisiae are acid-tolerant microorganisms that are able to spoil citrus juices before and after pasteurization. The growth of these microorganisms in orange juice with and without pasteurization was investigated. Two samples of orange juice were inoculated with ca. 10(5) CFU/ml of each microorganism. Others were inoculated with ca. 10(7) CFU/ml of each microorganism and then thermally treated. L. plantarum populations were reduced by 2.5 and 6 and 2 log10 CFU/ml, respectively. Samples of heated and nonheated juice were incubated at 15 degrees C for 20 days. Injured populations of L. plantarum decreased by ca. 2 log10 CFU/ml during the first 70 h of storage, but those of S. cerevisiae did not decrease. The length of the lag phase after pasteurization increased 6.2-fold for L. plantarum and 1.9-fold for S. cerevisiae, and generation times increased by 41 and 86%, respectively. The results of this study demonstrate the differences in the capabilities of intact and injured cells of spoilage microorganisms to spoil citrus juice and the different thermal resistance levels of cells. While L. plantarum was more resistant to heat treatment than S. cerevisiae was, growth recovery after pasteurization was faster for the latter microorganism.

  1. Bioactive peptides released from Saccharomyces cerevisiae under accelerated autolysis in a wine model system.

    Alcaide-Hidalgo, J M; Pueyo, E; Polo, M C; Martínez-Rodríguez, A J

    2007-09-01

    The ACE inhibitory activity (IACE) and the oxygen radical absorbance capacity (ORAC-FL) values of yeast peptides isolated from a model wine during accelerated autolysis of Saccharomyces cerevisiae have been studied. Samples were taken at 6, 24, 48, 121, and 144 h of autolysis. Peptide concentration increased throughout autolysis process. Peptides were fractionated into 2 fractions: F1, constituted by hydrophilic peptides, and F2, containing hydrophobic peptides. Both IACE activity and ORAC-FL values increased during 121 h of autolysis, then decreased afterward. Peptide fraction F2 was the main fraction involved in IACE activity and ORAC-FL.

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

    Mortimer, R.K.; Contopoulou, R.; Schild, D.

    1981-01-01

    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

  3. Saccharomyces cerevisiae SSB1 protein and its relationship to nucleolar RNA-binding proteins.

    Jong, A Y; Clark, M W; Gilbert, M; Oehm, A; Campbell, J L

    1987-01-01

    To better define the function of Saccharomyces cerevisiae SSB1, an abundant single-stranded nucleic acid-binding protein, we determined the nucleotide sequence of the SSB1 gene and compared it with those of other proteins of known function. The amino acid sequence contains 293 amino acid residues and has an Mr of 32,853. There are several stretches of sequence characteristic of other eucaryotic single-stranded nucleic acid-binding proteins. At the amino terminus, residues 39 to 54 are highly ...

  4. Cell cycle phases in the unequal mother/daughter cell cycles of Saccharomyces cerevisiae.

    Brewer, B J; Chlebowicz-Sledziewska, E; Fangman, W L

    1984-11-01

    During cell division in the yeast Saccharomyces cerevisiae mother cells produce buds (daughter cells) which are smaller and have longer cell cycles. We performed experiments to compare the lengths of cell cycle phases in mothers and daughters. As anticipated from earlier indirect observations, the longer cell cycle time of daughter cells is accounted for by a longer G1 interval. The S-phase and the G2-phase are of the same duration in mother and daughter cells. An analysis of five isogenic strains shows that cell cycle phase lengths are independent of cell ploidy and mating type.

  5. Tight Coupling of Metabolic Oscillations and Intracellular Water Dynamics in Saccharomyces cerevisiae

    Thoke, Henrik Seir; Tobiesen, Asger; Brewer, Jonathan R.

    2015-01-01

    We detected very strong coupling between the oscillating concentration of ATP and the dynamics of intracellular water during glycolysis in Saccharomyces cerevisiae. Our results indicate that: i) dipolar relaxation of intracellular water is heterogeneous within the cell and different from dilute...... conditions, ii) water dipolar relaxation oscillates with glycolysis and in phase with ATP concentration, iii) this phenomenon is scale-invariant from the subcellular to the ensemble of synchronized cells and, iv) the periodicity of both glycolytic oscillations and dipolar relaxation are equally affected by D...

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

    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...... rates were 2.5-3.3-fold higher on glucose than on galactose for all the strains examined, and hence, ethanol production was pronounced on glucose due to respiro-fermentative metabolism. The T256 strain and the T200 strain having the MEL1 gene inserted in the HXK2 locus and the LEU2 locus, respectively...

  7. Quantification of Saccharomyces cerevisiae flocculation by contaminant bacteria from alcoholic fermentation

    LUDWIG, K.M.; OLIVA-NETO, P.; ANGELIS, D.F. de

    2001-01-01

    O assentamento de células de leveduras no fundo das dornas e perdas de células nas centrífugas podem ser causadas por bactérias floculantes, contaminantes naturais da fermentação alcoólica industrial. Estes problemas levam a queda no rendimento e produtividade do etanol. O presente trabalho visa a caracterização da floculação de Saccharomyces cerevisiae por Lactobacillus fermentum CCT 1396. As células de leveduras e bactérias foram misturadas e a floculação das células quantificadas por espec...

  8. Inicio de replicación y estabilidad genómica en Saccharomyces cerevisiae

    Ayuda Durán, Pilar

    2014-01-01

    [ES]La levadura de gemación Saccharomyces cerevisiae constituye un buen sistema modelo para el estudio de los mecanismos que dirigen la replicación del material genético y controlan su estabilidad en organismos eucariotas. Utilizando este modelo se ha estudiado el efecto de la desregulación de actividad CDK durante la fase G1 del ciclo celular de esta levadura, a causa de la carencia de sus reguladores específicos (las proteínas Cdh1, Sic1 y el dominio N-terminal de la proteína Cdc6). L...

  9. Implementation of communication-mediating domains for non-ribosomal peptide production in Saccharomyces cerevisiae

    Siewers, Verena; San-Bento, Rita; Nielsen, Jens

    2010-01-01

    Saccharomyces cerevisiae has in several cases been proven to be a suitable host for the production of natural products and was recently exploited for the production of non-ribosomal peptides. Synthesis of non-ribosomal peptides (NRPs) is mediated by NRP synthetases (NRPSs), modular enzymes, which...... are often organized in enzyme complexes. In these complexes, partner NRPSs interact via communication-mediating domains (COM domains). In order to test whether functional interaction between separate NRPS modules is possible in yeast we constructed a yeast strain expressing two modules with compatible COM...

  10. Ethanol from hydrolyzed whey permeate using Saccharomyces cerevisiae in a membrane recycle bioreactor

    Mehaia, M A [King Saud Univ., Buriedah (Saudi Arabia). Dairy Technology Lab.; Cheryan, M [Illinois Univ., Urbana, IL (USA). Agricultural Bioprocess Lab.

    1990-02-13

    A diauxic fermentation was observed during batch fermentation of enzyme-hydrolyzed whey permeate to ethanol by Saccharomyces cerevisiae. Glucose was consumed before and much faster than galactose. In the continuous membrane recycle bioreactor (MRB), sugar utilization was a function of dilution rate and concentration of sugars. At a cell concentration of 160 kg/m{sup 3}, optimum productivity was 31 kg/(m{sup 3}.h) at ethanol concentration of 65 kg/m{sup 3}. Low levels of acetate (0.05-0.1 M) reduced cell growth during continuous fermentation, but also reduced galactose utilization. (orig.).

  11. Functional screening of aldehyde decarbonylases for long-chain alkane production by Saccharomyces cerevisiae

    Kang, Min-Kyoung; Zhou, Yongjin J.; Buijs, Nicolaas A.

    2017-01-01

    Background: Low catalytic activities of pathway enzymes are often a limitation when using microbial based chemical production. Recent studies indicated that the enzyme activity of aldehyde decarbonylase (AD) is a critical bottleneck for alkane biosynthesis in Saccharomyces cerevisiae. We therefore...... detected in other AD expressed yeast strains. Dynamic expression of SeADO and CwADO under GAL promoters increased alkane production to 0.20 mg/L/OD600 and no fatty alcohols, with even number chain lengths from C8 to C14, were detected in the cells. Conclusions: We demonstrated in vivo enzyme activities...

  12. Oxygen requirements for formation and activity of the squalene expoxidase in Saccharomyces cerevisiae

    Jahnke, L.; Klein, H. P.

    1983-01-01

    The effect of oxygen on squalene epoxidase activity in Saccharomyces cerevisiae was investigated. In cells grown in standing cultures, the epoxidase was localized mainly in the 'mitochondrial' fraction. Upon aeration, enzyme activity increased and the newly formed enzyme was associated with the 'microsomal' fraction. At 0.03 percent (vol/vol) oxygen, epoxidase levels doubled, whereas the ergosterol level was only slightly increased. Cycloheximide inhibited the increase in epoxidase under these conditions. An apparent K sub m for oxygen of 0.38 percent (vol/vol) was determined from a crude particulate preparation for the epoxidase.

  13. Regularities of ''rapid'' repair in radiosensitive mutants of diploid yeasts Saccharomyces cerevisiae

    Glazunov, A.V.; Kapul'tsevich, Yu.G.

    1982-01-01

    A study was made of ''rapid'' repair in radiosensitive mutants of diploid yeast Saccharomyces cerevisiae after irradiation with ν-quanta and α-particles. It was shown that the capacity of ''rapid'' repair does not always correlate with the ability of ''slow'' postirradiation repair of viability of yeast cells. A conclusion is made that ''rapid'' and ''slow'' repair are independent processes. It was found that ''rapid'' repair of the studied strains of diploid yeast is more effective after exposure to ν-quanta than α-particles

  14. The DNA repair capability of cdc9, the saccharomyces cerevisiae mutant defective in DNA ligase

    Johnston, L.H.

    1979-01-01

    The cell cycle mutant, cdc9, in the yeast Saccharomyces cerevisiae is defective in DNA ligase with the consequence to be deficient in the repair of DNA damaged by methyl methane sulphonate. On the other hand survival of cdc9 after irradiation by γ-rays is little different from that of the wild-type, even after a period of stress at the restrictive temperature. The mutant cdc9 is not allelic with any known rad or mms mutants. (orig./AJ) [de

  15. Attempt to stimulate cell division in Saccharomyces cerevisiae with weak ultraviolet light

    Quickenden, T.I.; Matich, A.J.; Pung, S.H.; Tilbury, R.N.

    1989-01-01

    Liquid cultures of the yeast Saccharomyces cerevisiae were irradiated with weak light having irradiances ranging from ca. 1 X 10(2) to 5 X 10(9) photons cm-2 s-1 and at wavelengths ranging from 200 to 700 nm. When particular care was taken to control the temperature of the cultures and the flow rate of oxygen, no evidence was obtained for stimulation of either yeast growth or division by the incident light. These results do not support the claims of early workers that very low intensity uv light can stimulate cell division in living organisms

  16. Repair of UV-irradiated plasmid DNA in excision repair deficient mutants of Saccharomyces cerevisiae

    Ikai, K.; Tano, K.; Ohnishi, T.; Nozu, K.

    1985-01-01

    The repair of UV-irradiated DNA of plasmid YEp13 was studied in the incision defective strains by measurement of cell transformation frequency. In Saccharomyces cerevisiae, rad1,2,3 and 4 mutants could repair UV-damaged plasmid DNA. In Escherichia coli, uvrA mutant was unable to repair UV-damaged plasmid DNA; however, pretreatment of the plasmid with Micrococcus luteus endonuclease increased repair. It was concluded that all the mutations of yeast were probably limited only to the nuclear DNA. (author)

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

    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...

  18. Effects of Infrared Optical Trapping on Saccharomyces cerevisiae in a Microfluidic System

    Pilát, Zdeněk; Jonáš, A.; Ježek, Jan; Zemánek, Pavel

    2017-01-01

    Roč. 17, NOV (2017), s. 1-12, č. článku 2640. ISSN 1424-8220 R&D Projects: GA MŠk(CZ) LO1212; GA MŠk ED0017/01/01 Institutional support: RVO:68081731 Keywords : optical trapping * microfluidics * phototoxicity * laser * Saccharomyces cerevisiae Subject RIV: BH - Optics, Masers, Lasers OBOR OECD: Optics (including laser optics and quantum optics) Impact factor: 2.677, year: 2016 http://www.mdpi.com/1424-8220/17/11/2640

  19. Inorganic polyphosphate in the yeast Saccharomyces cerevisiae with a mutation disturbing the function of vacuolar ATPase.

    Tomaschevsky, A A; Ryasanova, L P; Kulakovskaya, T V; Kulaev, I S

    2010-08-01

    A mutation in the vma2 gene disturbing V-ATPase function in the yeast Saccharomyces cerevisiae results in a five- and threefold decrease in inorganic polyphosphate content in the stationary and active phases of growth on glucose, respectively. The average polyphosphate chain length in the mutant cells is decreased. The mutation does not prevent polyphosphate utilization during cultivation in a phosphate-deficient medium and recovery of its level on reinoculation in complete medium after phosphate deficiency. The content of short chain acid-soluble polyphosphates is recovered first. It is supposed that these polyphosphates are less dependent on the electrochemical gradient on the vacuolar membrane.

  20. Increasing ethanol productivity during xylose fermentation by cell recycling of recombinant Saccharomyces cerevisiae

    Roca, Christophe Francois Aime; Olsson, Lisbeth

    2003-01-01

    The influence of cell recycling of xylose-fermenting Saccharomyces cerevisiae TMB3001 was investigated during continuous cultivation on a xylose-glucose mixture. By using cell recycling at the dilution rate (D) of 0.05 h(-1), the cell-mass concentration could be increased from 2.2 g l(-1) to 22 g l...... ethanol productivity was in the range of 0.23-0.26 g g(-1) h(-1) with or without cell recycling, showing that an increased cell-mass concentration did not influence the efficiency of the yeast....

  1. Effects of lovastatin (mevinolin) on sterol levels and on activity of azoles in Saccharomyces cerevisiae.

    Lorenz, R T; Parks, L W

    1990-01-01

    The hypocholesterolemic drug lovastatin (mevinolin) was found to be very effective in lowering the sterol levels of the wild-type yeast Saccharomyces cerevisiae. Lovastatin dramatically decreased the steryl ester content from 2.62 to 0.8 micrograms/mg (dry weight), whereas the free sterol content decreased only from 2.79 to 2.24 micrograms/mg (dry weight) when lovastatin was present in the medium at 10 micrograms/ml. At higher concentrations (100 micrograms/ml), lovastatin nearly abolished th...

  2. Increased isobutanol production in Saccharomyces cerevisiae by overexpression of genes in valine metabolism

    Chen, Xiao; Nielsen, Kristian Fog; Borodina, Irina

    2011-01-01

    BACKGROUND: Isobutanol can be a better biofuel than ethanol due to its higher energy density and lower hygroscopicity. Furthermore, the branched-chain structure of isobutanol gives a higher octane number than the isomeric n-butanol. Saccharomyces cerevisiae was chosen as the production host because...... of its relative tolerance to alcohols, robustness in industrial fermentations, and the possibility for future combination of isobutanol production with fermentation of lignocellulosic materials. RESULTS: The yield of isobutanol was improved from 0.16 to 0.97 mg per g glucose by simultaneous...

  3. Kinetics of volatile metabolites during alcoholic fermentation of cane molasses by Saccharomyces cerevisiae

    Cachot, T; Mueller, M; Pons, M N [Centre National de la Recherche Scientifique, 54 - Nancy (France). Lab. des Sciences du Genie Chimique

    1991-07-01

    The kinetics of ethanol, acetaldehyde, ethyl acetate and fusel alcohols during alcoholic fermentations on cane molasses by Saccharomyces cerevisiae have been obtained via an in-situ gas membrane sensor connected to a gas chromatograph. Various operation parameters have been investigated such as inoculum rate, molasses concentration, operation mode (batch, fed-batch). The modification of fusel alcohols kinetics in response to addition of amino acids has been studied as well as the assimilation of two intermediary aldehydes (isovaleraldehyde and isobutyraldehyde) in the fusel alcohol synthesis pathway. (orig.).

  4. Isolation and characterization of PEP3, a gene required for vacuolar biogenesis in Saccharomyces cerevisiae.

    Preston, R A; Manolson, M F; Becherer, K; Weidenhammer, E; Kirkpatrick, D; Wright, R; Jones, E W

    1991-01-01

    The Saccharomyces cerevisiae PEP3 gene was cloned from a wild-type genomic library by complementation of the carboxypeptidase Y deficiency in a pep3-12 strain. Subclone complementation results localized the PEP3 gene to a 3.8-kb DNA fragment. The DNA sequence of the fragment was determined; a 2,754-bp open reading frame predicts that the PEP3 gene product is a hydrophilic, 107-kDa protein that has no significant similarity to any known protein. The PEP3 predicted protein has a zinc finger (CX...

  5. Overexpression of the truncated version of ILV2 enhances glycerol production in Saccharomyces cerevisiae.

    Murashchenko, Lidiia; Abbas, Charles; Dmytruk, Kostyantyn; Sibirny, Andriy

    2016-08-01

    Acetolactate synthase is a mitochondrial enzyme that catalyses the conversion of two pyruvate molecules to an acetolactate molecule with release of carbon dioxide. The overexpression of the truncated version of the corresponding gene, ILV2, that codes for presumably cytosolic acetolactate synthase in the yeast Saccharomyces cerevisiae, led to a decrease in intracellular pyruvate concentration. This recombinant strain was also characterized by a four-fold increase in glycerol production, with a concomitant 1.8-fold reduction in ethanol production, when compared to that of the wild-type strain under anaerobic conditions in a glucose alcoholic fermentation. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  6. Bioconversion of lignocellulose-derived sugars to ethanol by engineered Saccharomyces cerevisiae.

    Madhavan, Anjali; Srivastava, Aradhana; Kondo, Akihiko; Bisaria, Virendra S

    2012-03-01

    Lignocellulosic biomass from agricultural and agro-industrial residues represents one of the most important renewable resources that can be utilized for the biological production of ethanol. The yeast Saccharomyces cerevisiae is widely used for the commercial production of bioethanol from sucrose or starch-derived glucose. While glucose and other hexose sugars like galactose and mannose can be fermented to ethanol by S. cerevisiae, the major pentose sugars D-xylose and L-arabinose remain unutilized. Nevertheless, D-xylulose, the keto isomer of xylose, can be fermented slowly by the yeast and thus, the incorporation of functional routes for the conversion of xylose and arabinose to xylulose or xylulose-5-phosphate in Saccharomyces cerevisiae can help to improve the ethanol productivity and make the fermentation process more cost-effective. Other crucial bottlenecks in pentose fermentation include low activity of the pentose phosphate pathway enzymes and competitive inhibition of xylose and arabinose transport into the cell cytoplasm by glucose and other hexose sugars. Along with a brief introduction of the pretreatment of lignocellulose and detoxification of the hydrolysate, this review provides an updated overview of (a) the key steps involved in the uptake and metabolism of the hexose sugars: glucose, galactose, and mannose, together with the pentose sugars: xylose and arabinose, (b) various factors that play a major role in the efficient fermentation of pentose sugars along with hexose sugars, and (c) the approaches used to overcome the metabolic constraints in the production of bioethanol from lignocellulose-derived sugars by developing recombinant S. cerevisiae strains.

  7. Biosorption of heavy metal copper (Cu2+) by Saccharomyces cerevisiae

    Ririhena, S. A. J.; Astuti, A. D.; Fachrul, M. F.; Silalahi, M. D. S.; Hadisoebroto, R.; Rinanti, A.

    2018-01-01

    This research aims to study the optimum effect of contact time and pH adsorption of copper (Cu2+) from electroplating industry waste by dried beer waste S.cerevisiae. This research conducted using batch culture with pH variation 2,3,4,5, and 6, contact time variation 60, 90, 120, 150, 180 minutes, 150 rpm at room temperature (± 28°C), initial Cu2+ concentration 33,746 mg/l, and biosorbent mass 200 mg & 500 mg. The adsorption of heavy metal ions Cu2+ occurs in all variations of pH and contact time at optimum pH. The optimum adsorption occurs at pH 4 with contact time 120 minutes for both 200 mg (41.60%) and 500 mg (61.04%) beer waste biosorbent. Cell morphology seen with Scanning Electron Microscope (SEM) analysis shows the change of cell wall that gets damaged from Cu2+ adsorption. It also proved by the decreased concentration of initial high concentration carboxyl groups. The adsorption process of this research complies to Freundlich Isotherm with R2 value closest to 1 and followed first order kinetic.

  8. Drug resistance is conferred on the model yeast Saccharomyces cerevisiae by expression of full-length melanoma-associated human ATP-binding cassette transporter ABCB5.

    Keniya, Mikhail V; Holmes, Ann R; Niimi, Masakazu; Lamping, Erwin; Gillet, Jean-Pierre; Gottesman, Michael M; Cannon, Richard D

    2014-10-06

    ABCB5, an ATP-binding cassette (ABC) transporter, is highly expressed in melanoma cells, and may contribute to the extreme resistance of melanomas to chemotherapy by efflux of anti-cancer drugs. Our goal was to determine whether we could functionally express human ABCB5 in the model yeast Saccharomyces cerevisiae, in order to demonstrate an efflux function for ABCB5 in the absence of background pump activity from other human transporters. Heterologous expression would also facilitate drug discovery for this important target. DNAs encoding ABCB5 sequences were cloned into the chromosomal PDR5 locus of a S. cerevisiae strain in which seven endogenous ABC transporters have been deleted. Protein expression in the yeast cells was monitored by immunodetection using both a specific anti-ABCB5 antibody and a cross-reactive anti-ABCB1 antibody. ABCB5 function in recombinant yeast cells was measured by determining whether the cells possessed increased resistance to known pump substrates, compared to the host yeast strain, in assays of yeast growth. Three ABCB5 constructs were made in yeast. One was derived from the ABCB5-β mRNA, which is highly expressed in human tissues but is a truncation of a canonical full-size ABC transporter. Two constructs contained full-length ABCB5 sequences: either a native sequence from cDNA or a synthetic sequence codon-harmonized for S. cerevisiae. Expression of all three constructs in yeast was confirmed by immunodetection. Expression of the codon-harmonized full-length ABCB5 DNA conferred increased resistance, relative to the host yeast strain, to the putative substrates rhodamine 123, daunorubicin, tetramethylrhodamine, FK506, or clorgyline. We conclude that full-length ABCB5 can be functionally expressed in S. cerevisiae and confers drug resistance.

  9. Gene Amplification on Demand Accelerates Cellobiose Utilization in Engineered Saccharomyces cerevisiae.

    Oh, Eun Joong; Skerker, Jeffrey M; Kim, Soo Rin; Wei, Na; Turner, Timothy L; Maurer, Matthew J; Arkin, Adam P; Jin, Yong-Su

    2016-06-15

    Efficient microbial utilization of cellulosic sugars is essential for the economic production of biofuels and chemicals. Although the yeast Saccharomyces cerevisiae is a robust microbial platform widely used in ethanol plants using sugar cane and corn starch in large-scale operations, glucose repression is one of the significant barriers to the efficient fermentation of cellulosic sugar mixtures. A recent study demonstrated that intracellular utilization of cellobiose by engineered yeast expressing a cellobiose transporter (encoded by cdt-1) and an intracellular β-glucosidase (encoded by gh1-1) can alleviate glucose repression, resulting in the simultaneous cofermentation of cellobiose and nonglucose sugars. Here we report enhanced cellobiose fermentation by engineered yeast expressing cdt-1 and gh1-1 through laboratory evolution. When cdt-1 and gh1-1 were integrated into the genome of yeast, the single copy integrant showed a low cellobiose consumption rate. However, cellobiose fermentation rates by engineered yeast increased gradually during serial subcultures on cellobiose. Finally, an evolved strain exhibited a 15-fold-higher cellobiose fermentation rate. To identify the responsible mutations in the evolved strain, genome sequencing was performed. Interestingly, no mutations affecting cellobiose fermentation were identified, but the evolved strain contained 9 copies of cdt-1 and 23 copies of gh1-1 We also traced the copy numbers of cdt-1 and gh1-1 of mixed populations during the serial subcultures. The copy numbers of cdt-1 and gh1-1 in the cultures increased gradually with similar ratios as cellobiose fermentation rates of the cultures increased. These results suggest that the cellobiose assimilation pathway (transport and hydrolysis) might be a rate-limiting step in engineered yeast and copies of genes coding for metabolic enzymes might be amplified in yeast if there is a growth advantage. This study indicates that on-demand gene amplification might be an

  10. Impact of acute metal stress in Saccharomyces cerevisiae.

    Dagmar Hosiner

    Full Text Available Although considered as essential cofactors for a variety of enzymatic reactions and for important structural and functional roles in cell metabolism, metals at high concentrations are potent toxic pollutants and pose complex biochemical problems for cells. We report results of single dose acute toxicity testing in the model organism S. cerevisiae. The effects of moderate toxic concentrations of 10 different human health relevant metals, Ag(+, Al(3+, As(3+, Cd(2+, Co(2+, Hg(2+, Mn(2+, Ni(2+, V(3+, and Zn(2+, following short-term exposure were analyzed by transcription profiling to provide the identification of early-on target genes or pathways. In contrast to common acute toxicity tests where defined endpoints are monitored we focused on the entire genomic response. We provide evidence that the induction of central elements of the oxidative stress response by the majority of investigated metals is the basic detoxification process against short-term metal exposure. General detoxification mechanisms also comprised the induction of genes coding for chaperones and those for chelation of metal ions via siderophores and amino acids. Hierarchical clustering, transcription factor analyses, and gene ontology data further revealed activation of genes involved in metal-specific protein catabolism along with repression of growth-related processes such as protein synthesis. Metal ion group specific differences in the expression responses with shared transcriptional regulators for both, up-regulation and repression were also observed. Additionally, some processes unique for individual metals were evident as well. In view of current concerns regarding environmental pollution our results may support ongoing attempts to develop methods to monitor potentially hazardous areas or liquids and to establish standardized tests using suitable eukaryotic a model organism.

  11. Construction of a novel kind of expression plasmid by homologous recombination in Saccharomyces cerevisiae

    CHEN; Xiangling

    2005-01-01

    [1]Brunelli, J. P., Pall, M. L., A series of yeast vectors for expression of cDNAs and other DNA sequences, Yeast, 1993, 9: 1299―1308.[2]Sikorski, R. S., Hieter, P., A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae, Genetics, 1989, 122: 19―27.[3]Bonneaud, N., Ozier-Kalogerogoulos, O., Li, G. et al., A family of low and high copy replicative, integrative and single-stranded S. cerevisiae /E. coli shuttle vector, Yeast, 1991, 7: 609―615.[4]Huo, K. K., Yu, L. L., Chen, X. J., Li, Y. Y., A stable vector for high-level expression and secretion of human interferon alpha A in yeast, Science in China, Ser. B, 1993, 36(5): 557―567.[5]Zhou, Z. X., Yuan, H. Y., He, W. et al., Expression of the modified HBsAg gene SA-28 directed by a constitutive promoter, Journal of Fudan university (Natural Science), 2000, 39(3): 264―268.[6]Paques, F., Haber, J. E., Multiple pathways of recombination induces by double-strand breaks in Saccharomyces cerevisiae, Microbiology and Molecular Biology Reviews, 1999, 63(2): 349―404.[7]Martin, K., Damage-induced recombination in the yeast Saccharomyces cerevisiae, Mutation Research, 2000, 451: 91―105.[8]Alira, S., Tomoko, O., Homologous recombination and the roles of double-strand breaks, TIBS, 1995, 20: 387―391.[9]Patrick, S., Kelly, M. T., Stephen, V. K., Recombination factor of Saccharomyces cerevisiae, Mutation Research, 2000, 451: 257―275.[10]Manivasakam, P., Weber, S. C., McElver, J., Schiestl, R. H., Micro-homology mediated PCR targeting in Saccharomyces cerevisiae, Nucleic Acids Res., 1995, 23(14): 2799―2800.[11]Baudin, A., Lacroute, F., Cullin, C., A simple and efficient method for direct gene deletion in Saccharomyces cerevisiae, Nucleic Acids Res., 1993, 21(14): 3329―3330.[12]Hua, S. B., Qiu, M., Chan, E., Zhu, L., Luo, Y., Minimum length of sequence homology required for in vivo cloning by homolo-gous recombination in yeast, Plasmid, 1997, 38

  12. Phosphorylation of Dgk1 Diacylglycerol Kinase by Casein Kinase II Regulates Phosphatidic Acid Production in Saccharomyces cerevisiae.

    Qiu, Yixuan; Hassaninasab, Azam; Han, Gil-Soo; Carman, George M

    2016-12-16

    In the yeast Saccharomyces cerevisiae, Dgk1 diacylglycerol (DAG) kinase catalyzes the CTP-dependent phosphorylation of DAG to form phosphatidic acid (PA). The enzyme in conjunction with Pah1 PA phosphatase controls the levels of PA and DAG for the synthesis of triacylglycerol and membrane phospholipids, the growth of the nuclear/endoplasmic reticulum membrane, and the formation of lipid droplets. Little is known about how DAG kinase activity is regulated by posttranslational modification. In this work, we examined the phosphorylation of Dgk1 DAG kinase by casein kinase II (CKII). When phosphate groups were globally reduced using nonspecific alkaline phosphatase, Triton X-100-solubilized membranes from DGK1-overexpressing cells showed a 7.7-fold reduction in DAG kinase activity; the reduced enzyme activity could be increased 5.5-fold by treatment with CKII. Dgk1(1-77) expressed heterologously in Escherichia coli was phosphorylated by CKII on a serine residue, and its phosphorylation was dependent on time as well as on the concentrations of CKII, ATP, and Dgk1(1-77). We used site-specific mutagenesis, coupled with phosphorylation analysis and phosphopeptide mapping, to identify Ser-45 and Ser-46 of Dgk1 as the CKII target sites, with Ser-46 being the major phosphorylation site. In vivo, the S46A and S45A/S46A mutations of Dgk1 abolished the stationary phase-dependent stimulation of DAG kinase activity. In addition, the phosphorylation-deficient mutations decreased Dgk1 function in PA production and in eliciting pah1Δ phenotypes, such as the expansion of the nuclear/endoplasmic reticulum membrane, reduced lipid droplet formation, and temperature sensitivity. This work demonstrates that the CKII-mediated phosphorylation of Dgk1 regulates its function in the production of PA. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Phosphorylation of Dgk1 Diacylglycerol Kinase by Casein Kinase II Regulates Phosphatidic Acid Production in Saccharomyces cerevisiae*

    Qiu, Yixuan; Hassaninasab, Azam; Han, Gil-Soo; Carman, George M.

    2016-01-01

    In the yeast Saccharomyces cerevisiae, Dgk1 diacylglycerol (DAG) kinase catalyzes the CTP-dependent phosphorylation of DAG to form phosphatidic acid (PA). The enzyme in conjunction with Pah1 PA phosphatase controls the levels of PA and DAG for the synthesis of triacylglycerol and membrane phospholipids, the growth of the nuclear/endoplasmic reticulum membrane, and the formation of lipid droplets. Little is known about how DAG kinase activity is regulated by posttranslational modification. In this work, we examined the phosphorylation of Dgk1 DAG kinase by casein kinase II (CKII). When phosphate groups were globally reduced using nonspecific alkaline phosphatase, Triton X-100-solubilized membranes from DGK1-overexpressing cells showed a 7.7-fold reduction in DAG kinase activity; the reduced enzyme activity could be increased 5.5-fold by treatment with CKII. Dgk1(1–77) expressed heterologously in Escherichia coli was phosphorylated by CKII on a serine residue, and its phosphorylation was dependent on time as well as on the concentrations of CKII, ATP, and Dgk1(1–77). We used site-specific mutagenesis, coupled with phosphorylation analysis and phosphopeptide mapping, to identify Ser-45 and Ser-46 of Dgk1 as the CKII target sites, with Ser-46 being the major phosphorylation site. In vivo, the S46A and S45A/S46A mutations of Dgk1 abolished the stationary phase-dependent stimulation of DAG kinase activity. In addition, the phosphorylation-deficient mutations decreased Dgk1 function in PA production and in eliciting pah1Δ phenotypes, such as the expansion of the nuclear/endoplasmic reticulum membrane, reduced lipid droplet formation, and temperature sensitivity. This work demonstrates that the CKII-mediated phosphorylation of Dgk1 regulates its function in the production of PA. PMID:27834677

  14. Comparison of the xylose reductase-xylitol dehydrogenase and the xylose isomerase pathways for xylose fermentation by recombinant Saccharomyces cerevisiae

    Hahn-Hägerdal Bärbel

    2007-02-01

    Full Text Available Abstract Background Two heterologous pathways have been used to construct recombinant xylose-fermenting Saccharomyces cerevisiae strains: i the xylose reductase (XR and xylitol dehydrogenase (XDH pathway and ii the xylose isomerase (XI pathway. In the present study, the Pichia stipitis XR-XDH pathway and the Piromyces XI pathway were compared in an isogenic strain background, using a laboratory host strain with genetic modifications known to improve xylose fermentation (overexpressed xylulokinase, overexpressed non-oxidative pentose phosphate pathway and deletion of the aldose reductase gene GRE3. The two isogenic strains and the industrial xylose-fermenting strain TMB 3400 were studied regarding their xylose fermentation capacity in defined mineral medium and in undetoxified lignocellulosic hydrolysate. Results In defined mineral medium, the xylose consumption rate, the specific ethanol productivity, and the final ethanol concentration were significantly higher in the XR- and XDH-carrying strain, whereas the highest ethanol yield was achieved with the strain carrying XI. While the laboratory strains only fermented a minor fraction of glucose in the undetoxified lignocellulose hydrolysate, the industrial strain TMB 3400 fermented nearly all the sugar available. Xylitol was formed by the XR-XDH-carrying strains only in mineral medium, whereas in lignocellulose hydrolysate no xylitol formation was detected. Conclusion Despite by-product formation, the XR-XDH xylose utilization pathway resulted in faster ethanol production than using the best presently reported XI pathway in the strain background investigated. The need for robust industrial yeast strains for fermentation of undetoxified spruce hydrolysates was also confirmed.

  15. Optimization of the Production of Polygalacturonase from Aspergillus kawachii Cloned in Saccharomyces cerevisiae in Batch and Fed-Batch Cultures

    Diego Jorge Baruque

    2011-01-01

    Full Text Available Polygalacturonases (PG; EC 3.2.1.15 catalyze the hydrolysis of pectin and/or pectic acid and are useful for industrial applications such as juice clarification and pectin extraction. Growth and heterologous expression of recombinant Saccharomyces cerevisiae which expresses an acidic PG from Aspergillus kawachii has been studied in batch and fed-batch cultures. Kinetics and stoichiometric parameters of the recombinant yeast were determined in batch cultures in a synthetic medium. In these cultures, the total biomass concentration, protein concentration, and enzyme activity achieved were 2.2 g/L, 10 mg/L, and 3 U/mL, respectively, to give a productivity of 0.06 U/(mL·h. In fed-batch cultures, various strategies for galactose feeding were used: (i after a glucose growth phase, the addition of a single pulse of galactose which gave a productivity of 0.19 U/(mL·h; (ii after a glucose growth phase, a double pulse of galactose at the same final concentration was added, resulting in a productivity of 0.21 U/(mL·h; (iii a simultaneous feeding of glucose and galactose, yielding a productivity of 1.32 U/(mL·h. Based on these results, the simultaneous feeding of glucose and galactose was by far the most suitable strategy for the production of this enzyme. Moreover, some biochemical characteristics of the recombinant enzyme such as a molecular mass of ~60 kDa, an isoelectric point of 3.7 and its ability to hydrolyze polygalacturonic acid at pH=2.5 were determined.

  16. Engineering of a mammalian O-glycosylation pathway in the yeast Saccharomyces cerevisiae: production of O-fucosylated epidermal growth factor domains.

    Chigira, Yuko; Oka, Takuji; Okajima, Tetsuya; Jigami, Yoshifumi

    2008-04-01

    Development of a heterologous system for the production of homogeneous sugar structures has the potential to elucidate structure-function relationships of glycoproteins. In the current study, we used an artificial O-glycosylation pathway to produce an O-fucosylated epidermal growth factor (EGF) domain in Saccharomyces cerevisiae. The in vivo O-fucosylation system was constructed via expression of genes that encode protein O-fucosyltransferase 1 and the EGF domain, along with genes whose protein products convert cytoplasmic GDP-mannose to GDP-fucose. This system allowed identification of an endogenous ability of S. cerevisiae to transport GDP-fucose. Moreover, expression of EGF domain mutants in this system revealed the different contribution of three disulfide bonds to in vivo O-fucosylation. In addition, lectin blotting revealed differences in the ability of fucose-specific lectin to bind the O-fucosylated structure of EGF domains from human factors VII and IX. Further introduction of the human fringe gene into yeast equipped with the in vivo O-fucosylation system facilitated the addition of N-acetylglucosamine to the EGF domain from factor IX but not from factor VII. The results suggest that engineering of an O-fucosylation system in yeast provides a powerful tool for producing proteins with homogenous carbohydrate chains. Such proteins can be used for the analysis of substrate specificity and the production of antibodies that recognize O-glycosylated EGF domains.

  17. Saccharomyces cerevisiae Boulardii Reduces the Deoxynivalenol-Induced Alteration of the Intestinal Transcriptome

    Imourana Alassane-Kpembi

    2018-05-01

    Full Text Available Type B trichothecene mycotoxin deoxynivalenol (DON is one of the most frequently occurring food contaminants. By inducing trans-activation of a number of pro-inflammatory cytokines and increasing the stability of their mRNA, trichothecene can impair intestinal health. Several yeast products, especially Saccharomyces cerevisiae, have the potential for improving the enteric health of piglets, but little is known about the mechanisms by which the administration of yeast counteracts the DON-induced intestinal alterations. Using a pig jejunum explant model, a whole-transcriptome analysis was performed to decipher the early response of the small intestine to the deleterious effects of DON after administration of S. cerevisiae boulardii strain CNCM I-1079. Compared to the control condition, no differentially expressed gene (DE was observed after treatment by yeast only. By contrast, 3619 probes—corresponding to 2771 genes—were differentially expressed following exposure to DON, and 32 signaling pathways were identified from the IPA software functional analysis of the set of DE genes. When the intestinal explants were treated with S. cerevisiae boulardii prior to DON exposure, the number of DE genes decreased by half (1718 probes corresponding to 1384 genes. Prototypical inflammation signaling pathways triggered by DON, including NF-κB and p38 MAPK, were reversed, although the yeast demonstrated limited efficacy toward some other pathways. S. cerevisiae boulardii also restored the lipid metabolism signaling pathway, and reversed the down-regulation of the antioxidant action of vitamin C signaling pathway. The latter effect could reduce the burden of DON-induced oxidative stress. Altogether, the results show that S. cerevisiae boulardii reduces the DON-induced alteration of intestinal transcriptome, and point to new mechanisms for the healing of tissue injury by yeast.

  18. COCOA (Theobroma cacao) Polyphenol-Rich Extract Increases the Chronological Lifespan of Saccharomyces cerevisiae.

    Baiges, I; Arola, L

    2016-01-01

    BACKGROUND: Saccharomyces cerevisiae is a model organism with conserved aging pathways. Yeast chronological lifespan experiments mimic the processes involved in human non-dividing tissues, such as the nervous system or skeletal muscle, and can speed up the search for biomolecules with potential anti-aging effects before proceeding to animal studies. OBJECTIVE: To test the effectiveness of a cocoa polyphenol-rich extract (CPE) in expanding the S. cerevisiae chronological lifespan in two conditions: in the stationary phase reached after glucose depletion and under severe caloric restriction. MEASUREMENTS: Using a high-throughput method, wild-type S. cerevisiae and its mitochondrial manganese-dependent superoxide dismutase null mutant (sod2Δ) were cultured in synthetic complete dextrose medium. After 2 days, 0, 5 and 20 mg/ml of CPE were added, and viability was measured throughout the stationary phase. The effects of the major components of CPE were also evaluated. To determine yeast lifespan under severe caloric restriction conditions, cultures were washed with water 24 h after the addition of 0 and 20 mg/ml of CPE, and viability was followed over time. RESULTS : CPE increased the chronological lifespan of S. cerevisiae during the stationary phase in a dose-dependent manner. A similar increase was also observed in (sod2Δ). None of the major CPE components (theobromine, caffeine, maltodextrin, (-)-epicatechin, (+)-catechin and procyanidin B2) was able to increase the yeast lifespan. CPE further increased the yeast lifespan under severe caloric restriction. CONCLUSION: CPE increases the chronological lifespan of S. cerevisiae through a SOD2-independent mechanism. The extract also extends yeast lifespan under severe caloric restriction conditions. The high-throughput assay used makes it possible to simply and rapidly test the efficacy of a large number of compounds on yeast aging, requiring only small amounts, and is thus a convenient screening assay to accelerate

  19. Transcriptome response to alkane biofuels in Saccharomyces cerevisiae: identification of efflux pumps involved in alkane tolerance

    2013-01-01

    Background Hydrocarbon alkanes have been recently considered as important next-generation biofuels because microbial production of alkane biofuels was demonstrated. However, the toxicity of alkanes to microbial hosts can possibly be a bottleneck for high productivity of alkane biofuels. To tackle this toxicity issue, it is essential to understand molecular mechanisms of interactions between alkanes and microbial hosts, and to harness these mechanisms to develop microbial host strains with improved tolerance against alkanes. In this study, we aimed to improve the tolerance of Saccharomyces cerevisiae, a model eukaryotic host of industrial significance, to alkane biofuels by exploiting cellular mechanisms underlying alkane response. Results To this end, we first confirmed that nonane (C9), decane (C10), and undecane (C11) were significantly toxic and accumulated in S. cerevisiae. Transcriptome analyses suggested that C9 and C10 induced a range of cellular mechanisms such as efflux pumps, membrane modification, radical detoxification, and energy supply. Since efflux pumps could possibly aid in alkane secretion, thereby reducing the cytotoxicity, we formed the hypothesis that those induced efflux pumps could contribute to alkane export and tolerance. In support of this hypothesis, we demonstrated the roles of the efflux pumps Snq2p and Pdr5p in reducing intracellular levels of C10 and C11, as well as enhancing tolerance levels against C10 and C11. This result provided the evidence that Snq2p and Pdr5p were associated with alkane export and tolerance in S. cerevisiae. Conclusions Here, we investigated the cellular mechanisms of S. cerevisiae response to alkane biofuels at a systems level through transcriptome analyses. Based on these mechanisms, we identified efflux pumps involved in alkane export and tolerance in S. cerevisiae. We believe that the results here provide valuable insights into designing microbial engineering strategies to improve cellular tolerance for

  20. Characterization of the Viable but Nonculturable (VBNC State in Saccharomyces cerevisiae.

    Mohammad Salma

    Full Text Available The Viable But Non Culturable (VBNC state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to "resuscitate". The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO2-induced VBNC state in S. cerevisiae and that the stress removal allows the "resuscitation" of VBNC cells during the VBNC state.