WorldWideScience

Sample records for human yeast escherichia

  1. Yeast redoxyendonuclease, a DNA repair enzyme similar to Escherichia coli endonuclease III

    International Nuclear Information System (INIS)

    Gossett, J.; Lee, K.; Cunningham, R.P.; Doetsch, P.W.

    1988-01-01

    A DNA repair endonuclease (redoxyendonuclease) was isolated from bakers' yeast (Saccharomyces cerevisiae). The enzyme has been purified by a series of column chromatography steps and cleaves OsO 4 -damaged, double-stranded DNA at sites of thymine glycol and heavily UV-irradiated DNA at sites of cytosine, thymine, and guanine photoproducts. The base specificity and mechanism of phosphodiester bond cleavage for the yeast redoxyendonuclease appear to be identical with those of Escherichia coli endonuclease III when thymine glycol containing, end-labeled DNA fragments of defined sequence are employed as substrates. Yeast redoxyendonuclease has an apparent molecular size of 38,000-42,000 daltons and is active in the absence of divalent metal cations. The identification of such an enzyme in yeast may be of value in the elucidation of the biochemical basis for radiation sensitivity in certain yeast mutants

  2. Functional conservation of coenzyme Q biosynthetic genes among yeasts, plants, and humans.

    Directory of Open Access Journals (Sweden)

    Kazuhiro Hayashi

    Full Text Available Coenzyme Q (CoQ is an essential factor for aerobic growth and oxidative phosphorylation in the electron transport system. The biosynthetic pathway for CoQ has been proposed mainly from biochemical and genetic analyses of Escherichia coli and Saccharomyces cerevisiae; however, the biosynthetic pathway in higher eukaryotes has been explored in only a limited number of studies. We previously reported the roles of several genes involved in CoQ synthesis in the fission yeast Schizosaccharomyces pombe. Here, we expand these findings by identifying ten genes (dps1, dlp1, ppt1, and coq3-9 that are required for CoQ synthesis. CoQ10-deficient S. pombe coq deletion strains were generated and characterized. All mutant fission yeast strains were sensitive to oxidative stress, produced a large amount of sulfide, required an antioxidant to grow on minimal medium, and did not survive at the stationary phase. To compare the biosynthetic pathway of CoQ in fission yeast with that in higher eukaryotes, the ability of CoQ biosynthetic genes from humans and plants (Arabidopsis thaliana to functionally complement the S. pombe coq deletion strains was determined. With the exception of COQ9, expression of all other human and plant COQ genes recovered CoQ10 production by the fission yeast coq deletion strains, although the addition of a mitochondrial targeting sequence was required for human COQ3 and COQ7, as well as A. thaliana COQ6. In summary, this study describes the functional conservation of CoQ biosynthetic genes between yeasts, humans, and plants.

  3. Increased biological activity of deglycosylated recombinant human granulocyte/macrophage colony-stimulating factor produced by yeast or animal cells

    International Nuclear Information System (INIS)

    Moonen, P.; Mermod, J.J.; Ernst, J.F.; Hirschi, M.; DeLamarter, J.F.

    1987-01-01

    Human granulocyte/macrophage colony-stimulating factor (hGM-CSF) produced by several recombinant sources including Escherichia coli, yeast, and animal cells was studied. Recombinant animal cells produced hGM-CSF in low quantities and in multiple forms of varying size. Mammalian hGM-CSF was purified 200,000-fold using immunoaffinity and lectin chromatography. Partially purified proteins produced in yeast and mammalian cells were assayed for the effects of deglycosylation. Following enzymatic deglycosylation, immunoreactivity was measured by radioimmunoassay and biological activity was measured in vitro on responsive human primary cells. Removal of N-linked oligosaccharides from both proteins increased their immunoreactivities by 4- to 8-fold. Removal of these oligosaccharides also increased their specific biological activities about 20-fold, to reach approximately the specific activity of recombinant hGM-CSF from E. coli. The E. coli produced-protein-lacking any carbohydrate- had by far the highest specific activity observed for the recombinant hGM-CSFs

  4. Yeast modulation of human dendritic cell cytokine secretion: an in vitro study.

    Directory of Open Access Journals (Sweden)

    Ida M Smith

    Full Text Available Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. The concept of individual microorganisms influencing the makeup of T cell subsets via interactions with intestinal dendritic cells (DCs appears to constitute the foundation for immunoregulatory effects of probiotics, and several studies have reported probiotic strains resulting in reduction of intestinal inflammation through modulation of DC function. Consequent to a focus on Saccharomyces boulardii as the fundamental probiotic yeast, very little is known about hundreds of non-Saccharomyces yeasts in terms of their interaction with the human gastrointestinal immune system. The aim of the present study was to evaluate 170 yeast strains representing 75 diverse species for modulation of inflammatory cytokine secretion by human DCs in vitro, as compared to cytokine responses induced by a S. boulardii reference strain with probiotic properties documented in clinical trials. Furthermore, we investigated whether cytokine inducing interactions between yeasts and human DCs are dependent upon yeast viability or rather a product of membrane interactions regardless of yeast metabolic function. We demonstrate high diversity in yeast induced cytokine profiles and employ multivariate data analysis to reveal distinct clustering of yeasts inducing similar cytokine profiles in DCs, highlighting clear species distinction within specific yeast genera. The observed differences in induced DC cytokine profiles add to the currently very limited knowledge of the cross-talk between yeasts and human immune cells and provide a foundation for selecting yeast strains for further characterization and development toward potentially novel yeast probiotics. Additionally, we present data to support a hypothesis that the interaction between yeasts and human DCs does not solely depend on yeast viability, a concept which may suggest a need for further classifications

  5. Yeast Modulation of Human Dendritic Cell Cytokine Secretion: An In Vitro Study

    Science.gov (United States)

    Smith, Ida M.; Christensen, Jeffrey E.; Arneborg, Nils; Jespersen, Lene

    2014-01-01

    Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. The concept of individual microorganisms influencing the makeup of T cell subsets via interactions with intestinal dendritic cells (DCs) appears to constitute the foundation for immunoregulatory effects of probiotics, and several studies have reported probiotic strains resulting in reduction of intestinal inflammation through modulation of DC function. Consequent to a focus on Saccharomyces boulardii as the fundamental probiotic yeast, very little is known about hundreds of non-Saccharomyces yeasts in terms of their interaction with the human gastrointestinal immune system. The aim of the present study was to evaluate 170 yeast strains representing 75 diverse species for modulation of inflammatory cytokine secretion by human DCs in vitro, as compared to cytokine responses induced by a S. boulardii reference strain with probiotic properties documented in clinical trials. Furthermore, we investigated whether cytokine inducing interactions between yeasts and human DCs are dependent upon yeast viability or rather a product of membrane interactions regardless of yeast metabolic function. We demonstrate high diversity in yeast induced cytokine profiles and employ multivariate data analysis to reveal distinct clustering of yeasts inducing similar cytokine profiles in DCs, highlighting clear species distinction within specific yeast genera. The observed differences in induced DC cytokine profiles add to the currently very limited knowledge of the cross-talk between yeasts and human immune cells and provide a foundation for selecting yeast strains for further characterization and development toward potentially novel yeast probiotics. Additionally, we present data to support a hypothesis that the interaction between yeasts and human DCs does not solely depend on yeast viability, a concept which may suggest a need for further classifications beyond the current

  6. The ecology of insect-yeast relationships and its relevance to human industry.

    Science.gov (United States)

    Madden, Anne A; Epps, Mary Jane; Fukami, Tadashi; Irwin, Rebecca E; Sheppard, John; Sorger, D Magdalena; Dunn, Robert R

    2018-03-28

    Many species of yeast are integral to human society. They produce many of our foods, beverages and industrial chemicals, challenge us as pathogens, and provide models for the study of our own biology. However, few species are regularly studied and much of their ecology remains unclear, hindering the development of knowledge that is needed to improve the relationships between humans and yeasts. There is increasing evidence that insects are an essential component of ascomycetous yeast ecology. We propose a 'dispersal-encounter hypothesis' whereby yeasts are dispersed by insects between ephemeral, spatially disparate sugar resources, and insects, in turn, obtain the benefits of an honest signal from yeasts for the sugar resources. We review the relationship between yeasts and insects through three main examples: social wasps, social bees and beetles, with some additional examples from fruit flies. Ultimately, we suggest that over the next decades, consideration of these ecological and evolutionary relationships between insects and yeasts will allow prediction of where new yeast diversity is most likely to be discovered, particularly yeasts with traits of interest to human industry. © 2018 The Author(s).

  7. Expression of human α-fetoprotein in yeast

    International Nuclear Information System (INIS)

    Yamamoto, Ritsu; Sakamoto, Takashi; Nishi, Shinzo; Sakai, Masaharu; Morinaga, Tomonori; Tamaoki, Taiki

    1990-01-01

    Human α-fetoprotein (AFP) was expressed in Saccharomyces cerevisiae, with a plasmid containing the cDNA sequence for human AFP fused with the rat AFP signal peptide. The recombinant AFP was purified from the yeast lysate by DEAE-cellulose and immunoaffinity chromatography. The amino acid composition and the molecular weight of the recombinant AFP were similar to those of hepatoma AFP. N-terminal amino acids sequence analysis indicated that the signal peptide had been processed. The recombinant and hepatoma AFP reacted identically in Ouchterlony immunodiffusion and radioimmunoassay tests. These observations indicated that the yeast recombinant protein had the properties of native AFP

  8. Overexpression of functional human oxidosqualene cyclase in Escherichia coli

    DEFF Research Database (Denmark)

    Kürten, Charlotte; Uhlén, Mathias; Syrén, Per-Olof

    2015-01-01

    The generation of multicyclic scaffolds from linear oxidosqualene by enzymatic polycyclization catalysis constitutes a cornerstone in biology for the generation of bioactive compounds. Human oxidosqualene cyclase (hOSC) is a membrane-bound triterpene cyclase that catalyzes the formation of the te......The generation of multicyclic scaffolds from linear oxidosqualene by enzymatic polycyclization catalysis constitutes a cornerstone in biology for the generation of bioactive compounds. Human oxidosqualene cyclase (hOSC) is a membrane-bound triterpene cyclase that catalyzes the formation...... of the tetracyclic steroidal backbone, a key step in cholesterol biosynthesis. Protein expression of hOSC and other eukaryotic oxidosqualene cyclases has traditionally been performed in yeast and insect cells, which has resulted in protein yields of 2.7mg protein/g cells (hOSC in Pichia pastoris) after 48h...... of expression. Herein we present, to the best of our knowledge, the first functional expression of hOSC in the model organism Escherichia coli. Using a codon-optimized gene and a membrane extraction procedure for which detergent is immediately added after cell lysis, a protein yield of 2.9mg/g bacterial cells...

  9. Yeast prions and human prion-like proteins: sequence features and prediction methods.

    Science.gov (United States)

    Cascarina, Sean M; Ross, Eric D

    2014-06-01

    Prions are self-propagating infectious protein isoforms. A growing number of prions have been identified in yeast, each resulting from the conversion of soluble proteins into an insoluble amyloid form. These yeast prions have served as a powerful model system for studying the causes and consequences of prion aggregation. Remarkably, a number of human proteins containing prion-like domains, defined as domains with compositional similarity to yeast prion domains, have recently been linked to various human degenerative diseases, including amyotrophic lateral sclerosis. This suggests that the lessons learned from yeast prions may help in understanding these human diseases. In this review, we examine what has been learned about the amino acid sequence basis for prion aggregation in yeast, and how this information has been used to develop methods to predict aggregation propensity. We then discuss how this information is being applied to understand human disease, and the challenges involved in applying yeast prediction methods to higher organisms.

  10. The Escherichia coli Tus-Ter replication fork barrier causes site-specific DNA replication perturbation in yeast.

    Science.gov (United States)

    Larsen, Nicolai B; Sass, Ehud; Suski, Catherine; Mankouri, Hocine W; Hickson, Ian D

    2014-04-07

    Replication fork (RF) pausing occurs at both 'programmed' sites and non-physiological barriers (for example, DNA adducts). Programmed RF pausing is required for site-specific DNA replication termination in Escherichia coli, and this process requires the binding of the polar terminator protein, Tus, to specific DNA sequences called Ter. Here, we demonstrate that Tus-Ter modules also induce polar RF pausing when engineered into the Saccharomyces cerevisiae genome. This heterologous RF barrier is distinct from a number of previously characterized, protein-mediated, RF pause sites in yeast, as it is neither Tof1-dependent nor counteracted by the Rrm3 helicase. Although the yeast replisome can overcome RF pausing at Tus-Ter modules, this event triggers site-specific homologous recombination that requires the RecQ helicase, Sgs1, for its timely resolution. We propose that Tus-Ter can be utilized as a versatile, site-specific, heterologous DNA replication-perturbing system, with a variety of potential applications.

  11. The truth about mouse, human, worms and yeast

    Directory of Open Access Journals (Sweden)

    Nelson David R

    2004-01-01

    Full Text Available Abstract Genome comparisons are behind the powerful new annotation methods being developed to find all human genes, as well as genes from other genomes. Genomes are now frequently being studied in pairs to provide cross-comparison datasets. This 'Noah's Ark' approach often reveals unsuspected genes and may support the deletion of false-positive predictions. Joining mouse and human as the cross-comparison dataset for the first two mammals are: two Drosophila species, D. melanogaster and D. pseudoobscura; two sea squirts, Ciona intestinalis and Ciona savignyi; four yeast (Saccharomyces species; two nematodes, Caenorhabditis elegans and Caenorhabditis briggsae; and two pufferfish (Takefugu rubripes and Tetraodon nigroviridis. Even genomes like yeast and C. elegans, which have been known for more than five years, are now being significantly improved. Methods developed for yeast or nematodes will now be applied to mouse and human, and soon to additional mammals such as rat and dog, to identify all the mammalian protein-coding genes. Current large disparities between human Unigene predictions (127,835 genes and gene-scanning methods (45,000 genes still need to be resolved. This will be the challenge during the next few years.

  12. Yeast Biodiversity in Vineyard Environments Is Increased by Human Intervention.

    Directory of Open Access Journals (Sweden)

    João Drumonde-Neves

    Full Text Available One hundred and five grape samples were collected during two consecutive years from 33 locations on seven oceanic islands of the Azores Archipelago. Grape samples were obtained from vineyards that were either abandoned or under regular cultivation involving common viticultural interventions, to evaluate the impact of regular human intervention on grape yeast biota diversity in vineyards. A total of 3150 yeast isolates were obtained and 23 yeast species were identified. The predominant species were Hanseniaspora uvarum, Pichia terricola, Starmerella bacillaris and Issatchenkia hanoiensis. The species Barnettozyma californica, Candida azymoides and Pichia cecembensis were reported in grapes or wine-associated environments for the first time. A higher biodiversity was found in active vineyards where regular human intervention takes place (Shannon index: 1.89 and 1.53 in the first and second years, respectively when compared to the abandoned ones (Shannon index: 0.76 and 0.31. This finding goes against the assumptions that human intervention can destroy biodiversity and lead to homogeneity in the environment. Biodiversity indices were considerably lower in the year with the heaviest rainfall. This study is the first to report on the grape yeast communities from several abandoned vineyards that have undergone no human intervention.

  13. Pharmaceutical protein production by yeast: towards production of human blood proteins by microbial fermentation

    DEFF Research Database (Denmark)

    Martinez Ruiz, José Luis; Liu, Lifang; Petranovic, Dina

    2012-01-01

    Since the approval of recombinant insulin from Escherichia coli for its clinical use in the early 1980s, the amount of recombinant pharmaceutical proteins obtained by microbial fermentations has significantly increased. The recent advances in genomics together with high throughput analysis...... of recombinant therapeutics using yeast Saccharomyces cerevisiae as a model platform, and discusses the future potential of this platform for production of blood proteins and substitutes....

  14. High-yield expression in Escherichia coli, purification and application of budding yeast K2 killer protein.

    Science.gov (United States)

    Podoliankaitė, Monika; Lukša, Juliana; Vyšniauskis, Gintautas; Sereikaitė, Jolanta; Melvydas, Vytautas; Serva, Saulius; Servienė, Elena

    2014-07-01

    Saccharomyces cerevisiae K2 toxin is a highly active extracellular protein, important as a biocontrol agent for biotechnological applications in the wine industry. This protein is produced at negligible levels in yeast, making difficult to isolate it in amounts sufficient for investigation and generation of analysis tools. In this work, we demonstrate the use of a bacterial system for expression of the recombinant K2 protein, suitable for generation of antibodies specific for toxin of the yeast origin. Synthesis of the full-length S. cerevisiae K2 preprotoxin in Escherichia coli was found to be toxic to the host cell, resulting in diminished growth. Such effect was abolished by the introduction of the C-terminal truncation into K2 protein, directing it into non-toxic inclusion body fraction. The obtained protein is of limited solubility thus, facilitating the purification by simple and efficient chromatography-free procedure. The protein aggregates were successfully refolded into a soluble form yielding sufficient amounts of a tag-less truncated K2 protein suitable for polyclonal antibody production. Antibodies were raised in rabbit and found to be specific for detection of both antigen and native S. cerevisiae K2 toxin.

  15. Identifying pathogenicity of human variants via paralog-based yeast complementation.

    Directory of Open Access Journals (Sweden)

    Fan Yang

    2017-05-01

    Full Text Available To better understand the health implications of personal genomes, we now face a largely unmet challenge to identify functional variants within disease-associated genes. Functional variants can be identified by trans-species complementation, e.g., by failure to rescue a yeast strain bearing a mutation in an orthologous human gene. Although orthologous complementation assays are powerful predictors of pathogenic variation, they are available for only a few percent of human disease genes. Here we systematically examine the question of whether complementation assays based on paralogy relationships can expand the number of human disease genes with functional variant detection assays. We tested over 1,000 paralogous human-yeast gene pairs for complementation, yielding 34 complementation relationships, of which 33 (97% were novel. We found that paralog-based assays identified disease variants with success on par with that of orthology-based assays. Combining all homology-based assay results, we found that complementation can often identify pathogenic variants outside the homologous sequence region, presumably because of global effects on protein folding or stability. Within our search space, paralogy-based complementation more than doubled the number of human disease genes with a yeast-based complementation assay for disease variation.

  16. Beneficial Effects of Probiotic and Food Borne Yeasts on Human Health

    Directory of Open Access Journals (Sweden)

    Saloomeh Moslehi-Jenabian

    2010-04-01

    Full Text Available Besides being important in the fermentation of foods and beverages, yeasts have shown numerous beneficial effects on human health. Among these, probiotic effects are the most well known health effects including prevention and treatment of intestinal diseases and immunomodulatory effects. Other beneficial functions of yeasts are improvement of bioavailability of minerals through the hydrolysis of phytate, folate biofortification and detoxification of mycotoxins due to surface binding to the yeast cell wall.

  17. Mitochondrial import of human and yeast fumarase in live mammalian cells: Retrograde translocation of the yeast enzyme is mainly caused by its poor targeting sequence

    International Nuclear Information System (INIS)

    Singh, Bhag; Gupta, Radhey S.

    2006-01-01

    Studies on yeast fumarase provide the main evidence for dual localization of a protein in mitochondria and cytosol by means of retrograde translocation. We have examined the subcellular targeting of yeast and human fumarase in live cells to identify factors responsible for this. The cDNAs for mature yeast or human fumarase were fused to the gene for enhanced green fluorescent protein (eGFP) and they contained, at their N-terminus, a mitochondrial targeting sequence (MTS) derived from either yeast fumarase, human fumarase, or cytochrome c oxidase subunit VIII (COX) protein. Two nuclear localization sequences (2x NLS) were also added to these constructs to facilitate detection of any cytosolic protein by its targeting to nucleus. In Cos-1 cells transfected with these constructs, human fumarase with either the native or COX MTSs was detected exclusively in mitochondria in >98% of the cells, while the remainder 1-2% of the cells showed varying amounts of nuclear labeling. In contrast, when human fumarase was fused to the yeast MTS, >50% of the cells showed nuclear labeling. Similar studies with yeast fumarase showed that with its native MTS, nuclear labeling was seen in 80-85% of the cells, but upon fusion to either human or COX MTS, nuclear labeling was observed in only 10-15% of the cells. These results provide evidence that extramitochondrial presence of yeast fumarase is mainly caused by the poor mitochondrial targeting characteristics of its MTS (but also affected by its primary sequence), and that the retrograde translocation mechanism does not play a significant role in the extramitochondrial presence of mammalian fumarase

  18. Active role of a human genomic insert in replication of a yeast artificial chromosome.

    Science.gov (United States)

    van Brabant, A J; Fangman, W L; Brewer, B J

    1999-06-01

    Yeast artificial chromosomes (YACs) are a common tool for cloning eukaryotic DNA. The manner by which large pieces of foreign DNA are assimilated by yeast cells into a functional chromosome is poorly understood, as is the reason why some of them are stably maintained and some are not. We examined the replication of a stable YAC containing a 240-kb insert of DNA from the human T-cell receptor beta locus. The human insert contains multiple sites that serve as origins of replication. The activity of these origins appears to require the yeast ARS consensus sequence and, as with yeast origins, additional flanking sequences. In addition, the origins in the human insert exhibit a spacing, a range of activation efficiencies, and a variation in times of activation during S phase similar to those found for normal yeast chromosomes. We propose that an appropriate combination of replication origin density, activation times, and initiation efficiencies is necessary for the successful maintenance of YAC inserts.

  19. Distant homology between yeast photoreactivating gene fragment and human genomic digests

    International Nuclear Information System (INIS)

    Meechan, P.J.; Milam, K.M.; Cleaver, J.E.

    1985-01-01

    Hybridization of DNA coding for the yeast DNA photolyase to human genomic DNA appears to allow one to determine whether a conserved enzyme is coded for in human cells. Under stringent conditions (68 0 C), hybridization is not found between the cloned yeast fragment (YEp13-phr1) and human or chick genomic digests. At less stringent conditions (60 0 C), hybridization is observed with chick digests, indicating evolutionary divergence even among organisms capable of photo-reactivation. At 50 0 C, weak hybridization with human digests was observed, indicating further divergence from the cloned gene. Data concerning the precise extent of homology and methods to clone the chick gene for use as another probe are discussed

  20. Phosphorylation and cellular function of the human Rpa2 N-terminus in the budding yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Ghospurkar, Padmaja L; Wilson, Timothy M; Liu, Shengqin; Herauf, Anna; Steffes, Jenna; Mueller, Erica N; Oakley, Gregory G; Haring, Stuart J

    2015-02-01

    Maintenance of genome integrity is critical for proper cell growth. This occurs through accurate DNA replication and repair of DNA lesions. A key factor involved in both DNA replication and the DNA damage response is the heterotrimeric single-stranded DNA (ssDNA) binding complex Replication Protein A (RPA). Although the RPA complex appears to be structurally conserved throughout eukaryotes, the primary amino acid sequence of each subunit can vary considerably. Examination of sequence differences along with the functional interchangeability of orthologous RPA subunits or regions could provide insight into important regions and their functions. This might also allow for study in simpler systems. We determined that substitution of yeast Replication Factor A (RFA) with human RPA does not support yeast cell viability. Exchange of a single yeast RFA subunit with the corresponding human RPA subunit does not function due to lack of inter-species subunit interactions. Substitution of yeast Rfa2 with domains/regions of human Rpa2 important for Rpa2 function (i.e., the N-terminus and the loop 3-4 region) supports viability in yeast cells, and hybrid proteins containing human Rpa2 N-terminal phospho-mutations result in similar DNA damage phenotypes to analogous yeast Rfa2 N-terminal phospho-mutants. Finally, the human Rpa2 N-terminus (NT) fused to yeast Rfa2 is phosphorylated in a manner similar to human Rpa2 in human cells, indicating that conserved kinases recognize the human domain in yeast. The implication is that budding yeast represents a potential model system for studying not only human Rpa2 N-terminal phosphorylation, but also phosphorylation of Rpa2 N-termini from other eukaryotic organisms. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  1. The Yeast Saccharomyces cerevisiae as a Model for Understanding RAS Proteins and Their Role in Human Tumorigenesis

    Science.gov (United States)

    Cazzanelli, Giulia; Francisco, Rita; Azevedo, Luísa; Carvalho, Patrícia Dias; Almeida, Ana; Côrte-Real, Manuela; Oliveira, Maria José; Lucas, Cândida; Sousa, Maria João

    2018-01-01

    The exploitation of the yeast Saccharomyces cerevisiae as a biological model for the investigation of complex molecular processes conserved in multicellular organisms, such as humans, has allowed fundamental biological discoveries. When comparing yeast and human proteins, it is clear that both amino acid sequences and protein functions are often very well conserved. One example of the high degree of conservation between human and yeast proteins is highlighted by the members of the RAS family. Indeed, the study of the signaling pathways regulated by RAS in yeast cells led to the discovery of properties that were often found interchangeable with RAS proto-oncogenes in human pathways, and vice versa. In this work, we performed an updated critical literature review on human and yeast RAS pathways, specifically highlighting the similarities and differences between them. Moreover, we emphasized the contribution of studying yeast RAS pathways for the understanding of human RAS and how this model organism can contribute to unveil the roles of RAS oncoproteins in the regulation of mechanisms important in the tumorigenic process, like autophagy. PMID:29463063

  2. [Genetic system for maintaining the mitochondrial human genome in yeast Yarrowia lipolytica].

    Science.gov (United States)

    Isakova, E P; Deryabina, Yu I; Velyakova, A V; Biryukova, J K; Teplova, V V; Shevelev, A B

    2016-01-01

    For the first time, the possibility of maintaining an intact human mitochondrial genome in a heterologous system in the mitochondria of yeast Yarrowia lipolytica is shown. A method for introducing directional changes into the structure of the mitochondrial human genome replicating in Y. lipolytica by an artificially induced ability of yeast mitochondria for homologous recombination is proposed. A method of introducing and using phenotypic selection markers for the presence or absence of defects in genes tRNA-Lys and tRNA-Leu of the mitochondrial genome is developed. The proposed system can be used to correct harmful mutations of the human mitochondrial genome associated with mitochondrial diseases and for preparative amplification of intact mitochondrial DNA with an adjusted sequence in yeast cells. The applicability of the new system for the correction of mutations in the genes of Lys- and Leu-specific tRNAs of the human mitochondrial genome associated with serious and widespread human mitochondrial diseases such as myoclonic epilepsy with lactic acidosis (MELAS) and myoclonic epilepsy with ragged-red fibers (MERRF) is shown.

  3. Network Thermodynamic Curation of Human and Yeast Genome-Scale Metabolic Models

    Science.gov (United States)

    Martínez, Verónica S.; Quek, Lake-Ee; Nielsen, Lars K.

    2014-01-01

    Genome-scale models are used for an ever-widening range of applications. Although there has been much focus on specifying the stoichiometric matrix, the predictive power of genome-scale models equally depends on reaction directions. Two-thirds of reactions in the two eukaryotic reconstructions Homo sapiens Recon 1 and Yeast 5 are specified as irreversible. However, these specifications are mainly based on biochemical textbooks or on their similarity to other organisms and are rarely underpinned by detailed thermodynamic analysis. In this study, a to our knowledge new workflow combining network-embedded thermodynamic and flux variability analysis was used to evaluate existing irreversibility constraints in Recon 1 and Yeast 5 and to identify new ones. A total of 27 and 16 new irreversible reactions were identified in Recon 1 and Yeast 5, respectively, whereas only four reactions were found with directions incorrectly specified against thermodynamics (three in Yeast 5 and one in Recon 1). The workflow further identified for both models several isolated internal loops that require further curation. The framework also highlighted the need for substrate channeling (in human) and ATP hydrolysis (in yeast) for the essential reaction catalyzed by phosphoribosylaminoimidazole carboxylase in purine metabolism. Finally, the framework highlighted differences in proline metabolism between yeast (cytosolic anabolism and mitochondrial catabolism) and humans (exclusively mitochondrial metabolism). We conclude that network-embedded thermodynamics facilitates the specification and validation of irreversibility constraints in compartmentalized metabolic models, at the same time providing further insight into network properties. PMID:25028891

  4. Cell wall trapping of autocrine peptides for human G-protein-coupled receptors on the yeast cell surface.

    Directory of Open Access Journals (Sweden)

    Jun Ishii

    Full Text Available G-protein-coupled receptors (GPCRs regulate a wide variety of physiological processes and are important pharmaceutical targets for drug discovery. Here, we describe a unique concept based on yeast cell-surface display technology to selectively track eligible peptides with agonistic activity for human GPCRs (Cell Wall Trapping of Autocrine Peptides (CWTrAP strategy. In our strategy, individual recombinant yeast cells are able to report autocrine-positive activity for human GPCRs by expressing a candidate peptide fused to an anchoring motif. Following expression and activation, yeast cells trap autocrine peptides onto their cell walls. Because captured peptides are incapable of diffusion, they have no impact on surrounding yeast cells that express the target human GPCR and non-signaling peptides. Therefore, individual yeast cells can assemble the autonomous signaling complex and allow single-cell screening of a yeast population. Our strategy may be applied to identify eligible peptides with agonistic activity for target human GPCRs.

  5. Inducibility of error-prone DNA repair in yeast

    International Nuclear Information System (INIS)

    Siede, W.; Eckardt, F.

    1984-01-01

    Whereas some experimental evidence suggests that mutagenesis in yeast after treatment with DNA-damaging agents involves inducible functions, a general-acting error-prone repair activity analogous to the SOS system of Escherichia coli has not yet been demonstrated. The current literature on the problem of inducibility of mutagenic repair in yeast is reviewed with emphasis on the differences in the experimental procedures applied. (orig.)

  6. Network thermodynamic curation of human and yeast genome-scale metabolic models.

    Science.gov (United States)

    Martínez, Verónica S; Quek, Lake-Ee; Nielsen, Lars K

    2014-07-15

    Genome-scale models are used for an ever-widening range of applications. Although there has been much focus on specifying the stoichiometric matrix, the predictive power of genome-scale models equally depends on reaction directions. Two-thirds of reactions in the two eukaryotic reconstructions Homo sapiens Recon 1 and Yeast 5 are specified as irreversible. However, these specifications are mainly based on biochemical textbooks or on their similarity to other organisms and are rarely underpinned by detailed thermodynamic analysis. In this study, a to our knowledge new workflow combining network-embedded thermodynamic and flux variability analysis was used to evaluate existing irreversibility constraints in Recon 1 and Yeast 5 and to identify new ones. A total of 27 and 16 new irreversible reactions were identified in Recon 1 and Yeast 5, respectively, whereas only four reactions were found with directions incorrectly specified against thermodynamics (three in Yeast 5 and one in Recon 1). The workflow further identified for both models several isolated internal loops that require further curation. The framework also highlighted the need for substrate channeling (in human) and ATP hydrolysis (in yeast) for the essential reaction catalyzed by phosphoribosylaminoimidazole carboxylase in purine metabolism. Finally, the framework highlighted differences in proline metabolism between yeast (cytosolic anabolism and mitochondrial catabolism) and humans (exclusively mitochondrial metabolism). We conclude that network-embedded thermodynamics facilitates the specification and validation of irreversibility constraints in compartmentalized metabolic models, at the same time providing further insight into network properties. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  7. An improved yeast transformation method for the generation of very large human antibody libraries.

    Science.gov (United States)

    Benatuil, Lorenzo; Perez, Jennifer M; Belk, Jonathan; Hsieh, Chung-Ming

    2010-04-01

    Antibody library selection by yeast display technology is an efficient and highly sensitive method to identify binders to target antigens. This powerful selection tool, however, is often hampered by the typically modest size of yeast libraries (approximately 10(7)) due to the limited yeast transformation efficiency, and the full potential of the yeast display technology for antibody discovery and engineering can only be realized if it can be coupled with a mean to generate very large yeast libraries. We describe here a yeast transformation method by electroporation that allows for the efficient generation of large antibody libraries up to 10(10) in size. Multiple components and conditions including CaCl(2), MgCl(2), sucrose, sorbitol, lithium acetate, dithiothreitol, electroporation voltage, DNA input and cell volume have been tested to identify the best combination. By applying this developed protocol, we have constructed a 1.4 x 10(10) human spleen antibody library essentially in 1 day with a transformation efficiency of 1-1.5 x 10(8) transformants/microg vector DNA. Taken together, we have developed a highly efficient yeast transformation method that enables the generation of very large and productive human antibody libraries for antibody discovery, and we are now routinely making 10(9) libraries in a day for antibody engineering purposes.

  8. Computational determination of the effects of virulent Escherichia coli and salmonella bacteriophages on human gut.

    Science.gov (United States)

    Mostafa, Marwa Mostafa; Nassef, Mohammad; Badr, Amr

    2016-10-01

    Salmonella and Escherichia coli are different types of bacteria that cause food poisoning in humans. In the elderly, infants and people with chronic conditions, it is very dangerous if Salmonella or E. coli gets into the bloodstream and then they must be treated by phage therapy. Treating Salmonella and E. coli by phage therapy affects the gut flora. This research paper presents a system for detecting the effects of virulent E. coli and Salmonella bacteriophages on human gut. A method based on Domain-Domain Interactions (DDIs) model is implemented in the proposed system to determine the interactions between the proteins of human gut bacteria and the proteins of bacteriophages that infect virulent E. coli and Salmonella. The system helps gastroenterologists to realize the effect of injecting bacteriophages that infect virulent E. coli and Salmonella on the human gut. By testing the system over Enterobacteria phage 933W, Enterobacteria phage VT2-Sa and Enterobacteria phage P22, it resulted in four interactions between the proteins of the bacteriophages that infect E. coli O157:H7, E. coli O104:H4 and Salmonella typhimurium and the proteins of human gut bacterium strains. Several effects were detected such as: antibacterial activity against a number of bacterial species in human gut, regulation of cellular differentiation and organogenesis during gut, lung, and heart development, ammonia assimilation in bacteria, yeasts, and plants, energizing defense system and its function in the detoxification of lipopolysaccharide, and in the prevention of bacterial translocation in human gut. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Phospholipids chiral at phosphorus. Steric course of the reactions catalyzed by phosphatidylserine synthase from Escherichia coli and yeast

    International Nuclear Information System (INIS)

    Raetz, C.R.H.; Carman, G.M.; Dowhan, W.; Jiang, R.T.; Waszkuc, W.; Loffredo, W.; Tsai, M.D.

    1987-01-01

    The steric courses of the reactions catalyzed by phosphatidylserine (PS) synthase from Escherichia coli and yeast were elucidated by the following procedure. R/sub P/ and S/sub P/ isomers of 1,2-dipalmitoyl-sn-glycero-3-[ 17 O, 18 O]phosphoethanolamine ([ 17 O, 18 O]DPPE) were synthesized and converted to (R/sub P/)- and (S/sub P/)-1,2-dipalmitoyl-sn-glycero-3-[ 16 O, 17 O, 18 O]DPPA), respectively, by incubating with phospholipase D. Condensation of [ 16 O, 17 O, 18 O]DPPA with cytidine 5'-monophosphomorpholidate in pyridine gave the desired substrate for PS synthase, [ 17 O, 18 O]cytidine 5'-diphospho-1,2-dipalmitoyl-sn-glycerol ([ 17 O, 18 O]CDP-DPG), as a mixture of several isotopic and configurational isomers. Incubation of [ 17 O, 18 O]CDP-DPG), as a mixture of several isotopic and configurational isomers. Incubation of [ 17 O, 18 O] CDP-DPG with a mixture of L-serine, PS synthase and PS decarboxylase gave [ 17 O, 18 O]DPPE. The configuration and isotopic enrichments of the starting [ 17 O, 18 O]DPPE and the product were analyzed by 31 P NMR following trimethylsilylation of the DPPE. The results indicate that the reaction of E. coli PS synthase proceeds with retention of configuration at phosphorus, which suggests a two-step mechanism involving a phosphatidyl-enzyme intermediate, while the yeast PS synthase catalyzes the reaction with inversion of configuration, which suggests a single-displacement mechanism. Such results lend strong support to the ping-pong mechanism proposed for the E. coli enzyme and the sequential Bi-Bi mechanism proposed for the yeast enzyme, both based on previous isotopic exchange experiments

  10. Human gut Bacteroidetes can utilize yeast mannan through a selfish mechanism

    Science.gov (United States)

    The architecture of the human distal gut microbiota (microbiota) is sculpted by the complex carbohydrates delivered in the diet. Yeasts, which are among the earliest domesticated microorganisms and have been a component of the human diet for at least 7000 years, possess an elaborate cell wall alpha-...

  11. Bacterial toxin-antitoxin gene system as containment control in yeast cells

    DEFF Research Database (Denmark)

    Kristoffersen, P.; Jensen, G. B.; Gerdes, K.

    2000-01-01

    The potential of a bacterial toxin-antitoxin gene system for use in containment control in eukaryotes was explored. The Escherichia coli relE and relB genes were expressed in the yeast Saccharomyces cerevisiae, Expression of the relE gene was highly toxic to yeast cells. However, expression...... fermentation processes in which the escape of genetically modified cells would be considered highly risky....

  12. Triosephosphate isomerase: energetics of the reaction catalyzed by the yeast enzyme expressed in Escherichia coli

    International Nuclear Information System (INIS)

    Nickbarg, E.B.; Knowles, J.R.

    1988-01-01

    Triosephosphate isomerase from bakers' yeast, expressed in Escherichia coli strain DF502(p12), has been purified to homogeneity. The kinetics of the reaction in each direction have been determined at pH 7.5 and 30 degrees C. Deuterium substitution at the C-2 position of substrate (R)-glyceraldehyde phosphate and at the 1-pro-R position of substrate dihydroxyacetone phosphate results in kinetic isotope effects on kcat of 1.6 and 3.4, respectively. The extent of transfer of tritium from [1(R)- 3 H]dihydroxyacetone phosphate to product (R)-glyceraldehyde phosphate during the catalyzed reaction is only 3% after 66% conversion to product, indicating that the enzymic base that mediates proton transfer is in rapid exchange with solvent protons. When the isomerase-catalyzed reaction is run in tritiated water in each direction, radioactivity is incorporated both into the remaining substrate and into the product. In the exchange-conversion experiment with dihydroxyacetone phosphate as substrate, the specific radioactivity of remaining dihydroxyacetone phosphate rises as a function of the extent of reaction with a slope of about 0.3, while the specific radioactivity of the products is 54% that of the solvent. In the reverse direction with (R)-glyceraldehyde phosphate as substrate, the specific radioactivity of the product formed is only 11% that of the solvent, while the radioactivity incorporated into the remaining substrate (R)-glyceraldehyde phosphate also rises as a function of the extent of reaction with a slope of 0.3. These results have been analyzed according to the protocol described earlier to yield the free energy profile of the reaction catalyzed by the yeast isomerase

  13. Over-expression in Escherichia coli and characterization of two recombinant isoforms of human FAD synthetase

    International Nuclear Information System (INIS)

    Brizio, Carmen; Galluccio, Michele; Wait, Robin; Torchetti, Enza Maria; Bafunno, Valeria; Accardi, Rosita; Gianazza, Elisabetta; Indiveri, Cesare; Barile, Maria

    2006-01-01

    FAD synthetase (FADS) (EC 2.7.7.2) is a key enzyme in the metabolic pathway that converts riboflavin into the redox cofactor FAD. Two hypothetical human FADSs, which are the products of FLAD1 gene, were over-expressed in Escherichia coli and identified by ESI-MS/MS. Isoform 1 was over-expressed as a T7-tagged protein which had a molecular mass of 63 kDa on SDS-PAGE. Isoform 2 was over-expressed as a 6-His-tagged fusion protein, carrying an extra 84 amino acids at the N-terminal with an apparent molecular mass of 60 kDa on SDS-PAGE. It was purified near to homogeneity from the soluble cell fraction by one-step affinity chromatography. Both isoforms possessed FADS activity and had a strict requirement for MgCl 2 , as demonstrated using both spectrophotometric and chromatographic methods. The purified recombinant isoform 2 showed a specific activity of 6.8 ± 1.3 nmol of FAD synthesized/min/mg protein and exhibited a K M value for FMN of 1.5 ± 0.3 μM. This is First report on characterization of human FADS, and First cloning and over-expression of FADS from an organism higher than yeast

  14. Functional expression of parasite drug targets and their human orthologs in yeast.

    Directory of Open Access Journals (Sweden)

    Elizabeth Bilsland

    2011-10-01

    Full Text Available The exacting nutritional requirements and complicated life cycles of parasites mean that they are not always amenable to high-throughput drug screening using automated procedures. Therefore, we have engineered the yeast Saccharomyces cerevisiae to act as a surrogate for expressing anti-parasitic targets from a range of biomedically important pathogens, to facilitate the rapid identification of new therapeutic agents.Using pyrimethamine/dihydrofolate reductase (DHFR as a model parasite drug/drug target system, we explore the potential of engineered yeast strains (expressing DHFR enzymes from Plasmodium falciparum, P. vivax, Homo sapiens, Schistosoma mansoni, Leishmania major, Trypanosoma brucei and T. cruzi to exhibit appropriate differential sensitivity to pyrimethamine. Here, we demonstrate that yeast strains (lacking the major drug efflux pump, Pdr5p expressing yeast ((ScDFR1, human ((HsDHFR, Schistosoma ((SmDHFR, and Trypanosoma ((TbDHFR and (TcDHFR DHFRs are insensitive to pyrimethamine treatment, whereas yeast strains producing Plasmodium ((PfDHFR and (PvDHFR DHFRs are hypersensitive. Reassuringly, yeast strains expressing field-verified, drug-resistant mutants of P. falciparum DHFR ((Pfdhfr(51I,59R,108N are completely insensitive to pyrimethamine, further validating our approach to drug screening. We further show the versatility of the approach by replacing yeast essential genes with other potential drug targets, namely phosphoglycerate kinases (PGKs and N-myristoyl transferases (NMTs.We have generated a number of yeast strains that can be successfully harnessed for the rapid and selective identification of urgently needed anti-parasitic agents.

  15. One-pot green synthesis of carbon dots by using Saccharum officinarum juice for fluorescent imaging of bacteria (Escherichia coli) and yeast (Saccharomyces cerevisiae) cells

    Energy Technology Data Exchange (ETDEWEB)

    Mehta, Vaibhavkumar N. [Applied Chemistry Department, S. V. National Institute of Technology, Surat, 395 007 (India); Jha, Sanjay [Gujarat Agricultural Biotechnology Institute, Navsari Agricultural University, Surat, 395007 (India); Kailasa, Suresh Kumar, E-mail: sureshkumarchem@gmail.com [Applied Chemistry Department, S. V. National Institute of Technology, Surat, 395 007 (India)

    2014-05-01

    We are reporting highly economical plant-based hydrothermal method for one-pot green synthesis of water-dispersible fluorescent carbon dots (CDs) by using Saccharum officinarum juice as precursor. The synthesized CDs were characterized by UV-visible, fluorescence, Fourier transform infrared (FT-IR), dynamic light scattering (DLS), high-resolution transmission electron microscopic (HR-TEM), and laser scanning confocal microscopic techniques. The CDs are well dispersed in water with an average size of ∼ 3 nm and showed bright blue fluorescence under UV-light (λ{sub ex} = 365 nm). These CDs acted as excellent fluorescent probes in cellular imaging of bacteria (Escherichia coli) and yeast (Saccharomyces cerevisiae). - Highlights: • One-pot green synthesis was used for fluorescent CDs. • FT-IR, DLS, and TEM were used for the characterization of CDs. • The CDs are well dispersed in water with an average size of ∼ 3 nm. • The CDs acted as fluorescent probes for imaging of bacteria and yeast cells.

  16. Effects of recombinant human collagen VI from Escherichia coli on ...

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... In this study, we reported the cloning and over expression of a gene coding for human collagen peptide. (CP6) in Escherichia coli and investigated the protective effects of CP6 on UVA-irradiated human skin fibroblasts cells. The collagen peptide (CP6) was highly soluble and the expression level was.

  17. Beneficial effects of probiotic and food borne yeasts on human health

    DEFF Research Database (Denmark)

    Moslehi Jenabian, Saloomeh; Pedersen, Line Lindegaard; Jespersen, Lene

    2010-01-01

    Besides being important in the fermentation of foods and beverages, yeasts have shown numerous beneficial effects on human health. Among these, propiotic effects are the most well known health effects including prevention and treatment of intestinal diseases and immunomodulatory effects. Other...

  18. Filament formation of the Escherichia coli actin-related protein, MreB, in fission yeast.

    Science.gov (United States)

    Srinivasan, Ramanujam; Mishra, Mithilesh; Murata-Hori, Maki; Balasubramanian, Mohan K

    2007-02-06

    Proteins structurally related to eukaryotic actins have recently been identified in several prokaryotic organisms. These actin-like proteins (MreB and ParM) and the deviant Walker A ATPase (SopA) play a key role in DNA segregation and assemble into polymers in vitro and in vivo. MreB also plays a role in cellular morphogenesis. Whereas the dynamic properties of eukaryotic actins have been extensively characterized, those of bacterial actins are only beginning to emerge. We have established the fission yeast Schizosaccharomyces pombe as a cellular model for the functional analysis of the Escherichia coli actin-related protein MreB. We show that MreB organizes into linear bundles that grow in a symmetrically bidirectional manner at 0.46 +/- 0.03 microm/min, with new monomers and/or oligomers being added along the entire length of the bundle. Organization of linear arrays was dependent on the ATPase activity of MreB, and their alignment along the cellular long axis was achieved by sliding along the cortex of the cylindrical part of the cell. The cell ends appeared to provide a physical barrier for bundle elongation. These experiments provide new insights into the mechanism of assembly and organization of the bacterial actin cytoskeleton.

  19. Human COQ9 Rescues a coq9 Yeast Mutant by Enhancing Coenzyme Q Biosynthesis from 4-Hydroxybenzoic Acid and Stabilizing the CoQ-Synthome

    Directory of Open Access Journals (Sweden)

    Cuiwen H. He

    2017-07-01

    Full Text Available Coq9 is required for the stability of a mitochondrial multi-subunit complex, termed the CoQ-synthome, and the deamination step of Q intermediates that derive from para-aminobenzoic acid (pABA in yeast. In human, mutations in the COQ9 gene cause neonatal-onset primary Q10 deficiency. In this study, we determined whether expression of human COQ9 could complement yeast coq9 point or null mutants. We found that expression of human COQ9 rescues the growth of the temperature-sensitive yeast mutant, coq9-ts19, on a non-fermentable carbon source and increases the content of Q6, by enhancing Q biosynthesis from 4-hydroxybenzoic acid (4HB. To study the mechanism for the rescue by human COQ9, we determined the steady-state levels of yeast Coq polypeptides in the mitochondria of the temperature-sensitive yeast coq9 mutant expressing human COQ9. We show that the expression of human COQ9 significantly increased steady-state levels of yeast Coq4, Coq6, Coq7, and Coq9 at permissive temperature. Human COQ9 polypeptide levels persisted at non-permissive temperature. A small amount of the human COQ9 co-purified with tagged Coq6, Coq6-CNAP, indicating that human COQ9 interacts with the yeast Q-biosynthetic complex. These findings suggest that human COQ9 rescues the yeast coq9 temperature-sensitive mutant by stabilizing the CoQ-synthome and increasing Q biosynthesis from 4HB. This finding provides a powerful approach to studying the function of human COQ9 using yeast as a model.

  20. NetPhosYeast: prediction of protein phosphorylation sites in yeast

    DEFF Research Database (Denmark)

    Ingrell, C.R.; Miller, Martin Lee; Jensen, O.N.

    2007-01-01

    sites compared to those in humans, suggesting the need for an yeast-specific phosphorylation site predictor. NetPhosYeast achieves a correlation coefficient close to 0.75 with a sensitivity of 0.84 and specificity of 0.90 and outperforms existing predictors in the identification of phosphorylation sites...

  1. The Protein Interaction Network of Bacteriophage Lambda with Its Host, Escherichia coli

    Science.gov (United States)

    Blasche, Sonja; Wuchty, Stefan; Rajagopala, Seesandra V.

    2013-01-01

    Although most of the 73 open reading frames (ORFs) in bacteriophage λ have been investigated intensively, the function of many genes in host-phage interactions remains poorly understood. Using yeast two-hybrid screens of all lambda ORFs for interactions with its host Escherichia coli, we determined a raw data set of 631 host-phage interactions resulting in a set of 62 high-confidence interactions after multiple rounds of retesting. These links suggest novel regulatory interactions between the E. coli transcriptional network and lambda proteins. Targeted host proteins and genes required for lambda infection are enriched among highly connected proteins, suggesting that bacteriophages resemble interaction patterns of human viruses. Lambda tail proteins interact with both bacterial fimbrial proteins and E. coli proteins homologous to other phage proteins. Lambda appears to dramatically differ from other phages, such as T7, because of its unusually large number of modified and processed proteins, which reduces the number of host-virus interactions detectable by yeast two-hybrid screens. PMID:24049175

  2. Small heat shock proteins potentiate amyloid dissolution by protein disaggregases from yeast and humans.

    Directory of Open Access Journals (Sweden)

    Martin L Duennwald

    Full Text Available How small heat shock proteins (sHsps might empower proteostasis networks to control beneficial prions or disassemble pathological amyloid is unknown. Here, we establish that yeast sHsps, Hsp26 and Hsp42, inhibit prionogenesis by the [PSI+] prion protein, Sup35, via distinct and synergistic mechanisms. Hsp42 prevents conformational rearrangements within molten oligomers that enable de novo prionogenesis and collaborates with Hsp70 to attenuate self-templating. By contrast, Hsp26 inhibits self-templating upon binding assembled prions. sHsp binding destabilizes Sup35 prions and promotes their disaggregation by Hsp104, Hsp70, and Hsp40. In yeast, Hsp26 or Hsp42 overexpression prevents [PSI+] induction, cures [PSI+], and potentiates [PSI+]-curing by Hsp104 overexpression. In vitro, sHsps enhance Hsp104-catalyzed disaggregation of pathological amyloid forms of α-synuclein and polyglutamine. Unexpectedly, in the absence of Hsp104, sHsps promote an unprecedented, gradual depolymerization of Sup35 prions by Hsp110, Hsp70, and Hsp40. This unanticipated amyloid-depolymerase activity is conserved from yeast to humans, which lack Hsp104 orthologues. A human sHsp, HspB5, stimulates depolymerization of α-synuclein amyloid by human Hsp110, Hsp70, and Hsp40. Thus, we elucidate a heretofore-unrecognized human amyloid-depolymerase system that could have applications in various neurodegenerative disorders.

  3. Introduction of the yeast DNA repair gene PHR1 into normal and xeroderma pigmentosum human cells

    International Nuclear Information System (INIS)

    Whyte, D.B.

    1988-01-01

    The goal of the work described herein is to determine how UV light kills and mutates human cells. Specifically, the hypothesis to be tested states that the major cause of cell death is the cyclobutane dimer. The yeast (S. cerevisiae) enzyme photolyase provides an elegant means of dissecting the biological effects of the two lesions. Photolyase, the product of the PHR1 gene, catalyzes the visible light-dependent reversal of cyclobutane pyrimidine dimers. Introducing the gene for photolyase into human cells, which do not have a functional photoreactivation mechanism, should allow specific repair of cyclobutane pyrimidine dimers. To express the yeast DNA repair gene in human cells, the yeast PHR1 coding sequence was cloned into the mammalian expression vector pRSV4NEO-I. The resulting plasmid, pRSVPHR1, contains the coding sequence of the yeast gene, under control of transcription signals recognized by mammalian cells, and the dominant selectable gene neo. pRSVPHR1 was introduced into normal and XP SV40-transformed fibroblasts by the calcium phosphate coprecipitation technique, and G418-resistant clones were isolated. The level of PHR1 expression was determined by cytoplasmic RNA dot blots. Two clones, XP-3B and GM-20A, had high levels of expression

  4. Methylation of mercuric chloride by human intestinal bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Rowland, I R; Grasso, P; Davies, M J

    1975-01-01

    There is now evidence that ingested mercuric chloride (HgCl/sub 2/) may be methylated, in vivo, in the rat intestine and, in vitro, by human feces. However, one cannot infer from these experiments that the microbial flora of the intestine is responsible for the methylation reaction, since the gut contents contain several sources of metabolic activity other than bacteria. Data are presented on the ability of pure cultures of bacteria and yeasts, isolated from human feces, to convert HgCl/sub 2/ to methylmercury. Strains of Escherichia coli, streptococci, staphylococci, bacteriodes and bifidobacteria were inoculated into a medium containing 0.1 M potassium phosphate buffer, pH 7.0, Bacto-tryptone, yeast extract and D-glucose, each at 0.5% (w/v). Results indicate that most strains of staphylococci, streptococci, yeasts and E. coli isolated from human feces, could synthesize methylmercury compounds. In contrast, few strains of obligate anaerobes could do so. Up to 6 ng methylmercury/ml were formed in 44 h from 2 ..mu..g mercuric chloride.

  5. Screening for proteins interacting with MCM7 in human lung cancer library using yeast two hybrid system

    Directory of Open Access Journals (Sweden)

    Yuchen HAN

    2008-08-01

    Full Text Available Background and objective MCM7 is a subunit of the MCM complex that plays a key role in DNA replication initiation. But little is known about its interaction proteins. In this study yeast two hybrid screening was used to identify the MCM7 interacting proteins. Methods Yeast expression vector containing human full length MCM7-pGBKT7 plasmid was constructed, and with a library of cDNAs from human lung cancer-pACT2 plasmid was transformed into yeast strain AH109, and was electively grew in X-a-gal auxotrophy medium SD/-Trp-Leu-His-Ade, and the blue colonies were picked up, the plasmid of the yeast colonies was extracted , and transformed into E. Coli to extract DNA and performed sequence analysis. Results Eleven proteins were identified which could specifically interact with MCM7 proteins, among these five were cytoskeleton proteins, six were enzymes, kinases and related receptors. Conclusion The investigation provides functional clues for further exploration of MCM7 gene.

  6. High-level secretion of native recombinant human calreticulin in yeast

    DEFF Research Database (Denmark)

    Čiplys, Evaldas; Žitkus, Eimantas; Gold, Leslie I.

    2015-01-01

    , Saccharomyces cerevisiae and Pichia pastoris. RESULTS: Expression of a full-length human CRT precursor including its native signal sequence resulted in high-level secretion of mature recombinant protein into the culture medium by both S. cerevisiae and P. pastoris. To ensure the structural and functional...... by non-denaturing PAGE. Moreover, limited trypsin digestion yielded identical fragment patterns of calcium-binding recombinant and native CRT suggesting that the yeast-derived CRT was correctly folded. Furthermore, both native and recombinant CRT induced cellular proliferation (MTS assay) and migration...... recombinant CRT protein with yields reaching 75 % of total secreted protein and with production levels of 60 and 200 mg/l from S. cerevisiae and P. pastoris, respectively. Finally, cultivation of P. pastoris in a bioreactor yielded CRT secretion titer to exceed 1.5 g/l of culture medium. CONCLUSIONS: Yeasts...

  7. Studies of the expression of human poly(ADP-ribose) polymerase-1 in Saccharomyces cerevisiae and identification of PARP-1 substrates by yeast proteome microarray screening.

    Science.gov (United States)

    Tao, Zhihua; Gao, Peng; Liu, Hung-Wen

    2009-12-15

    Poly(ADP-ribosyl)ation of various nuclear proteins catalyzed by a family of NAD(+)-dependent enzymes, poly(ADP-ribose) polymerases (PARPs), is an important posttranslational modification reaction. PARP activity has been demonstrated in all types of eukaryotic cells with the exception of yeast, in which the expression of human PARP-1 was shown to lead to retarded cell growth. We investigated the yeast growth inhibition caused by human PARP-1 expression in Saccharomyces cerevisiae. Flow cytometry analysis reveals that PARP-1-expressing yeast cells accumulate in the G(2)/M stage of the cell cycle. Confocal microscopy analysis shows that human PARP-1 is distributed throughout the nucleus of yeast cells but is enriched in the nucleolus. Utilizing yeast proteome microarray screening, we identified 33 putative PARP-1 substrates, six of which are known to be involved in ribosome biogenesis. The poly(ADP-ribosyl)ation of three of these yeast proteins, together with two human homologues, was confirmed by an in vitro PARP-1 assay. Finally, a polysome profile analysis using sucrose gradient ultracentrifugation demonstrated that the ribosome levels in yeast cells expressing PARP-1 are lower than those in control yeast cells. Overall, our data suggest that human PARP-1 may affect ribosome biogenesis by modifying certain nucleolar proteins in yeast. The artificial PARP-1 pathway in yeast may be used as a simple platform to identify substrates and verify function of this important enzyme.

  8. Emergence of Antimicrobial-Resistant Escherichia coli of Animal Origin Spreading in Humans

    Science.gov (United States)

    Skurnik, David; Clermont, Olivier; Guillard, Thomas; Launay, Adrien; Danilchanka, Olga; Pons, Stéphanie; Diancourt, Laure; Lebreton, François; Kadlec, Kristina; Roux, Damien; Jiang, Deming; Dion, Sara; Aschard, Hugues; Denamur, Maurice; Cywes-Bentley, Colette; Schwarz, Stefan; Tenaillon, Olivier; Andremont, Antoine; Picard, Bertrand; Mekalanos, John; Brisse, Sylvain; Denamur, Erick

    2016-01-01

    In the context of the great concern about the impact of human activities on the environment, we studied 403 commensal Escherichia coli/Escherichia clade strains isolated from several animal and human populations that have variable contacts to one another. Multilocus sequence typing (MLST) showed a decrease of diversity 1) in strains isolated from animals that had an increasing contact with humans and 2) in all strains that had increased antimicrobial resistance. A specific B1 phylogroup clonal complex (CC87, Institut Pasteur schema nomenclature) of animal origin was identified and characterized as being responsible for the increased antimicrobial resistance prevalence observed in strains from the environments with a high human-mediated antimicrobial pressure. CC87 strains have a high capacity of acquiring and disseminating resistance genes with specific metabolic and genetic determinants as demonstrated by high-throughput sequencing and phenotyping. They are good mouse gut colonizers but are not virulent. Our data confirm the predominant role of human activities in the emergence of antimicrobial resistance in the environmental bacterial strains and unveil a particular E. coli clonal complex of animal origin capable of spreading antimicrobial resistance to other members of microbial communities. PMID:26613786

  9. Structural analysis of the core COMPASS family of histone H3K4 methylases from yeast to human.

    Science.gov (United States)

    Takahashi, Yoh-hei; Westfield, Gerwin H; Oleskie, Austin N; Trievel, Raymond C; Shilatifard, Ali; Skiniotis, Georgios

    2011-12-20

    Histone H3 lysine 4 (H3K4) methylation is catalyzed by the highly evolutionarily conserved multiprotein complex known as Set1/COMPASS or MLL/COMPASS-like complexes from yeast to human, respectively. Here we have reconstituted fully functional yeast Set1/COMPASS and human MLL/COMPASS-like complex in vitro and have identified the minimum subunit composition required for histone H3K4 methylation. These subunits include the methyltransferase C-terminal SET domain of Set1/MLL, Cps60/Ash2L, Cps50/RbBP5, Cps30/WDR5, and Cps25/Dpy30, which are all common components of the COMPASS family from yeast to human. Three-dimensional (3D) cryo-EM reconstructions of the core yeast complex, combined with immunolabeling and two-dimensional (2D) EM analysis of the individual subcomplexes reveal a Y-shaped architecture with Cps50 and Cps30 localizing on the top two adjacent lobes and Cps60-Cps25 forming the base at the bottom. EM analysis of the human complex reveals a striking similarity to its yeast counterpart, suggesting a common subunit organization. The SET domain of Set1 is located at the juncture of Cps50, Cps30, and the Cps60-Cps25 module, lining the walls of a central channel that may act as the platform for catalysis and regulative processing of various degrees of H3K4 methylation. This structural arrangement suggested that COMPASS family members function as exo-methylases, which we have confirmed by in vitro and in vivo studies.

  10. 21 CFR 172.325 - Bakers yeast protein.

    Science.gov (United States)

    2010-04-01

    ... and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) FOOD ADDITIVES PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Special Dietary and Nutritional Additives § 172.325 Bakers yeast protein. Bakers yeast protein may be...

  11. Antimicrobial activity of yeasts against some pathogenic bacteria

    Directory of Open Access Journals (Sweden)

    Gamal Younis

    2017-08-01

    Full Text Available Aim: This study was designed to isolate and identify yeast species from milk and meat products, and to test their antimicrobial activity against some bacterial species. Materials and Methods: A total of 160 milk and meat products samples were collected from random sellers and super markets in New Damietta city, Damietta, Egypt. Samples were subjected to yeast isolation procedures and tested for its antimicrobial activity against Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. In addition, all yeast species isolates were subjected to polymerase chain reaction (PCR for detection of khs (kievitone hydratase and pelA (pectate degrading enzyme genes. Results: The recovery rate of yeasts from sausage was 20% (2/10 followed by kareish cheese, processed cheese, and butter 10% (1/10 each as well as raw milk 9% (9/100, and fruit yoghurt 30% (6/20. Different yeast species were recovered, namely, Candida kefyr (5 isolates, Saccharomyces cerevisiae (4 isolates, Candida intermedia (3 isolates, Candida tropicalis (2 isolates, Candida lusitaniae (2 isolates, and Candida krusei (1 isolate. khs gene was detected in all S. cerevisiae isolates, however, pelA gene was not detected in all identified yeast species. Antimicrobial activity of recovered yeasts against the selected bacterial species showed high activity with C. intermedia against S. aureus and E. coli, C. kefyr against E. coli, and C. lusitaniae against S. aureus. Moderate activities were obtained with C. tropicalis, C. lusitaniae, and S. cerevisiae against E. coli; meanwhile, all the tested yeasts revealed a very low antimicrobial activity against P. aeruginosa. Conclusion: The obtained results confirmed that some kinds of yeasts have the ability to produce antimicrobial compounds that could inhibit some pathogenic and spoilage bacteria and these antimicrobial activity of yeasts enables them to be one of the novel agents in controlling spoilage of food.

  12. Heterologously expressed bacterial and human multidrug resistance proteins confer cadmium resistance to Escherichia coli

    NARCIS (Netherlands)

    Achard-Joris, M; van Saparoea, HBV; Driessen, AJM; Bourdineaud, JP; Bourdineaud, Jean-Paul

    2005-01-01

    The human MDR1 gene is induced by cadmium exposure although no resistance to this metal is observed in human cells overexpressing hMDR1. To access the role of MDR proteins in cadmium resistance, human MDR1, Lactococcus lactis lmrA, and Oenococcus oeni omrA were expressed in an Escherichia coli tolC

  13. Genetic Structure and Antimicrobial Resistance of Escherichia coli and Cryptic Clades in Birds with Diverse Human Associations.

    Science.gov (United States)

    Blyton, Michaela D J; Pi, Hongfei; Vangchhia, Belinda; Abraham, Sam; Trott, Darren J; Johnson, James R; Gordon, David M

    2015-08-01

    The manner and extent to which birds associate with humans may influence the genetic attributes and antimicrobial resistance of their commensal Escherichia communities through strain transmission and altered selection pressures. In this study, we determined whether the distribution of the different Escherichia coli phylogenetic groups and cryptic clades, the occurrence of 49 virulence associated genes, and/or the prevalence of resistance to 12 antimicrobials differed between four groups of birds from Australia with contrasting types of human association. We found that birds sampled in suburban and wilderness areas had similar Escherichia communities. The Escherichia communities of backyard domestic poultry were phylogenetically distinct from the Escherichia communities sourced from all other birds, with a large proportion (46%) of poultry strains belonging to phylogenetic group A and a significant minority (17%) belonging to the cryptic clades. Wild birds sampled from veterinary and wildlife rehabilitation centers (in-care birds) carried Escherichia isolates that possessed particular virulence-associated genes more often than Escherichia isolates from birds sampled in suburban and wilderness areas. The Escherichia isolates from both the backyard poultry and in-care birds were more likely to be multidrug resistant than the Escherichia isolates from wild birds. We also detected a multidrug-resistant E. coli strain circulating in a wildlife rehabilitation center, reinforcing the importance of adequate hygiene practices when handling and caring for wildlife. We suggest that the relatively high frequency of antimicrobial resistance in the in-care birds and backyard poultry is due primarily to the use of antimicrobials in these animals, and we recommend that the treatment protocols used for these birds be reviewed. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  14. Overexpression screens identify conserved dosage chromosome instability genes in yeast and human cancer

    Science.gov (United States)

    Duffy, Supipi; Fam, Hok Khim; Wang, Yi Kan; Styles, Erin B.; Kim, Jung-Hyun; Ang, J. Sidney; Singh, Tejomayee; Larionov, Vladimir; Shah, Sohrab P.; Andrews, Brenda; Boerkoel, Cornelius F.; Hieter, Philip

    2016-01-01

    Somatic copy number amplification and gene overexpression are common features of many cancers. To determine the role of gene overexpression on chromosome instability (CIN), we performed genome-wide screens in the budding yeast for yeast genes that cause CIN when overexpressed, a phenotype we refer to as dosage CIN (dCIN), and identified 245 dCIN genes. This catalog of genes reveals human orthologs known to be recurrently overexpressed and/or amplified in tumors. We show that two genes, TDP1, a tyrosyl-DNA-phosphdiesterase, and TAF12, an RNA polymerase II TATA-box binding factor, cause CIN when overexpressed in human cells. Rhabdomyosarcoma lines with elevated human Tdp1 levels also exhibit CIN that can be partially rescued by siRNA-mediated knockdown of TDP1. Overexpression of dCIN genes represents a genetic vulnerability that could be leveraged for selective killing of cancer cells through targeting of an unlinked synthetic dosage lethal (SDL) partner. Using SDL screens in yeast, we identified a set of genes that when deleted specifically kill cells with high levels of Tdp1. One gene was the histone deacetylase RPD3, for which there are known inhibitors. Both HT1080 cells overexpressing hTDP1 and rhabdomyosarcoma cells with elevated levels of hTdp1 were more sensitive to histone deacetylase inhibitors valproic acid (VPA) and trichostatin A (TSA), recapitulating the SDL interaction in human cells and suggesting VPA and TSA as potential therapeutic agents for tumors with elevated levels of hTdp1. The catalog of dCIN genes presented here provides a candidate list to identify genes that cause CIN when overexpressed in cancer, which can then be leveraged through SDL to selectively target tumors. PMID:27551064

  15. Expression and function of the human estrogen receptor in yeast

    International Nuclear Information System (INIS)

    White, J.H.; Metzger, D.; Chambon, P.

    1988-01-01

    Gene expression in eukaryotes is regulated at many levels. Moreover, there is increasing evidence that the basic control mechanisms of transcription initiation have been conserved across the range of eukaryotes from yeast to man. In vertebrates, the nuclear receptors, whose activity is dependent on the binding of specific ligands, stimulate transcription by interacting with specific cis-acting sequences and display all of the hallmarks of inducible enhancer factors. Alignment of their amino acid sequences indicates that they are composed of a series of conserved domains. The domain structure of the human estrogen receptor (hER) is typical of receptor proteins. Region C, containing two putative zinc fingers, comprises the DNA-binding domain responsible for specific recognition of estrogen response elements (ERE). Region E contains the hormone-binding domain and domain(s) responsible for transcription activation. A mutant of the hER, called HE15, which lacks the hormone-binding domain, binds DNA in vivo and in vitro but activates transcription only poorly in a constitutive manner in vivo in HeLa cells. A series of studies have demonstrated that the hormone- and DNA-binding domains of the nuclear receptors function independently. Chimeric proteins consisting of the DNA-binding domain of yeast GAL4 coupled to the hormone-binding domains of either the hER or glucocorticoid receptor element (GRE) will stimulate transcription in HeLa cells when bound to a UAS. Taken together, these results demonstrate that the hER and other nuclear receptors, as well as GAL4 and GCN4 proteins of yeast, consist of discrete and separable DNA-binding and transcription-activation functions. To investigate these striking parallels further, the authors have expressed the hER in the yeast Saccharomyces cerevisiae and have analyzed its hormone- and DNA-binding properties in vitro and its ability to stimulate transcription in vivo

  16. Fungal spore germination into yeast or mycelium: possible implications of dimorphism in evolution and human pathogenesis

    Science.gov (United States)

    Ghormade, Vandana; Deshpande, M. V.

    The ability of dimorphism in fungi is conventionally regarded as a reversible change between the two vegetative forms, yeast and mycelium, in response to environmental change. A zygomycetous isolate, Benjaminiella poitrasii, exhibited yeast-mycelium transition in response to the change in temperature (37-28 °C) and decrease in glucose concentration. For the first time the presence of dimorphic response during asexual and sexual spore germination is reported under the dimorphism-triggering conditions in B. poitrasii. The zygospores germinated into budding yeast when subjected to yeast-form supporting conditions. The mycelium-form favoring conditions gave rise to true mycelium. Similarly, the asexual spores displayed a dimorphic response during germination. Our observations suggest that dimorphism is an intrinsic ability present in the vegetative, asexual, and sexual forms of the fungus. As dimorphic fungi are intermediate to the unicellular yeast and the filamentous forms, understanding of the dimorphic character could be useful to trace the evolutionary relationships among taxonomically different fungi. Moreover, the implications of spore germination during the onset of pathogenesis and in drug development for human health care are discussed.

  17. Does Probiotic Yeast Act as Antigenotoxin?

    Directory of Open Access Journals (Sweden)

    Jekabs Raipulis

    2005-01-01

    Full Text Available The effect of probiotic yeast Saccharomyces boulardii on genotoxicity induced by the well-known mutagen 4-nitroquinoline-N-oxide (4-NQO, as well as antibacterial (furazolidone and antibiotic (nalidixic acid drugs, has been studied using the short-term bacterial assay, SOS chromotest, with Escherichia coli PQ 37 as the test organism. It has been shown that S. boulardii possesses antigenotoxic activity, revealed by SOS chromotest, when coincubated with these genotoxins. A weaker antigenotoxic activity against the same compounds was observed with S. carlsbergensis, too.

  18. Yeast cell factories on the horizon

    DEFF Research Database (Denmark)

    Nielsen, Jens

    2015-01-01

    For thousands of years, yeast has been used for making beer, bread, and wine. In modern times, it has become a commercial workhorse for producing fuels, chemicals, and pharmaceuticals such as insulin, human serum albumin, and vaccines against hepatitis virus and human papillomavirus. Yeast has also...... been engineered to make chemicals at industrial scale (e.g., succinic acid, lactic acid, resveratrol) and advanced biofuels (e.g., isobutanol) (1). On page 1095 of this issue, Galanie et al. (2) demonstrate that yeast can now be engineered to produce opioids (2), a major class of compounds used...

  19. The Genome of the Basidiomycetous Yeast and Human Pathogen Cryptococcus neoformans

    OpenAIRE

    Loftus, Brendan J.; Fung, Eula; Roncaglia, Paola; Rowley, Don; Amedeo, Paolo; Bruno, Dan; Vamathevan, Jessica; Miranda, Molly; Anderson, Iain J.; Fraser, James A.; Allen, Jonathan E.; Bosdet, Ian E.; Brent, Michael R.; Chiu, Readman; Doering, Tamara L.

    2005-01-01

    Cryptococcus neoformans is a basidiomycetous yeast ubiquitous in the environment, a model for fungal pathogenesis, and an opportunistic human pathogen of global importance. We have sequenced its ~20-megabase genome, which contains ~6500 intron-rich gene structures and encodes a transcriptome abundant in alternatively spliced and antisense messages. The genome is rich in transposons, many of which cluster at candidate centromeric regions. The presence of these transposons may drive karyotype i...

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

    Science.gov (United States)

    Zhang, Yueping; Nielsen, Jens; Liu, Zihe

    2017-12-01

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

  1. Emergence of Antimicrobial-Resistant Escherichia coli of Animal Origin Spreading in Humans.

    Science.gov (United States)

    Skurnik, David; Clermont, Olivier; Guillard, Thomas; Launay, Adrien; Danilchanka, Olga; Pons, Stéphanie; Diancourt, Laure; Lebreton, François; Kadlec, Kristina; Roux, Damien; Jiang, Deming; Dion, Sara; Aschard, Hugues; Denamur, Maurice; Cywes-Bentley, Colette; Schwarz, Stefan; Tenaillon, Olivier; Andremont, Antoine; Picard, Bertrand; Mekalanos, John; Brisse, Sylvain; Denamur, Erick

    2016-04-01

    In the context of the great concern about the impact of human activities on the environment, we studied 403 commensal Escherichia coli/Escherichia clade strains isolated from several animal and human populations that have variable contacts to one another. Multilocus sequence typing (MLST) showed a decrease of diversity 1) in strains isolated from animals that had an increasing contact with humans and 2) in all strains that had increased antimicrobial resistance. A specific B1 phylogroup clonal complex (CC87, Institut Pasteur schema nomenclature) of animal origin was identified and characterized as being responsible for the increased antimicrobial resistance prevalence observed in strains from the environments with a high human-mediated antimicrobial pressure. CC87 strains have a high capacity of acquiring and disseminating resistance genes with specific metabolic and genetic determinants as demonstrated by high-throughput sequencing and phenotyping. They are good mouse gut colonizers but are not virulent. Our data confirm the predominant role of human activities in the emergence of antimicrobial resistance in the environmental bacterial strains and unveil a particular E. coli clonal complex of animal origin capable of spreading antimicrobial resistance to other members of microbial communities. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Proteolytic activities in yeast after UV irradiation. Pt. 1

    International Nuclear Information System (INIS)

    Schwencke, J.; Moustacchi, E.

    1982-01-01

    Specific proteolytic activities are known to be induced in Escherichia coli following irradiation. Consequently it seemed of interest to investigate whether variations in proteinase activities occur in yeast. Among the five most well known proteinases of Saccharomyces cerevisiae, we have found that proteinase B activity increases up to three times in wild-type RAD + yeast cells after a dose of 50 Jm -2 of 254 nm ultraviolet light (40% survival). Carboxypeptidase Y and aminopeptidase I (leucin aminopeptidase) activities were only moderately increased. Proteinase A activity was only slightly enhanced, while aminopeptidase II (lysin aminopeptidase) was unaffected in both RAD + strains studied. The observed post UV-increase in proteinase B activity was inhibited by cycloheximide and was dose dependent. Increases in proteinase B levels were independent of the activation method used to destroy the proteinase B-inhibitor complex present in the crude yeast extracts. A standard method for comparison of the postirradiation levels among different proteinases, strains and methods of activation is presented. (orig.)

  3. Proteolytic activities in yeast after UV irradiation. Pt. 1

    Energy Technology Data Exchange (ETDEWEB)

    Schwencke, J.; Moustacchi, E.

    1982-04-01

    Specific proteolytic activities are known to be induced in Escherichia coli following irradiation. Consequently it seemed of interest to investigate whether variations in proteinase activities occur in yeast. Among the five most well known proteinases of Saccharomyces cerevisiae, we have found that proteinase B activity increases up to three times in wild-type RAD/sup +/ yeast cells after a dose of 50 Jm/sup -2/ of 254 nm ultraviolet light (40% survival). Carboxypeptidase Y and aminopeptidase I (leucin aminopeptidase) activities were only moderately increased. Proteinase A activity was only slightly enhanced, while aminopeptidase II (lysin aminopeptidase) was unaffected in both RAD/sup +/ strains studied. The observed post UV-increase in proteinase B activity was inhibited by cycloheximide and was dose dependent. Increases in proteinase B levels were independent of the activation method used to destroy the proteinase B-inhibitor complex present in the crude yeast extracts. A standard method for comparison of the postirradiation levels among different proteinases, strains and methods of activation is presented.

  4. Multiplex PCR Assay for Identification of Human Diarrheagenic Escherichia coli

    OpenAIRE

    Toma, Claudia; Lu, Yan; Higa, Naomi; Nakasone, Noboru; Chinen, Isabel; Baschkier, Ariela; Rivas, Marta; Iwanaga, Masaaki

    2003-01-01

    A multiplex PCR assay for the identification of human diarrheagenic Escherichia coli was developed. The targets selected for each category were eae for enteropathogenic E. coli, stx for Shiga toxin-producing E. coli, elt and est for enterotoxigenic E. coli, ipaH for enteroinvasive E. coli, and aggR for enteroaggregative E. coli. This assay allowed the categorization of a diarrheagenic E. coli strain in a single reaction tube.

  5. Rationalization of paclitaxel insensitivity of yeast β-tubulin and human βIII-tubulin isotype using principal component analysis

    Directory of Open Access Journals (Sweden)

    Das Lalita

    2012-08-01

    Full Text Available Abstract Background The chemotherapeutic agent paclitaxel arrests cell division by binding to the hetero-dimeric protein tubulin. Subtle differences in tubulin sequences, across eukaryotes and among β-tubulin isotypes, can have profound impact on paclitaxel-tubulin binding. To capture the experimentally observed paclitaxel-resistance of human βIII tubulin isotype and yeast β-tubulin, within a common theoretical framework, we have performed structural principal component analyses of β-tubulin sequences across eukaryotes. Results The paclitaxel-resistance of human βIII tubulin isotype and yeast β-tubulin uniquely mapped on to the lowest two principal components, defining the paclitaxel-binding site residues of β-tubulin. The molecular mechanisms behind paclitaxel-resistance, mediated through key residues, were identified from structural consequences of characteristic mutations that confer paclitaxel-resistance. Specifically, Ala277 in βIII isotype was shown to be crucial for paclitaxel-resistance. Conclusions The present analysis captures the origin of two apparently unrelated events, paclitaxel-insensitivity of yeast tubulin and human βIII tubulin isotype, through two common collective sequence vectors.

  6. Genomics and the making of yeast biodiversity.

    Science.gov (United States)

    Hittinger, Chris Todd; Rokas, Antonis; Bai, Feng-Yan; Boekhout, Teun; Gonçalves, Paula; Jeffries, Thomas W; Kominek, Jacek; Lachance, Marc-André; Libkind, Diego; Rosa, Carlos A; Sampaio, José Paulo; Kurtzman, Cletus P

    2015-12-01

    Yeasts are unicellular fungi that do not form fruiting bodies. Although the yeast lifestyle has evolved multiple times, most known species belong to the subphylum Saccharomycotina (syn. Hemiascomycota, hereafter yeasts). This diverse group includes the premier eukaryotic model system, Saccharomyces cerevisiae; the common human commensal and opportunistic pathogen, Candida albicans; and over 1000 other known species (with more continuing to be discovered). Yeasts are found in every biome and continent and are more genetically diverse than angiosperms or chordates. Ease of culture, simple life cycles, and small genomes (∼10-20Mbp) have made yeasts exceptional models for molecular genetics, biotechnology, and evolutionary genomics. Here we discuss recent developments in understanding the genomic underpinnings of the making of yeast biodiversity, comparing and contrasting natural and human-associated evolutionary processes. Only a tiny fraction of yeast biodiversity and metabolic capabilities has been tapped by industry and science. Expanding the taxonomic breadth of deep genomic investigations will further illuminate how genome function evolves to encode their diverse metabolisms and ecologies. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. The central domain of yeast transcription factor Rpn4 facilitates degradation of reporter protein in human cells.

    Science.gov (United States)

    Morozov, A V; Spasskaya, D S; Karpov, D S; Karpov, V L

    2014-10-16

    Despite high interest in the cellular degradation machinery and protein degradation signals (degrons), few degrons with universal activity along species have been identified. It has been shown that fusion of a target protein with a degradation signal from mammalian ornithine decarboxylase (ODC) induces fast proteasomal degradation of the chimera in both mammalian and yeast cells. However, no degrons from yeast-encoded proteins capable to function in mammalian cells were identified so far. Here, we demonstrate that the yeast transcription factor Rpn4 undergoes fast proteasomal degradation and its central domain can destabilize green fluorescent protein and Alpha-fetoprotein in human HEK 293T cells. Furthermore, we confirm the activity of this degron in yeast. Thus, the Rpn4 central domain is an effective interspecies degradation signal. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  8. Yeast cell wall chitin reduces wine haze formation.

    Science.gov (United States)

    Ndlovu, Thulile; Divol, Benoit; Bauer, Florian F

    2018-04-27

    Protein haze formation in bottled wines is a significant concern for the global wine industry and wine clarification before bottling is therefore a common but expensive practice. Previous studies have shown that wine yeast strains can reduce haze formation through the secretion of certain mannoproteins, but it has been suggested that other yeast-dependent haze protective mechanisms exist. On the other hand, addition of chitin has been shown to reduce haze formation, likely because grape chitinases have been shown to be the major contributors to haze. In this study, Chardonnay grape must fermented by various yeast strains resulted in wines with different protein haze levels indicating differences in haze protective capacities of the strains. The cell wall chitin levels of these strains were determined, and a strong correlation between cell wall chitin levels and haze protection capability was observed. To further evaluate the mechanism of haze protection, Escherichia coli -produced GFP-tagged grape chitinase was shown to bind efficiently to yeast cell walls in a cell wall chitin concentration-dependent manner, while commercial chitinase was removed from synthetic wine in quantities also correlated with the cell wall chitin levels of the strains. Our findings suggest a new mechanism of reducing wine haze, and propose a strategy for optimizing wine yeast strains to improve wine clarification. Importance In this study, we establish a new mechanism by which wine yeast strains can impact on the protein haze formation of wines, and demonstrate that yeast cell wall chitin binds grape chitinase in a chitin-concentration dependent manner. We also show that yeast can remove this haze-forming protein from wine. Chitin has in the past been shown to efficiently reduce wine haze formation when added to the wine in high concentration as a clarifying agent. Our data suggest that the selection of yeast strains with high levels of cell wall chitin can reduce protein haze. We also

  9. Yeast synthetic biology toolbox and applications for biofuel production.

    Science.gov (United States)

    Tsai, Ching-Sung; Kwak, Suryang; Turner, Timothy L; Jin, Yong-Su

    2015-02-01

    Yeasts are efficient biofuel producers with numerous advantages outcompeting bacterial counterparts. While most synthetic biology tools have been developed and customized for bacteria especially for Escherichia coli, yeast synthetic biological tools have been exploited for improving yeast to produce fuels and chemicals from renewable biomass. Here we review the current status of synthetic biological tools and their applications for biofuel production, focusing on the model strain Saccharomyces cerevisiae We describe assembly techniques that have been developed for constructing genes, pathways, and genomes in yeast. Moreover, we discuss synthetic parts for allowing precise control of gene expression at both transcriptional and translational levels. Applications of these synthetic biological approaches have led to identification of effective gene targets that are responsible for desirable traits, such as cellulosic sugar utilization, advanced biofuel production, and enhanced tolerance against toxic products for biofuel production from renewable biomass. Although an array of synthetic biology tools and devices are available, we observed some gaps existing in tool development to achieve industrial utilization. Looking forward, future tool development should focus on industrial cultivation conditions utilizing industrial strains. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  10. Assessing pathogenicity of MLH1 variants by co-expression of human MLH1 and PMS2 genes in yeast

    Energy Technology Data Exchange (ETDEWEB)

    Vogelsang, Matjaz; Comino, Aleksandra; Zupanec, Neja [Department for Biosynthesis and Biotransformation, National Institute of Chemistry, Hajdrihova 19, SI-1001 Ljubljana (Slovenia); Hudler, Petra [Medical Center for Molecular Biology, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, SI-1000 Ljubljana (Slovenia); Komel, Radovan [Department for Biosynthesis and Biotransformation, National Institute of Chemistry, Hajdrihova 19, SI-1001 Ljubljana (Slovenia); Medical Center for Molecular Biology, Faculty of Medicine, University of Ljubljana, Vrazov trg 2, SI-1000 Ljubljana (Slovenia)

    2009-10-28

    Loss of DNA mismatch repair (MMR) in humans, mainly due to mutations in the hMLH1 gene, is linked to hereditary nonpolyposis colorectal cancer (HNPCC). Because not all MLH1 alterations result in loss of MMR function, accurate characterization of variants and their classification in terms of their effect on MMR function is essential for reliable genetic testing and effective treatment. To date, in vivo assays for functional characterization of MLH1 mutations performed in various model systems have used episomal expression of the modified MMR genes. We describe here a novel approach to determine accurately the functional significance of hMLH1 mutations in vivo, based on co-expression of human MLH1 and PMS2 in yeast cells. Yeast MLH1 and PMS1 genes, whose protein products form the MutLα complex, were replaced by human orthologs directly on yeast chromosomes by homologous recombination, and the resulting MMR activity was tested. The yeast strain co-expressing hMLH1 and hPMS2 exhibited the same mutation rate as the wild-type. Eight cancer-related MLH1 variants were introduced, using the same approach, into the prepared yeast model, and their effect on MMR function was determined. Five variants (A92P, S93G, I219V, K618R and K618T) were classified as non-pathogenic, whereas variants T117M, Y646C and R659Q were characterized as pathogenic. Results of our in vivo yeast-based approach correlate well with clinical data in five out of seven hMLH1 variants and the described model was thus shown to be useful for functional characterization of MLH1 variants in cancer patients found throughout the entire coding region of the gene.

  11. Assessing pathogenicity of MLH1 variants by co-expression of human MLH1 and PMS2 genes in yeast

    Directory of Open Access Journals (Sweden)

    Hudler Petra

    2009-10-01

    Full Text Available Abstract Background Loss of DNA mismatch repair (MMR in humans, mainly due to mutations in the hMLH1 gene, is linked to hereditary nonpolyposis colorectal cancer (HNPCC. Because not all MLH1 alterations result in loss of MMR function, accurate characterization of variants and their classification in terms of their effect on MMR function is essential for reliable genetic testing and effective treatment. To date, in vivo assays for functional characterization of MLH1 mutations performed in various model systems have used episomal expression of the modified MMR genes. We describe here a novel approach to determine accurately the functional significance of hMLH1 mutations in vivo, based on co-expression of human MLH1 and PMS2 in yeast cells. Methods Yeast MLH1 and PMS1 genes, whose protein products form the MutLα complex, were replaced by human orthologs directly on yeast chromosomes by homologous recombination, and the resulting MMR activity was tested. Results The yeast strain co-expressing hMLH1 and hPMS2 exhibited the same mutation rate as the wild-type. Eight cancer-related MLH1 variants were introduced, using the same approach, into the prepared yeast model, and their effect on MMR function was determined. Five variants (A92P, S93G, I219V, K618R and K618T were classified as non-pathogenic, whereas variants T117M, Y646C and R659Q were characterized as pathogenic. Conclusion Results of our in vivo yeast-based approach correlate well with clinical data in five out of seven hMLH1 variants and the described model was thus shown to be useful for functional characterization of MLH1 variants in cancer patients found throughout the entire coding region of the gene.

  12. Assessing pathogenicity of MLH1 variants by co-expression of human MLH1 and PMS2 genes in yeast

    International Nuclear Information System (INIS)

    Vogelsang, Matjaz; Comino, Aleksandra; Zupanec, Neja; Hudler, Petra; Komel, Radovan

    2009-01-01

    Loss of DNA mismatch repair (MMR) in humans, mainly due to mutations in the hMLH1 gene, is linked to hereditary nonpolyposis colorectal cancer (HNPCC). Because not all MLH1 alterations result in loss of MMR function, accurate characterization of variants and their classification in terms of their effect on MMR function is essential for reliable genetic testing and effective treatment. To date, in vivo assays for functional characterization of MLH1 mutations performed in various model systems have used episomal expression of the modified MMR genes. We describe here a novel approach to determine accurately the functional significance of hMLH1 mutations in vivo, based on co-expression of human MLH1 and PMS2 in yeast cells. Yeast MLH1 and PMS1 genes, whose protein products form the MutLα complex, were replaced by human orthologs directly on yeast chromosomes by homologous recombination, and the resulting MMR activity was tested. The yeast strain co-expressing hMLH1 and hPMS2 exhibited the same mutation rate as the wild-type. Eight cancer-related MLH1 variants were introduced, using the same approach, into the prepared yeast model, and their effect on MMR function was determined. Five variants (A92P, S93G, I219V, K618R and K618T) were classified as non-pathogenic, whereas variants T117M, Y646C and R659Q were characterized as pathogenic. Results of our in vivo yeast-based approach correlate well with clinical data in five out of seven hMLH1 variants and the described model was thus shown to be useful for functional characterization of MLH1 variants in cancer patients found throughout the entire coding region of the gene

  13. Homology of yeast photoreactivating gene fragment with human genomic digests

    International Nuclear Information System (INIS)

    Meechan, P.J.; Milam, K.M.; Cleaver, J.E.

    1984-01-01

    Enzymatic photoreactivation of UV-induced DNA lesions has been demonstrated for a variety of prokaryotic and eukaryotic organisms. Its presence in placental mammals, however, has not been clearly established. The authors attempted to resolve this question by assaying for the presence (or absence) of sequences in human DNA complimentary to a fragment of the photoreactivating gene from S. cerevisiae that has recently been cloned. In another study, DNA from human, chick E. coli and yeast cells was digested with either HindIII of BglII, electrophoresed on a 0.5% agarose gel, transferred (Southern blot) to a nylon membrane and probed for homology against a Sau3A restriction fragment from S. cerevisiae that compliments phr/sup -/ cells. Hybridization to human DNA digests was observed only under relatively non-stringent conditions indicating the gene is not conserved in placental mammals. These results are correlated with current literature data concerning photoreactivating enzymes

  14. Yeast for virus research

    Science.gov (United States)

    Zhao, Richard Yuqi

    2017-01-01

    Budding yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe) are two popular model organisms for virus research. They are natural hosts for viruses as they carry their own indigenous viruses. Both yeasts have been used for studies of plant, animal and human viruses. Many positive sense (+) RNA viruses and some DNA viruses replicate with various levels in yeasts, thus allowing study of those viral activities during viral life cycle. Yeasts are single cell eukaryotic organisms. Hence, many of the fundamental cellular functions such as cell cycle regulation or programed cell death are highly conserved from yeasts to higher eukaryotes. Therefore, they are particularly suited to study the impact of those viral activities on related cellular activities during virus-host interactions. Yeasts present many unique advantages in virus research over high eukaryotes. Yeast cells are easy to maintain in the laboratory with relative short doubling time. They are non-biohazardous, genetically amendable with small genomes that permit genome-wide analysis of virologic and cellular functions. In this review, similarities and differences of these two yeasts are described. Studies of virologic activities such as viral translation, viral replication and genome-wide study of virus-cell interactions in yeasts are highlighted. Impacts of viral proteins on basic cellular functions such as cell cycle regulation and programed cell death are discussed. Potential applications of using yeasts as hosts to carry out functional analysis of small viral genome and to develop high throughput drug screening platform for the discovery of antiviral drugs are presented. PMID:29082230

  15. Probiotic Properties of Non-Saccharomyces Yeasts

    DEFF Research Database (Denmark)

    Smith, Ida Mosbech

    to harmless luminal substances is a key feature of the intestinal immune system. In this context, dendritic cells (DCs) present in the tissues lining the human gut are central players involved in microbial sensing and shaping of appropriate adaptive immune responses. Probiotics are live microorganisms which...... when administered in adequate amounts confer a health benefit on the host. While the majority of probiotic microorganisms studied to date are lactic acid bacteria, research in yeasts with potentially beneficial influences on human health has mainly revolved around Saccharomyces boulardii. This yeast...... has shown a positive impact on disease outcome in clinical studies of inflammatory bowel disease, indicating an ability of S. boulardii to influence human immune responses underlying intestinal inflammation. Consequent to this focus on S. boulardii as the fundamental probiotic yeast, very little...

  16. Yeast cell differentiation: Lessons from pathogenic and non-pathogenic yeasts.

    Science.gov (United States)

    Palková, Zdena; Váchová, Libuše

    2016-09-01

    Yeasts, historically considered to be single-cell organisms, are able to activate different differentiation processes. Individual yeast cells can change their life-styles by processes of phenotypic switching such as the switch from yeast-shaped cells to filamentous cells (pseudohyphae or true hyphae) and the transition among opaque, white and gray cell-types. Yeasts can also create organized multicellular structures such as colonies and biofilms, and the latter are often observed as contaminants on surfaces in industry and medical care and are formed during infections of the human body. Multicellular structures are formed mostly of stationary-phase or slow-growing cells that diversify into specific cell subpopulations that have unique metabolic properties and can fulfill specific tasks. In addition to the development of multiple protective mechanisms, processes of metabolic reprogramming that reflect a changed environment help differentiated individual cells and/or community cell constituents to survive harmful environmental attacks and/or to escape the host immune system. This review aims to provide an overview of differentiation processes so far identified in individual yeast cells as well as in multicellular communities of yeast pathogens of the Candida and Cryptococcus spp. and the Candida albicans close relative, Saccharomyces cerevisiae. Molecular mechanisms and extracellular signals potentially involved in differentiation processes are also briefly mentioned. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Human Thyroid Cancer-1 (TC-1 is a vertebrate specific oncogenic protein that protects against copper and pro-apoptotic genes in yeast

    Directory of Open Access Journals (Sweden)

    Natalie K. Jones

    2015-07-01

    Full Text Available The human Thyroid Cancer-1 (hTC-1 protein, also known as C8orf4 was initially identified as a gene that was up-regulated in human thyroid cancer. Here we show that hTC-1 is a peptide that prevents the effects of over-expressing Bax in yeast. Analysis of the 106 residues of hTC-1 in available protein databases revealed direct orthologues in jawed-vertebrates, including mammals, frogs, fish and sharks. No TC-1 orthologue was detected in lower organisms, including yeast. Here we show that TC-1 is a general pro-survival peptide since it prevents the growth- and cell death-inducing effects of copper in yeast. Human TC-1 also prevented the deleterious effects that occur due to the over-expression of a number of key pro-apoptotic peptides, including YCA1, YBH3, NUC1, and AIF1. Even though the protective effects were more pronounced with the over-expression of YBH3 and YCA1, hTC-1 could still protect yeast mutants lacking YBH3 and YCA1 from the effects of copper sulfate. This suggests that the protective effects of TC-1 are not limited to specific pathways or processes. Taken together, our results indicate that hTC-1 is a pro-survival protein that retains its function when heterologously expressed in yeast. Thus yeast is a useful model to characterize the potential roles in cell death and survival of cancer related genes.

  18. Molecular relatedness of ESBL/AmpC-producing Escherichia coli from humans, animals, food and the enviroment

    NARCIS (Netherlands)

    Dorado-Garcia, Alejandro; Smid, J.H.; Pelt, Van Wilfrid; Bonten, M.J.M.; Fluit, A.C.; Bunt, van den Gerrita; Wagenaar, J.A.; Hordijk, J.; Dierikx, C.M.; Veldman, K.T.; Koeijer, de A.A.; Dohmen, W.; Schmitt, H.; Liakopoulos, A.; Pacholewicz, Ewa; Lam, T.J.G.M.; Velthuis, Annet; Heuvelink, A.; Gonggrijp, Maaike; Duijkeren, van E.; Hoek, van A.H.A.M.; Roda Husman, de A.N.; Blaak, H.; Havelaar, A.H.; Mevius, D.J.; Heederik, D.J.J.

    2018-01-01

    Background: In recent years, ESBL/AmpC-producing Escherichia coli ESBL/AmpC-EC) have been isolated with increasing frequency from animals, food, environmental sources and humans. With incomplete and scattered evidence, the contribution to the human carriage burden from these reservoirs remains

  19. Production of Food Grade Yeasts

    Directory of Open Access Journals (Sweden)

    Argyro Bekatorou

    2006-01-01

    Full Text Available Yeasts have been known to humans for thousands of years as they have been used in traditional fermentation processes like wine, beer and bread making. Today, yeasts are also used as alternative sources of high nutritional value proteins, enzymes and vitamins, and have numerous applications in the health food industry as food additives, conditioners and flavouring agents, for the production of microbiology media and extracts, as well as livestock feeds. Modern scientific advances allow the isolation, construction and industrial production of new yeast strains to satisfy the specific demands of the food industry. Types of commercial food grade yeasts, industrial production processes and raw materials are highlighted. Aspects of yeast metabolism, with respect to carbohydrate utilization, nutritional aspects and recent research advances are also discussed.

  20. UV-dependent production of 25-hydroxyvitamin D2 in the recombinant yeast cells expressing human CYP2R1

    International Nuclear Information System (INIS)

    Yasuda, Kaori; Endo, Mariko; Ikushiro, Shinichi; Kamakura, Masaki; Ohta, Miho; Sakaki, Toshiyuki

    2013-01-01

    Highlights: •We produce 25-hydroxyvitamin D in the recombinant yeast expressing human CYP2R1. •Vitamin D2 is produced in yeast from endogenous ergosterol with UV irradiation. •We produce 25-hydroxyvitamin D2 in the recombinant yeast without added substrate. -- Abstract: CYP2R1 is known to be a physiologically important vitamin D 25-hydroxylase. We have successfully expressed human CYP2R1 in Saccharomyces cerevisiae to reveal its enzymatic properties. In this study, we examined production of 25-hydroxylated vitamin D using whole recombinant yeast cells that expressed CYP2R1. When vitamin D 3 or vitamin D 2 was added to the cell suspension of CYP2R1-expressing yeast cells in a buffer containing glucose and β-cyclodextrin, the vitamins were converted into their 25-hydroxylated products. Next, we irradiated the cell suspension with UVB and incubated at 37 °C. Surprisingly, the 25-hydroxy vitamin D 2 was produced without additional vitamin D 2 . Endogenous ergosterol was likely converted into vitamin D 2 by UV irradiation and thermal isomerization, and then the resulting vitamin D 2 was converted to 25-hydroxyvitamin D 2 by CYP2R1. This novel method for producing 25-hydroxyvitamin D 2 without a substrate could be useful for practical purposes

  1. Problem-Solving Test: Analysis of DNA Damage Recognizing Proteins in Yeast and Human Cells

    Science.gov (United States)

    Szeberenyi, Jozsef

    2013-01-01

    The experiment described in this test was aimed at identifying DNA repair proteins in human and yeast cells. Terms to be familiar with before you start to solve the test: DNA repair, germline mutation, somatic mutation, inherited disease, cancer, restriction endonuclease, radioactive labeling, [alpha-[superscript 32]P]ATP, [gamma-[superscript…

  2. A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast

    DEFF Research Database (Denmark)

    Kampranis, S C; Damianova, R; Atallah, M

    2000-01-01

    The mammalian inducer of apoptosis Bax is lethal when expressed in yeast and plant cells. To identify potential inhibitors of Bax in plants we transformed yeast cells expressing Bax with a tomato cDNA library and we selected for cells surviving after the induction of Bax. This genetic screen allows...... for the identification of plant genes, which inhibit either directly or indirectly the lethal phenotype of Bax. Using this method a number of cDNA clones were isolated, the more potent of which encodes a protein homologous to the class theta glutathione S-transferases. This Bax-inhibiting (BI) protein was expressed...... in Escherichia coli and found to possess glutathione S-transferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glutathione, causes a substantial reduction of total cellular phospholipids, diminishes the mitochondrial membrane...

  3. Defining the pathogenesis of the human Atp12p W94R mutation using a Saccharomyces cerevisiae yeast model.

    Science.gov (United States)

    Meulemans, Ann; Seneca, Sara; Pribyl, Thomas; Smet, Joel; Alderweirldt, Valerie; Waeytens, Anouk; Lissens, Willy; Van Coster, Rudy; De Meirleir, Linda; di Rago, Jean-Paul; Gatti, Domenico L; Ackerman, Sharon H

    2010-02-05

    Studies in yeast have shown that a deficiency in Atp12p prevents assembly of the extrinsic domain (F(1)) of complex V and renders cells unable to make ATP through oxidative phosphorylation. De Meirleir et al. (De Meirleir, L., Seneca, S., Lissens, W., De Clercq, I., Eyskens, F., Gerlo, E., Smet, J., and Van Coster, R. (2004) J. Med. Genet. 41, 120-124) have reported that a homozygous missense mutation in the gene for human Atp12p (HuAtp12p), which replaces Trp-94 with Arg, was linked to the death of a 14-month-old patient. We have investigated the impact of the pathogenic W94R mutation on Atp12p structure/function. Plasmid-borne wild type human Atp12p rescues the respiratory defect of a yeast ATP12 deletion mutant (Deltaatp12). The W94R mutation alters the protein at the most highly conserved position in the Pfam sequence and renders HuAtp12p insoluble in the background of Deltaatp12. In contrast, the yeast protein harboring the corresponding mutation, ScAtp12p(W103R), is soluble in the background of Deltaatp12 but not in the background of Deltaatp12Deltafmc1, a strain that also lacks Fmc1p. Fmc1p is a yeast mitochondrial protein not found in higher eukaryotes. Tryptophan 94 (human) or 103 (yeast) is located in a positively charged region of Atp12p, and hence its mutation to arginine does not alter significantly the electrostatic properties of the protein. Instead, we provide evidence that the primary effect of the substitution is on the dynamic properties of Atp12p.

  4. Inaccurate DNA synthesis in cell extracts of yeast producing active human DNA polymerase iota.

    Science.gov (United States)

    Makarova, Alena V; Grabow, Corinn; Gening, Leonid V; Tarantul, Vyacheslav Z; Tahirov, Tahir H; Bessho, Tadayoshi; Pavlov, Youri I

    2011-01-31

    Mammalian Pol ι has an unusual combination of properties: it is stimulated by Mn(2+) ions, can bypass some DNA lesions and misincorporates "G" opposite template "T" more frequently than incorporates the correct "A." We recently proposed a method of detection of Pol ι activity in animal cell extracts, based on primer extension opposite the template T with a high concentration of only two nucleotides, dGTP and dATP (incorporation of "G" versus "A" method of Gening, abbreviated as "misGvA"). We provide unambiguous proof of the "misGvA" approach concept and extend the applicability of the method for the studies of variants of Pol ι in the yeast model system with different cation cofactors. We produced human Pol ι in baker's yeast, which do not have a POLI ortholog. The "misGvA" activity is absent in cell extracts containing an empty vector, or producing catalytically dead Pol ι, or Pol ι lacking exon 2, but is robust in the strain producing wild-type Pol ι or its catalytic core, or protein with the active center L62I mutant. The signature pattern of primer extension products resulting from inaccurate DNA synthesis by extracts of cells producing either Pol ι or human Pol η is different. The DNA sequence of the template is critical for the detection of the infidelity of DNA synthesis attributed to DNA Pol ι. The primer/template and composition of the exogenous DNA precursor pool can be adapted to monitor replication fidelity in cell extracts expressing various error-prone Pols or mutator variants of accurate Pols. Finally, we demonstrate that the mutation rates in yeast strains producing human DNA Pols ι and η are not elevated over the control strain, despite highly inaccurate DNA synthesis by their extracts.

  5. Inaccurate DNA synthesis in cell extracts of yeast producing active human DNA polymerase iota.

    Directory of Open Access Journals (Sweden)

    Alena V Makarova

    2011-01-01

    Full Text Available Mammalian Pol ι has an unusual combination of properties: it is stimulated by Mn(2+ ions, can bypass some DNA lesions and misincorporates "G" opposite template "T" more frequently than incorporates the correct "A." We recently proposed a method of detection of Pol ι activity in animal cell extracts, based on primer extension opposite the template T with a high concentration of only two nucleotides, dGTP and dATP (incorporation of "G" versus "A" method of Gening, abbreviated as "misGvA". We provide unambiguous proof of the "misGvA" approach concept and extend the applicability of the method for the studies of variants of Pol ι in the yeast model system with different cation cofactors. We produced human Pol ι in baker's yeast, which do not have a POLI ortholog. The "misGvA" activity is absent in cell extracts containing an empty vector, or producing catalytically dead Pol ι, or Pol ι lacking exon 2, but is robust in the strain producing wild-type Pol ι or its catalytic core, or protein with the active center L62I mutant. The signature pattern of primer extension products resulting from inaccurate DNA synthesis by extracts of cells producing either Pol ι or human Pol η is different. The DNA sequence of the template is critical for the detection of the infidelity of DNA synthesis attributed to DNA Pol ι. The primer/template and composition of the exogenous DNA precursor pool can be adapted to monitor replication fidelity in cell extracts expressing various error-prone Pols or mutator variants of accurate Pols. Finally, we demonstrate that the mutation rates in yeast strains producing human DNA Pols ι and η are not elevated over the control strain, despite highly inaccurate DNA synthesis by their extracts.

  6. [Engineering of a System for the Production of Mutant Human Alpha-Fetoprotein in the Methylotrophic Yeast Pichia pastoris].

    Science.gov (United States)

    Morozkina, E V; Vavilova, E A; Zatsepin, S S; Klyachko, E V; Yagudin, T A; Chulkin, A M; Dudich, I V; Semenkova, L N; Churilova, I V; Benevolenskii, S V

    2016-01-01

    A system for the production of mutant recombinant human alpha-fetoprotein (rhAFPO) lacking the glycosylation site has been engineered in the yeast Pichia pastoris. A strain of the methylotrophic yeast Pichia pastoris GS 115/pPICZ?A/rhAFP0, which produces unglycosylated rhAFPO and secretes it to the culture medium, has been constructed. Optimization and scale-up of the fermentation technology have resulted in an increase in the rhAFP0 yield to 20 mg/L. A scheme of isolation and purification of biologically active rhAFP0 has been developed. The synthesized protein has the antitumor activity, which is analogous to the activity of natural human embryonic alpha-fetoprotein.

  7. Detection of Protein Interactions in T3S Systems Using Yeast Two-Hybrid Analysis.

    Science.gov (United States)

    Nilles, Matthew L

    2017-01-01

    Two-hybrid systems, sometimes termed interaction traps, are genetic systems designed to find and analyze interactions between proteins. The most common systems are yeast based (commonly Saccharomyces cerevisae) and rely on the functional reconstitution of the GAL4 transcriptional activator. Reporter genes, such as the lacZ gene of Escherichia coli (encodes β-galactosidase), are placed under GAL4-dependent transcriptional control to provide quick and reliable detection of protein interactions. In this method the use of a yeast-based two-hybrid system is described to study protein interactions between components of type III secretion systems.

  8. Bioavailability of selenium from fish, yeast and selenate: A comparative study in humans using stable isotopes

    NARCIS (Netherlands)

    Fox, T.E.; Heuvel, E.G.H.M. van den; Atherton, C.A.; Dainty, J.R.; Lewis, D.J.; Langford, N.J.; Crews, H.M.; Luten, J.B.; Lorentzen, M.; Sieling, F.W.; Aken-Schneyder, P. van; Hoek, M.; Kotterman, M.J.J.; Dael, P. van; Firweather-Tail, S.J.

    2004-01-01

    Objective: To measure the bioavailability of selenium from cooked and raw fish in humans by estimating and comparing apparent absorption and retention of selenium in biosynthetically labelled fish with labelled selenate and biosynthetically labelled selenium in brewers yeast. Design: The

  9. Bioavailibility of selenium from fish, yeast and selenate: a comparative study in humans using stable isotopes

    NARCIS (Netherlands)

    Fox, T.E.; Heuvel, van den E.G.H.M.; Atherton, C.A.; Luten, J.B.; Hoek-van Nieuwenhuizen, van M.; Kotterman, M.J.J.

    2004-01-01

    Objective: To measure the bioavailability of selenium from cooked and raw fish in humans by estimating and comparing apparent absorption and retention of selenium in biosynthetically labelled fish with labelled selenate and biosynthetically labelled selenium in brewers yeast. Design: The

  10. UV-dependent production of 25-hydroxyvitamin D{sub 2} in the recombinant yeast cells expressing human CYP2R1

    Energy Technology Data Exchange (ETDEWEB)

    Yasuda, Kaori; Endo, Mariko; Ikushiro, Shinichi; Kamakura, Masaki [Department of Biotechnology, Faculty of Engineering, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398 (Japan); Ohta, Miho [Department of Food and Nutrition Management Studies, Faculty of Human Development, Soai University, 4-4-1 Nanko-naka, Suminoe-ku, Osaka 559-0033 (Japan); Sakaki, Toshiyuki, E-mail: tsakaki@pu-toyama.ac.jp [Department of Biotechnology, Faculty of Engineering, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398 (Japan)

    2013-05-03

    Highlights: •We produce 25-hydroxyvitamin D in the recombinant yeast expressing human CYP2R1. •Vitamin D2 is produced in yeast from endogenous ergosterol with UV irradiation. •We produce 25-hydroxyvitamin D2 in the recombinant yeast without added substrate. -- Abstract: CYP2R1 is known to be a physiologically important vitamin D 25-hydroxylase. We have successfully expressed human CYP2R1 in Saccharomyces cerevisiae to reveal its enzymatic properties. In this study, we examined production of 25-hydroxylated vitamin D using whole recombinant yeast cells that expressed CYP2R1. When vitamin D{sub 3} or vitamin D{sub 2} was added to the cell suspension of CYP2R1-expressing yeast cells in a buffer containing glucose and β-cyclodextrin, the vitamins were converted into their 25-hydroxylated products. Next, we irradiated the cell suspension with UVB and incubated at 37 °C. Surprisingly, the 25-hydroxy vitamin D{sub 2} was produced without additional vitamin D{sub 2}. Endogenous ergosterol was likely converted into vitamin D{sub 2} by UV irradiation and thermal isomerization, and then the resulting vitamin D{sub 2} was converted to 25-hydroxyvitamin D{sub 2} by CYP2R1. This novel method for producing 25-hydroxyvitamin D{sub 2} without a substrate could be useful for practical purposes.

  11. Protein and Amino Acid Restriction, Aging and Disease: from yeast to humans

    OpenAIRE

    Mirzaei, Hamed; Suarez, Jorge A.; Longo, Valter D.

    2014-01-01

    Many of the effects of dietary restriction (DR) on longevity and health span in model organisms have been linked to reduced protein and amino acid (AA) intake and the stimulation of specific nutrient signaling pathways. Studies in yeast have shown that addition of serine, threonine, and valine in media promotes cellular sensitization and aging by activating different but connected pathways. Protein or essential AA restriction extends both lifespan and healthspan in rodent models. In humans, p...

  12. Nitrile Metabolizing Yeasts

    Science.gov (United States)

    Bhalla, Tek Chand; Sharma, Monica; Sharma, Nitya Nand

    Nitriles and amides are widely distributed in the biotic and abiotic components of our ecosystem. Nitrile form an important group of organic compounds which find their applications in the synthesis of a large number of compounds used as/in pharmaceutical, cosmetics, plastics, dyes, etc>. Nitriles are mainly hydro-lyzed to corresponding amide/acid in organic chemistry. Industrial and agricultural activities have also lead to release of nitriles and amides into the environment and some of them pose threat to human health. Biocatalysis and biotransformations are increasingly replacing chemical routes of synthesis in organic chemistry as a part of ‘green chemistry’. Nitrile metabolizing organisms or enzymes thus has assumed greater significance in all these years to convert nitriles to amides/ acids. The nitrile metabolizing enzymes are widely present in bacteria, fungi and yeasts. Yeasts metabolize nitriles through nitrilase and/or nitrile hydratase and amidase enzymes. Only few yeasts have been reported to possess aldoxime dehydratase. More than sixty nitrile metabolizing yeast strains have been hither to isolated from cyanide treatment bioreactor, fermented foods and soil. Most of the yeasts contain nitrile hydratase-amidase system for metabolizing nitriles. Transformations of nitriles to amides/acids have been carried out with free and immobilized yeast cells. The nitrilases of Torulopsis candida>and Exophiala oligosperma>R1 are enantioselec-tive and regiospecific respectively. Geotrichum>sp. JR1 grows in the presence of 2M acetonitrile and may have potential for application in bioremediation of nitrile contaminated soil/water. The nitrilase of E. oligosperma>R1 being active at low pH (3-6) has shown promise for the hydroxy acids. Immobilized yeast cells hydrolyze some additional nitriles in comparison to free cells. It is expected that more focus in future will be on purification, characterization, cloning, expression and immobilization of nitrile metabolizing

  13. Drug resistance is conferred on the model yeast Saccharomyces cerevisiae by expression of full-length melanoma-associated human ATP-binding cassette transporter ABCB5.

    Science.gov (United States)

    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.

  14. Stalled replication forks generate a distinct mutational signature in yeast

    DEFF Research Database (Denmark)

    Larsen, Nicolai B.; Liberti, Sascha E.; Vogel, Ivan

    2017-01-01

    Proliferating cells acquire genome alterations during the act of DNA replication. This leads to mutation accumulation and somatic cell mosaicism in multicellular organisms, and is also implicated as an underlying cause of aging and tumorigenesis. The molecular mechanisms of DNA replication...... Escherichia coli Tus/Ter complex) engineered into the yeast genome. We demonstrate that transient stalling at this barrier induces a distinct pattern of genome rearrangements in the newly replicated region behind the stalled fork, which primarily consist of localized losses and duplications of DNA sequences....... These genetic alterations arise through the aberrant repair of a single-stranded DNA gap, in a process that is dependent on Exo1- and Shu1-dependent homologous recombination repair (HRR). Furthermore, aberrant processing of HRR intermediates, and elevated HRR-associated mutagenesis, is detectable in a yeast...

  15. The single-strand DNA binding activity of human PC4 preventsmutagenesis and killing by oxidative DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jen-Yeu; Sarker, Altaf Hossain; Cooper, Priscilla K.; Volkert, Michael R.

    2004-02-01

    Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Yeast mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub l{Delta} mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide-resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show XPG recruits PC4 to a bubble-containing DNA substrate with resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate.

  16. Aggregation and retention of human urokinase type plasminogen activator in the yeast endoplasmic reticulum

    Directory of Open Access Journals (Sweden)

    Smirnov Vladimir N

    2002-10-01

    Full Text Available Abstract Background Secretion of recombinant proteins in yeast can be affected by their improper folding in the endoplasmic reticulum and subsequent elimination of the misfolded molecules via the endoplasmic reticulum associated protein degradation pathway. Recombinant proteins can also be degraded by the vacuolar protease complex. Human urokinase type plasminogen activator (uPA is poorly secreted by yeast but the mechanisms interfering with its secretion are largely unknown. Results We show that in Hansenula polymorpha overexpression worsens uPA secretion and stimulates its intracellular aggregation. The absence of the Golgi modifications in accumulated uPA suggests that aggregation occurs within the endoplasmic reticulum. Deletion analysis has shown that the N-terminal domains were responsible for poor uPA secretion and propensity to aggregate. Mutation abolishing N-glycosylation decreased the efficiency of uPA secretion and increased its aggregation degree. Retention of uPA in the endoplasmic reticulum stimulates its aggregation. Conclusions The data obtained demonstrate that defect of uPA secretion in yeast is related to its retention in the endoplasmic reticulum. Accumulation of uPA within the endoplasmic reticulum disturbs its proper folding and leads to formation of high molecular weight aggregates.

  17. Yeast Genomics for Bread, Beer, Biology, Bucks and Breath

    Science.gov (United States)

    Sakharkar, Kishore R.; Sakharkar, Meena K.

    The rapid advances and scale up of projects in DNA sequencing dur ing the past two decades have produced complete genome sequences of several eukaryotic species. The versatile genetic malleability of the yeast, and the high degree of conservation between its cellular processes and those of human cells have made it a model of choice for pioneering research in molecular and cell biology. The complete sequence of yeast genome has proven to be extremely useful as a reference towards the sequences of human and for providing systems to explore key gene functions. Yeast has been a ‘legendary model’ for new technologies and gaining new biological insights into basic biological sciences and biotechnology. This chapter describes the awesome power of yeast genetics, genomics and proteomics in understanding of biological function. The applications of yeast as a screening tool to the field of drug discovery and development are highlighted and the traditional importance of yeast for bakers and brewers is discussed.

  18. Caspase inhibitors of the P35 family are more active when purified from yeast than bacteria.

    Directory of Open Access Journals (Sweden)

    Ingo L Brand

    Full Text Available Many insect viruses express caspase inhibitors of the P35 superfamily, which prevent defensive host apoptosis to enable viral propagation. The prototypical P35 family member, AcP35 from Autographa californica M nucleopolyhedrovirus, has been extensively studied. Bacterially purified AcP35 has been previously shown to inhibit caspases from insect, mammalian and nematode species. This inhibition occurs via a pseudosubstrate mechanism involving caspase-mediated cleavage of a "reactive site loop" within the P35 protein, which ultimately leaves cleaved P35 covalently bound to the caspase's active site. We observed that AcP35 purifed from Saccharomyces cerevisae inhibited caspase activity more efficiently than AcP35 purified from Escherichia coli. This differential potency was more dramatic for another P35 family member, MaviP35, which inhibited human caspase 3 almost 300-fold more potently when purified from yeast than bacteria. Biophysical assays revealed that MaviP35 proteins produced in bacteria and yeast had similar primary and secondary structures. However, bacterially produced MaviP35 possessed greater thermal stability and propensity to form higher order oligomers than its counterpart purified from yeast. Caspase 3 could process yeast-purified MaviP35, but failed to detectably cleave bacterially purified MaviP35. These data suggest that bacterially produced P35 proteins adopt subtly different conformations from their yeast-expressed counterparts, which hinder caspase access to the reactive site loop to reduce the potency of caspase inhibition, and promote aggregation. These data highlight the differential caspase inhibition by recombinant P35 proteins purified from different sources, and caution that analyses of bacterially produced P35 family members (and perhaps other types of proteins may underestimate their activity.

  19. History of genome editing in yeast.

    Science.gov (United States)

    Fraczek, Marcin G; Naseeb, Samina; Delneri, Daniela

    2018-05-01

    For thousands of years humans have used the budding yeast Saccharomyces cerevisiae for the production of bread and alcohol; however, in the last 30-40 years our understanding of the yeast biology has dramatically increased, enabling us to modify its genome. Although S. cerevisiae has been the main focus of many research groups, other non-conventional yeasts have also been studied and exploited for biotechnological purposes. Our experiments and knowledge have evolved from recombination to high-throughput PCR-based transformations to highly accurate CRISPR methods in order to alter yeast traits for either research or industrial purposes. Since the release of the genome sequence of S. cerevisiae in 1996, the precise and targeted genome editing has increased significantly. In this 'Budding topic' we discuss the significant developments of genome editing in yeast, mainly focusing on Cre-loxP mediated recombination, delitto perfetto and CRISPR/Cas. © 2018 The Authors. Yeast published by John Wiley & Sons, Ltd.

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

    Directory of Open Access Journals (Sweden)

    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.

  1. Small-angle neutron scattering study of recombinant yeast-derived human hepatitis B virus surface antigen vaccine particle

    Science.gov (United States)

    Sato, M.; Ito, Y.; Kameyama, K.; Imai, M.; Ishikawa, N.; Takagi, T.

    1995-02-01

    The overall and internal structure of recombinant yeast-derived human hepatitis B virus surface antigen vaccine particles was investigated by small-angle neutron scattering using the contrast variation method. The vaccine is a nearly spherical particle, and its contrast-matching point was determined to be at about 24% D 2O content, indicating that a large part of the vaccine particle is occupied by lipids and carbohydrates from the yeast. The Stuhrmann plot suggests that the surface antigens exist predominantly in the peripheral region of the particle, which is favorable to the induction of anti-virus antibodies.

  2. Characterization of Human and Yeast Mitochondrial Glycine Carriers with Implications for Heme Biosynthesis and Anemia.

    Science.gov (United States)

    Lunetti, Paola; Damiano, Fabrizio; De Benedetto, Giuseppe; Siculella, Luisa; Pennetta, Antonio; Muto, Luigina; Paradies, Eleonora; Marobbio, Carlo Marya Thomas; Dolce, Vincenza; Capobianco, Loredana

    2016-09-16

    Heme is an essential molecule in many biological processes, such as transport and storage of oxygen and electron transfer as well as a structural component of hemoproteins. Defects of heme biosynthesis in developing erythroblasts have profound medical implications, as represented by sideroblastic anemia. The synthesis of heme requires the uptake of glycine into the mitochondrial matrix where glycine is condensed with succinyl coenzyme A to yield δ-aminolevulinic acid. Herein we describe the biochemical and molecular characterization of yeast Hem25p and human SLC25A38, providing evidence that they are mitochondrial carriers for glycine. In particular, the hem25Δ mutant manifests a defect in the biosynthesis of δ-aminolevulinic acid and displays reduced levels of downstream heme and mitochondrial cytochromes. The observed defects are rescued by complementation with yeast HEM25 or human SLC25A38 genes. Our results identify new proteins in the heme biosynthetic pathway and demonstrate that Hem25p and its human orthologue SLC25A38 are the main mitochondrial glycine transporters required for heme synthesis, providing definitive evidence of their previously proposed glycine transport function. Furthermore, our work may suggest new therapeutic approaches for the treatment of congenital sideroblastic anemia. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Human Health Hazards from Antimicrobial-Resistant Escherichia coli of Animal Origin

    DEFF Research Database (Denmark)

    Hammerum, A. M.; Heuer, Ole Eske

    2009-01-01

    of antimicrobial agents in food animals may add to the burden of antimicrobial resistance in humans. Bacteria from the animal reservoir that carry resistance to antimicrobial agents that are regarded as highly or critically important in human therapy (e.g., aminoglycosides, fluoroquinolones, and third- and fourth......Because of the intensive use of antimicrobial agents in food animal production, meat is frequently contaminated with antimicrobial-resistant Escherichia coli. Humans can be colonized with E. coli of animal origin, and because of resistance to commonly used antimicrobial agents, these bacteria may...... cause infections for which limited therapeutic options are available. This may lead to treatment failure and can have serious consequences for the patient. Furthermore, E. coli of animal origin may act as a donor of antimicrobial resistance genes for other pathogenic E. coli. Thus, the intensive use...

  4. Systematic high-yield production of human secreted proteins in Escherichia coli

    International Nuclear Information System (INIS)

    Dai Xueyu; Chen Qiang; Lian Min; Zhou Yanfeng; Zhou Mo; Lu Shanyun; Chen Yunjia; Luo Jingchu; Gu Xiaocheng; Jiang Ying; Luo Ming; Zheng Xiaofeng

    2005-01-01

    Human secreted proteins play a very important role in signal transduction. In order to study all potential secreted proteins identified from the human genome sequence, systematic production of large amounts of biologically active secreted proteins is a prerequisite. We selected 25 novel genes as a trial case for establishing a reliable expression system to produce active human secreted proteins in Escherichia coli. Expression of proteins with or without signal peptides was examined and compared in E. coli strains. The results indicated that deletion of signal peptides, to a certain extent, can improve the expression of these proteins and their solubilities. More importantly, under expression conditions such as induction temperature, N-terminus fusion peptides need to be optimized in order to express adequate amounts of soluble proteins. These recombinant proteins were characterized as well-folded proteins. This system enables us to rapidly obtain soluble and highly purified human secreted proteins for further functional studies

  5. Human NKCC2 cation–Cl– co-transporter complements lack of Vhc1 transporter in yeast vacuolar membranes.

    Science.gov (United States)

    Petrezselyova, Silvia; Dominguez, Angel; Herynkova, Pavla; Macias, Juan F; Sychrova, Hana

    2013-10-01

    Cation–chloride co-transporters serve to transport Cl– and alkali metal cations. Whereas a large family of these exists in higher eukaryotes, yeasts only possess one cation–chloride co-transporter, Vhc1, localized to the vacuolar membrane. In this study, the human cation–chloride co-transporter NKCC2 complemented the phenotype of VHC1 deletion in Saccharomyces cerevisiae and its activity controlled the growth of salt-sensitive yeast cells in the presence of high KCl, NaCl and LiCl. A S. cerevisiae mutant lacking plasma-membrane alkali–metal cation exporters Nha1 and Ena1-5 and the vacuolar cation–chloride co-transporter Vhc1 is highly sensitive to increased concentrations of alkali–metal cations, and it proved to be a suitable model for characterizing the substrate specificity and transport activity of human wild-type and mutated cation–chloride co-transporters. Copyright © 2013 John Wiley & Sons, Ltd.

  6. Pathogenic Potential to Humans of Bovine Escherichia coli O26, Scotland

    Science.gov (United States)

    Rosser, Tracy; Allison, Lesley J.; Courcier, Emily; Evans, Judith; McKendrick, Iain J.; Pearce, Michael C.; Handel, Ian; Caprioli, Alfredo; Karch, Helge; Hanson, Mary F.; Pollock, Kevin G.J.; Locking, Mary E.; Woolhouse, Mark E.J.; Matthews, Louise; Low, J. Chris; Gally, David L.

    2012-01-01

    Escherichia coli O26 and O157 have similar overall prevalences in cattle in Scotland, but in humans, Shiga toxin–producing E. coli O26 infections are fewer and clinically less severe than E. coli O157 infections. To investigate this discrepancy, we genotyped E. coli O26 isolates from cattle and humans in Scotland and continental Europe. The genetic background of some strains from Scotland was closely related to that of strains causing severe infections in Europe. Nonmetric multidimensional scaling found an association between hemolytic uremic syndrome (HUS) and multilocus sequence type 21 strains and confirmed the role of stx2 in severe human disease. Although the prevalences of E. coli O26 and O157 on cattle farms in Scotland are equivalent, prevalence of more virulent strains is low, reducing human infection risk. However, new data on E. coli O26–associated HUS in humans highlight the need for surveillance of non-O157 enterohemorrhagic E. coli and for understanding stx2 phage acquisition. PMID:22377426

  7. Lager Yeast Comes of Age

    Science.gov (United States)

    2014-01-01

    Alcoholic fermentations have accompanied human civilizations throughout our history. Lager yeasts have a several-century-long tradition of providing fresh beer with clean taste. The yeast strains used for lager beer fermentation have long been recognized as hybrids between two Saccharomyces species. We summarize the initial findings on this hybrid nature, the genomics/transcriptomics of lager yeasts, and established targets of strain improvements. Next-generation sequencing has provided fast access to yeast genomes. Its use in population genomics has uncovered many more hybridization events within Saccharomyces species, so that lager yeast hybrids are no longer the exception from the rule. These findings have led us to propose network evolution within Saccharomyces species. This “web of life” recognizes the ability of closely related species to exchange DNA and thus drain from a combined gene pool rather than be limited to a gene pool restricted by speciation. Within the domesticated lager yeasts, two groups, the Saaz and Frohberg groups, can be distinguished based on fermentation characteristics. Recent evidence suggests that these groups share an evolutionary history. We thus propose to refer to the Saaz group as Saccharomyces carlsbergensis and to the Frohberg group as Saccharomyces pastorianus based on their distinct genomes. New insight into the hybrid nature of lager yeast will provide novel directions for future strain improvement. PMID:25084862

  8. Biomedical applications of yeast- a patent view, part one: yeasts as workhorses for the production of therapeutics and vaccines.

    Science.gov (United States)

    Roohvand, Farzin; Shokri, Mehdi; Abdollahpour-Alitappeh, Meghdad; Ehsani, Parastoo

    2017-08-01

    Yeasts, as Eukaryotes, offer unique features for ease of growth and genetic manipulation possibilities, making it an exceptional microbial host. Areas covered: This review provides general and patent-oriented insights into production of biopharmaceuticals by yeasts. Patents, wherever possible, were correlated to the original or review articles. The review describes applications of major GRAS (generally regarded as safe) yeasts for the production of therapeutic proteins and subunit vaccines; additionally, immunomodulatory properties of yeast cell wall components were reviewed for use of whole yeast cells as a new vaccine platform. The second part of the review will discuss yeast- humanization strategies and innovative applications. Expert opinion: Biomedical applications of yeasts were initiated by utilization of Saccharomyces cerevisiae, for production of leavened (fermented) products, and advanced to serve to produce biopharmaceuticals. Higher biomass production and expression/secretion yields, more similarity of glycosylation patterns to mammals and possibility of host-improvement strategies through application of synthetic biology might enhance selection of Pichia pastoris (instead of S. cerevisiae) as a host for production of biopharmaceutical in future. Immunomodulatory properties of yeast cell wall β-glucans and possibility of intracellular expression of heterologous pathogen/tumor antigens in yeast cells have expanded their application as a new platform, 'Whole Yeast Vaccines'.

  9. Yeast Augmented Network Analysis (YANA: a new systems approach to identify therapeutic targets for human genetic diseases [v1; ref status: indexed, http://f1000r.es/3gk

    Directory of Open Access Journals (Sweden)

    David J. Wiley

    2014-06-01

    Full Text Available Genetic interaction networks that underlie most human diseases are highly complex and poorly defined. Better-defined networks will allow identification of a greater number of therapeutic targets. Here we introduce our Yeast Augmented Network Analysis (YANA approach and test it with the X-linked spinal muscular atrophy (SMA disease gene UBA1. First, we express UBA1 and a mutant variant in fission yeast and use high-throughput methods to identify fission yeast genetic modifiers of UBA1. Second, we analyze available protein-protein interaction network databases in both fission yeast and human to construct UBA1 genetic networks. Third, from these networks we identified potential therapeutic targets for SMA. Finally, we validate one of these targets in a vertebrate (zebrafish SMA model. This study demonstrates the power of combining synthetic and chemical genetics with a simple model system to identify human disease gene networks that can be exploited for treating human diseases.

  10. Fermentation of sweet whey by recombinant Escherichia coli K011 Fermentação de soro de leite por Escherichia coli KO11 recombinante

    Directory of Open Access Journals (Sweden)

    Amarildo Ricardo Leite

    2000-09-01

    Full Text Available The production of ethanol from sweet whey using the recombinant Escherichia coli KO11, in batch fermentation, was tested. The maximum ethanol yield was reached after 96h, representing only 38% of the theoretical yield. The supplementation of whey with components of LB broth increased the maximum yield to 96% in 72h. The addition of 0.5% yeast extract to whey resulted in maximum yield of 74% at 36h and it increased to over 100% when yeast extract and trace metals solution (Fe++, Mn++ and Zn++ were added.A produção de etanol a partir de soro de leite empregando a cepa Escherichia coli KO11 recombinante, em fermentação de batelada, foi testada. O rendimento máximo de etanol foi obtido em 96h, representando apenas 38% do rendimento teórico. A suplementação do soro com os componentes do caldo LB aumentou o rendimento para 96% em 72h. A adição de 0,5% de extrato de levedura ao soro resultou em um rendimento máximo de 74% em 36h que aumentou para acima de 100% quando se adicionou extrato de levedura e uma solução de metais traço (Fe++, Mn++ e Zn++.

  11. The Genome of the Basidiomycetous Yeast and Human Pathogen Cryptococcus neoformans

    Science.gov (United States)

    Loftus, Brendan J.; Fung, Eula; Roncaglia, Paola; Rowley, Don; Amedeo, Paolo; Bruno, Dan; Vamathevan, Jessica; Miranda, Molly; Anderson, Iain J.; Fraser, James A.; Allen, Jonathan E.; Bosdet, Ian E.; Brent, Michael R.; Chiu, Readman; Doering, Tamara L.; Donlin, Maureen J.; D’Souza, Cletus A.; Fox, Deborah S.; Grinberg, Viktoriya; Fu, Jianmin; Fukushima, Marilyn; Haas, Brian J.; Huang, James C.; Janbon, Guilhem; Jones, Steven J. M.; Koo, Hean L.; Krzywinski, Martin I.; Kwon-Chung, June K.; Lengeler, Klaus B.; Maiti, Rama; Marra, Marco A.; Marra, Robert E.; Mathewson, Carrie A.; Mitchell, Thomas G.; Pertea, Mihaela; Riggs, Florenta R.; Salzberg, Steven L.; Schein, Jacqueline E.; Shvartsbeyn, Alla; Shin, Heesun; Shumway, Martin; Specht, Charles A.; Suh, Bernard B.; Tenney, Aaron; Utterback, Terry R.; Wickes, Brian L.; Wortman, Jennifer R.; Wye, Natasja H.; Kronstad, James W.; Lodge, Jennifer K.; Heitman, Joseph; Davis, Ronald W.; Fraser, Claire M.; Hyman, Richard W.

    2012-01-01

    Cryptococcus neoformans is a basidiomycetous yeast ubiquitous in the environment, a model for fungal pathogenesis, and an opportunistic human pathogen of global importance. We have sequenced its ~20-megabase genome, which contains ~6500 intron-rich gene structures and encodes a transcriptome abundant in alternatively spliced and antisense messages. The genome is rich in transposons, many of which cluster at candidate centromeric regions. The presence of these transposons may drive karyotype instability and phenotypic variation. C. neoformans encodes unique genes that may contribute to its unusual virulence properties, and comparison of two phenotypically distinct strains reveals variation in gene content in addition to sequence polymorphisms between the genomes. PMID:15653466

  12. Accelerating Yeast Prion Biology using Droplet Microfluidics

    Science.gov (United States)

    Ung, Lloyd; Rotem, Assaf; Jarosz, Daniel; Datta, Manoshi; Lindquist, Susan; Weitz, David

    2012-02-01

    Prions are infectious proteins in a misfolded form, that can induce normal proteins to take the misfolded state. Yeast prions are relevant, as a model of human prion diseases, and interesting from an evolutionary standpoint. Prions may also be a form of epigenetic inheritance, which allow yeast to adapt to stressful conditions at rates exceeding those of random mutations and propagate that adaptation to their offspring. Encapsulation of yeast in droplet microfluidic devices enables high-throughput measurements with single cell resolution, which would not be feasible using bulk methods. Millions of populations of yeast can be screened to obtain reliable measurements of prion induction and loss rates. The population dynamics of clonal yeast, when a fraction of the cells are prion expressing, can be elucidated. Furthermore, the mechanism by which certain strains of bacteria induce yeast to express prions in the wild can be deduced. Integrating the disparate fields of prion biology and droplet microfluidics reveals a more complete picture of how prions may be more than just diseases and play a functional role in yeast.

  13. Molecular relatedness of ESBL/AmpC-producing Escherichia coli from humans, animals, food and the environment : a pooled analysis

    NARCIS (Netherlands)

    Dorado-García, Alejandro|info:eu-repo/dai/nl/372621023; Smid, Joost H|info:eu-repo/dai/nl/313996458; van Pelt, Wilfrid; Bonten, Marc J M; Fluit, Ad C; van den Bunt, Gerrita; Wagenaar, Jaap A|info:eu-repo/dai/nl/126613354; Hordijk, Joost|info:eu-repo/dai/nl/314839542; Dierikx, Cindy M; Veldman, Kees T; de Koeijer, Aline; Dohmen, Wietske|info:eu-repo/dai/nl/333690451; Schmitt, Heike|info:eu-repo/dai/nl/304831042; Liakopoulos, Apostolos; Pacholewicz, Ewa; Lam, Theo J G M|info:eu-repo/dai/nl/14686820X; Velthuis, Annet G J; Heuvelink, Annet; Gonggrijp, Maaike A; van Duijkeren, Engeline; van Hoek, Angela H A M; de Roda Husman, Ana Maria|info:eu-repo/dai/nl/139498281; Blaak, Hetty; Havelaar, Arie H|info:eu-repo/dai/nl/072306122; Mevius, Dik J|info:eu-repo/dai/nl/079677347; Heederik, Dick J J|info:eu-repo/dai/nl/072910542

    Background: In recent years, ESBL/AmpC-producing Escherichia coli (ESBL/AmpC-EC) have been isolated with increasing frequency from animals, food, environmental sources and humans. With incomplete and scattered evidence, the contribution to the human carriage burden from these reservoirs remains

  14. Biodegradation of dioxins by recombinant Escherichia coli expressing rat CYP1A1 or its mutant

    Energy Technology Data Exchange (ETDEWEB)

    Shinkyo, Raku; Inouye, Kuniyo [Kyoto Univ. (Japan). Div. of Food Science and Biotechnology; Kamakura, Masaki; Ikushiro, Shin-ichi; Sakaki, Toshiyuki [Toyama Prefectural Univ. (Japan). Biotechnology Research Center

    2006-09-15

    Among polychlorinated dibenzo-p-dioxins (PCDDs), 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TetraCDD) is the most toxic one. Recently, we reported that rat CYP1A1 mutant, F240A, expressed in yeast showed metabolic activity toward 2,3,7,8-TetraCDD. In this study, we successfully expressed N-terminal truncated P450s ({delta}1A1 and {delta}F240A) in Escherichia coli cells. Kinetic analysis using membrane fractions prepared from the recombinant E. coli cells revealed that {delta}F240A has enzymatic properties similar to F240A expressed in yeast. The metabolism of PCDDs by recombinant E. coli cells expressing both {delta}F240A and human NADPH-P450 reductase was also examined. When 2,3,7-TriCDD was added to the E. coli cell culture at a final concentration of 10 {mu}M, approximately 90% of the 2,3,7-TriCDD was converted into multiple metabolites within 8 h. These results indicate the possible application of prokaryotic cells expressing {delta}F240A to the bioremediation of PCDD-contaminated soil. (orig.)

  15. Mitochondrial fission proteins regulate programmed cell death in yeast.

    Science.gov (United States)

    Fannjiang, Yihru; Cheng, Wen-Chih; Lee, Sarah J; Qi, Bing; Pevsner, Jonathan; McCaffery, J Michael; Hill, R Blake; Basañez, Gorka; Hardwick, J Marie

    2004-11-15

    The possibility that single-cell organisms undergo programmed cell death has been questioned in part because they lack several key components of the mammalian cell death machinery. However, yeast encode a homolog of human Drp1, a mitochondrial fission protein that was shown previously to promote mammalian cell death and the excessive mitochondrial fragmentation characteristic of apoptotic mammalian cells. In support of a primordial origin of programmed cell death involving mitochondria, we found that the Saccharomyces cerevisiae homolog of human Drp1, Dnm1, promotes mitochondrial fragmentation/degradation and cell death following treatment with several death stimuli. Two Dnm1-interacting factors also regulate yeast cell death. The WD40 repeat protein Mdv1/Net2 promotes cell death, consistent with its role in mitochondrial fission. In contrast to its fission function in healthy cells, Fis1 unexpectedly inhibits Dnm1-mediated mitochondrial fission and cysteine protease-dependent cell death in yeast. Furthermore, the ability of yeast Fis1 to inhibit mitochondrial fission and cell death can be functionally replaced by human Bcl-2 and Bcl-xL. Together, these findings indicate that yeast and mammalian cells have a conserved programmed death pathway regulated by a common molecular component, Drp1/Dnm1, that is inhibited by a Bcl-2-like function.

  16. Bioadsorption of cadmium ion by cell surface-engineered yeasts displaying metallothionein and hexa-His

    Energy Technology Data Exchange (ETDEWEB)

    Kuroda, K.; Ueda, M. [Lab. of Applied Biological Chemistry, Kyoto Univ., Yoshida, Kyoto (Japan)

    2004-07-01

    The Cd{sup 2+}-chelating abilities of yeast metallothionein (YMT) and hexa-His displayed on the yeast-cell surface were compared. Display of YMT and hexa-His by {alpha}-agglutinin-based cell-surface engineering was confirmed by immunofluorescent labeling. Surface-engineered yeast cells with YMT and hexa-His fused in tandem showed superior cell-surface adsorption and recovery of Cd{sup 2+} under EDTA treatment on the cell surface than hexa-His-displaying cells. YMT was demonstrated to be more effective than hexa-His for the adsorption of Cd{sup 2+}. Yeast cells displaying YMT and/or hexa-His exhibited a higher potential for the adsorption of Cd{sup 2+} than Escherichia coli cells displaying these molecules. In order to investigate the effect of the displayed YMT and hexa-His on sensitivity to toxic Cd{sup 2+}, growth in Cd{sup 2+}-containing liquid medium was monitored. Unlike hexa-His-displaying cells, cells displaying YMT and hexa-His fused in tandem induced resistance to Cd{sup 2+} through active and enhanced adsorption of toxic Cd{sup 2+}. These results indicate that YMT-displaying yeast cells are a unique bioadsorbent with a functional chelating ability superior to that of E. coli. (orig.)

  17. Killer toxin from a novel killer yeast Pichia kudriavzevii RY55 with idiosyncratic antibacterial activity.

    Science.gov (United States)

    Bajaj, Bijender Kumar; Raina, Sandeepu; Singh, Satbir

    2013-08-01

    The killer phenomenon of yeast may have technological implications in many areas like beverage fermentation, food technology, biological control in agriculture, and in medicine. In the present study the killer phenomenon in Pichia kudriavzevii (P. kudriavzevii RY55) is being reported for the first time. The P. kudriavzevii RY55 toxin exhibited excellent antibacterial activity against several pathogens of human health significance such as Escherichia coli, Enterococcus faecalis, Klebsiella sp., Staphylococcus aureus, Pseudomonas aeruginosa and Pseudomonas alcaligenes. Killer toxin was purified to homogeneity by using ammonium sulphate precipitation and ion exchange chromatography and characterized for few properties. P. kudriavzevii RY55 killer toxin may be of vast significance in the development of novel antimicrobial chemotherapeutic agents, new bio-based safer candidates for food preservation and biocontrol, and starter cultures for fermentation industries. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Lipid raft involvement in yeast cell growth and death

    Energy Technology Data Exchange (ETDEWEB)

    Mollinedo, Faustino, E-mail: fmollin@usal.es [Instituto de Biología Molecular y Celular del Cáncer, Centro de Investigación del Cáncer, Consejo Superior de Investigaciones Científicas - Universidad de Salamanca, Salamanca (Spain)

    2012-10-10

    The notion that cellular membranes contain distinct microdomains, acting as scaffolds for signal transduction processes, has gained considerable momentum. In particular, a class of such domains that is rich in sphingolipids and cholesterol, termed as lipid rafts, is thought to compartmentalize the plasma membrane, and to have important roles in survival and cell death signaling in mammalian cells. Likewise, yeast lipid rafts are membrane domains enriched in sphingolipids and ergosterol, the yeast counterpart of mammalian cholesterol. Sterol-rich membrane domains have been identified in several fungal species, including the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe as well as the pathogens Candida albicans and Cryptococcus neoformans. Yeast rafts have been mainly involved in membrane trafficking, but increasing evidence implicates rafts in a wide range of additional cellular processes. Yeast lipid rafts house biologically important proteins involved in the proper function of yeast, such as proteins that control Na{sup +}, K{sup +}, and pH homeostasis, which influence many cellular processes, including cell growth and death. Membrane raft constituents affect drug susceptibility, and drugs interacting with sterols alter raft composition and membrane integrity, leading to yeast cell death. Because of the genetic tractability of yeast, analysis of yeast rafts could be an excellent model to approach unanswered questions of mammalian raft biology, and to understand the role of lipid rafts in the regulation of cell death and survival in human cells. A better insight in raft biology might lead to envisage new raft-mediated approaches to the treatment of human diseases where regulation of cell death and survival is critical, such as cancer and neurodegenerative diseases.

  19. Lipid raft involvement in yeast cell growth and death

    International Nuclear Information System (INIS)

    Mollinedo, Faustino

    2012-01-01

    The notion that cellular membranes contain distinct microdomains, acting as scaffolds for signal transduction processes, has gained considerable momentum. In particular, a class of such domains that is rich in sphingolipids and cholesterol, termed as lipid rafts, is thought to compartmentalize the plasma membrane, and to have important roles in survival and cell death signaling in mammalian cells. Likewise, yeast lipid rafts are membrane domains enriched in sphingolipids and ergosterol, the yeast counterpart of mammalian cholesterol. Sterol-rich membrane domains have been identified in several fungal species, including the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe as well as the pathogens Candida albicans and Cryptococcus neoformans. Yeast rafts have been mainly involved in membrane trafficking, but increasing evidence implicates rafts in a wide range of additional cellular processes. Yeast lipid rafts house biologically important proteins involved in the proper function of yeast, such as proteins that control Na + , K + , and pH homeostasis, which influence many cellular processes, including cell growth and death. Membrane raft constituents affect drug susceptibility, and drugs interacting with sterols alter raft composition and membrane integrity, leading to yeast cell death. Because of the genetic tractability of yeast, analysis of yeast rafts could be an excellent model to approach unanswered questions of mammalian raft biology, and to understand the role of lipid rafts in the regulation of cell death and survival in human cells. A better insight in raft biology might lead to envisage new raft-mediated approaches to the treatment of human diseases where regulation of cell death and survival is critical, such as cancer and neurodegenerative diseases.

  20. Mammalian amyloidogenic proteins promote prion nucleation in yeast.

    Science.gov (United States)

    Chandramowlishwaran, Pavithra; Sun, Meng; Casey, Kristin L; Romanyuk, Andrey V; Grizel, Anastasiya V; Sopova, Julia V; Rubel, Aleksandr A; Nussbaum-Krammer, Carmen; Vorberg, Ina M; Chernoff, Yury O

    2018-03-02

    Fibrous cross-β aggregates (amyloids) and their transmissible forms (prions) cause diseases in mammals (including humans) and control heritable traits in yeast. Initial nucleation of a yeast prion by transiently overproduced prion-forming protein or its (typically, QN-rich) prion domain is efficient only in the presence of another aggregated (in most cases, QN-rich) protein. Here, we demonstrate that a fusion of the prion domain of yeast protein Sup35 to some non-QN-rich mammalian proteins, associated with amyloid diseases, promotes nucleation of Sup35 prions in the absence of pre-existing aggregates. In contrast, both a fusion of the Sup35 prion domain to a multimeric non-amyloidogenic protein and the expression of a mammalian amyloidogenic protein that is not fused to the Sup35 prion domain failed to promote prion nucleation, further indicating that physical linkage of a mammalian amyloidogenic protein to the prion domain of a yeast protein is required for the nucleation of a yeast prion. Biochemical and cytological approaches confirmed the nucleation of protein aggregates in the yeast cell. Sequence alterations antagonizing or enhancing amyloidogenicity of human amyloid-β (associated with Alzheimer's disease) and mouse prion protein (associated with prion diseases), respectively, antagonized or enhanced nucleation of a yeast prion by these proteins. The yeast-based prion nucleation assay, developed in our work, can be employed for mutational dissection of amyloidogenic proteins. We anticipate that it will aid in the identification of chemicals that influence initial amyloid nucleation and in searching for new amyloidogenic proteins in a variety of proteomes. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Identification of the yeast gene encoding the tRNA m1G methyltransferase responsible for modification at position 9.

    Science.gov (United States)

    Jackman, Jane E; Montange, Rebecca K; Malik, Harmit S; Phizicky, Eric M

    2003-05-01

    Methylation of tRNA at the N-1 position of guanosine to form m(1)G occurs widely in nature. It occurs at position 37 in tRNAs from all three kingdoms, and the methyltransferase that catalyzes this reaction is known from previous work of others to be critically important for cell growth in Escherichia coli and the yeast Saccharomyces cerevisiae. m(1)G is also widely found at position 9 in eukaryotic tRNAs, but the corresponding methyltransferase was unknown. We have used a biochemical genomics approach with a collection of purified yeast GST-ORF fusion proteins to show that m(1)G(9) formation of yeast tRNA(Gly) is associated with ORF YOL093w, named TRM10. Extracts lacking Trm10p have undetectable levels of m(1)G(9) methyltransferase activity but retain normal m(1)G(37) methyltransferase activity. Yeast Trm10p purified from E. coli quantitatively modifies the G(9) position of tRNA(Gly) in an S-adenosylmethionine-dependent fashion. Trm10p is responsible in vivo for most if not all m(1)G(9) modification of tRNAs, based on two results: tRNA(Gly) purified from a trm10-Delta/trm10-Delta strain is lacking detectable m(1)G; and a primer extension block occurring at m(1)G(9) is removed in trm10-Delta/trm10-Delta-derived tRNAs for all 9 m(1)G(9)-containing species that were testable by this method. There is no obvious growth defect of trm10-Delta/trm10-Delta strains. Trm10p bears no detectable resemblance to the yeast m(1)G(37) methyltransferase, Trm5p, or its orthologs. Trm10p homologs are found widely in eukaryotes and many archaea, with multiple homologs in several metazoans, including at least three in humans.

  2. Production and Purification of the Native Saccharomyces cerevisiae Hsp12 in Escherichia coli.

    Science.gov (United States)

    Léger, Antoine; Hocquellet, Agnès; Dieryck, Wilfrid; Moine, Virginie; Marchal, Axel; Marullo, Philippe; Josseaume, Annabelle; Cabanne, Charlotte

    2017-09-20

    Hsp12 is a small heat shock protein produced in many organisms, including the yeast Saccharomyces cerevisiae. It has been described as an indicator of yeast stress rate and has also been linked to the sweetness sensation of wine. To obtain a sufficient amount of protein, we produced and purified Hsp12 without tag in Escherichia coli. A simple fast two-step process was developed using a microplate approach and a design of experiments. A capture step on an anion-exchange salt-tolerant resin was followed by size exclusion chromatography for polishing, leading to a purity of 97%. Thereafter, specific anti-Hsp12 antibodies were obtained by rabbit immunization. An ELISA was developed to quantify Hsp12 in various strains of Saccharomyces cerevisiae. The antibodies showed high specificity and allowed the quantitation of Hsp12 in the yeast. The quantities of Hsp12 measured in the strains differed in direct proportion to the level of expression found in previous studies.

  3. Pentose utilization in yeasts: Physiology and biochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Jeppson, H.

    1996-04-01

    The fermentive performance of bacteria, yeasts, and filamentous fungi was investigated in a pentose (xylose)-rich lignocellulosic hydrolyzate. The filamentous fungus Fusarium oxysporum and the xylose-fermenting yeast Pichia stipitis were found to be very sensitive to the inhibiting hydrolyzate. Recombinant xylose-utilizing Saccharomyces cerevisiae showed very poor ethanol formation from xylose; xylitol being the major product formed. The highest ethanol yields were obtained with recombinant Escherichia coli KO11, however, for maximal ethanol yield detoxification of the hydrolyzate was required. The influence of oxygen on the regulation of carbohydrate metabolism in the xylose-fermenting yeast P. stipitis CBS 6054 was investigated. A low and well-controlled level of oxygenation has been found to be required for efficient ethanol formation from xylose by the xylose-fermenting yeasts. The requirement of oxygen is frequently ascribed to the apparent redox imbalance which develops under anaerobic conditions due to the difference in co-factor utilization of the two first enzymes in the xylose metabolism, further reflected in xylitol excretion. However, a low and well controlled level of oxygenation for maximal ethanol production from glucose was also demonstrated, suggesting that the oxygen requirement is not only due to the dual co-factor utilization, but also serves other purposes. Cyanide-insensitive and salicyl hydroxamic acid-sensitive respiration (CIR) was found in P. stipitis. CIR is suggested to act as a redox sink preventing xylitol formation in P. stipitis under oxygen-limited xylose fermentations. Xylitol metabolism by P. stipitis CBS 6054 was strictly respiratory and ethanol was not formed under any conditions. The absence of ethanol formation was not due to a lack of fermentative enzymes, since the addition of glucose to xylitol-pregrown cells resulted in ethanol formation. 277 refs, 5 figs, 7 tabs

  4. Metabolic Modeling of Common Escherichia coli Strains in Human Gut Microbiome

    Directory of Open Access Journals (Sweden)

    Yue-Dong Gao

    2014-01-01

    Full Text Available The recent high-throughput sequencing has enabled the composition of Escherichia coli strains in the human microbial community to be profiled en masse. However, there are two challenges to address: (1 exploring the genetic differences between E. coli strains in human gut and (2 dynamic responses of E. coli to diverse stress conditions. As a result, we investigated the E. coli strains in human gut microbiome using deep sequencing data and reconstructed genome-wide metabolic networks for the three most common E. coli strains, including E. coli HS, UTI89, and CFT073. The metabolic models show obvious strain-specific characteristics, both in network contents and in behaviors. We predicted optimal biomass production for three models on four different carbon sources (acetate, ethanol, glucose, and succinate and found that these stress-associated genes were involved in host-microbial interactions and increased in human obesity. Besides, it shows that the growth rates are similar among the models, but the flux distributions are different, even in E. coli core reactions. The correlations between human diabetes-associated metabolic reactions in the E. coli models were also predicted. The study provides a systems perspective on E. coli strains in human gut microbiome and will be helpful in integrating diverse data sources in the following study.

  5. Yeast tRNAPhe expressed in human cells can be selected by HIV-1 for use as a reverse transcription primer

    International Nuclear Information System (INIS)

    Kelly, Nathan J.; Morrow, Casey D.

    2003-01-01

    All naturally occurring human immune deficiency viruses (HIV-1) select and use tRNA Lys,3 as the primer for reverse transcription. Studies to elucidate the mechanism of tRNA selection from the intracellular milieu have been hampered due to the difficulties in manipulating the endogenous levels of tRNA Lys,3 . We have previously described a mutant HIV-1 with a primer binding site (PBS) complementary to yeast tRNA Phe (psHIV-Phe) that relies on transfection of yeast tRNA Phe for infectivity. To more accurately recapitulate the selection process, a cDNA was designed for the intracellular expression of the yeast tRNA Phe . Increasing amounts of the plasmid encoding tRNA Phe resulted in a corresponding increase in levels of yeast tRNA Phe in the cell. The yeast tRNA Phe isolated from cells transfected with the cDNA for yeast tRNA Phe , or in the cell lines expressing yeast tRNA Phe , were aminoacylated, indicating that the expressed yeast tRNA Phe was incorporated into tRNA biogenesis pathways and translation. Increasing the cytoplasmic levels of tRNA Phe resulted in increased encapsidation of tRNA Phe in viruses with a PBS complementary to tRNA Phe (psHIV-Phe) or tRNA Lys,3 (wild-type HIV-1). Production of infectious psHIV-Phe was dependent on the amount of cotransfected tRNA Phe cDNA. Increasing amounts of plasmids encoding yeast tRNA Phe produced an increase of infectious psHIV-Phe that plateaued at a level lower than that from the transfection of the wild-type genome, which uses tRNA Lys,3 as the primer for reverse transcription. Cell lines were generated that expressed yeast tRNA Phe at levels approximately 0.1% of that for tRNA Lys,3 . Even with this reduced level of yeast tRNA Phe , the cell lines complemented psHIV-Phe over background levels. The results of these studies demonstrate that intracellular levels of primer tRNA can have a direct effect on HIV-1 infectivity and further support the role for PBS-tRNA complementarity in the primer selection process

  6. Yeasts: providing questions and answers for modern biology.

    Science.gov (United States)

    Dickinson, J R

    2000-01-01

    Yeasts are to be found in virtually every conceivable niche on this planet and are amazingly varied in their shapes ('morphologies'), life cycles, metabolic capabilities, potentials for use in industrial processes, abilities to spoil food and drink or to act as dangerous human pathogens. This review describes four very different species of yeast to illustrate some of the diversity which exists and, in the case of one of them, Saccharomyces cerevisiae (the familiar baker's or brewer's yeast), the extent of both our knowledge and ignorance.

  7. Whole genome sequencing of Escherichia coli encoding blaNDM isolated from humans and companion animals in Egypt

    Science.gov (United States)

    Companion animals are a source of zoonotic infections and especially important considering the potential of companion animals to harbor antibiotic resistant pathogens. In this study, blaNDM positive Escherichia coli from companion animals, humans, and the environment from Mansoura, Egypt were charac...

  8. Building Better Biosensors for Exploration into Deep-Space, Using Humanized Yeast

    Science.gov (United States)

    Liddell, Lauren; Santa Maria, Sergio; Tieze, Sofia; Bhattacharya, Sharmila

    2017-01-01

    1.BioSentinel is 1 of 13 secondary payloads hitching a ride beyond Low Earth Orbit on Exploration Mission 1 (EM-1), set to launch from NASAs Space Launch System in 2019. EM-1 is our first opportunity to investigate the effects of the deep space environment on a eukaryotic biological system, the budding yeast S. cerevisiae. Though separated by a billion years of evolution we share hundreds of genes important for basic cell function, including responses to DNA damage. Thus, yeast is an ideal biosensor for detecting typesextent of damage induced by deep-space radiation.We will fly desiccated cells, then rehydrate to wake them up when the automated payload is ready to initiate the experiment. Rehydration solution contains SC (Synthetic Complete) media and alamarBlue, an indicator for changes in growth and metabolism. Telemetry of LED readings will then allow us to detect how cells respond throughout the mission. The desiccation-rehydration process can be extremely damaging to cells, and can severely diminish our ability to accurately measure and model cellular responses to deep-space radiation. The aim of this study is to develop a better biosensor: yeast strains that are more resistant to desiccation stress. We will over-express known cellular protectants, including hydrophilin Sip18, the protein disaggregase Hsp104, and thioredoxin Trx2, a responder to oxidative stress, then measure cell viability after desiccation to determine which factors improve stress tolerance. Over-expression of SIP18 in wine yeast starter cultures was previously reported to increase viability following desiccation stress by up to 70. Thus, we expect similar improvements in our space-yeast strains. By designing better yeast biosensors we can better prepare for and mitigate the potential dangers of deep-space radiation for future missions.This work is funded by NASAs AES program.

  9. Oxidative Stress and Programmed Cell Death in Yeast

    International Nuclear Information System (INIS)

    Farrugia, Gianluca; Balzan, Rena

    2012-01-01

    Yeasts, such as Saccharomyces cerevisiae, have long served as useful models for the study of oxidative stress, an event associated with cell death and severe human pathologies. This review will discuss oxidative stress in yeast, in terms of sources of reactive oxygen species (ROS), their molecular targets, and the metabolic responses elicited by cellular ROS accumulation. Responses of yeast to accumulated ROS include upregulation of antioxidants mediated by complex transcriptional changes, activation of pro-survival pathways such as mitophagy, and programmed cell death (PCD) which, apart from apoptosis, includes pathways such as autophagy and necrosis, a form of cell death long considered accidental and uncoordinated. The role of ROS in yeast aging will also be discussed.

  10. Probiotic properties of yeasts occurring in fermented food and beverages

    DEFF Research Database (Denmark)

    Jespersen, Lene

    Besides being able to improve the quality and safety of many fermented food and beverages some yeasts offer a number of probiotic traits. Especially a group of yeast referred to as "Saccharomyces boulardii", though taxonomically belonging to Saccharomyces cerevisiae, has been claimed to have...... probiotic properties. Besides, yeasts naturally occurring globally in food and beverages will have traits that might have a positive impact on human health....

  11. Discovery of a nucleocytoplasmic O-mannose glycoproteome in yeast

    DEFF Research Database (Denmark)

    Halim, Adnan; Larsen, Ida Signe Bohse; Neubert, Patrick

    2015-01-01

    developed a sensitive lectin enrichment and mass spectrometry workflow for identification of the human O-linked mannose (O-Man) glycoproteome and used this to identify a pleothora of O-Man glycoproteins in human cell lines including the large family of cadherins and protocadherins. Here, we applied...... the workflow to yeast with the aim to characterize the yeast O-Man glycoproteome, and in doing so, we discovered hitherto unknown O-Man glycosites on nuclear, cytoplasmic, and mitochondrial proteins in S. cerevisiae and S. pombe. Such O-Man glycoproteins were not found in our analysis of human cell lines....... However, the type of yeast O-Man nucleocytoplasmic proteins and the localization of identified O-Man residues mirror that of the O-GlcNAc glycoproteome found in other eukaryotic cells, indicating that the two different types of O-glycosylations serve the same important biological functions. The discovery...

  12. Yeast biotechnology: teaching the old dog new tricks.

    Science.gov (United States)

    Mattanovich, Diethard; Sauer, Michael; Gasser, Brigitte

    2014-03-06

    Yeasts are regarded as the first microorganisms used by humans to process food and alcoholic beverages. The technology developed out of these ancient processes has been the basis for modern industrial biotechnology. Yeast biotechnology has gained great interest again in the last decades. Joining the potentials of genomics, metabolic engineering, systems and synthetic biology enables the production of numerous valuable products of primary and secondary metabolism, technical enzymes and biopharmaceutical proteins. An overview of emerging and established substrates and products of yeast biotechnology is provided and discussed in the light of the recent literature.

  13. Escherichia coli pathotypes

    Science.gov (United States)

    Escherichia coli strains are important commensals of the intestinal tract of humans and animals; however, pathogenic strains, including diarrhea-inducing E. coli and extraintestinal pathogenic E. coli. Intestinal E. coli pathotypes may cause a dehydrating watery diarrhea, or more severe diseases su...

  14. Yeasts and coliform bacteria of water accumulated in bromeliads of mangrove and sand dune ecosystems of southeast Brazil.

    Science.gov (United States)

    Hagler, A N; Rosa, C A; Morais, P B; Mendonça-Hagler, L C; Franco, G M; Araujo, F V; Soares, C A

    1993-10-01

    Yeasts and coliform bacteria were isolated from water that accumulated in the central cups and adjacent leaf axilae of two bromeliads, Neoregelia cruenta of a coastal sand dune and Quesnelia quesneliana of a mangrove ecosystem near the city of Rio de Janeiro, Brazil. The mean total coliform counts were above 10,000 per 100 mL for waters of both plants, but the mean fecal coliform counts were only 74 per 100 mL for Q. quesneliana and mostly undetected in water from N. cruenta. Of 90 fecal coliform isolates, 51 were typical of Escherichia coli in colony morphology and indol, methyl red, Volges-Proskauer, and citrate (IMViC) tests. Seven representatives of the typical E. coli cultures were identified as this species, but the identifications of nine other coliform bacteria were mostly dubious. The yeast community of N. cruenta was typical of plant surfaces with basidiomycetous yeasts anamorphs, and the black yeast Aureobasidium pullulans was prevalent. Quesnelia quesneliana had a substantial proportion of ascomycetous yeasts and their anamorphs, including a probable new biotype of Saccharomyces unisporus. Our results suggested that the microbial communities in bromeliad waters are typically autochtonous and not contaminants.

  15. Biotechnology of non-Saccharomyces yeasts-the basidiomycetes.

    Science.gov (United States)

    Johnson, Eric A

    2013-09-01

    Yeasts are the major producer of biotechnology products worldwide, exceeding production in capacity and economic revenues of other groups of industrial microorganisms. Yeasts have wide-ranging fundamental and industrial importance in scientific, food, medical, and agricultural disciplines (Fig. 1). Saccharomyces is the most important genus of yeast from fundamental and applied perspectives and has been expansively studied. Non-Saccharomyces yeasts (non-conventional yeasts) including members of the Ascomycetes and Basidiomycetes also have substantial current utility and potential applicability in biotechnology. In an earlier mini-review, "Biotechnology of non-Saccharomyces yeasts-the ascomycetes" (Johnson Appl Microb Biotechnol 97: 503-517, 2013), the extensive biotechnological utility and potential of ascomycetous yeasts are described. Ascomycetous yeasts are particularly important in food and ethanol formation, production of single-cell protein, feeds and fodder, heterologous production of proteins and enzymes, and as model and fundamental organisms for the delineation of genes and their function in mammalian and human metabolism and disease processes. In contrast, the roles of basidiomycetous yeasts in biotechnology have mainly been evaluated only in the past few decades and compared to the ascomycetous yeasts and currently have limited industrial utility. From a biotechnology perspective, the basidiomycetous yeasts are known mainly for the production of enzymes used in pharmaceutical and chemical synthesis, for production of certain classes of primary and secondary metabolites such as terpenoids and carotenoids, for aerobic catabolism of complex carbon sources, and for bioremediation of environmental pollutants and xenotoxicants. Notwithstanding, the basidiomycetous yeasts appear to have considerable potential in biotechnology owing to their catabolic utilities, formation of enzymes acting on recalcitrant substrates, and through the production of unique primary

  16. p53 inhibits autophagy by interacting with the human ortholog of yeast Atg17, RB1CC1/FIP200.

    Science.gov (United States)

    Morselli, Eugenia; Shen, Shensi; Ruckenstuhl, Christoph; Bauer, Maria Anna; Mariño, Guillermo; Galluzzi, Lorenzo; Criollo, Alfredo; Michaud, Mickael; Maiuri, Maria Chiara; Chano, Tokuhiro; Madeo, Frank; Kroemer, Guido

    2011-08-15

    The tumor suppressor protein p53 tonically suppresses autophagy when it is present in the cytoplasm. This effect is phylogenetically conserved from mammals to nematodes, and human p53 can inhibit autophagy in yeast, as we show here. Bioinformatic investigations of the p53 interactome in relationship to the autophagy-relevant protein network underscored the possible relevance of a direct molecular interaction between p53 and the mammalian ortholog of the essential yeast autophagy protein Atg17, namely RB1-inducible coiled-coil protein 1 (RB1CC1), also called FAK family kinase-interacting protein of 200 KDa (FIP200). Mutational analyses revealed that a single point mutation in p53 (K382R) abolished its capacity to inhibit autophagy upon transfection into p53-deficient human colon cancer or yeast cells. In conditions in which wild-type p53 co-immunoprecipitated with RB1CC1/FIP200, p53 (K382R) failed to do so, underscoring the importance of the physical interaction between these proteins for the control of autophagy. In conclusion, p53 regulates autophagy through a direct molecular interaction with RB1CC1/FIP200, a protein that is essential for the very apical step of autophagy initiation.

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

    DEFF Research Database (Denmark)

    Nielsen, Jens

    2013-01-01

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

  18. Global gene expression profiling of the asymptomatic bacteriuria Escherichia coli strain 83972 in the human urinary tract

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Klemm, Per

    2006-01-01

    Urinary tract infections (UTIs) are an important health problem worldwide, with many million cases each year. Escherichia coli is the most common organism causing UTIs in humans. The asymptomatic bacteriuria E. coli strain 83972 is an excellent colonizer of the human urinary tract, where it causes...... long-term bladder colonization. The strain has been used for prophylactic purposes in patients prone to more severe and recurrent UTIs. For this study, we used DNA microarrays to monitor the expression profile of strain 83972 in the human urinary tract. Significant differences in expression levels were...

  19. Bread, beer and wine: yeast domestication in the Saccharomyces sensu stricto complex.

    Science.gov (United States)

    Sicard, Delphine; Legras, Jean-Luc

    2011-03-01

    Yeasts of the Saccharomyces sensu stricto species complex are able to convert sugar into ethanol and CO(2) via fermentation. They have been used for thousands years by mankind for fermenting food and beverages. In the Neolithic times, fermentations were probably initiated by naturally occurring yeasts, and it is unknown when humans started to consciously add selected yeast to make beer, wine or bread. Interestingly, such human activities gave rise to the creation of new species in the Saccharomyces sensu stricto complex by interspecies hybridization or polyploidization. Within the S. cerevisiae species, they have led to the differentiation of genetically distinct groups according to the food process origin. Although the evolutionary history of wine yeast populations has been well described, the histories of other domesticated yeasts need further investigation. Copyright © 2011 Académie des sciences. Published by Elsevier SAS. All rights reserved.

  20. Untangling the Roles of Anti-Apoptosis in Regulating Programmed Cell Death using Humanized Yeast Cells

    International Nuclear Information System (INIS)

    Clapp, Caitlin; Portt, Liam; Khoury, Chamel; Sheibani, Sara; Eid, Rawan; Greenwood, Matthew; Vali, Hojatollah; Mandato, Craig A.; Greenwood, Michael T.

    2012-01-01

    -apoptosis, we screened a human heart cDNA expression library in yeast cells undergoing PCD due to the conditional expression of a mammalian pro-apoptotic Bax cDNA. Analysis of the multiple Bax suppressors identified revealed several previously known as well as a large number of clones representing potential novel anti-apoptotic sequences. The focus of this review is to report on recent achievements in the use of humanized yeast in genetic screens to identify novel stress-induced PCD suppressors, supporting the use of yeast as a unicellular model organism to elucidate anti-apoptotic and cell survival mechanisms.

  1. Yeast biotechnology: teaching the old dog new tricks

    Science.gov (United States)

    2014-01-01

    Yeasts are regarded as the first microorganisms used by humans to process food and alcoholic beverages. The technology developed out of these ancient processes has been the basis for modern industrial biotechnology. Yeast biotechnology has gained great interest again in the last decades. Joining the potentials of genomics, metabolic engineering, systems and synthetic biology enables the production of numerous valuable products of primary and secondary metabolism, technical enzymes and biopharmaceutical proteins. An overview of emerging and established substrates and products of yeast biotechnology is provided and discussed in the light of the recent literature. PMID:24602262

  2. SNPs altering ammonium transport activity of human Rhesus factors characterized by a yeast-based functional assay.

    Directory of Open Access Journals (Sweden)

    Aude Deschuyteneer

    Full Text Available Proteins of the conserved Mep-Amt-Rh family, including mammalian Rhesus factors, mediate transmembrane ammonium transport. Ammonium is an important nitrogen source for the biosynthesis of amino acids but is also a metabolic waste product. Its disposal in urine plays a critical role in the regulation of the acid/base homeostasis, especially with an acid diet, a trait of Western countries. Ammonium accumulation above a certain concentration is however pathologic, the cytotoxicity causing fatal cerebral paralysis in acute cases. Alteration in ammonium transport via human Rh proteins could have clinical outcomes. We used a yeast-based expression assay to characterize human Rh variants resulting from non synonymous single nucleotide polymorphisms (nsSNPs with known or unknown clinical phenotypes and assessed their ammonium transport efficiency, protein level, localization and potential trans-dominant impact. The HsRhAG variants (I61R, F65S associated to overhydrated hereditary stomatocytosis (OHSt, a disease affecting erythrocytes, proved affected in intrinsic bidirectional ammonium transport. Moreover, this study reveals that the R202C variant of HsRhCG, the orthologue of mouse MmRhcg required for optimal urinary ammonium excretion and blood pH control, shows an impaired inherent ammonium transport activity. Urinary ammonium excretion was RHcg gene-dose dependent in mouse, highlighting MmRhcg as a limiting factor. HsRhCG(R202C may confer susceptibility to disorders leading to metabolic acidosis for instance. Finally, the analogous R211C mutation in the yeast ScMep2 homologue also impaired intrinsic activity consistent with a conserved functional role of the preserved arginine residue. The yeast expression assay used here constitutes an inexpensive, fast and easy tool to screen nsSNPs reported by high throughput sequencing or individual cases for functional alterations in Rh factors revealing potential causal variants.

  3. Genome-wide screening for genes whose deletions confer sensitivity to mutagenic purine base analogs in yeast

    Directory of Open Access Journals (Sweden)

    Kozmin Stanislav G

    2005-06-01

    Full Text Available Abstract Background N-hydroxylated base analogs, such as 6-hydroxylaminopurine (HAP and 2-amino-6-hydroxylaminopurine (AHA, are strong mutagens in various organisms due to their ambiguous base-pairing properties. The systems protecting cells from HAP and related noncanonical purines in Escherichia coli include specialized deoxyribonucleoside triphosphatase RdgB, DNA repair endonuclease V, and a molybdenum cofactor-dependent system. Fewer HAP-detoxification systems have been identified in yeast Saccharomyces cerevisiae and other eukaryotes. Cellular systems protecting from AHA are unknown. In the present study, we performed a genome-wide search for genes whose deletions confer sensitivity to HAP and AHA in yeast. Results We screened the library of yeast deletion mutants for sensitivity to the toxic and mutagenic action of HAP and AHA. We identified novel genes involved in the genetic control of base analogs sensitivity, including genes controlling purine metabolism, cytoskeleton organization, and amino acid metabolism. Conclusion We developed a method for screening the yeast deletion library for sensitivity to the mutagenic and toxic action of base analogs and identified 16 novel genes controlling pathways of protection from HAP. Three of them also protect from AHA.

  4. Lactoferrin release and interleukin-1, interleukin-6, and tumor necrosis factor production by human polymorphonuclear cells stimulated by various lipopolysaccharides: relationship to growth inhibition of Candida albicans.

    OpenAIRE

    Palma, C; Cassone, A; Serbousek, D; Pearson, C A; Djeu, J Y

    1992-01-01

    Lipopolysaccharides (LPSs) from Escherichia coli, Serratia marcescens, and Salmonella typhimurium, at doses from 1 to 100 ng/ml, strongly enhanced growth inhibition of Candida albicans by human polymorphonuclear leukocytes (PMN) in vitro. Flow cytometry analysis demonstrated that LPS markedly augmented phagocytosis of Candida cells by increasing the number of yeasts ingested per neutrophil as well as the number of neutrophils capable of ingesting fungal cells. LPS activation caused augmented ...

  5. Automated thermometric enzyme immunoassay of human proinsulin produced by Escherichia coli.

    Science.gov (United States)

    Birnbaum, S; Bülow, L; Hardy, K; Danielsson, B; Mosbach, K

    1986-10-01

    We have determined and monitored the production and release of human proinsulin by genetically engineered Escherichia coli cells. Several M9 media samples were analyzed sequentially after centrifugation with the aid of a rapid automated flow-through thermometric enzyme-linked immunosorbent assay (TELISA) system. The response time was 7 min after sample injection and a single assay was complete after 13 min. Insulin concentrations in the range of 0.1-50 micrograms/ml could be determined. The TELISA method correlated well with conventional radioimmunoassay determinations. Standard curves were reproducible over a period of several days even when the immobilized antibody column was stored at 25 degrees C in the enzyme thermistor unit. Thus, immediate assay start up was possible.

  6. Human Intestinal Cells Modulate Conjugational Transfer of Multidrug Resistance Plasmids between Clinical Escherichia coli Isolates

    DEFF Research Database (Denmark)

    Machado, Ana Manuel; Sommer, Morten

    2014-01-01

    Bacterial conjugation in the human gut microbiota is believed to play a major role in the dissemination of antibiotic resistance genes and virulence plasmids. However, the modulation of bacterial conjugation by the human host remains poorly understood and there is a need for controlled systems...... to study this process. We established an in vitro co-culture system to study the interaction between human intestinal cells and bacteria. We show that the conjugation efficiency of a plasmid encoding an extended spectrum beta-lactamase is reduced when clinical isolates of Escherichia coli are co...... of the intestinal cells exposed to bacteria leading to a two-fold reduction in conjugation efficiency. These results show that human gut epithelial cells can modulate bacterial conjugation and may have relevance to gene exchange in the gut....

  7. Enhanced Adsorption and Recovery of Uranyl Ions by NikR Mutant-Displaying Yeast

    Directory of Open Access Journals (Sweden)

    Kouichi Kuroda

    2014-04-01

    Full Text Available Uranium is one of the most important metal resources, and the technology for the recovery of uranyl ions (UO22+ from aqueous solutions is required to ensure a semi-permanent supply of uranium. The NikR protein is a Ni2+-dependent transcriptional repressor of the nickel-ion uptake system in Escherichia coli, but its mutant protein (NikRm is able to selectively bind uranyl ions in the interface of the two monomers. In this study, NikRm protein with ability to adsorb uranyl ions was displayed on the cell surface of Saccharomyces cerevisiae. To perform the binding of metal ions in the interface of the two monomers, two metal-binding domains (MBDs of NikRm were tandemly fused via linker peptides and displayed on the yeast cell surface by fusion with the cell wall-anchoring domain of yeast α-agglutinin. The NikRm-MBD-displaying yeast cells with particular linker lengths showed the enhanced adsorption of uranyl ions in comparison to the control strain. By treating cells with citrate buffer (pH 4.3, the uranyl ions adsorbed on the cell surface were recovered. Our results indicate that the adsorption system by yeast cells displaying tandemly fused MBDs of NikRm is effective for simple and concentrated recovery of uranyl ions, as well as adsorption of uranyl ions.

  8. Identification of small molecule inhibitors of Pseudomonas aeruginosa exoenzyme S using a yeast phenotypic screen.

    Directory of Open Access Journals (Sweden)

    Anthony Arnoldo

    2008-02-01

    Full Text Available Pseudomonas aeruginosa is an opportunistic human pathogen that is a key factor in the mortality of cystic fibrosis patients, and infection represents an increased threat for human health worldwide. Because resistance of Pseudomonas aeruginosa to antibiotics is increasing, new inhibitors of pharmacologically validated targets of this bacterium are needed. Here we demonstrate that a cell-based yeast phenotypic assay, combined with a large-scale inhibitor screen, identified small molecule inhibitors that can suppress the toxicity caused by heterologous expression of selected Pseudomonas aeruginosa ORFs. We identified the first small molecule inhibitor of Exoenzyme S (ExoS, a toxin involved in Type III secretion. We show that this inhibitor, exosin, modulates ExoS ADP-ribosyltransferase activity in vitro, suggesting the inhibition is direct. Moreover, exosin and two of its analogues display a significant protective effect against Pseudomonas infection in vivo. Furthermore, because the assay was performed in yeast, we were able to demonstrate that several yeast homologues of the known human ExoS targets are likely ADP-ribosylated by the toxin. For example, using an in vitro enzymatic assay, we demonstrate that yeast Ras2p is directly modified by ExoS. Lastly, by surveying a collection of yeast deletion mutants, we identified Bmh1p, a yeast homologue of the human FAS, as an ExoS cofactor, revealing that portions of the bacterial toxin mode of action are conserved from yeast to human. Taken together, our integrated cell-based, chemical-genetic approach demonstrates that such screens can augment traditional drug screening approaches and facilitate the discovery of new compounds against a broad range of human pathogens.

  9. Antifungal susceptibility profiles of 1698 yeast reference strains revealing potential emerging human pathogens.

    Directory of Open Access Journals (Sweden)

    Marie Desnos-Ollivier

    Full Text Available New molecular identification techniques and the increased number of patients with various immune defects or underlying conditions lead to the emergence and/or the description of novel species of human and animal fungal opportunistic pathogens. Antifungal susceptibility provides important information for ecological, epidemiological and therapeutic issues. The aim of this study was to assess the potential risk of the various species based on their antifungal drug resistance, keeping in mind the methodological limitations. Antifungal susceptibility profiles to the five classes of antifungal drugs (polyens, azoles, echinocandins, allylamines and antimetabolites were determined for 1698 yeast reference strains belonging to 992 species (634 Ascomycetes and 358 Basidiomycetes. Interestingly, geometric mean minimum inhibitory concentrations (MICs of all antifungal drugs tested were significantly higher for Basidiomycetes compared to Ascomycetes (p<0.001. Twenty four strains belonging to 23 species of which 19 were Basidiomycetes seem to be intrinsically "resistant" to all drugs. Comparison of the antifungal susceptibility profiles of the 4240 clinical isolates and the 315 reference strains belonging to 53 shared species showed similar results. Even in the absence of demonstrated in vitro/in vivo correlation, knowing the in vitro susceptibility to systemic antifungal agents and the putative intrinsic resistance of yeast species present in the environment is important because they could become opportunistic pathogens.

  10. Warburg effect and translocation-induced genomic instability: two yeast models for cancer cells

    International Nuclear Information System (INIS)

    Tosato, Valentina; Grüning, Nana-Maria; Breitenbach, Michael; Arnak, Remigiusz; Ralser, Markus; Bruschi, Carlo V.

    2013-01-01

    Yeast has been established as an efficient model system to study biological principles underpinning human health. In this review we focus on yeast models covering two aspects of cancer formation and progression (i) the activity of pyruvate kinase (PK), which recapitulates metabolic features of cancer cells, including the Warburg effect, and (ii) chromosome bridge-induced translocation (BIT) mimiking genome instability in cancer. Saccharomyces cerevisiae is an excellent model to study cancer cell metabolism, as exponentially growing yeast cells exhibit many metabolic similarities with rapidly proliferating cancer cells. The metabolic reconfiguration includes an increase in glucose uptake and fermentation, at the expense of respiration and oxidative phosphorylation (the Warburg effect), and involves a broad reconfiguration of nucleotide and amino acid metabolism. Both in yeast and humans, the regulation of this process seems to have a central player, PK, which is up-regulated in cancer, and to occur mostly on a post-transcriptional and post-translational basis. Furthermore, BIT allows to generate selectable translocation-derived recombinants (“translocants”), between any two desired chromosomal locations, in wild-type yeast strains transformed with a linear DNA cassette carrying a selectable marker flanked by two DNA sequences homologous to different chromosomes. Using the BIT system, targeted non-reciprocal translocations in mitosis are easily inducible. An extensive collection of different yeast translocants exhibiting genome instability and aberrant phenotypes similar to cancer cells has been produced and subjected to analysis. In this review, we hence provide an overview upon two yeast cancer models, and extrapolate general principles for mimicking human disease mechanisms in yeast.

  11. Warburg effect and translocation-induced genomic instability: two yeast models for cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Tosato, Valentina [International Centre for Genetic Engineering and Biotechnology, Trieste (Italy); Grüning, Nana-Maria [Cambridge System Biology Center, Department of Biochemistry, University of Cambridge, Cambridge (United Kingdom); Breitenbach, Michael [Division of Genetics, Department of Cell Biology, University of Salzburg, Salzburg (Austria); Arnak, Remigiusz [International Centre for Genetic Engineering and Biotechnology, Trieste (Italy); Ralser, Markus [Cambridge System Biology Center, Department of Biochemistry, University of Cambridge, Cambridge (United Kingdom); Bruschi, Carlo V., E-mail: bruschi@icgeb.org [International Centre for Genetic Engineering and Biotechnology, Trieste (Italy)

    2013-01-18

    Yeast has been established as an efficient model system to study biological principles underpinning human health. In this review we focus on yeast models covering two aspects of cancer formation and progression (i) the activity of pyruvate kinase (PK), which recapitulates metabolic features of cancer cells, including the Warburg effect, and (ii) chromosome bridge-induced translocation (BIT) mimiking genome instability in cancer. Saccharomyces cerevisiae is an excellent model to study cancer cell metabolism, as exponentially growing yeast cells exhibit many metabolic similarities with rapidly proliferating cancer cells. The metabolic reconfiguration includes an increase in glucose uptake and fermentation, at the expense of respiration and oxidative phosphorylation (the Warburg effect), and involves a broad reconfiguration of nucleotide and amino acid metabolism. Both in yeast and humans, the regulation of this process seems to have a central player, PK, which is up-regulated in cancer, and to occur mostly on a post-transcriptional and post-translational basis. Furthermore, BIT allows to generate selectable translocation-derived recombinants (“translocants”), between any two desired chromosomal locations, in wild-type yeast strains transformed with a linear DNA cassette carrying a selectable marker flanked by two DNA sequences homologous to different chromosomes. Using the BIT system, targeted non-reciprocal translocations in mitosis are easily inducible. An extensive collection of different yeast translocants exhibiting genome instability and aberrant phenotypes similar to cancer cells has been produced and subjected to analysis. In this review, we hence provide an overview upon two yeast cancer models, and extrapolate general principles for mimicking human disease mechanisms in yeast.

  12. WARBURG EFFECT AND TRANSLOCATION-INDUCED GENOMIC INSTABILITY: TWO YEAST MODELS FOR CANCER CELLS

    Directory of Open Access Journals (Sweden)

    Valentina eTosato

    2013-01-01

    Full Text Available Yeast has been established as an efficient model system to study biological principles underpinning human health. In this review we focus on yeast models covering two aspects of cancer formation and progression i the activity of pyruvate kinase (PK, which recapitulates metabolic features of cancer cells, including the Warburg effect, and ii Bridge-Induced chromosome Translocation (BIT mimicking genome instability in cancer. Saccharomyces cerevisiae is an excellent model to study cancer cell metabolism, as exponentially growing yeast cells exhibit many metabolic similarities with rapidly proliferating cancer cells. The metabolic reconfiguration includes an increase in glucose uptake and fermentation, at the expense of respiration and oxidative phosphorylation (the Warburg effect, and involves a broad reconfiguration of nucleotide and amino acid metabolism. Both in yeast and humans, the regulation of this process seems to have a central player, pyruvate kinase, which is up-regulated in cancer, and to occur mostly on a post-transcriptional and posttranslational basis. Furthermore, BIT allows to generate selectable translocation-derived recombinants (translocants, between any two desired chromosomal locations, in wild-type yeast strains transformed with a linear DNA cassette carrying a selectable marker flanked by two DNA sequences homologous to different chromosomes. Using the Bridge-Induced Translocation system, targeted non-reciprocal translocations in mitosis are easily inducible. An extensive collection of different yeast translocants exhibiting genome instability and aberrant phenotypes similar to cancer cells has been produced and subjected to analysis. In this review, we hence provide an overview upon two yeast cancer models, and extrapolate general principles for mimicking human disease mechanisms in yeast.

  13. The splicing mutant of the human tumor suppressor protein DFNA5 induces programmed cell death when expressed in the yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Van Rossom, Sofie; Op de Beeck, Ken; Franssens, Vanessa; Swinnen, Erwin; Schepers, Anne; Ghillebert, Ruben; Caldara, Marina; Van Camp, Guy; Winderickx, Joris

    2012-01-01

    DFNA5 was first identified as a gene responsible for autosomal dominant deafness. Different mutations were found, but they all resulted in exon 8 skipping during splicing and premature termination of the protein. Later, it became clear that the protein also has a tumor suppression function and that it can induce apoptosis. Epigenetic silencing of the DFNA5 gene is associated with different types of cancers, including gastric and colorectal cancers as well as breast tumors. We introduced the wild-type and mutant DFNA5 allele in the yeast Saccharomyces cerevisiae. The expression of the wild-type protein was well tolerated by the yeast cells, although the protein was subject of degradation and often deposited in distinct foci when cells entered the diauxic shift. In contrast, cells had problems to cope with mutant DFNA5 and despite an apparent compensatory reduction in expression levels, the mutant protein still triggered a marked growth defect, which in part can be ascribed to its interaction with mitochondria. Consistently, cells with mutant DFNA5 displayed significantly increased levels of ROS and signs of programmed cell death. The latter occurred independently of the yeast caspase, Mca1, but involved the mitochondrial fission protein, Fis1, the voltage-dependent anion channel protein, Por1 and the mitochondrial adenine nucleotide translocators, Aac1 and Aac3. Recent data proposed DFNA5 toxicity to be associated to a globular domain encoded by exon 2–6. We confirmed these data by showing that expression of solely this domain confers a strong growth phenotype. In addition, we identified a point mutant in this domain that completely abrogated its cytotoxicity in yeast as well as human Human Embryonic Kidney 293T cells (HEK293T). Combined, our data underscore that the yeast system offers a valuable tool to further dissect the apoptotic properties of DFNA5.

  14. The splicing mutant of the human tumor suppressor protein DFNA5 induces programmed cell death when expressed in the yeast Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Van Rossom, Sofie [Department of Biology, Functional Biology, KU Leuven, Leuven-Heverlee (Belgium); Department of Biomedical Sciences, Center of Medical Genetics, University of Antwerp, Wilrijk-Antwerp (Belgium); Op de Beeck, Ken [Department of Biomedical Sciences, Center of Medical Genetics, University of Antwerp, Wilrijk-Antwerp (Belgium); Franssens, Vanessa; Swinnen, Erwin [Department of Biology, Functional Biology, KU Leuven, Leuven-Heverlee (Belgium); Schepers, Anne [Department of Biomedical Sciences, Center of Medical Genetics, University of Antwerp, Wilrijk-Antwerp (Belgium); Ghillebert, Ruben; Caldara, Marina [Department of Biology, Functional Biology, KU Leuven, Leuven-Heverlee (Belgium); Van Camp, Guy [Department of Biomedical Sciences, Center of Medical Genetics, University of Antwerp, Wilrijk-Antwerp (Belgium); Winderickx, Joris, E-mail: guy.vancamp@ua.ac.be, E-mail: joris.winderickx@bio.kuleuven.be [Department of Biology, Functional Biology, KU Leuven, Leuven-Heverlee (Belgium)

    2012-07-25

    DFNA5 was first identified as a gene responsible for autosomal dominant deafness. Different mutations were found, but they all resulted in exon 8 skipping during splicing and premature termination of the protein. Later, it became clear that the protein also has a tumor suppression function and that it can induce apoptosis. Epigenetic silencing of the DFNA5 gene is associated with different types of cancers, including gastric and colorectal cancers as well as breast tumors. We introduced the wild-type and mutant DFNA5 allele in the yeast Saccharomyces cerevisiae. The expression of the wild-type protein was well tolerated by the yeast cells, although the protein was subject of degradation and often deposited in distinct foci when cells entered the diauxic shift. In contrast, cells had problems to cope with mutant DFNA5 and despite an apparent compensatory reduction in expression levels, the mutant protein still triggered a marked growth defect, which in part can be ascribed to its interaction with mitochondria. Consistently, cells with mutant DFNA5 displayed significantly increased levels of ROS and signs of programmed cell death. The latter occurred independently of the yeast caspase, Mca1, but involved the mitochondrial fission protein, Fis1, the voltage-dependent anion channel protein, Por1 and the mitochondrial adenine nucleotide translocators, Aac1 and Aac3. Recent data proposed DFNA5 toxicity to be associated to a globular domain encoded by exon 2–6. We confirmed these data by showing that expression of solely this domain confers a strong growth phenotype. In addition, we identified a point mutant in this domain that completely abrogated its cytotoxicity in yeast as well as human Human Embryonic Kidney 293T cells (HEK293T). Combined, our data underscore that the yeast system offers a valuable tool to further dissect the apoptotic properties of DFNA5.

  15. Bridge-Induced Translocation between NUP145 and TOP2 Yeast Genes Models the Genetic Fusion between the Human Orthologs Associated With Acute Myeloid Leukemia

    Directory of Open Access Journals (Sweden)

    Valentina Tosato

    2017-09-01

    Full Text Available In mammalian organisms liquid tumors such as acute myeloid leukemia (AML are related to spontaneous chromosomal translocations ensuing in gene fusions. We previously developed a system named bridge-induced translocation (BIT that allows linking together two different chromosomes exploiting the strong endogenous homologous recombination system of the yeast Saccharomyces cerevisiae. The BIT system generates a heterogeneous population of cells with different aneuploidies and severe aberrant phenotypes reminiscent of a cancerogenic transformation. In this work, thanks to a complex pop-out methodology of the marker used for the selection of translocants, we succeeded by BIT technology to precisely reproduce in yeast the peculiar chromosome translocation that has been associated with AML, characterized by the fusion between the human genes NUP98 and TOP2B. To shed light on the origin of the DNA fragility within NUP98, an extensive analysis of the curvature, bending, thermostability, and B-Z transition aptitude of the breakpoint region of NUP98 and of its yeast ortholog NUP145 has been performed. On this basis, a DNA cassette carrying homologous tails to the two genes was amplified by PCR and allowed the targeted fusion between NUP145 and TOP2, leading to reproduce the chimeric transcript in a diploid strain of S. cerevisiae. The resulting translocated yeast obtained through BIT appears characterized by abnormal spherical bodies of nearly 500 nm of diameter, absence of external membrane and defined cytoplasmic localization. Since Nup98 is a well-known regulator of the post-transcriptional modification of P53 target genes, and P53 mutations are occasionally reported in AML, this translocant yeast strain can be used as a model to test the constitutive expression of human P53. Although the abnormal phenotype of the translocant yeast was never rescued by its expression, an exogenous P53 was recognized to confer increased vitality to the translocants, in

  16. Construction and characterization of a yeast artificial chromosome library containing seven haploid human genome equivalents

    International Nuclear Information System (INIS)

    Albertsen, H.M.; Abderrahim, H.; Cann, H.M.; Dausset, J.; Le Paslier, D.; Cohen, D.

    1990-01-01

    Prior to constructing a library of yeast artificial chromosomes (YACs) containing very large human DNA fragments, the authors performed a series of preliminary experiments aimed at developing a suitable protocol. They found an inverse relationship between YAC insert size and transformation efficiency. Evidence of occasional rearrangement within YAC inserts was found resulting in clonally stable internal deletions or clonally unstable size variations. A protocol was developed for preparative electrophoretic enrichment of high molecular mass human DNA fragments from partial restriction digests and ligation with the YAC vector in agarose. A YAC library has been constructed from large fragments of DNA from an Epstein-Barr virus-transformed human lymphoblastoid cell line. The library presently contains 50,000 clones, 95% of which are greater than 250 kilobase pairs in size. The mean YAC size of the library, calculated from 132 randomly isolated clones, is 430 kilobase pairs. The library thus contains the equivalent of approximately seven haploid human genomes

  17. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium.

    Science.gov (United States)

    Smith, I M; Baker, A; Arneborg, N; Jespersen, L

    2015-11-01

    The human gastrointestinal epithelium makes up the largest barrier separating the body from the external environment. Whereas invasive pathogens cause epithelial barrier disruption, probiotic micro-organisms modulate tight junction regulation and improve epithelial barrier function. In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study demonstrates distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Further, our data demonstrate significant yeast-mediated modulation of Salmonella Typhimurium-induced epithelial cell barrier disruption and identify Kluyveromyces marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study is the first to demonstrate significant non-Saccharomyces yeast

  18. Yeasts in foods and beverages: impact on product quality and safety.

    Science.gov (United States)

    Fleet, Graham H

    2007-04-01

    The role of yeasts in food and beverage production extends beyond the well-known bread, beer and wine fermentations. Molecular analytical technologies have led to a major revision of yeast taxonomy, and have facilitated the ecological study of yeasts in many other products. The mechanisms by which yeasts grow in these ecosystems and impact on product quality can now be studied at the level of gene expression. Their growth and metabolic activities are moderated by a network of strain and species interactions, including interactions with bacteria and other fungi. Some yeasts have been developed as agents for the biocontrol of food spoilage fungi, and others are being considered as novel probiotic organisms. The association of yeasts with opportunistic infections and other adverse responses in humans raises new issues in the field of food safety.

  19. The H159A mutant of yeast enolase 1 has significant activity.

    Science.gov (United States)

    Brewer, J M; Holland, M J; Lebioda, L

    2000-10-05

    The function of His159 in the enolase mechanism is disputed. Recently, Vinarov and Nowak (Biochemistry (1999) 38, 12138-12149) prepared the H159A mutant of yeast enolase 1 and expressed this in Escherichia coli. They reported minimal (ca. 0.01% of the native value) activity, though the protein appeared to be correctly folded, according to its CD spectrum, tryptophan fluorescence, and binding of metal ion and substrate. We prepared H159A enolase using a multicopy plasmid and expressed the enzyme in yeast. Our preparations of H159A enolase have 0.2-0.4% of the native activity under standard assay conditions and are further activated by Mg(2+) concentrations above 1 mM to 1-1.5% of the native activity. Native enolase 1 (and enolase 2) are inhibited by such Mg(2+) concentrations. It is possible that His159 is necessary for correct folding of the enzyme and that expression in E. coli leads to largely misfolded protein. Copyright 2000 Academic Press.

  20. Flux control through protein phosphorylation in yeast

    DEFF Research Database (Denmark)

    Chen, Yu; Nielsen, Jens

    2016-01-01

    Protein phosphorylation is one of the most important mechanisms regulating metabolism as it can directly modify metabolic enzymes by the addition of phosphate groups. Attributed to such a rapid and reversible mechanism, cells can adjust metabolism rapidly in response to temporal changes. The yeast...... as well as identify mechanisms underlying human metabolic diseases. Here we collect functional phosphorylation events of 41 enzymes involved in yeast metabolism and demonstrate functional mechanisms and the application of this information in metabolic engineering. From a systems biology perspective, we...... describe the development of phosphoproteomics in yeast as well as approaches to analysing the phosphoproteomics data. Finally, we focus on integrated analyses with other omics data sets and genome-scale metabolic models. Despite the advances, future studies improving both experimental technologies...

  1. Molecular cloning and nucleotide sequence of cDNA for human liver arginase

    International Nuclear Information System (INIS)

    Haraguchi, Y.; Takiguchi, M.; Amaya, Y.; Kawamoto, S.; Matsuda, I.; Mori, M.

    1987-01-01

    Arginase (EC3.5.3.1) catalyzes the last step of the urea cycle in the liver of ureotelic animals. Inherited deficiency of the enzyme results in argininemia, an autosomal recessive disorder characterized by hyperammonemia. To facilitate investigation of the enzyme and gene structures and to elucidate the nature of the mutation in argininemia, the authors isolated cDNA clones for human liver arginase. Oligo(dT)-primed and random primer human liver cDNA libraries in λ gt11 were screened using isolated rat arginase cDNA as a probe. Two of the positive clones, designated λ hARG6 and λ hARG109, contained an overlapping cDNA sequence with an open reading frame encoding a polypeptide of 322 amino acid residues (predicted M/sub r/, 34,732), a 5'-untranslated sequence of 56 base pairs, a 3'-untranslated sequence of 423 base pairs, and a poly(A) segment. Arginase activity was detected in Escherichia coli cells transformed with the plasmid carrying λ hARG6 cDNA insert. RNA gel blot analysis of human liver RNA showed a single mRNA of 1.6 kilobases. The predicted amino acid sequence of human liver arginase is 87% and 41% identical with those of the rat liver and yeast enzymes, respectively. There are several highly conserved segments among the human, rat, and yeast enzymes

  2. Analysis of the human intestinal epithelial cell transcriptional response to Lactobacillus acidophilus, Lactobacillus salivarius, Bifidobacterium lactis and Escherichia coli

    DEFF Research Database (Denmark)

    Putaala, H; Barrangou, R; Leyer, G J

    2010-01-01

    a comparative analysis of the global in vitro transcriptional response of human intestinal epithelial cells to Lactobacillus acidophilus NCFM™, Lactobacillus salivarius Ls-33, Bifidobacterium animalis subsp. lactis 420, and enterohaemorrhagic Escherichia coli O157:H7 (EHEC). Interestingly, L. salivarius Ls-33...

  3. Comparative analysis of antibiotic resistance and phylogenetic group patterns in human and porcine urinary tract infectious Escherichia coli

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Nielsen, E.M.; Krag, L.

    2009-01-01

    Urinary tract infections (UTIs) are one of the most common infectious diseases in humans and domestic animals such as pigs. The most frequent infectious agent in such infections is Escherichia coli. Virulence characteristics of E. coli UTI strains range from highly virulent pyelonephritis strains...... to relatively benign asymptomatic bacteriuria strains. Here we analyse a spectrum of porcine and human UTI E. coli strains with respect to their antibiotic resistance patterns and their phylogenetic groups, determined by multiplex PCR. The clonal profiles of the strains differed profoundly; whereas human...

  4. Expression of fully functional tetrameric human hemoglobin in Escherichia coli

    International Nuclear Information System (INIS)

    Hoffman, S.J.; Looker, D.L.; Roehrich, J.M.; Cozart, P.E.; Durfee, S.L.; Tedesco, J.L.; Stetler, G.L.

    1990-01-01

    Synthesis genes encoding the human α- and β-globin polypeptides have been expressed from a single operon in Escherichia coli. The α- and β-globin polypeptides associate into soluble tetramers, incorporate heme, and accumulate to >5% of the total cellular protein. Purified recombinant hemoglobin has the correct stoichiometry of α- and β-globin chains and contains a full complement of heme. Each globin chain also contains an additional methionine as an extension to the amino terminus. The recombinant hemoglobin has a C 4 reversed-phase HPLC profile essentially identical to that of human hemoglobin A 0 and comigrates with hemoglobin A 0 on SDS/PAGE. The visible spectrum and oxygen affinity are similar to that of native human hemoglobin A 0 . The authors have also expressed the α- and β-globin genes separately and found that the expression of the α-globin gene alone results in a marked decrease in the accumulation of α-globin in the cell. Separate expression of the β-globin gene results in high levels of insoluble β-globin. These observations suggest that the presence of α- and β-globin in the same cell stabilizes α-globin and aids the correct folding of β-globin. This system provides a simple method for expressing large quantities of recombinant hemoglobin and allows facile manipulation of the genes encoding hemoglobin to produce functionally altered forms of this protein

  5. Fermentative performance of bacteria and yeasts in lignocellulose hydrolysates

    Energy Technology Data Exchange (ETDEWEB)

    Olsson, Lisbeth; Hahn-Haegerdal, B. (Lund Univ. (Sweden). Dept. of Applied Microbiology)

    1993-01-01

    The sugar consumption rates and the product formation of yeasts (Saccharomyces cidri NCYC 775, S. cerevisiae NCYC 1047, S.cerevisiae ATCC 4132) and bacteria (Lactobacillus brevis DSM 20054, Lactococcus lactis ssp. lactis ATCC 19435, Escherichia coli ATCC 11303, Zymomonas mobilis ATCC 31821) were investigated in spent sulphite liquor and an enzymatic hydrolysate of steam-pretreated Salix caprea at different pH values in order to elucidate the suitability of the organisms with respect to future genetic engineering approaches. The possible inhibitory action of the two substrates on the investigated microorganisms was also considered. S.cerevisiae emerged as one of the better candidates, owing to its fast sugar consumption rate and efficient ethanol production. (author)

  6. Nucleotide excision repair in yeast

    NARCIS (Netherlands)

    Eijk, Patrick van

    2012-01-01

    Nucleotide Excision Repair (NER) is a conserved DNA repair pathway capable of removing a broad spectrum of DNA damage. In human cells a defect in NER leads to the disorder Xeroderma pigmentosum (XP). The yeast Saccharomyces cerevisiae is an excellent model organism to study the mechanism of NER. The

  7. Automated thermometric enzyme immunoassay of human proinsulin produced by Escherichia coli

    International Nuclear Information System (INIS)

    Birnbaum, S.; Buelow, L.; Hardy, K.; Danielsson, B.; Mosbach, K.

    1986-01-01

    The authors have determined and monitored the production and release of human proinsulin by genetically engineered Escherichia coli cells. Several M9 media samples were analyzed sequentially after centrifugation with the aid of a rapid automated flow-through thermometric enzyme-linked immunosorbent assay (TELISA) system. The response time was 7 min after after sample injection and a single assay was complete after 13 min. Insulin concentrations in the range of 0.1-50 μg/ml could be determined. The TELISA method correlated well with conventional radioimmunoassay determinations. Standard curves were reproducible over a period of several days even when the immobilized antibody column was stored at 25 0 C in the enzyme thermistor unit. Thus, immediate assay start up was possible

  8. The relationship between salivary histatin levels and oral yeast carriage.

    Science.gov (United States)

    Jainkittivong, A; Johnson, D A; Yeh, C K

    1998-06-01

    Candida species are common commensal inhabitants of the oral cavity. Human saliva contains antifungal proteins called histatins. We tested the hypothesis that oral yeast status is related to salivary histatin levels. Thirty subjects were divided into two groups based on the presence (n = 15) or absence (n = 15) of yeast on oral mucosa surfaces. Unstimulated and stimulated submandibular and sublingual and parotid saliva was collected from each subject. Salivary flow rates were measured and histatin concentrations were determined in the stimulated saliva samples. The yeast colony positive group showed lower median unstimulated parotid saliva flow rates as well as lower median concentrations of total histatins in submandibular and sublingual saliva. There was a negative correlation between yeast colony-forming units and unstimulated parotid saliva flow rates and between yeast colony-forming units and submandibular and sublingual saliva histatin concentration and secretion. The results suggest that oral yeast status may be influenced by unstimulated parotid saliva flow rates and by submandibular and sublingual histatin concentration and secretion.

  9. Spent yeast as natural source of functional food additives

    Science.gov (United States)

    Rakowska, Rita; Sadowska, Anna; Dybkowska, Ewa; Świderski, Franciszek

    Spent yeasts are by-products arising from beer and wine production which over many years have been chiefly used as feed additives for livestock. They contain many valuable and bioactive substances which has thereby generated much interest in their exploitation. Up till now, the main products obtained from beer-brewing yeasts are β-glucans and yeast extracts. Other like foodstuffs include dried brewer’s yeast, where this is dried and the bitterness removed to be fit for human consumption as well as mannan-oligosaccharides hitherto used in the feed industry. β-glucans constitute the building blocks of yeast cell walls and can thus be used in human nutrition as dietary supplements or serving as food additives in functional foods. β-glucans products obtained via post-fermentation of beer also exhibit a high and multi-faceted biological activity where they improve the blood’s lipid profile, enhance immunological status and have both prebiotic and anti-oxidant properties. Yeast extracts are currently being used more and more to enhance flavour in foodstuffs, particularly for meat and its products. Depending on how autolysis is carried out, it is possible to design extracts of various meat flavours characteristic of specific meats. Many different flavour profiles can be created which may be additionally increased in combination with vegetable extracts. Within the food market, yeast extracts can appear in various guises such as liquids, pastes or powders. They all contain significant amounts of glutamic acid, 5’-GMP and 5’-IMP nucleotides together with various amino acids and peptides that act synergistically for enhancing the flavour of foodstuff products. Recent studies have demonstrated additional benefits of yeast extracts as valuable sources of amino acids and peptides which can be used in functional foods and dietary supplements. These products possess GRAS status (Generally Recognised As Safe) which thereby also adds further as to why they should be used

  10. Architecture of the human and yeast general transcription and DNA repair factor TFIIH

    Science.gov (United States)

    Luo, Jie; Cimermancic, Peter; Viswanath, Shruthi; Ebmeier, Christopher C.; Kim, Bong; Dehecq, Marine; Raman, Vishnu; Greenberg, Charles H.; Pellarin, Riccardo; Sali, Andrej; Taatjes, Dylan J.; Hahn, Steven; Ranish, Jeff

    2015-01-01

    Summary TFIIH is essential for both RNA polymerase II transcription and DNA repair, and mutations in TFIIH can result in human disease. Here, we determine the molecular architecture of human and yeast TFIIH by an integrative approach using chemical crosslinking/mass spectrometry (CXMS) data, biochemical analyses, and previously published electron microscopy maps. We identified four new conserved “topological regions” that function as hubs for TFIIH assembly and more than 35 conserved topological features within TFIIH, illuminating a network of interactions involved in TFIIH assembly and regulation of its activities. We show that one of these conserved regions, the p62/Tfb1 Anchor region, directly interacts with the DNA helicase subunit XPD/Rad3 in native TFIIH and is required for the integrity and function of TFIIH. We also reveal the structural basis for defects in patients with Xeroderma pigmentosum and Trichothiodystrophy, with mutations found at the interface between the p62 Anchor region and the XPD subunit. PMID:26340423

  11. Distinct Domestication Trajectories in Top-Fermenting Beer Yeasts and Wine Yeasts.

    Science.gov (United States)

    Gonçalves, Margarida; Pontes, Ana; Almeida, Pedro; Barbosa, Raquel; Serra, Marta; Libkind, Diego; Hutzler, Mathias; Gonçalves, Paula; Sampaio, José Paulo

    2016-10-24

    Beer is one of the oldest alcoholic beverages and is produced by the fermentation of sugars derived from starches present in cereal grains. Contrary to lager beers, made by bottom-fermenting strains of Saccharomyces pastorianus, a hybrid yeast, ale beers are closer to the ancient beer type and are fermented by S. cerevisiae, a top-fermenting yeast. Here, we use population genomics to investigate (1) the closest relatives of top-fermenting beer yeasts; (2) whether top-fermenting yeasts represent an independent domestication event separate from those already described; (3) whether single or multiple beer yeast domestication events can be inferred; and (4) whether top-fermenting yeasts represent non-recombinant or recombinant lineages. Our results revealed that top-fermenting beer yeasts are polyphyletic, with a main clade composed of at least three subgroups, dominantly represented by the German, British, and wheat beer strains. Other beer strains were phylogenetically close to sake, wine, or bread yeasts. We detected genetic signatures of beer yeast domestication by investigating genes previously linked to brewing and using genome-wide scans. We propose that the emergence of the main clade of beer yeasts is related with a domestication event distinct from the previously known cases of wine and sake yeast domestication. The nucleotide diversity of the main beer clade more than doubled that of wine yeasts, which might be a consequence of fundamental differences in the modes of beer and wine yeast domestication. The higher diversity of beer strains could be due to the more intense and different selection regimes associated to brewing. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Prions in yeast

    OpenAIRE

    Bezdíčka, Martin

    2013-01-01

    The thesis describes yeast prions and their biological effects on yeast in general. It defines the basic characteristics of yeast prions, that distinguish prions from other proteins. The thesis introduces various possibilities of prion formation, and propagation as well as specific types of yeast prions, including various functions of most studied types of prions. The thesis also focuses on chaperones that affect the state of yeast prions in cells. Lastly, the thesis indicates similarities be...

  13. Selection of Co-Substrate and Aeration Conditions for Vanillin Production by Escherichia coli JM109/pBB1

    Directory of Open Access Journals (Sweden)

    Paolo Torre

    2004-01-01

    Full Text Available Yeast extract, Luria-Bertani medium and tryptone were tested as co-substrates for vanillin production from ferulic acid by resting cells of Escherichia coli JM109/pBB1. Yeast extract proved to be the best component for sustaining such a bioconversion, which is not self-sustained from the bioenergetic point of view. Tests were also performed under variable aeration conditions by simultaneously varying the ratio of medium to vessel volume and the agitation speed. The results of these tests suggest that, under excess aeration, a non-specific oxidase activity was very likely responsible for the oxidation of a significant portion of vanillin to vanillic acid, thus reducing the vanillin yield.

  14. Crystallization and preliminary crystallographic study of the yeast Malassezia sympodialis allergen Mala s 1

    International Nuclear Information System (INIS)

    Vilhelmsson, Monica; Hallberg, B. Martin; Rasool, Omid; Zargari, Arezou; Scheynius, Annika; Achour, Adnane

    2006-01-01

    Crystals of the M. sympodialis allergen Mala s 1 have been obtained using the hanging-drop vapour-diffusion method. A diffraction data set has been collected from native crystals to 1.35 Å resolution. The opportunistic yeast Malassezia sympodialis can act as an allergen and elicit specific IgE- and T-cell reactivity in patients with atopic eczema. The first identified major allergen from M. sympodialis, Mala s 1, is present on the cell surface of the yeast. Recombinant Mala s 1 was expressed in Escherichia coli, purified and refolded in a soluble form. Crystals of Mala s 1 were obtained in 25% PEG 8K, 0.2 M (NH 4 ) 2 SO 4 . Crystals belong to space group P2 1 2 1 2, with unit-cell parameters a = 44.4, b = 163.7, c = 50.6 Å, and diffract to 1.35 Å resolution

  15. Crystallization and preliminary crystallographic study of the yeast Malassezia sympodialis allergen Mala s 1

    Energy Technology Data Exchange (ETDEWEB)

    Vilhelmsson, Monica, E-mail: monica.vilhelmsson@medks.ki.se [Department of Medicine, Clinical Allergy Research Unit, Karolinska Institutet and Karolinska University Hospital, Stockholm (Sweden); Center for Infectious Medicine, Department of Medicine, Karolinska University Hospital, Huddinge, Stockholm (Sweden); Hallberg, B. Martin [Department of Biochemistry and Biophysics, Stockholm University, Stockholm (Sweden); Rasool, Omid; Zargari, Arezou; Scheynius, Annika [Department of Medicine, Clinical Allergy Research Unit, Karolinska Institutet and Karolinska University Hospital, Stockholm (Sweden); Achour, Adnane [Center for Infectious Medicine, Department of Medicine, Karolinska University Hospital, Huddinge, Stockholm (Sweden); Department of Medicine, Clinical Allergy Research Unit, Karolinska Institutet and Karolinska University Hospital, Stockholm (Sweden)

    2006-02-01

    Crystals of the M. sympodialis allergen Mala s 1 have been obtained using the hanging-drop vapour-diffusion method. A diffraction data set has been collected from native crystals to 1.35 Å resolution. The opportunistic yeast Malassezia sympodialis can act as an allergen and elicit specific IgE- and T-cell reactivity in patients with atopic eczema. The first identified major allergen from M. sympodialis, Mala s 1, is present on the cell surface of the yeast. Recombinant Mala s 1 was expressed in Escherichia coli, purified and refolded in a soluble form. Crystals of Mala s 1 were obtained in 25% PEG 8K, 0.2 M (NH{sub 4}){sub 2}SO{sub 4}. Crystals belong to space group P2{sub 1}2{sub 1}2, with unit-cell parameters a = 44.4, b = 163.7, c = 50.6 Å, and diffract to 1.35 Å resolution.

  16. Genomic diversity of Escherichia isolates from diverse habitats.

    Directory of Open Access Journals (Sweden)

    Seungdae Oh

    Full Text Available Our understanding of the Escherichia genus is heavily biased toward pathogenic or commensal isolates from human or animal hosts. Recent studies have recovered Escherichia isolates that persist, and even grow, outside these hosts. Although the environmental isolates are typically phylogenetically distinct, they are highly related to and phenotypically indistinguishable from their human counterparts, including for the coliform test. To gain insights into the genomic diversity of Escherichia isolates from diverse habitats, including freshwater, soil, animal, and human sources, we carried out comparative DNA-DNA hybridizations using a multi-genome E. coli DNA microarray. The microarray was validated based on hybridizations with selected strains whose genome sequences were available and used to assess the frequency of microarray false positive and negative signals. Our results showed that human fecal isolates share two sets of genes (n>90 that are rarely found among environmental isolates, including genes presumably important for evading host immune mechanisms (e.g., a multi-drug transporter for acids and antimicrobials and adhering to epithelial cells (e.g., hemolysin E and fimbrial-like adhesin protein. These results imply that environmental isolates are characterized by decreased ability to colonize host cells relative to human isolates. Our study also provides gene markers that can distinguish human isolates from those of warm-blooded animal and environmental origins, and thus can be used to more reliably assess fecal contamination in natural ecosystems.

  17. Prevalence and characterization of plasmids carrying sulfonamide resistance genes among Escherichia coli from pigs, pig carcasses and human

    OpenAIRE

    Wu, Shuyu; Dalsgaard, Anders; Hammerum, Anette M; Porsbo, Lone J; Jensen, Lars B

    2010-01-01

    Abstract Background Sulfonamide resistance is very common in Escherichia coli. The aim of this study was to characterize plasmids carrying sulfonamide resistance genes (sul1, sul2 and sul3) in E. coli isolated from pigs and humans with a specific objective to assess the genetic diversity of plasmids involved in the mobility of sul genes. Methods A total of 501 E. coli isolates from pig feces, pig carcasses and human stools were tested for their susceptibility to selected antimicrobial. Multip...

  18. Identification of an evolutionary conserved SURF-6 domain in a family of nucleolar proteins extending from human to yeast

    International Nuclear Information System (INIS)

    Polzikov, Mikhail; Zatsepina, Olga; Magoulas, Charalambos

    2005-01-01

    The mammalian SURF-6 protein is localized in the nucleolus, yet its function remains elusive in the recently characterized nucleolar proteome. We discovered by searching the Protein families database that a unique evolutionary conserved SURF-6 domain is present in the carboxy-terminal of a novel family of eukaryotic proteins extending from human to yeast. By using the enhanced green fluorescent protein as a fusion protein marker in mammalian cells, we show that proteins from distantly related taxonomic groups containing the SURF-6 domain are localized in the nucleolus. Deletion sequence analysis shows that multiple regions of the SURF-6 protein are capable of nucleolar targeting independently of the evolutionary conserved domain. We identified that the Saccharomyces cerevisiae member of the SURF-6 family, named rrp14 or ykl082c, has been categorized in yeast databases to interact with proteins involved in ribosomal biogenesis and cell polarity. These results classify SURF-6 as a new family of nucleolar proteins in the eukaryotic kingdom and point out that SURF-6 has a distinct domain within the known nucleolar proteome that may mediate complex protein-protein interactions for analogous processes between yeast and mammalian cells

  19. Population FBA predicts metabolic phenotypes in yeast.

    Directory of Open Access Journals (Sweden)

    Piyush Labhsetwar

    2017-09-01

    Full Text Available Using protein counts sampled from single cell proteomics distributions to constrain fluxes through a genome-scale model of metabolism, Population flux balance analysis (Population FBA successfully described metabolic heterogeneity in a population of independent Escherichia coli cells growing in a defined medium. We extend the methodology to account for correlations in protein expression arising from the co-regulation of genes and apply it to study the growth of independent Saccharomyces cerevisiae cells in two different growth media. We find the partitioning of flux between fermentation and respiration predicted by our model agrees with recent 13C fluxomics experiments, and that our model largely recovers the Crabtree effect (the experimentally known bias among certain yeast species toward fermentation with the production of ethanol even in the presence of oxygen, while FBA without proteomics constraints predicts respirative metabolism almost exclusively. The comparisons to the 13C study showed improvement upon inclusion of the correlations and motivated a technique to systematically identify inconsistent kinetic parameters in the literature. The minor secretion fluxes for glycerol and acetate are underestimated by our method, which indicate a need for further refinements to the metabolic model. For yeast cells grown in synthetic defined (SD medium, the calculated broad distribution of growth rates matches experimental observations from single cell studies, and we characterize several metabolic phenotypes within our modeled populations that make use of diverse pathways. Fast growing yeast cells are predicted to perform significant amount of respiration, use serine-glycine cycle and produce ethanol in mitochondria as opposed to slow growing cells. We use a genetic algorithm to determine the proteomics constraints necessary to reproduce the growth rate distributions seen experimentally. We find that a core set of 51 constraints are essential but

  20. Yeast-2-Hybrid data file showing progranulin interactions in human fetal brain and bone marrow libraries.

    Science.gov (United States)

    Tegeder, Irmgard

    2016-12-01

    Progranulin deficiency in humans is associated with neurodegeneration. Its mechanisms are not yet fully understood. We performed a Yeast-2-Hybrid screen using human full-length progranulin as bait to assess the interactions of progranulin. Progranulin was screened against human fetal brain and human bone marrow libraries using the standard Matchmaker technology (Clontech). This article contains the full Y2H data table, including blast results and sequences, a sorted table according to selection criteria for likely positive, putatively positive, likely false and false preys, and tables showing the gene ontology terms associated with the likely and putative preys of the brain and bone marrow libraries. The interactions with autophagy proteins were confirmed and functionally analyzed in "Progranulin overexpression in sensory neurons attenuates neuropathic pain in mice: Role of autophagy" (C. Altmann, S. Hardt, C. Fischer, J. Heidler, H.Y. Lim, A. Haussler, B. Albuquerque, B. Zimmer, C. Moser, C. Behrends, F. Koentgen, I. Wittig, M.H. Schmidt, A.M. Clement, T. Deller, I. Tegeder, 2016) [1].

  1. ELECTROPHORETIC MOBILITIES OF ESCHERICHIA COLI 0157:H7 AND WILD-TYPE ESCHERICHIA COLI STRAINS

    Science.gov (United States)

    The electrophoretic mobility (EPM) of a number of human-virulent and "wild-type" Escherichia coli strains in phosphate buffered water was measured. The impact of pH, ionic strength, cation type (valence) and concentration, and bacterial strain on the EPM was investigated. Resul...

  2. Pro-Apoptotic Role of the Human YPEL5 Gene Identified by Functional Complementation of a Yeast moh1Δ Mutation.

    Science.gov (United States)

    Lee, Ji Young; Jun, Do Youn; Park, Ju Eun; Kwon, Gi Hyun; Kim, Jong-Sik; Kim, Young Ho

    2017-03-28

    To examine the pro-apoptotic role of the human ortholog (YPEL5) of the Drosophila Yippee protein, the cell viability of Saccharomyces cerevisiae mutant strain with deleted MOH1 , the yeast ortholog, was compared with that of the wild-type (WT)- MOH1 strain after exposure to different apoptogenic stimulants, including UV irradiation, methyl methanesulfonate (MMS), camptothecin (CPT), heat shock, and hyperosmotic shock. The moh1 Δ mutant exhibited enhanced cell viability compared with the WT- MOH1 strain when treated with lethal UV irradiation, 1.8 mM MMS, 100 µ CPT, heat shock at 50°C, or 1.2 M KCl. At the same time, the level of Moh1 protein was commonly up-regulated in the WT- MOH1 strain as was that of Ynk1 protein, which is known as a marker for DNA damage. Although the enhanced UV resistance of the moh1 Δ mutant largely disappeared following transformation with the yeast MOH1 gene or one of the human YPEL1-YPEL5 genes, the transformant bearing pYES2- YPEL5 was more sensitive to lethal UV irradiation and its UV sensitivity was similar to that of the WT- MOH1 strain. Under these conditions, the UV irradiation-induced apoptotic events, such as FITC-Annexin V stainability, mitochondrial membrane potential (ΔΨm) loss, and metacaspase activation, occurred to a much lesser extent in the moh1 Δ mutant compared with the WT- MOH1 strain and the mutant strain bearing pYES2- MOH1 or pYES2- YPEL5 . These results demonstrate the functional conservation between yeast Moh1 and human YPEL5, and their involvement in mitochondria-dependent apoptosis induced by DNA damage.

  3. Identification of human ferritin, heavy polypeptide 1 (FTH1) and yeast RGI1 (YER067W) as pro-survival sequences that counteract the effects of Bax and copper in Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Eid, Rawan [Department of Chemistry and Chemical Engineering, Royal Military College, Kingston, Ontario (Canada); Department of Biology, Queen' s University, Kingston, Ontario (Canada); Boucher, Eric [Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec (Canada); Gharib, Nada [Department of Chemistry and Chemical Engineering, Royal Military College, Kingston, Ontario (Canada); Khoury, Chamel [Department of Chemistry and Chemical Engineering, Royal Military College, Kingston, Ontario (Canada); Department of Biology, Queen' s University, Kingston, Ontario (Canada); Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec (Canada); Arab, Nagla T.T. [Department of Chemistry and Chemical Engineering, Royal Military College, Kingston, Ontario (Canada); Department of Biology, Queen' s University, Kingston, Ontario (Canada); Murray, Alistair [Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec (Canada); Young, Paul G. [Department of Biology, Queen' s University, Kingston, Ontario (Canada); Mandato, Craig A. [Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec (Canada); Greenwood, Michael T., E-mail: michael.greenwood@rmc.ca [Department of Chemistry and Chemical Engineering, Royal Military College, Kingston, Ontario (Canada)

    2016-03-01

    Ferritin is a sub-family of iron binding proteins that form multi-subunit nanotype iron storage structures and prevent oxidative stress induced apoptosis. Here we describe the identification and characterization of human ferritin, heavy polypeptide 1 (FTH1) as a suppressor of the pro-apoptotic murine Bax sequence in yeast. In addition we demonstrate that FTH1 is a general pro-survival sequence since it also prevents the cell death inducing effects of copper when heterologously expressed in yeast. Although ferritins are phylogenetically widely distributed and are present in most species of Bacteria, Archaea and Eukarya, ferritin is conspicuously absent in most fungal species including Saccharomyces cerevisiae. An in silico analysis of the yeast proteome lead to the identification of the 161 residue RGI1 (YER067W) encoded protein as a candidate for being a yeast ferritin. In addition to sharing 20% sequence identity with the 183 residue FTH1, RGI1 also has similar pro-survival properties as ferritin when overexpressed in yeast. Analysis of recombinant protein by SDS-PAGE and by electron microscopy revealed the expected formation of higher-order structures for FTH1 that was not observed with Rgi1p. Further analysis revealed that cells overexpressing RGI1 do not show increased resistance to iron toxicity and do not have enhanced capacity to store iron. In contrast, cells lacking RGI1 were found to be hypersensitive to the toxic effects of iron. Overall, our results suggest that Rgi1p is a novel pro-survival protein whose function is not related to ferritin but nevertheless it may have a role in regulating yeast sensitivity to iron stress. - Highlights: • Human ferritin, heavy polypeptide 1 (FTH1) was identified as a suppressor of the pro-apoptotic Bax in yeast. • Based on its similarity to ferritin we examined Rgi1p/YER067W for potential ferritin like functions. • Like human H-ferritin, RGI1 confers increased resistance to apoptotic inducing stresses in yeast.

  4. Identification of human ferritin, heavy polypeptide 1 (FTH1) and yeast RGI1 (YER067W) as pro-survival sequences that counteract the effects of Bax and copper in Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Eid, Rawan; Boucher, Eric; Gharib, Nada; Khoury, Chamel; Arab, Nagla T.T.; Murray, Alistair; Young, Paul G.; Mandato, Craig A.; Greenwood, Michael T.

    2016-01-01

    Ferritin is a sub-family of iron binding proteins that form multi-subunit nanotype iron storage structures and prevent oxidative stress induced apoptosis. Here we describe the identification and characterization of human ferritin, heavy polypeptide 1 (FTH1) as a suppressor of the pro-apoptotic murine Bax sequence in yeast. In addition we demonstrate that FTH1 is a general pro-survival sequence since it also prevents the cell death inducing effects of copper when heterologously expressed in yeast. Although ferritins are phylogenetically widely distributed and are present in most species of Bacteria, Archaea and Eukarya, ferritin is conspicuously absent in most fungal species including Saccharomyces cerevisiae. An in silico analysis of the yeast proteome lead to the identification of the 161 residue RGI1 (YER067W) encoded protein as a candidate for being a yeast ferritin. In addition to sharing 20% sequence identity with the 183 residue FTH1, RGI1 also has similar pro-survival properties as ferritin when overexpressed in yeast. Analysis of recombinant protein by SDS-PAGE and by electron microscopy revealed the expected formation of higher-order structures for FTH1 that was not observed with Rgi1p. Further analysis revealed that cells overexpressing RGI1 do not show increased resistance to iron toxicity and do not have enhanced capacity to store iron. In contrast, cells lacking RGI1 were found to be hypersensitive to the toxic effects of iron. Overall, our results suggest that Rgi1p is a novel pro-survival protein whose function is not related to ferritin but nevertheless it may have a role in regulating yeast sensitivity to iron stress. - Highlights: • Human ferritin, heavy polypeptide 1 (FTH1) was identified as a suppressor of the pro-apoptotic Bax in yeast. • Based on its similarity to ferritin we examined Rgi1p/YER067W for potential ferritin like functions. • Like human H-ferritin, RGI1 confers increased resistance to apoptotic inducing stresses in yeast.

  5. Measuring strand discontinuity-directed mismatch repair in yeast Saccharomyces cerevisiae by cell-free nuclear extracts.

    Science.gov (United States)

    Yuan, Fenghua; Lai, Fangfang; Gu, Liya; Zhou, Wen; El Hokayem, Jimmy; Zhang, Yanbin

    2009-05-01

    Mismatch repair corrects biosynthetic errors generated during DNA replication, whose deficiency causes a mutator phenotype and directly underlies hereditary non-polyposis colorectal cancer and sporadic cancers. Because of remarkably high conservation of the mismatch repair machinery between the budding yeast (Saccharomyces cerevisiae) and humans, the study of mismatch repair in yeast has provided tremendous insights into the mechanisms of this repair pathway in humans. In addition, yeast cells possess an unbeatable advantage over human cells in terms of the easy genetic manipulation, the availability of whole genome deletion strains, and the relatively low cost for setting up the system. Although many components of eukaryotic mismatch repair have been identified, it remains unclear if additional factors, such as DNA helicase(s) and redundant nuclease(s) besides EXO1, participate in eukaryotic mismatch repair. To facilitate the discovery of novel mismatch repair factors, we developed a straightforward in vitro cell-free repair system. Here, we describe the practical protocols for preparation of yeast cell-free nuclear extracts and DNA mismatch substrates, and the in vitro mismatch repair assay. The validity of the cell-free system was confirmed by the mismatch repair deficient yeast strain (Deltamsh2) and the complementation assay with purified yeast MSH2-MSH6.

  6. Ecological structuring of yeasts associated with trees around Hamilton, Ontario, Canada.

    Science.gov (United States)

    Maganti, Harinad; Bartfai, David; Xu, Jianping

    2012-02-01

    This study seeks to determine the distribution and diversity of yeasts in and around the Hamilton area in Canada. In light of the increasing number of fungal infections along with rising morbidity and mortality rates, especially among the immunocompromised, understanding the diversity and distribution of yeasts in natural environments close to human habitations has become an increasingly relevant topic. In this study, we analyzed 1110 samples obtained from the hollows of trees, shrubs and avian droppings at 8 geographical sites in and around Hamilton, Ontario, Canada. A total of 88 positive yeast strains were isolated and identified belonging to 20 yeast species. Despite the relative proximity of the sampling sites, our DNA fingerprinting results showed that the yeast populations were highly heterogenous. Among the 14 tree species sampled, cedar, cottonwood and basswood hollows had relatively high yeast colonization rates. Interestingly, Candida parapsilosis was isolated almost exclusively from Pine trees only. Our results are consistent with microgeographic and ecological differentiation of yeast species in and around an urban environment. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  7. Global gene expression profiling of asymptomatic bacteriuria Escherichia coli during biofilm growth in human urine

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Klemm, Per

    2007-01-01

    Urinary tract infection (UTI) is an important health problem worldwide, with many millions of cases each year, and Escherichia coli is the most common organism causing UTI in humans. Also, E. coli is responsible for most infections in patients with chronic indwelling bladder catheter. The two...... asymptomatic bacteriuria (ABU) E. coli strains 83972 and VR50 are significantly better biofilm formers in their natural growth medium, human urine, than the two uropathogenic E. coli isolates CFT073 and 536. We used DNA microarrays to monitor the expression profile during biofilm growth in urine of the two ABU...... strains 83972 and VR50. Significant differences in expression levels were seen between the biofilm expression profiles of the two strains with the corresponding planktonic expression profiles in morpholinepropanesulfonic acid minimal laboratory medium and human urine; 417 and 355 genes were up- and down...

  8. Radiation stimulation of yeast crops for increasing output of alcohol and baker yeasts

    International Nuclear Information System (INIS)

    Vlad, E.; Marsheu, P.

    1974-01-01

    The purpose of this study was to stimulate by gamma radiation the existing commercial types of yeast so as to obtain yeasts that would better reflect the substrate and have improved reproductive capacity. The experiments were conducted under ordinary conditions using commercial yeasts received from one factory producing alcohol and bakery yeasts and isolated as pure cultures. Irradiating yeast cultures with small doses (up to 10 krad) was found to stimulate the reproduction and fermenting activity of yeast cells as manifested in increased accumulation of yeast biomass and greater yield of ethyl alcohol. (E.T.)

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

    Directory of Open Access Journals (Sweden)

    Zhang Tingting

    2012-12-01

    Full Text Available Abstract Background Yeast Saccharomyces cerevisiae is a widely-used system for protein expression. We previously showed that heat-killed whole recombinant yeast vaccine expressing mammalian myostatin can modulate myostatin function in mice, resulting in increase of body weight and muscle composition in these animals. Foreign DNA introduced into yeast cells can be lost soon unless cells are continuously cultured in selection media, which usually contain antibiotics. For cost and safety concerns, it is essential to optimize conditions to produce quality food and pharmaceutical products. Results We developed a simple but effective method to engineer a yeast strain stably expressing mammalian myostatin. This method utilized high-copy-number integration of myostatin gene into the ribosomal DNA of Saccharomyces cerevisiae. In the final step, antibiotic selection marker was removed using the Cre-LoxP system to minimize any possible side-effects for animals. The resulting yeast strain can be maintained in rich culture media and stably express mammalian myostatin for two years. Oral administration of the recombinant yeast was able to induce immune response to myostatin and modulated the body weight of mice. Conclusions Establishment of such yeast strain is a step further toward transformation of yeast cells into edible vaccine to improve meat production in farm animals and treat human muscle-wasting diseases in the future.

  10. Molecular basis of antifungal drug resistance in yeasts

    DEFF Research Database (Denmark)

    Morio, Florent; Jensen, Rasmus Hare; Le Pape, Patrice

    2017-01-01

    Besides inherent differences in in vitro susceptibilities, clinically-relevant yeast species may acquire resistance upon exposure to most antifungal drugs used in the clinic. In recent years, major fundamental research studies have been conducted to improve our understanding of the molecular basis...... of antifungal resistance. This topic is of major interest as antifungal resistance in yeast is clearly evolving and is correlated with clinical failure. This minireview is an overview of the most recent findings about key molecular mechanisms evolving in human pathogenic yeasts, particularly Candida spp......., in the context of antifungal drug resistance. Also included are the methods currently available for in vitro antifungal susceptibility testing and for molecular detection of mutations associated with resistance. Finally, the genetic drivers of antifungal resistance are discussed in light of the spectra...

  11. Permeation of iodide from iodine-enriched yeast through porcine intestine.

    Science.gov (United States)

    Ryszka, Florian; Dolińska, Barbara; Zieliński, Michał; Chyra, Dagmara; Dobrzański, Zbigniew

    2013-01-01

    Iodine deficiency is a common phenomenon, threatening the whole global human population. Recommended daily intake of iodine is 150 μg for adults and 250 μg for pregnant and breastfeeding women. About 50% of human population can be at risk of moderate iodine deficiency. Due to this fact, increased iodine supplementation is recommended, through intake of iodized mineral water and salt iodization. The aim of this study was to investigate permeation and absorption of iodide from iodine bioplex (experimental group) in comparison with potassium iodide (controls). Permeation and absorption processes were investigated in vitro using a porcine intestine. The experimental model was based on a standard Franz diffusion cell (FD-Cell). The iodine bioplex was produced using Saccharomyces cerevisiae yeast and whey powder: iodine content - 388 μg/g, total protein - 28.5%, total fat - 0.9%., glutamic acid - 41.2%, asparaginic acid - 29.4%, lysine - 24.8%; purchased from: F.Z.N.P. Biochefa, Sosnowiec, Poland. Potassium iodide was used as controls, at 388 μg iodine concentration, which was the same as in iodine-enriched yeast bioplex. A statistically significant increase in iodide permeation was observed for iodine-enriched yeast bioplex in comparison with controls - potassium iodide. After 5h the total amount of permeated iodide from iodine-enriched yeast bioplex was 85%, which is ~ 2-fold higher than controls - 37%. Iodide absorption was by contrast statistically significantly higher in controls - 7.3%, in comparison with 4.5% in experimental group with iodine-enriched yeast bioplex. Presented results show that iodide permeation process dominates over absorption in case of iodine-enriched yeast bioplex.

  12. Human exposure assessment to antibiotic-resistant Escherichia coli through drinking water.

    Science.gov (United States)

    O'Flaherty, E; Borrego, C M; Balcázar, J L; Cummins, E

    2018-03-01

    Antibiotic-resistant bacteria (ARB) are a potential threat to human health through drinking water with strong evidence of ARB presence in post treated tap water around the world. This study examines potential human exposure to antibiotic-resistant (AR) Escherichia coli (E. coli) through drinking water, the effect of different drinking water treatments on AR E. coli and the concentration of AR E. coli required in the source water for the EU Drinking Water Directive (DWD) (Council Directive 98/83/EC, 0CFU/100ml of E. coli in drinking water) to be exceeded. A number of scenarios were evaluated to examine different water treatment combinations and to reflect site specific conditions at a study site in Europe. A literature search was carried out to collate data on the effect of environmental conditions on AR E. coli, the effect of different water treatments on AR E. coli and typical human consumption levels of tap water. A human exposure assessment model was developed with probability distributions used to characterise uncertainty and variability in the input data. Overall results show the mean adult human exposure to AR E. coli from tap water consumption ranged between 3.44×10 -7 and 2.95×10 -1 cfu/day for the scenarios tested and varied depending on the water treatments used. The level of AR E. coli required in the source water pre-treatment to exceed the DWD varied between 1 and 5logcfu/ml, depending on the water treatments used. This can be used to set possible monitoring criteria in pre-treated water for potential ARB exposure in drinking water. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Role of urinary cathelicidin LL-37 and human β-defensin 1 in uncomplicated Escherichia coli urinary tract infections

    DEFF Research Database (Denmark)

    Nielsen, Karen L; Dynesen, Pia; Larsen, Preben

    2014-01-01

    Cathelicidin (LL-37) and human β-defensin 1 (hBD-1) are important components of the innate defense in the urinary tract. The aim of this study was to characterize whether these peptides are important for developing uncomplicated Escherichia coli urinary tract infections (UTIs). This was investiga......Cathelicidin (LL-37) and human β-defensin 1 (hBD-1) are important components of the innate defense in the urinary tract. The aim of this study was to characterize whether these peptides are important for developing uncomplicated Escherichia coli urinary tract infections (UTIs......). This was investigated by comparing urinary peptide levels of UTI patients during and after infection to those of controls, as well as characterizing the fecal flora of participants with respect to susceptibility to LL-37 and in vivo virulence. Forty-seven UTI patients and 50 controls who had never had a UTI were...... included. Participants were otherwise healthy, premenopausal, adult women. LL-37 MIC levels were compared for fecal E. coli clones from patients and controls and were also compared based on phylotypes (A, B1, B2, and D). In vivo virulence was investigated in the murine UTI model by use of selected fecal...

  14. Structure–function relations in the NTPase domain of the antiviral tRNA ribotoxin Escherichia coli PrrC

    International Nuclear Information System (INIS)

    Meineke, Birthe; Shuman, Stewart

    2012-01-01

    Breakage of tRNA by Escherichia coli anticodon nuclease PrrC (EcoPrrC) underlies a host antiviral response to phage T4 infection. Expression of EcoPrrC is cytocidal in yeast, signifying that PrrC ribotoxicity crosses phylogenetic domain boundaries. EcoPrrC consists of an N-terminal NTPase module that resembles ABC transporters and a C-terminal nuclease module that is sui generis. PrrC homologs are prevalent in many other bacteria. Here we report that Haemophilus influenzae PrrC is toxic in E. coli and yeast. To illuminate structure–activity relations, we conducted a new round of mutational analysis of EcoPrrC guided by primary structure conservation among toxic PrrC homologs. We indentify 17 candidate active site residues in the NTPase module that are essential for toxicity in yeast when EcoPrrC is expressed at high gene dosage. Their functions could be educed by integrating mutational data with the atomic structure of the transition-state complex of a homologous ABC protein.

  15. Structure-function relations in the NTPase domain of the antiviral tRNA ribotoxin Escherichia coli PrrC

    Energy Technology Data Exchange (ETDEWEB)

    Meineke, Birthe; Shuman, Stewart, E-mail: s-shuman@ski.mskcc.org

    2012-06-05

    Breakage of tRNA by Escherichia coli anticodon nuclease PrrC (EcoPrrC) underlies a host antiviral response to phage T4 infection. Expression of EcoPrrC is cytocidal in yeast, signifying that PrrC ribotoxicity crosses phylogenetic domain boundaries. EcoPrrC consists of an N-terminal NTPase module that resembles ABC transporters and a C-terminal nuclease module that is sui generis. PrrC homologs are prevalent in many other bacteria. Here we report that Haemophilus influenzae PrrC is toxic in E. coli and yeast. To illuminate structure-activity relations, we conducted a new round of mutational analysis of EcoPrrC guided by primary structure conservation among toxic PrrC homologs. We indentify 17 candidate active site residues in the NTPase module that are essential for toxicity in yeast when EcoPrrC is expressed at high gene dosage. Their functions could be educed by integrating mutational data with the atomic structure of the transition-state complex of a homologous ABC protein.

  16. Rec8p, a meiotic recombination and sister chromatid cohesion phosphoprotein of the Rad21p family conserved from fision yeast to humans.

    NARCIS (Netherlands)

    S. Parisi; M.J. McKay (Michael); M. Molnar; M.A. Thompson (Anne); P.J. van der Spek (Peter); E. van Drunen-Schoenmaker; R. Kanaar (Roland); E. Lehmann; J.H.J. Hoeijmakers (Jan); J. Kohli

    1999-01-01

    textabstractOur work and that of others defined mitosis-specific (Rad21 subfamily) and meiosis-specific (Rec8 subfamily) proteins involved in sister chromatid cohesion in several eukaryotes, including humans. Mutation of the fission yeast Schizosaccharomyces pombe rec8 gene was previously shown to

  17. Yeast-2-Hybrid data file showing progranulin interactions in human fetal brain and bone marrow libraries

    Directory of Open Access Journals (Sweden)

    Irmgard Tegeder

    2016-12-01

    Full Text Available Progranulin deficiency in humans is associated with neurodegeneration. Its mechanisms are not yet fully understood. We performed a Yeast-2-Hybrid screen using human full-length progranulin as bait to assess the interactions of progranulin. Progranulin was screened against human fetal brain and human bone marrow libraries using the standard Matchmaker technology (Clontech. This article contains the full Y2H data table, including blast results and sequences, a sorted table according to selection criteria for likely positive, putatively positive, likely false and false preys, and tables showing the gene ontology terms associated with the likely and putative preys of the brain and bone marrow libraries. The interactions with autophagy proteins were confirmed and functionally analyzed in "Progranulin overexpression in sensory neurons attenuates neuropathic pain in mice: Role of autophagy" (C. Altmann, S. Hardt, C. Fischer, J. Heidler, H.Y. Lim, A. Haussler, B. Albuquerque, B. Zimmer, C. Moser, C. Behrends, F. Koentgen, I. Wittig, M.H. Schmidt, A.M. Clement, T. Deller, I. Tegeder, 2016 [1].

  18. Fatty acids from oleaginous yeasts and yeast-like fungi and their potential applications.

    Science.gov (United States)

    Xue, Si-Jia; Chi, Zhe; Zhang, Yu; Li, Yan-Feng; Liu, Guang-Lei; Jiang, Hong; Hu, Zhong; Chi, Zhen-Ming

    2018-02-01

    Oleaginous yeasts, fatty acids biosynthesis and regulation in the oleaginous yeasts and the fatty acids from the oleaginous yeasts and their applications are reviewed in this article. Oleaginous yeasts such as Rhodosporidium toruloides, Yarrowia lipolytica, Rhodotorula mucilaginosa, and Aureobasidium melanogenum, which can accumulate over 50% lipid of their cell dry weight, have many advantages over other oleaginous microorganisms. The fatty acids from the oleaginous yeasts have many potential applications. Many oleaginous yeasts have now been genetically modified to over-produce fatty acids and their derivatives. The most important features of the oleaginous yeasts are that they have special enzymatic systems for enhanced biosynthesis and regulation of fatty acids in their lipid particles. Recently, some oleaginous yeasts such as R. toruloides have been found to have a unique fatty acids synthetase and other oleaginous yeasts such as A. melanogenum have a unique highly reducing polyketide synthase (HR-PKS) involved in the biosynthesis of hydroxyl fatty acids. It is necessary to further enhance lipid biosynthesis using metabolic engineering and explore new applications of fatty acids in biotechnology.

  19. Multiplex polymerase chain reaction for identification of Escherichia coli, Escherichia albertii and Escherichia fergusonii.

    Science.gov (United States)

    Lindsey, Rebecca L; Garcia-Toledo, L; Fasulo, D; Gladney, L M; Strockbine, N

    2017-09-01

    Escherichia coli, Escherichia albertii, and Escherichia fergusonii are closely related bacteria that can cause illness in humans, such as bacteremia, urinary tract infections and diarrhea. Current identification strategies for these three species vary in complexity and typically rely on the use of multiple phenotypic and genetic tests. To facilitate their rapid identification, we developed a multiplex PCR assay targeting conserved, species-specific genes. We used the Daydreamer™ (Pattern Genomics, USA) software platform to concurrently analyze whole genome sequence assemblies (WGS) from 150 Enterobacteriaceae genomes (107 E. coli, 5 Shigella spp., 21 E. albertii, 12 E. fergusonii and 5 other species) and design primers for the following species-specific regions: a 212bp region of the cyclic di-GMP regulator gene (cdgR, AW869_22935 from genome K-12 MG1655, CP014225) for E. coli/Shigella; a 393bp region of the DNA-binding transcriptional activator of cysteine biosynthesis gene (EAKF1_ch4033 from genome KF1, CP007025) for E. albertii; and a 575bp region of the palmitoleoyl-acyl carrier protein (ACP)-dependent acyltransferase (EFER_0790 from genome ATCC 35469, CU928158) for E. fergusonii. We incorporated the species-specific primers into a conventional multiplex PCR assay and assessed its performance with a collection of 97 Enterobacteriaceae strains. The assay was 100% sensitive and specific for detecting the expected species and offers a quick and accurate strategy for identifying E. coli, E. albertii, and E. fergusonii in either a single reaction or by in silico PCR with sequence assemblies. Published by Elsevier B.V.

  20. Eubiotics for Food Security at Farm Level: Yeast Cell Wall Products and Their Antimicrobial Potential Against Pathogenic Bacteria.

    Science.gov (United States)

    Santovito, Elisa; Greco, Donato; Logrieco, Antonio F; Avantaggiato, Giuseppina

    2018-06-06

    The population increase in the last century was the first cause of the industrialization of animal productions, together with the necessity to satisfy the high food demand and the lack of space and land for the husbandry practices. As a consequence, the farmers moved from extensive to intensive agricultural systems and introduced new practices, such as the administration of antimicrobial drugs. Antibiotics were then used as growth promoters and for disease prevention. The uncontrolled and continuous use of antibiotics contributed to the spread of antibiotic resistance in animals, and this had adverse impacts on human health. This emergence led the European Union, in 2003, to ban the marketing and use of antibiotics as growth promoters, and for prophylaxis purposes from January 2006. This ban caused problems in farms, due to the decrease in animal performances (weight gain, feed conversion ratio, reproduction, etc.), and the rise in the incidence of certain diseases, such as those induced by Clostridium perfringens, Salmonella, Escherichia coli, and Listeria monocytogenes. The economic losses due to the ban increased the interest in researching alternative strategies for the prophylaxis of infectious diseases and for health and growth promotion, such as feed additives. Yeast-based materials, such as cell wall extract, represent promising alternatives to antibiotics, on the base of their prebiotic activity and their claimed capacity to bind enteropathogenic bacteria. Several authors reported examples of the effectiveness of yeast cell wall products in adsorbing bacteria, but there is a lack of knowledge on the mechanisms involved in this interaction. The purpose of this review is to provide an overview of the current approaches used for the control of pathogenic bacteria in feed, with a particular focus on the use of yeast-derived materials proposed to control zoonoses at farm level, and on their effect on animal health.

  1. A Comparison of Two Yeast MnSODs: Mitochondrial Saccharomyces cerevisiae versus Cytosolic Candida albicans

    International Nuclear Information System (INIS)

    Sheng, Y.; Cabelli, D.; Stich, T.A.; Barnese, K.; Gralla, E.B.; Cascio, D.; Britt, R.D.; Valentine, J.S.

    2011-01-01

    Human MnSOD is significantly more product-inhibited than bacterial MnSODs at high concentrations of superoxide (O 2 - ). This behavior limits the amount of H 2 O 2 produced at high [O 2 - ]; its desirability can be explained by the multiple roles of H 2 O 2 in mammalian cells, particularly its role in signaling. To investigate the mechanism of product inhibition in MnSOD, two yeast MnSODs, one from Saccharomyces cerevisiae mitochondria (ScMnSOD) and the other from Candida albicans cytosol (CaMnSODc), were isolated and characterized. ScMnSOD and CaMnSODc are similar in catalytic kinetics, spectroscopy, and redox chemistry, and they both rest predominantly in the reduced state (unlike most other MnSODs). At high [O 2 - ], the dismutation efficiencies of the yeast MnSODs surpass those of human and bacterial MnSODs, due to very low level of product inhibition. Optical and parallel-mode electron paramagnetic resonance (EPR) spectra suggest the presence of two Mn 3+ species in yeast Mn 3+ SODs, including the well-characterized 5-coordinate Mn 3+ species and a 6-coordinate L-Mn 3+ species with hydroxide as the putative sixth ligand (L). The first and second coordination spheres of ScMnSOD are more similar to bacterial than to human MnSOD. Gln154, an H-bond donor to the Mn-coordinated solvent molecule, is slightly further away from Mn in yeast MnSODs, which may result in their unusual resting state. Mechanistically, the high efficiency of yeast MnSODs could be ascribed to putative translocation of an outer-sphere solvent molecule, which could destabilize the inhibited complex and enhance proton transfer from protein to peroxide. Our studies on yeast MnSODs indicate the unique nature of human MnSOD in that it predominantly undergoes the inhibited pathway at high [O 2 - ].

  2. Yeast-expressed human membrane protein aquaporin-1 yields excellent resolution of solid-state MAS NMR spectra

    International Nuclear Information System (INIS)

    Emami, Sanaz; Fan Ying; Munro, Rachel; Ladizhansky, Vladimir; Brown, Leonid S.

    2013-01-01

    One of the biggest challenges in solid-state NMR studies of membrane proteins is to obtain a homogeneous natively folded sample giving high spectral resolution sufficient for structural studies. Eukaryotic membrane proteins are especially difficult and expensive targets in this respect. Methylotrophic yeast Pichia pastoris is a reliable producer of eukaryotic membrane proteins for crystallography and a promising economical source of isotopically labeled proteins for NMR. We show that eukaryotic membrane protein human aquaporin 1 can be doubly ( 13 C/ 15 N) isotopically labeled in this system and functionally reconstituted into phospholipids, giving excellent resolution of solid-state magic angle spinning NMR spectra.

  3. Toward low-cost affinity reagents: lyophilized yeast-scFv probes specific for pathogen antigens.

    Directory of Open Access Journals (Sweden)

    Sean A Gray

    Full Text Available The generation of affinity reagents, usually monoclonal antibodies, remains a critical bottleneck in biomedical research and diagnostic test development. Recombinant antibody-like proteins such as scFv have yet to replace traditional monoclonal antibodies in antigen detection applications, in large part because of poor performance of scFv in solution. To address this limitation, we have developed assays that use whole yeast cells expressing scFv on their surfaces (yeast-scFv in place of soluble purified scFv or traditional monoclonal antibodies. In this study, a nonimmune library of human scFv displayed on the surfaces of yeast cells was screened for clones that bind to recombinant cyst proteins of Entamoeba histolytica, an enteric pathogen of humans. Selected yeast-scFv clones were stabilized by lyophilization and used in detection assay formats in which the yeast-scFv served as solid support-bound monoclonal antibodies. Specific binding of antigen to the yeast-scFv was detected by staining with rabbit polyclonal antibodies. In flow cytometry-based assays, lyophilized yeast-scFv reagents retained full binding activity and specificity for their cognate antigens after 4 weeks of storage at room temperature in the absence of desiccants or stabilizers. Because flow cytometry is not available to all potential assay users, an immunofluorescence assay was also developed that detects antigen with similar sensitivity and specificity. Antigen-specific whole-cell yeast-scFv reagents can be selected from nonimmune libraries in 2-3 weeks, produced in vast quantities, and packaged in lyophilized form for extended shelf life. Lyophilized yeast-scFv show promise as low cost, renewable alternatives to monoclonal antibodies for diagnosis and research.

  4. The flexible feedstock concept in Industrial Biotechnology: Metabolic engineering of Escherichia coli, Corynebacterium glutamicum, Pseudomonas, Bacillus and yeast strains for access to alternative carbon sources.

    Science.gov (United States)

    Wendisch, Volker F; Brito, Luciana Fernandes; Gil Lopez, Marina; Hennig, Guido; Pfeifenschneider, Johannes; Sgobba, Elvira; Veldmann, Kareen H

    2016-09-20

    Most biotechnological processes are based on glucose that is either present in molasses or generated from starch by enzymatic hydrolysis. At the very high, million-ton scale production volumes, for instance for fermentative production of the biofuel ethanol or of commodity chemicals such as organic acids and amino acids, competing uses of carbon sources e.g. in human and animal nutrition have to be taken into account. Thus, the biotechnological production hosts E. coli, C. glutamicum, pseudomonads, bacilli and Baker's yeast used in these large scale processes have been engineered for efficient utilization of alternative carbon sources. This flexible feedstock concept is central to the use of non-glucose second and third generation feedstocks in the emerging bioeconomy. The metabolic engineering efforts to broaden the substrate scope of E. coli, C. glutamicum, pseudomonads, B. subtilis and yeasts to include non-native carbon sources will be reviewed. Strategies to enable simultaneous consumption of mixtures of native and non-native carbon sources present in biomass hydrolysates will be summarized and a perspective on how to further increase feedstock flexibility for the realization of biorefinery processes will be given. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. L-arabinose fermenting yeast

    Science.gov (United States)

    Zhang, Min; Singh, Arjun; Suominen, Pirkko; Knoshaug, Eric; Franden, Mary Ann; Jarvis, Eric

    2013-02-12

    An L-arabinose utilizing yeast strain is provided for the production of ethanol by introducing and expressing bacterial araA, araB and araD genes. L-arabinose transporters are also introduced into the yeast to enhance the uptake of arabinose. The yeast carries additional genomic mutations enabling it to consume L-arabinose, even as the only carbon source, and to produce ethanol. A yeast strain engineered to metabolize arabinose through a novel pathway is also disclosed. Methods of producing ethanol include utilizing these modified yeast strains.

  6. Enterohemorrhagic Escherichia coli (EHEC

    Directory of Open Access Journals (Sweden)

    Abdullah Kilic

    2011-08-01

    Full Text Available Escherichia coli is a bacterium that is commonly found in the gut of humans and warm-blooded animals. Most strains of E. coli are harmless for human. E. coli O157:H7 is the most common member of a group of pathogenic E. coli strains known variously as enterohaemorrhagic, verocytotoxin-producing, or Shiga-toxin-producing organisms. EHEC bacterium is the major cause of haemorrhagic colitis and haemolytic uraemic syndrome. The reservoir of this pathogen appears to be mainly cattle and other ruminants such as camels. It is transmitted to humans primarily through consumption of contaminated foods. [TAF Prev Med Bull 2011; 10(4.000: 387-388

  7. Diversity of culturable yeasts associated with zoanthids from Brazilian reef and its relation with anthropogenic disturbance.

    Science.gov (United States)

    Paulino, Gustavo Vasconcelos Bastos; Félix, Ciro Ramon; Broetto, Leonardo; Landell, Melissa Fontes

    2017-10-15

    Some of the main threats to coral reefs come from human actions on marine environment, such as tourism, overfishing and pollution from urban development. While several studies have demonstrated an association between bacteria and corals, demonstrating how these communities react to different anthropogenic stressors, yeast communities associated with corals have received far less attention from researchers. The aim of this work was therefore to describe cultivable yeasts associated with three coral species and to evaluate the influence of sewage discharge on yeasts community. We obtained 130 isolates, mostly belonging to phylum Ascomycota and many of them had previously been isolated from human samples or are considered pathogens. The mycobiota was more similar among corals collected from the same reef, indicating that the composition of reef yeast community is more influenced by environmental conditions than host species. We suggest further studies to elucidate which factors are most influential on the composition of the coral-associated yeast community. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Yeast RNA viruses as indicators of exosome activity: human exosome hCsl4p participates in RNA degradation in Saccharomyces cerevisiae'.

    Science.gov (United States)

    Ramírez-Garrastacho, Manuel; Esteban, Rosa

    2011-12-01

    The exosome is an evolutionarily conserved 10-mer complex involved in RNA metabolism, located in both the nucleus and the cytoplasm. The cytoplasmic exosome plays an important role in mRNA turnover through its 3'→5' exonucleolytic activity. The superkiller (SKI) phenotype of yeast was originally identified as an increase of killer toxin production due to elevated levels of the L-A double-stranded RNA (dsRNA) Totivirus and its satellite toxin-encoding M dsRNA. Most SKI genes were later shown to be either components of the exosome or modulators of its activity. Variations in the amount of Totivirus are, thus, good indicators of yeast exosome activity, and can be used to analyse its components. Furthermore, if exosome proteins of higher eukaryotes were functional in S. cerevisiae, these viruses would provide a simple tool to analyse their function. In this work, we have found that hCSL4, the human orthologue of SKI4 in the yeast exosome, rescues the null phenotype of the deletion mutant. hCsl4p shares with Ski4p conserved S1 RNA-binding domains, but lacks the N-terminal third of Ski4p. Nevertheless, it interacts with the Dis3p exonuclease of yeast exosome, and partially complements the superkiller phenotype of ski4-1 mutation. The elimination of the N-terminal third of Ski4p does not affect its activity, indicating that it is dispensable for RNA degradation. We have also identified the point mutation G152E in hCSL4, equivalent to the ski4-1 mutation G253E, which impairs the activity of the protein, thus validating our approach of using yeast RNA virus to analyse the exosome of higher eukaryotes. Copyright © 2011 John Wiley & Sons, Ltd.

  9. Differentiation of the emerging human pathogens Trichosporon asahii and Trichosporon asteroides from other pathogenic yeasts and moulds by using species-specific monoclonal antibodies.

    Directory of Open Access Journals (Sweden)

    Genna E Davies

    Full Text Available The fungal genus Trichosporon contains emerging opportunistic pathogens of humans, and is the third most commonly isolated non-candidal yeast from humans. Trichosporon asahii and T. asteroides are the most important species causing disseminated disease in immunocompromised patients, while inhalation of T. asahii spores is the most important cause of summer-type hypersensitivity pneumonitis in healthy individuals. Trichosporonosis is misdiagnosed as candidiasis or cryptococcosis due to a lack of awareness and the ambiguity of diagnostic tests for these pathogens. In this study, hybridoma technology was used to produce two murine monoclonal antibodies (MAbs, CA7 and TH1, for detection and differentiation of Trichosporon from other human pathogenic yeasts and moulds. The MAbs react with extracellular antigens from T. asahii and T. asteroides, but do not recognise other related Trichosporon spp., or unrelated pathogenic yeasts and moulds including Candida, Cryptococcus, Aspergillus, Fusarium, and Scedosporium spp., or the etiologic agents of mucormycosis. Immunofluorescence and Western blotting studies show that MAb CA7, an immunoglobulin G1 (IgG1, binds to a major 60 kDa glycoprotein antigen produced on the surface of hyphae, while TH1, an immunoglobulin M (IgM, binds to an antigen produced on the surface of conidia. The MAbs were used in combination with a standard mycological growth medium (Sabouraud Dextrose Agar to develop an enzyme-linked immunosorbent assay (ELISA for differentiation of T. asahii from Candida albicans and Cryptococcus neoformans in single and mixed species cultures. The MAbs represent a major advance in the identification of T. asahii and T. asteroides using standard mycological identification methods.

  10. Functional toxicogenomic profiling expands insight into modulators of formaldehyde toxicity in yeast

    Directory of Open Access Journals (Sweden)

    Matthew North

    2016-11-01

    Full Text Available Formaldehyde (FA is a commercially important chemical with numerous and diverse uses. Accordingly, occupational and environmental exposure to FA is prevalent worldwide. Various adverse effects, including nasopharyngeal, sinonasal, and lymphohematopoietic cancers, have been linked to FA exposure, prompting designation of FA as a human carcinogen by U.S. and international scientific entities. Although the mechanism(s of FA toxicity have been well studied, additional insight is needed in regard to the genetic requirements for FA tolerance. In this study, a functional toxicogenomics approach was utilized in the model eukaryotic yeast Saccharomyces cerevisiae to identify genes and cellular processes modulating the cellular toxicity of FA. Our results demonstrate mutant strains deficient in multiple DNA repair pathways–including homologous recombination, single strand annealing, and postreplication repair–were sensitive to FA, indicating FA may cause various forms of DNA damage in yeast. The SKI complex and its associated factors, which regulate mRNA degradation by the exosome, were also required for FA tolerance, suggesting FA may have unappreciated effects on RNA stability. Furthermore, various strains involved in osmoregulation and stress response were sensitive to FA. Together, our results are generally consistent with FA-mediated damage to both DNA and RNA. Considering DNA repair and RNA degradation pathways are evolutionarily conserved from yeast to humans, mechanisms of FA toxicity identified in yeast may be relevant to human disease and genetic susceptibility.

  11. Expression of a Human Cytochrome P450 in Yeast Permits Analysis of Pathways for Response to and Repair of Aflatoxin-Induced DNA Damage†

    OpenAIRE

    Guo, Yingying; Breeden, Linda L.; Zarbl, Helmut; Preston, Bradley D.; Eaton, David L.

    2005-01-01

    Aflatoxin B1 (AFB1) is a human hepatotoxin and hepatocarcinogen produced by the mold Aspergillus flavus. In humans, AFB1 is primarily bioactivated by cytochrome P450 1A2 (CYP1A2) and 3A4 to a genotoxic epoxide that forms N7-guanine DNA adducts. A series of yeast haploid mutants defective in DNA repair and cell cycle checkpoints were transformed with human CYP1A2 to investigate how these DNA adducts are repaired. Cell survival and mutagenesis following aflatoxin B1 treatment was assayed in str...

  12. Yeast prions: structure, biology, and prion-handling systems.

    Science.gov (United States)

    Wickner, Reed B; Shewmaker, Frank P; Bateman, David A; Edskes, Herman K; Gorkovskiy, Anton; Dayani, Yaron; Bezsonov, Evgeny E

    2015-03-01

    A prion is an infectious protein horizontally transmitting a disease or trait without a required nucleic acid. Yeast and fungal prions are nonchromosomal genes composed of protein, generally an altered form of a protein that catalyzes the same alteration of the protein. Yeast prions are thus transmitted both vertically (as genes composed of protein) and horizontally (as infectious proteins, or prions). Formation of amyloids (linear ordered β-sheet-rich protein aggregates with β-strands perpendicular to the long axis of the filament) underlies most yeast and fungal prions, and a single prion protein can have any of several distinct self-propagating amyloid forms with different biological properties (prion variants). Here we review the mechanism of faithful templating of protein conformation, the biological roles of these prions, and their interactions with cellular chaperones, the Btn2 and Cur1 aggregate-handling systems, and other cellular factors governing prion generation and propagation. Human amyloidoses include the PrP-based prion conditions and many other, more common amyloid-based diseases, several of which show prion-like features. Yeast prions increasingly are serving as models for the understanding and treatment of many mammalian amyloidoses. Patients with different clinical pictures of the same amyloidosis may be the equivalent of yeasts with different prion variants. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Oxygen requirement for near-UV mediated cytotoxicity of α-terthienyl to Escherichia coli and Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Arnason, T.; Chan, G.F.Q.; Wat, C.K.; Downum, K.; Towers, G.H.N.

    1981-01-01

    The photosensitizing activity of α-terthienyl, a naturally occurring compound from Tagetes has been studied. Fluence-response curves for the effect of α-terthienyl and near-UV radiation on Escherichia coli have been prepared. Photosensitization of E. coli was enhanced in aerobic as compared to anaerobic conditions and exogenous superoxide dismutase had a slight protective effect. Sodium azide, a quencher of 1 O 2 , protected yeast cells from photosensitization with α-terthienyl. The available evidence suggests that α-terthienyl acts as a photodynamic sensitizer in vivo. (author)

  14. PREPARATION OF ZINC ENRICHED YEAST (SACCHAROMYCES CEREVISIAE BY CULTIVATION WITH DIFFERENT ZINC SALTS

    Directory of Open Access Journals (Sweden)

    Ľuboš Harangozo

    2012-02-01

    Full Text Available The yeast Saccharomyces cerevisiae is the best known microorganism and therefore widely used in many branches of industry. This study aims to investigate the accumulation of three inorganic zinc salts. Our research presents the ability of this yeast to absorb zinc from liquid medium and such enriched biomass use as a potential source of microelements in animal and/or human nutrition. It was found that the addition of different zinc forms, i.e. zinc nitrate, zinc sulphate and zinc chloride in fixed concentrations of 0, 25, 50 and 100 mg.100 ml-1 did not affect the amount of dry yeast biomass yielded, i.e. 1.0 – 1.2 g of yeast cells from 100 ml of cultivation medium, while higher presence of zinc solutions caused significantly lower yield of yeast biomass. The highest amount of zinc in yeast cells was achieved when added in the form of zinc nitrate in concentration of 200 mg.100 ml-1 YPD medium. The increment of intracellular zinc was up to 18.5 mg.g-1 of yeast biomass.

  15. [Expression, purification and immunogenicity of human papillomavirus type 11 virus-like particles from Escherichia coli].

    Science.gov (United States)

    Yan, Chunyan; Li, Shaowei; Wang, Jin; Wei, Minxi; Huang, Bo; Zhuang, Yudi; Li, Zhongyi; Pan, Huirong; Zhang, Jun; Xia, Ningshao

    2009-11-01

    To produce human papillomavirus type 11 virus-like particles (HPV11 VLPs) from Escherichia coli and to investigate its immunogenicity and type cross neutralization nature. We expressed the major capsid protein of HPV11 (HPV11-L1) in Escherichia coli ER2566 in non fusion fashion and purified by amino sulfate precipitation, ion-exchange chromatography and hydrophobic interaction chromatography, sequentially. Then we removed the reductant DTT to have the purified HPV11-L1 self-assemble into VLPs in vitro. We investigated the morphology of these VLPs with dynamic light scattering and transmission electron microscopy. We assayed the immunogenicity of the resultant HPV11 VLPs by vaccinations on mice and evaluated by HPV6/11/16/18 pseudovirion neutralization cell models. We expressed HPV11 L1 in Escherichia coli with two forms, soluble and inclusion body. The soluble HPV11 L1 with over 95% purity can self assemble to VLPs in high efficiency. Morphologically, these VLPs were globular, homogeneous and with a diameter of - 50 nm, which is quite similar with native HPV11 virions. The half effective dosage (ED50) of HPV11 VLPs is 0.031 microg, and the maximum titer of neutralizing antibody elicited is averaged to 10(6). The cross neutralization activity (against HPV6/16/18) of the anti-HPV11 serum was found to have exact correlation to the inter-type homology in amino acid alignment. We can provide HPV11 VLPs with highly immunogenicity from prokaryote expression system, which may pave a new way for research and development of prophylactic vaccine for HPV11.

  16. Prion-based memory of heat stress in yeast.

    Science.gov (United States)

    Chernova, Tatiana A; Chernoff, Yury O; Wilkinson, Keith D

    2017-05-04

    Amyloids and amyloid-based prions are self-perpetuating protein aggregates which can spread by converting a normal protein of the same sequence into a prion form. They are associated with diseases in humans and mammals, and control heritable traits in yeast and other fungi. Some amyloids are implicated in biologically beneficial processes. As prion formation generates reproducible memory of a conformational change, prions can be considered as molecular memory devices.  We have demonstrated that in yeast, stress-inducible cytoskeleton-associated protein Lsb2 forms a metastable prion in response to high temperature. This prion promotes conversion of other proteins into prions and can persist in a fraction of cells for a significant number of cell generations after stress, thus maintaining the memory of stress in a population of surviving cells. Acquisition of an amino acid substitution required for Lsb2 to form a prion coincides with acquisition of increased thermotolerance in the evolution of Saccharomyces yeast. Thus the ability to form an Lsb2 prion in response to stress coincides with yeast adaptation to growth at higher temperatures. These findings intimately connect prion formation to the cellular response to environmental stresses.

  17. The Escherichia coli Tus-Ter replication fork barrier causes site-specific DNA replication perturbation in yeast

    DEFF Research Database (Denmark)

    Larsen, Nicolai B; Sass, Ehud; Suski, Catherine

    2014-01-01

    Replication fork (RF) pausing occurs at both 'programmed' sites and non-physiological barriers (for example, DNA adducts). Programmed RF pausing is required for site-specific DNA replication termination in Escherichia coli, and this process requires the binding of the polar terminator protein, Tus...... as a versatile, site-specific, heterologous DNA replication-perturbing system, with a variety of potential applications....

  18. Fimbrial adhesins from extraintestinal Escherichia coli

    DEFF Research Database (Denmark)

    Klemm, Per; Hancock, Viktoria; Schembri, Mark A.

    2010-01-01

    Extraintestinal pathogenic Escherichia coli (ExPEC) represent an important subclass of E. coli that cause a wide spectrum of diseases in human and animal hosts. Fimbriae are key virulence factors of ExPEC strains. These long surface located rod-shaped organelles mediate receptor-specific attachment...

  19. Construction of high-quality Caco-2 three-frame cDNA library and its application to yeast two-hybrid for the human astrovirus protein-protein interaction.

    Science.gov (United States)

    Zhao, Wei; Li, Xin; Liu, Wen-Hui; Zhao, Jian; Jin, Yi-Ming; Sui, Ting-Ting

    2014-09-01

    Human epithelial colorectal adenocarcinoma (Caco-2) cells are widely used as an in vitro model of the human small intestinal mucosa. Caco-2 cells are host cells of the human astrovirus (HAstV) and other enteroviruses. High quality cDNA libraries are pertinent resources and critical tools for protein-protein interaction research, but are currently unavailable for Caco-2 cells. To construct a three-open reading frame, full length-expression cDNA library from the Caco-2 cell line for application to HAstV protein-protein interaction screening, total RNA was extracted from Caco-2 cells. The switching mechanism at the 5' end of the RNA transcript technique was used for cDNA synthesis. Double-stranded cDNA was digested by Sfi I and ligated to reconstruct a pGADT7-Sfi I three-frame vector. The ligation mixture was transformed into Escherichia coli HST08 premium electro cells by electroporation to construct the primary cDNA library. The library capacity was 1.0×10(6)clones. Gel electrophoresis results indicated that the fragments ranged from 0.5kb to 4.2kb. Randomly picked clones show that the recombination rate was 100%. The three-frame primary cDNA library plasmid mixture (5×10(5)cfu) was also transformed into E. coli HST08 premium electro cells, and all clones were harvested to amplify the cDNA library. To detect the sufficiency of the cDNA library, HAstV capsid protein as bait was screened and tested against the Caco-2 cDNA library by a yeast two-hybrid (Y2H) system. A total of 20 proteins were found to interact with the capsid protein. These results showed that a high-quality three-frame cDNA library from Caco-2 cells was successfully constructed. This library was efficient for the application to the Y2H system, and could be used for future research. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. The asymptomatic bacteriuria Escherichia coli strain 83972 outcompetes uropathogenic E. coli strains in human urine

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Ulett, G.C.; Schembri, M.A.

    2006-01-01

    Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU). In contrast to uropathogenic E. coli (UPEC), which causes symptomatic urinary tract infections (UTI), very little is known about the mechanisms by which these strains colonize the human urinary tract....... The prototype ABU E. coli strain 83972 was originally isolated from a girl who had carried it asymptomatically for 3 years. Deliberate colonization of UTI-susceptible individuals with E. coli 83972 has been used successfully as an alternative approach for the treatment of patients who are refractory...... to conventional therapy. Colonization with strain 83972 appears to prevent infection with UPEC strains in such patients despite the fact that this strain is unable to express the primary adhesins involved in UTI, viz. P and type 1 fimbriae. Here we investigated the growth characteristics of E. coli 83972 in human...

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

    Science.gov (United States)

    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.

  2. Yeast Flocculation—Sedimentation and Flotation

    Directory of Open Access Journals (Sweden)

    Graham G. Stewart

    2018-04-01

    Full Text Available Unlike most fermentation alcohol beverage production processes, brewers recycle their yeast. This is achieved by employing a yeast culture’s: flocculation, adhesion, sedimentation, flotation, and cropping characteristics. As a consequence of yeast recycling, the quality of the cropped yeast culture’s characteristics is critical. However, the other major function of brewer’s yeast is to metabolise wort into ethanol, carbon dioxide, glycerol, and other fermentation products, many of which contribute to beer’s overall flavour characteristics. This review will only focus on brewer’s yeast flocculation characteristics.

  3. Expression and structural and functional properties of human ferritin L-chain from Escherichia coli

    International Nuclear Information System (INIS)

    Levi, S.; Salfeld, J.; Franceschinelli, F.; Cozzi, A.; Dorner, M.H.; Arosio, P.

    1989-01-01

    The human ferritin L-chain cDNA was cloned into a vector for overproduction in Escherichia coli, under the regulation of a λ promoter. The plasmid obtained contains the full L-chain coding region modified at the first two codons. It is able to direct the synthesis of the L-chain which can constitute up to 15% of the total soluble protein of bacterial extract. The L-chains assemble to form a ferritin homopolymer with electrophoretic mobility, molecular weight, thermal stability, spectroscopic, and immunological properties analogous to natural ferritin from human liver (95% L-chain). This recombinant L-ferritin is able to incorporate and retain iron in solution at physiological pH values. At variance with the H-ferritin, the L form does not uptake iron at acidic pH values and does not show detectable ferroxidase activity. It is concluded that ferritin L-chain lacks the ferroxidase site present in the H-chain and that the two chains may have specialized functions in intracellular iron metabolism

  4. L-arabinose fermenting yeast

    Science.gov (United States)

    Zhang, Min; Singh, Arjun; Knoshaug, Eric; Franden, Mary Ann; Jarvis, Eric; Suominen, Pirkko

    2010-12-07

    An L-arabinose utilizing yeast strain is provided for the production of ethanol by introducing and expressing bacterial araA, araB and araD genes. L-arabinose transporters are also introduced into the yeast to enhance the uptake of arabinose. The yeast carries additional genomic mutations enabling it to consume L-arabinose, even as the only carbon source, and to produce ethanol. Methods of producing ethanol include utilizing these modified yeast strains. ##STR00001##

  5. A novel model to study neonatal Escherichia coli sepsis and the effect of treatment on the human immune system using humanized mice.

    Science.gov (United States)

    Schlieckau, Florian; Schulz, Daniela; Fill Malfertheiner, Sara; Entleutner, Kathrin; Seelbach-Goebel, Birgit; Ernst, Wolfgang

    2018-04-19

    Neonatal sepsis is a serious threat especially for preterm infants. As existing in vitro and in vivo models have limitations, we generated a novel neonatal sepsis model using humanized mice and tested the effect of Betamethasone and Indomethacin which are used in the clinic in case of premature birth. Humanized mice were infected with Escherichia coli (E. coli). Subsequently, the effect of the infection itself, and treatment with Betamethasone and Indomethacin on survival, recovery, bacterial burden, leukocyte populations, and cytokine production, was analyzed. The human immune system in the animals responded with leukocyte trafficking to the site of infection and granulopoiesis in the bone marrow. Treatment with Indomethacin had no pronounced effect on the immune system or bacterial burden. Betamethasone induced a decline of splenocytes. The human immune system in humanized mice responds to the infection, making them a suitable model to study neonatal E. coli sepsis and the immune response of the neonatal immune system. Treatment with Betamethasone could have potential negative long-term effects for the immune system of the child. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. Drosophila Regulate Yeast Density and Increase Yeast Community Similarity in a Natural Substrate

    OpenAIRE

    Stamps, Judy A.; Yang, Louie H.; Morales, Vanessa M.; Boundy-Mills, Kyria L.

    2012-01-01

    Drosophila melanogaster adults and larvae, but especially larvae, had profound effects on the densities and community structure of yeasts that developed in banana fruits. Pieces of fruit exposed to adult female flies previously fed fly-conditioned bananas developed higher yeast densities than pieces of the same fruits that were not exposed to flies, supporting previous suggestions that adult Drosophila vector yeasts to new substrates. However, larvae alone had dramatic effects on yeast densit...

  7. Characterization of TRZ1, a yeast homolog of the human candidate prostate cancer susceptibility gene ELAC2 encoding tRNase Z

    Directory of Open Access Journals (Sweden)

    Chen Yuan

    2005-05-01

    Full Text Available Abstract Background In humans, mutation of ELAC2 is associated with an increased risk of prostate cancer. ELAC2 has been shown to have tRNase Z activity and is associated with the γ-tubulin complex. Results In this work, we show that the yeast homolog of ELAC2, encoded by TRZ1 (tRNase Z 1, is involved genetically in RNA processing. The temperature sensitivity of a trz1 mutant can be rescued by multiple copies of REX2, which encodes a protein with RNA 3' processing activity, suggesting a role of Trz1p in RNA processing in vivo. Trz1p has two putative nucleotide triphosphate-binding motifs (P-loop and a conserved histidine motif. The histidine motif and the putative nucleotide binding motif at the C-domain are important for Trz1p function because mutant proteins bearing changes to the critical residues in these motifs are unable to rescue deletion of TRZ1. The growth defect exhibited by trz1 yeast is not complemented by the heterologous ELAC2, suggesting that Trz1p may have additional functions in yeast. Conclusion Our results provide genetic evidence that prostate cancer susceptibility gene ELAC2 may be involved in RNA processing, especially rRNA processing and mitochondrial function.

  8. MTO1 mutations are associated with hypertrophic cardiomyopathy and lactic acidosis and cause respiratory chain deficiency in humans and yeast.

    Science.gov (United States)

    Baruffini, Enrico; Dallabona, Cristina; Invernizzi, Federica; Yarham, John W; Melchionda, Laura; Blakely, Emma L; Lamantea, Eleonora; Donnini, Claudia; Santra, Saikat; Vijayaraghavan, Suresh; Roper, Helen P; Burlina, Alberto; Kopajtich, Robert; Walther, Anett; Strom, Tim M; Haack, Tobias B; Prokisch, Holger; Taylor, Robert W; Ferrero, Ileana; Zeviani, Massimo; Ghezzi, Daniele

    2013-11-01

    We report three families presenting with hypertrophic cardiomyopathy, lactic acidosis, and multiple defects of mitochondrial respiratory chain (MRC) activities. By direct sequencing of the candidate gene MTO1, encoding the mitochondrial-tRNA modifier 1, or whole exome sequencing analysis, we identified novel missense mutations. All MTO1 mutations were predicted to be deleterious on MTO1 function. Their pathogenic role was experimentally validated in a recombinant yeast model, by assessing oxidative growth, respiratory activity, mitochondrial protein synthesis, and complex IV activity. In one case, we also demonstrated that expression of wt MTO1 could rescue the respiratory defect in mutant fibroblasts. The severity of the yeast respiratory phenotypes partly correlated with the different clinical presentations observed in MTO1 mutant patients, although the clinical outcome was highly variable in patients with the same mutation and seemed also to depend on timely start of pharmacological treatment, centered on the control of lactic acidosis by dichloroacetate. Our results indicate that MTO1 mutations are commonly associated with a presentation of hypertrophic cardiomyopathy, lactic acidosis, and MRC deficiency, and that ad hoc recombinant yeast models represent a useful system to test the pathogenic potential of uncommon variants, and provide insight into their effects on the expression of a biochemical phenotype. © 2013 The Authors. *Human Mutation published by Wiley Periodicals, Inc.

  9. Budding yeast for budding geneticists: a primer on the Saccharomyces cerevisiae model system.

    Science.gov (United States)

    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.

  10. Recombinant yeast screen for new inhibitors of human acetyl-CoA carboxylase 2 identifies potential drugs to treat obesity

    Science.gov (United States)

    Marjanovic, Jasmina; Chalupska, Dominika; Patenode, Caroline; Coster, Adam; Arnold, Evan; Ye, Alice; Anesi, George; Lu, Ying; Okun, Ilya; Tkachenko, Sergey; Haselkorn, Robert; Gornicki, Piotr

    2010-01-01

    Acetyl-CoA carboxylase (ACC) is a key enzyme of fatty acid metabolism with multiple isozymes often expressed in different eukaryotic cellular compartments. ACC-made malonyl-CoA serves as a precursor for fatty acids; it also regulates fatty acid oxidation and feeding behavior in animals. ACC provides an important target for new drugs to treat human diseases. We have developed an inexpensive nonradioactive high-throughput screening system to identify new ACC inhibitors. The screen uses yeast gene-replacement strains depending for growth on cloned human ACC1 and ACC2. In “proof of concept” experiments, growth of such strains was inhibited by compounds known to target human ACCs. The screen is sensitive and robust. Medium-size chemical libraries yielded new specific inhibitors of human ACC2. The target of the best of these inhibitors was confirmed with in vitro enzymatic assays. This compound is a new drug chemotype inhibiting human ACC2 with 2.8 μM IC50 and having no effect on human ACC1 at 100 μM. PMID:20439761

  11. Characterization of RAD4 gene required for ultraviolet-induced excision repair of Saccharomyces cerevisiae propagated in Escherichia coli without inactivation

    International Nuclear Information System (INIS)

    Choi, I.S.; Kim, J.B.; Lee, K.N.; Park, S.D.

    1990-01-01

    The previously isolated RAD4 gene designated as pPC1 from the genomic library of Saccharomyces cerevisiae appeared to propagate in Escherichia coli and yet retained its complementing activity of rad4 mutants without inactivation. The subcloned RAD4 gene was found to be localized within a 2.5 kb DNA fragment flanking Bg/II and BamHI sites in the insert DNA, and was shown to have the same restriction map as a yeast chromosomal DNA, as determined by Southern hybridization. Tetrad analysis and pulse-field chromosome mapping have revealed that the cloned RAD4 gene can be mapped and integrated into the yeast chromosome V, the actual site of this gene. DNA-tRNA hybridization has shown that the isolated RAD4 gene did not contain a suppressor tRNA gene. These results have indicated that the pPC1 is a functional RAD4 gene playing a unique role involved in the nucleotide excision repair of yeast without any genetic change during amplification in E. coli. (author)

  12. Epidemiological importance of humans and domestic animals as reservoirs of verocytotoxin-producing Escherichia coli

    Directory of Open Access Journals (Sweden)

    Lazić Srđan

    2006-01-01

    Full Text Available Background/Aim. A "new" pathogenic agent, verocytotoxin - producing Escherichia coli (VTEC emerged in the last 20 years, causing an increased number of sporadic cases, as well as of outbreaks of diarrhoeal diseases. Humans and animals can be infected with VTEC, but their epidemiological importance as a reservoir of this agent is not quite clear, especially in the Balkan region. Therefore, the aim of this study was to investigate the frequency of isolation of VTEC from the intestinal tract of humans and animals and to determine the serogroups of the isolated strains. Methods. A total of, 3 401 stool samples from humans and 2 660 samples from five different species of domestic animals were tested for the presence of this pathogen. Results. VTEC was isolated from 20 (0.6% humans stools and from 431 (16.2% animal fecal samples (p < 0.001. Only 15 (3.3% VTEC strains belonged to human infection-associated serogroups (O26, O55, O111, O128 and O 157, designated as enterohaemorrhagic E. coli (EHEC. The most known serogroup- O157 was identified in 6 (1.3% of the isolated VTEC strains; of them, 1 (5% was of human origin and 5 (1.2% were animal strains. Conclusion. This study revealed that domestic animals were a more important reservoir of VTEC than humans, and that the isolated VTEC strains rarely belonged to O157, as well as to other EHEC serogroups that might explain rare sporadic cases and the absence of epidemic occurrence of diarrhoeal diseases caused by VTEC in this geographic region.

  13. Adding biological meaning to human protein-protein interactions identified by yeast two-hybrid screenings: A guide through bioinformatics tools.

    Science.gov (United States)

    Felgueiras, Juliana; Silva, Joana Vieira; Fardilha, Margarida

    2018-01-16

    "A man is known by the company he keeps" is a popular expression that perfectly fits proteins. A common approach to characterize the function of a target protein is to identify its interacting partners and thus infer its roles based on the known functions of the interactors. Protein-protein interaction networks (PPINs) have been created for several organisms, including humans, primarily as results of high-throughput screenings, such as yeast two-hybrid (Y2H). Their unequivocal use to understand events underlying human pathophysiology is promising in identifying genes and proteins associated with diseases. Therefore, numerous opportunities have emerged for PPINs as tools for clinical management of diseases: network-based disease classification systems, discovery of biomarkers and identification of therapeutic targets. Despite the great advantages of PPINs, their use is still unrecognised by several researchers who generate high-throughput data to generally characterize interactions in a certain model or to select an interaction to study in detail. We strongly believe that both approaches are not exclusive and that we can use PPINs as a complementary methodology and rich-source of information to the initial study proposal. Here, we suggest a pipeline to deal with Y2H results using bioinformatics tools freely available for academics. Yeast two-hybrid is widely-used to identify protein-protein interactions. Conventionally, the positive clones that result from a yeast two-hybrid screening are sequenced to identify the interactors of the protein of interest (also known as bait protein), and few interactions, thought as potentially relevant for the model in study, are selected for further validation using biochemical methods (e.g. co-immunoprecipitation and co-localization). The huge amount of data that is potentially lost during this conservative approach motivated us to write this tutorial-like review, so that researchers feel encouraged to take advantage of

  14. Independent Origins of Yeast Associated with Coffee and Cacao Fermentation.

    Science.gov (United States)

    Ludlow, Catherine L; Cromie, Gareth A; Garmendia-Torres, Cecilia; Sirr, Amy; Hays, Michelle; Field, Colburn; Jeffery, Eric W; Fay, Justin C; Dudley, Aimée M

    2016-04-04

    Modern transportation networks have facilitated the migration and mingling of previously isolated populations of plants, animals, and insects. Human activities can also influence the global distribution of microorganisms. The best-understood example is yeasts associated with winemaking. Humans began making wine in the Middle East over 9,000 years ago [1, 2]. Selecting favorable fermentation products created specialized strains of Saccharomyces cerevisiae [3, 4] that were transported along with grapevines. Today, S. cerevisiae strains residing in vineyards around the world are genetically similar, and their population structure suggests a common origin that followed the path of human migration [3-7]. Like wine, coffee and cacao depend on microbial fermentation [8, 9] and have been globally dispersed by humans. Theobroma cacao originated in the Amazon and Orinoco basins of Colombia and Venezuela [10], was cultivated in Central America by Mesoamerican peoples, and was introduced to Europeans by Hernán Cortés in 1530 [11]. Coffea, native to Ethiopia, was disseminated by Arab traders throughout the Middle East and North Africa in the 6(th) century and was introduced to European consumers in the 17(th) century [12]. Here, we tested whether the yeasts associated with coffee and cacao are genetically similar, crop-specific populations or genetically diverse, geography-specific populations. Our results uncovered populations that, while defined by niche and geography, also bear signatures of admixture between major populations in events independent of the transport of the plants. Thus, human-associated fermentation and migration may have affected the distribution of yeast involved in the production of coffee and chocolate. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. New yeasts-new brews: modern approaches to brewing yeast design and development.

    Science.gov (United States)

    Gibson, B; Geertman, J-M A; Hittinger, C T; Krogerus, K; Libkind, D; Louis, E J; Magalhães, F; Sampaio, J P

    2017-06-01

    The brewing industry is experiencing a period of change and experimentation largely driven by customer demand for product diversity. This has coincided with a greater appreciation of the role of yeast in determining the character of beer and the widespread availability of powerful tools for yeast research. Genome analysis in particular has helped clarify the processes leading to domestication of brewing yeast and has identified domestication signatures that may be exploited for further yeast development. The functional properties of non-conventional yeast (both Saccharomyces and non-Saccharomyces) are being assessed with a view to creating beers with new flavours as well as producing flavoursome non-alcoholic beers. The discovery of the psychrotolerant S. eubayanus has stimulated research on de novo S. cerevisiae × S. eubayanus hybrids for low-temperature lager brewing and has led to renewed interest in the functional importance of hybrid organisms and the mechanisms that determine hybrid genome function and stability. The greater diversity of yeast that can be applied in brewing, along with an improved understanding of yeasts' evolutionary history and biology, is expected to have a significant and direct impact on the brewing industry, with potential for improved brewing efficiency, product diversity and, above all, customer satisfaction. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  16. Vaginal yeast infection

    Science.gov (United States)

    Yeast infection - vagina; Vaginal candidiasis; Monilial vaginitis ... Most women have a vaginal yeast infection at some time. Candida albicans is a common type of fungus. It is often found in small amounts ...

  17. Yeast as a system for modeling mitochondrial disease mechanisms and discovering therapies

    Directory of Open Access Journals (Sweden)

    Jean-Paul Lasserre

    2015-06-01

    Full Text Available Mitochondrial diseases are severe and largely untreatable. Owing to the many essential processes carried out by mitochondria and the complex cellular systems that support these processes, these diseases are diverse, pleiotropic, and challenging to study. Much of our current understanding of mitochondrial function and dysfunction comes from studies in the baker's yeast Saccharomyces cerevisiae. Because of its good fermenting capacity, S. cerevisiae can survive mutations that inactivate oxidative phosphorylation, has the ability to tolerate the complete loss of mitochondrial DNA (a property referred to as ‘petite-positivity’, and is amenable to mitochondrial and nuclear genome manipulation. These attributes make it an excellent model system for studying and resolving the molecular basis of numerous mitochondrial diseases. Here, we review the invaluable insights this model organism has yielded about diseases caused by mitochondrial dysfunction, which ranges from primary defects in oxidative phosphorylation to metabolic disorders, as well as dysfunctions in maintaining the genome or in the dynamics of mitochondria. Owing to the high level of functional conservation between yeast and human mitochondrial genes, several yeast species have been instrumental in revealing the molecular mechanisms of pathogenic human mitochondrial gene mutations. Importantly, such insights have pointed to potential therapeutic targets, as have genetic and chemical screens using yeast.

  18. Antibiotic resistance profile of Escherichia coli isolated from five ...

    African Journals Online (AJOL)

    Information on the resistance profiles of clinical and non clinical human bacteria isolates in the developing countries can serve as important means of understanding the human pathogens drug resistance interactions in the zone. Escherichia coli isolated from five geopolitical zones of Nigeria were screened for anti-microbial ...

  19. A Yeast Artificial Chromosome Library Database: Design Considerations

    OpenAIRE

    Frisse, Mark E.; Ge, NengJie; Langenbacher, JulieM.; Kahn, Michael G.; Brownstein, Bernard H.

    1990-01-01

    This paper first describes a simple collection of HyperCard stacks created and used by genetics researchers to catalog information in a human yeast artificial chromosome (YAC) library. Although an intuitive human-computer interface made the HyperCard program easy to use, the program was neither an efficient nor a secure primary database for vital laboratory data. This paper subsequently describes a relational database implementation prototype that overcomes HyperCard's deficiencies as a datab...

  20. Schizosaccharomyces japonicus: the fission yeast is a fusion of yeast and hyphae.

    Science.gov (United States)

    Niki, Hironori

    2014-03-01

    The clade of Schizosaccharomyces includes 4 species: S. pombe, S. octosporus, S. cryophilus, and S. japonicus. Although all 4 species exhibit unicellular growth with a binary fission mode of cell division, S. japonicus alone is dimorphic yeast, which can transit from unicellular yeast to long filamentous hyphae. Recently it was found that the hyphal cells response to light and then synchronously activate cytokinesis of hyphae. In addition to hyphal growth, S. japonicas has many properties that aren't shared with other fission yeast. Mitosis of S. japonicas is referred to as semi-open mitosis because dynamics of nuclear membrane is an intermediate mode between open mitosis and closed mitosis. Novel genetic tools and the whole genomic sequencing of S. japonicas now provide us with an opportunity for revealing unique characters of the dimorphic yeast. © 2013 The Author. Yeast Published by John Wiley & Sons Ltd.

  1. Asymptomatic bacteriuria Escherichia coli strains

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Nielsen, E.M.; Klemm, Per

    2006-01-01

    Urinary tract infections (UTIs) affect millions of people each year. Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU) in humans. Persons affected by ABU may carry a particular E. coli strain for extended periods of time without any symptoms. In contrast...... to uropathogenic E. coli (UPEC) that cause symptomatic UTI, very little is known about the mechanisms by which these strains colonize the urinary tract. Here, we have investigated the growth characteristics in human urine as well as adhesin repertoire of nine ABU strains; the ability of ABU strains to compete...

  2. Structural studies of substrate and product complexes of 5-aminolaevulinic acid dehydratase from humans, Escherichia coli and the hyperthermophile Pyrobaculum calidifontis.

    Science.gov (United States)

    Mills-Davies, N; Butler, D; Norton, E; Thompson, D; Sarwar, M; Guo, J; Gill, R; Azim, N; Coker, A; Wood, S P; Erskine, P T; Coates, L; Cooper, J B; Rashid, N; Akhtar, M; Shoolingin-Jordan, P M

    2017-01-01

    A number of X-ray analyses of an enzyme involved in a key early stage of tetrapyrrole biosynthesis are reported. Two structures of human 5-aminolaevulinate dehydratase (ALAD), native and recombinant, have been determined at 2.8 Å resolution, showing that the enzyme adopts an octameric quaternary structure in accord with previously published analyses of the enzyme from a range of other species. However, this is in contrast to the finding that a disease-related F12L mutant of the human enzyme uniquely forms hexamers [Breinig et al. (2003), Nature Struct. Biol. 10, 757-763]. Monomers of all ALADs adopt the TIM-barrel fold; the subunit conformation that assembles into the octamer includes the N-terminal tail of one monomer curled around the (α/β) 8 barrel of a neighbouring monomer. Both crystal forms of the human enzyme possess two monomers per asymmetric unit, termed A and B. In the native enzyme there are a number of distinct structural differences between the A and B monomers, with the latter exhibiting greater disorder in a number of loop regions and in the active site. In contrast, the second monomer of the recombinant enzyme appears to be better defined and the active site of both monomers clearly possesses a zinc ion which is bound by three conserved cysteine residues. In native human ALAD, the A monomer also has a ligand resembling the substrate ALA which is covalently bound by a Schiff base to one of the active-site lysines (Lys252) and is held in place by an ordered active-site loop. In contrast, these features of the active-site structure are disordered or absent in the B subunit of the native human enzyme. The octameric structure of the zinc-dependent ALAD from the hyperthermophile Pyrobaculum calidifontis is also reported at a somewhat lower resolution of 3.5 Å. Finally, the details are presented of a high-resolution structure of the Escherichia coli ALAD enzyme co-crystallized with a noncovalently bound moiety of the product, porphobilinogen (PBG

  3. The Black Yeast Exophiala dermatitidis and Other Selected Opportunistic Human Fungal Pathogens Spread from Dishwashers to Kitchens.

    Directory of Open Access Journals (Sweden)

    Jerneja Zupančič

    Full Text Available We investigated the diversity and distribution of fungi in nine different sites inside 30 residential dishwashers. In total, 503 fungal strains were isolated, which belong to 10 genera and 84 species. Irrespective of the sampled site, 83% of the dishwashers were positive for fungi. The most frequent opportunistic pathogenic species were Exophiala dermatitidis, Candida parapsilosis sensu stricto, Exophiala phaeomuriformis, Fusarium dimerum, and the Saprochaete/Magnusiomyces clade. The black yeast E. dermatitidis was detected in 47% of the dishwashers, primarily at the dishwasher rubber seals, at up to 106 CFU/cm2; the other fungi detected were in the range of 102 to 105 CFU/cm2. The other most heavily contaminated dishwasher sites were side nozzles, doors and drains. Only F. dimerum was isolated from washed dishes, while dishwasher waste water contained E. dermatitidis, Exophiala oligosperma and Sarocladium killiense. Plumbing systems supplying water to household appliances represent the most probable route for contamination of dishwashers, as the fungi that represented the core dishwasher mycobiota were also detected in the tap water. Hot aerosols from dishwashers contained the human opportunistic yeast C. parapsilosis, Rhodotorula mucilaginosa and E. dermatitidis (as well as common air-borne genera such as Aspergillus, Penicillium, Trichoderma and Cladosporium. Comparison of fungal contamination of kitchens without and with dishwashers revealed that virtually all were contaminated with fungi. In both cases, the most contaminated sites were the kitchen drain and the dish drying rack. The most important difference was higher prevalence of black yeasts (E. dermatitidis in particular in kitchens with dishwashers. In kitchens without dishwashers, C. parapsilosis strongly prevailed with negligible occurrence of E. dermatitidis. F. dimerum was isolated only from kitchens with dishwashers, while Saprochaete/Magnusiomyces isolates were only found within

  4. [Mitochondria inheritance in yeast saccharomyces cerevisiae].

    Science.gov (United States)

    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.

  5. Isolation of Nicotiana plumbaginifolia cDNAs encoding isoforms of serine acetyltransferase and O-acetylserine (thiol) lyase in a yeast two-hybrid system with Escherichia coli cysE and cysK genes as baits.

    Science.gov (United States)

    Liszewska, Frantz; Gaganidze, Dali; Sirko, Agnieszka

    2005-01-01

    We applied the yeast two-hybrid system for screening of a cDNA library of Nicotiana plumbaginifolia for clones encoding plant proteins interacting with two proteins of Escherichia coli: serine acetyltransferase (SAT, the product of cysE gene) and O-acetylserine (thiol)lyase A, also termed cysteine synthase (OASTL-A, the product of cysK gene). Two plant cDNA clones were identified when using the cysE gene as a bait. These clones encode a probable cytosolic isoform of OASTL and an organellar isoform of SAT, respectively, as indicated by evolutionary trees. The second clone, encoding SAT, was identified independently also as a "prey" when using cysK as a bait. Our results reveal the possibility of applying the two-hybrid system for cloning of plant cDNAs encoding enzymes of the cysteine synthase complex in the two-hybrid system. Additionally, using genome walking sequences located upstream of the sat1 cDNA were identified. Subsequently, in silico analyses were performed aiming towards identification of the potential signal peptide and possible location of the deduced mature protein encoded by sat1.

  6. Stimulation of Egg Production in Japanese Quails by Enriching Feed with Residual Yeast

    Directory of Open Access Journals (Sweden)

    Letitia Oprean

    2010-05-01

    Full Text Available Quail eggs are more and more approved for consumers because they bring many benefits to the human body. Therefore, quails breeding for eggs production have become a very profitable business. Residual yeast may be a nutritional supplement, especially rich in vitamins and proteins. This article studies the influence of residual beer yeast on egg laying in Japanese quails. In order to be integrated into the diet of quails the yeast has undergone a process of autolysis; its influence has been examined on separate groups. The results were reported as a percentage compared with the control group, where the feed does not contain this supplement. Due to its content rich in vitamins and proteins, the residual beer yeast used in feeding the quails bred for eggs stimulates egg laying.

  7. Porcine aminopeptidase N binds to F4+ enterotoxigenic Escherichia coli fimbriae.

    Science.gov (United States)

    Xia, Pengpeng; Wang, Yiting; Zhu, Congrui; Zou, Yajie; Yang, Ying; Liu, Wei; Hardwidge, Philip R; Zhu, Guoqiang

    2016-02-09

    F4(+) enterotoxigenic Escherichia coli (ETEC) strains cause diarrheal disease in neonatal and post-weaned piglets. Several different host receptors for F4 fimbriae have been described, with porcine aminopeptidase N (APN) reported most recently. The FaeG subunit is essential for the binding of the three F4 variants to host cells. Here we show in both yeast two-hybrid and pulldown assays that APN binds directly to FaeG, the major subunit of F4 fimbriae, from three serotypes of F4(+) ETEC. Modulating APN gene expression in IPEC-J2 cells affected ETEC adherence. Antibodies raised against APN or F4 fimbriae both reduced ETEC adherence. Thus, APN mediates the attachment of F4(+) E. coli to intestinal epithelial cells.

  8. Colonization of Enteroaggregative Escherichia coli and Shiga toxin-producing Escherichia coli in chickens and humans in southern Vietnam

    NARCIS (Netherlands)

    Trung, Nguyen Vinh; Nhung, Hoang Ngoc; Carrique-Mas, Juan J.; Mai, Ho Huynh; Tuyen, Ha Thanh; Campbell, James; Nhung, Nguyen Thi; van Minh, Pham; Wagenaar, Jaap A.; Mai, Nguyen Thi Nhu; Hieu, Thai Quoc; Schultsz, Constance; Hoa, Ngo Thi

    2016-01-01

    Enteroaggregative (EAEC) and Shiga-toxin producing Escherichia coli (STEC) are a major cause of diarrhea worldwide. E. coli carrying both virulence factors characteristic for EAEC and STEC and producing extended-spectrum beta-lactamase caused severe and protracted disease during an outbreak of E.

  9. Studying Functions of All Yeast Genes Simultaneously

    Science.gov (United States)

    Stolc, Viktor; Eason, Robert G.; Poumand, Nader; Herman, Zelek S.; Davis, Ronald W.; Anthony Kevin; Jejelowo, Olufisayo

    2006-01-01

    A method of studying the functions of all the genes of a given species of microorganism simultaneously has been developed in experiments on Saccharomyces cerevisiae (commonly known as baker's or brewer's yeast). It is already known that many yeast genes perform functions similar to those of corresponding human genes; therefore, by facilitating understanding of yeast genes, the method may ultimately also contribute to the knowledge needed to treat some diseases in humans. Because of the complexity of the method and the highly specialized nature of the underlying knowledge, it is possible to give only a brief and sketchy summary here. The method involves the use of unique synthetic deoxyribonucleic acid (DNA) sequences that are denoted as DNA bar codes because of their utility as molecular labels. The method also involves the disruption of gene functions through deletion of genes. Saccharomyces cerevisiae is a particularly powerful experimental system in that multiple deletion strains easily can be pooled for parallel growth assays. Individual deletion strains recently have been created for 5,918 open reading frames, representing nearly all of the estimated 6,000 genetic loci of Saccharomyces cerevisiae. Tagging of each deletion strain with one or two unique 20-nucleotide sequences enables identification of genes affected by specific growth conditions, without prior knowledge of gene functions. Hybridization of bar-code DNA to oligonucleotide arrays can be used to measure the growth rate of each strain over several cell-division generations. The growth rate thus measured serves as an index of the fitness of the strain.

  10. Yeast ecology of Kombucha fermentation.

    Science.gov (United States)

    Teoh, Ai Leng; Heard, Gillian; Cox, Julian

    2004-09-01

    Kombucha is a traditional fermentation of sweetened tea, involving a symbiosis of yeast species and acetic acid bacteria. Despite reports of different yeast species being associated with the fermentation, little is known of the quantitative ecology of yeasts in Kombucha. Using oxytetracycline-supplemented malt extract agar, yeasts were isolated from four commercially available Kombucha products and identified using conventional biochemical and physiological tests. During the fermentation of each of the four products, yeasts were enumerated from both the cellulosic pellicle and liquor of the Kombucha. The number and diversity of species varied between products, but included Brettanomyces bruxellensis, Candida stellata, Schizosaccharomyces pombe, Torulaspora delbrueckii and Zygosaccharomyces bailii. While these yeast species are known to occur in Kombucha, the enumeration of each species present throughout fermentation of each of the four Kombucha cultures demonstrated for the first time the dynamic nature of the yeast ecology. Kombucha fermentation is, in general, initiated by osmotolerant species, succeeded and ultimately dominated by acid-tolerant species.

  11. Yeast genome sequencing:

    DEFF Research Database (Denmark)

    Piskur, Jure; Langkjær, Rikke Breinhold

    2004-01-01

    For decades, unicellular yeasts have been general models to help understand the eukaryotic cell and also our own biology. Recently, over a dozen yeast genomes have been sequenced, providing the basis to resolve several complex biological questions. Analysis of the novel sequence data has shown...... of closely related species helps in gene annotation and to answer how many genes there really are within the genomes. Analysis of non-coding regions among closely related species has provided an example of how to determine novel gene regulatory sequences, which were previously difficult to analyse because...... they are short and degenerate and occupy different positions. Comparative genomics helps to understand the origin of yeasts and points out crucial molecular events in yeast evolutionary history, such as whole-genome duplication and horizontal gene transfer(s). In addition, the accumulating sequence data provide...

  12. Heterologous transporter expression for improved fatty alcohol secretion in yeast

    DEFF Research Database (Denmark)

    Hu, Yating; Zhu, Zhiwei; Nielsen, Jens

    2017-01-01

    The yeast Saccharomyces cerevisiae is an attractive host for industrial scale production of biofuels including fatty alcohols due to its robustness and tolerance towards harsh fermentation conditions. Many metabolic engineering strategies have been applied to generate high fatty alcohol production...... transporters tested, human FATP1 was shown to mediate fatty alcohol export in a high fatty alcohol production yeast strain. An approximately five-fold increase of fatty alcohol secretion was achieved. The results indicate that the overall cell fitness benefited from fatty alcohol secretion and that the acyl......-CoA synthase activity of FATP1 contributed to increased cell growth as well. This is the first study that enabled an increased cell fitness for fatty alcohol production by heterologous transporter expression in yeast, and this investigation indicates a new potential function of FATP1, which has been known...

  13. Beer and bread to brains and beyond: can yeast cells teach us about neurodegenerative disease?

    Science.gov (United States)

    Gitler, Aaron D

    2008-01-01

    For millennia, humans have harnessed the astonishing power of yeast, producing such culinary masterpieces as bread, beer and wine. Therefore, in this new millennium, is it very farfetched to ask if we can also use yeast to unlock some of the modern day mysteries of human disease? Remarkably, these seemingly simple cells possess most of the same basic cellular machinery as the neurons in the brain. We and others have been using the baker's yeast, Saccharomyces cerevisiae, as a model system to study the mechanisms of devastating neurodegenerative diseases such as Parkinson's, Huntington's, Alzheimer's and amyotrophic lateral sclerosis. While very different in their pathophysiology, they are collectively referred to as protein-misfolding disorders because of the presence of misfolded and aggregated forms of various proteins in the brains of affected individuals. Using yeast genetics and the latest high-throughput screening technologies, we have identified some of the potential causes underpinning these disorders and discovered conserved genes that have proven effective in preventing neuron loss in animal models. Thus, these genes represent new potential drug targets. In this review, I highlight recent work investigating mechanisms of cellular toxicity in a yeast Parkinson's disease model and discuss how similar approaches are being applied to additional neurodegenerative diseases.

  14. Drying enhances immunoactivity of spent brewer's yeast cell wall β-D-glucans.

    Science.gov (United States)

    Liepins, Janis; Kovačova, Elena; Shvirksts, Karlis; Grube, Mara; Rapoport, Alexander; Kogan, Grigorij

    2015-07-20

    Due to immunological activity, microbial cell wall polysaccharides are defined as 'biological response modifiers' (BRM). Cell walls of spent brewer's yeast also have some BRM activity. However, up to date there is no consensus on the use of spent brewer's yeast D-glucan as specific BRM in humans or animals. The aim of this paper is to demonstrate the potential of spent brewer's yeast β-D-glucans as BRM, and drying as an efficient pretreatment to increase β-D-glucan's immunogenic activity. Our results revealed that drying does not change spent brewer's yeast biomass carbohydrate content as well as the chemical structure of purified β-D-glucan. However, drying increased purified β-D-glucan TNF-α induction activity in the murine macrophage model. We presume drying pretreatment enhances purity of extracted β-D-glucan. This is corroborated with FT-IR analyses of the β-D-glucan spectra. Based on our results, we suggest that dry spent brewer's yeast biomass can be used as a cheap source for high-quality β-D-glucan extraction. Drying in combination with carboxylmethylation (CM), endows spent brewer's yeast β-D-glucan with the immunoactivity similar or exceeding that of a well-characterized fungal BRM pleuran. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Antifungal activity of the extract of Curcuma zedoaria (Christm. Roscoe, Zingiberaceae, against yeasts of the genus Candida isolated from the oral cavity of patients infected with the human immunodeficiency virus

    Directory of Open Access Journals (Sweden)

    Cristiane S. Shinobu-Mesquita

    2011-02-01

    Full Text Available Oropharyngeal candidiasis is the most common fungal infection among patients infected with the human immunodeficiency virus (HIV, and is treated empirically with topical or systemic antifungals. The objective of the present study was to investigate the possible antifungal action of the hydroalcoholic extract of Curcuma zedoaria (Christm. Roscoe, Zingiberaceae, on yeasts in this population. Samples were collected from HIV-positive patients who attended the Laboratory for Teaching and Research in Clinical Analysis at the Universidade Estadual de Maringá for routine exams. The isolated yeasts were identified at the genus and species levels through classical methodology. Next, tests of microdilution in broth were carried out to determine the profile of susceptibility of these yeasts towards the hydroalcoholic extract of C. zedoaria, following methodology standardised by the CLSI (2002. A total of 53 yeasts were identified, 49 of them C. albicans, two C. tropicalis and two C. glabrata. These yeasts were inhibited by low concentrations of the extract of C. zedoaria (between 1.95 and 15.63 μg/mL. In addition, 7.82 μg/mL inhibited 90% of the yeasts. Our results indicate a potent antifungal action for C. zedoaria and suggest more detailed studies with a view towards the practical application of this phytomedicine in topical pharmaceutical forms for the treatment of oral candidosis or candidiasis.

  16. Shiga toxin-producing Escherichia coli in Central Greece: prevalence and virulence genes of O157:H7 and non-O157 in animal feces, vegetables, and humans.

    Science.gov (United States)

    Pinaka, O; Pournaras, S; Mouchtouri, V; Plakokefalos, E; Katsiaflaka, A; Kolokythopoulou, F; Barboutsi, E; Bitsolas, N; Hadjichristodoulou, C

    2013-11-01

    In Greece, Shiga toxin-producing Escherichia coli (STEC) have only been sporadically reported. The objective of this study was to estimate the prevalence of STEC and Escherichia coli O157:H7 in farm animals, vegetables, and humans in Greece. A total number of 1,010 fecal samples were collected from farm animals (sheep, goats, cattle, chickens, pigs), 667 diarrheal samples from humans, and 60 from vegetables, which were cultured in specific media for STEC isolates. Enzyme-linked immunosorbent assay (ELISA) was used to detect toxin-producing colonies, which, subsequently, were subjected to a multiplex polymerase chain reaction (PCR) for stx1, stx2, eae, rfbE O157, and fliC h7 genes. Eighty isolates (7.9 %) from animal samples were found to produce Shiga toxin by ELISA, while by PCR, O157 STEC isolates were detected from 8 (0.8 %) samples and non-O157 STEC isolates from 43 (4.2 %) samples. STEC isolates were recovered mainly from sheep and goats, rarely from cattle, and not from pigs and chickens, suggesting that small ruminants constitute a potential risk for human infections. However, only three human specimens (0.4 %) were positive for the detection of Shiga toxins and all were PCR-negative. Similarly, all 60 vegetable samples were negative for toxin production and for toxin genes, but three samples (two roman rockets and one spinach) were positive by PCR for rfbE O157 and fliC h7 genes. These findings indicate that sheep, goats, cattle, and leafy vegetables can be a reservoir of STEC and Escherichia coli O157:H7 isolates in Greece, which are still rarely detected among humans.

  17. Search for Enterohaemorrhagic Escherichia coli O157:H7 and ...

    African Journals Online (AJOL)

    Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 and Salmonella enterica are important zoonotic bacteria responsible for enteric infections in humans. The present study investigated the possible role of kittens in the zoonotic transmission of antimicrobial resistant EHEC O157 and Salmonella enterica to human using ...

  18. Siderophore biosynthesis coordinately modulated the virulence-associated interactive metabolome of uropathogenic Escherichia coli and human urine.

    Science.gov (United States)

    Su, Qiao; Guan, Tianbing; Lv, Haitao

    2016-04-14

    Uropathogenic Escherichia coli (UPEC) growth in women's bladders during urinary tract infection (UTI) incurs substantial chemical exchange, termed the "interactive metabolome", which primarily accounts for the metabolic costs (utilized metabolome) and metabolic donations (excreted metabolome) between UPEC and human urine. Here, we attempted to identify the individualized interactive metabolome between UPEC and human urine. We were able to distinguish UPEC from non-UPEC by employing a combination of metabolomics and genetics. Our results revealed that the interactive metabolome between UPEC and human urine was markedly different from that between non-UPEC and human urine, and that UPEC triggered much stronger perturbations in the interactive metabolome in human urine. Furthermore, siderophore biosynthesis coordinately modulated the individualized interactive metabolome, which we found to be a critical component of UPEC virulence. The individualized virulence-associated interactive metabolome contained 31 different metabolites and 17 central metabolic pathways that were annotated to host these different metabolites, including energetic metabolism, amino acid metabolism, and gut microbe metabolism. Changes in the activities of these pathways mechanistically pinpointed the virulent capability of siderophore biosynthesis. Together, our findings provide novel insights into UPEC virulence, and we propose that siderophores are potential targets for further discovery of drugs to treat UPEC-induced UTI.

  19. Expression of human ferredoxin and assembly of the [2Fe-2S] center in Escherichia coli

    International Nuclear Information System (INIS)

    Coghlan, V.M.; Vickery, L.E.

    1989-01-01

    A cDNA fragment encoding human ferredoxin, a mitochondrial [2Fe-2S] protein, was introduced into Escherichia coli by using an expression vector based on the approach of Nagai and Thogersen. Expression was under control of the λP L promoter and resulted in production of ferredoxin as a cleavable fusion protein with an amino-terminal fragment derived from bacteriophage λcII protein. The fusion protein was isolated from the soluble fraction of induced cells and was specifically cleaved to yield mature recombinant ferredoxin. The recombinant protein was shown to be identical in size to ferredoxin isolated from human placenta (13,546 Da) by NaDodSO 4 /PAGE and partial amino acid sequencing. E. coli cells expressing human ferredoxin were brown in color, and absorbance and electron paramagnetic resonance spectra of the purified recombinant protein established that the [2Fe-2S]center was assembled and incorporated into ferredoxin in vivo. Recombinant ferredoxin was active in steroid hydroxylations when reconstituted with cytochromes P-450 sec and P-450 11β and exhibited rates comparable to those observed for ferredoxin isolated from human placenta. This expression system should be useful in production of native and structurally altered forms of human ferredoxin for studies of ferredoxin structure and function

  20. High-level expression of human stem cell factor fused with erythropoietin mimetic peptide in Escherichia coli.

    Science.gov (United States)

    Su, Lin; Chen, Song-Sen; Yang, Ke-Gong; Liu, Chang-Zheng; Zhang, Yan-Li; Liang, Zhi-Quan

    2006-06-01

    Stem cell factor (SCF) and erythropoietin are essential for normal erythropoiesis and induce proliferation and differentiation synergistically for erythroid progenitor cells. Here, we report our work on construction of SCF/erythropoietin mimetic peptide (EMP) fusion protein gene, in which human SCF cDNA (1-165aa) and EMP sequence (20aa) were connected using a short (GGGGS) or long (GGGGSGGGGGS) linker sequence. The SCF/EMP gene was cloned into the pBV220 vector and expressed in the Escherichia coli DH5alpha strain. The expression level of the fusion protein was about 30% of total cell protein. The resulting inclusion bodies were solubilized with 8 M urea, followed by dilution refolding. The renatured protein was subsequently purified by Q-Sepharose FF column. The final product was >95% pure by SDS-PAGE and the yield of fusion protein was about 40 mg/L of culture. UT-7 cell proliferation and human cord blood cell colony-forming assays showed that the fusion proteins exhibited more potent activity than recombinant human SCF, suggesting a new strategy to enhance biological activities of growth factors.

  1. Developing a yeast-based assay protocol to monitor total ...

    African Journals Online (AJOL)

    A yeast-based assay protocol developed for detecting oestrogenic activity in activated sludge (AS) supernatant is described. The protocol used Saccharomyces cerevisiae construct RMY/ER-ERE with human oestrogen receptor (ERα) and lacZ reporter genes, and was developed by modifying existing assays for use with AS ...

  2. Differences between flocculating yeast and regular industrial yeast in transcription and metabolite profiling during ethanol fermentation

    Directory of Open Access Journals (Sweden)

    Lili Li

    2017-03-01

    Full Text Available Objectives: To improve ethanolic fermentation performance of self-flocculating yeast, difference between a flocculating yeast strain and a regular industrial yeast strain was analyzed by transcriptional and metabolic approaches. Results: The number of down-regulated (industrial yeast YIC10 vs. flocculating yeast GIM2.71 and up-regulated genes were 4503 and 228, respectively. It is the economic regulation for YIC10 that non-essential genes were down-regulated, and cells put more “energy” into growth and ethanol production. Hexose transport and phosphorylation were not the limiting-steps in ethanol fermentation for GIM2.71 compared to YIC10, whereas the reaction of 1,3-disphosphoglycerate to 3-phosphoglycerate, the decarboxylation of pyruvate to acetaldehyde and its subsequent reduction to ethanol were the most limiting steps. GIM2.71 had stronger stress response than non-flocculating yeast and much more carbohydrate was distributed to other bypass, such as glycerol, acetate and trehalose synthesis. Conclusions: Differences between flocculating yeast and regular industrial yeast in transcription and metabolite profiling will provide clues for improving the fermentation performance of GIM2.71.

  3. Epidemiology and clinical manifestations of enteroaggregative Escherichia coli

    DEFF Research Database (Denmark)

    Hebbelstrup Jensen, Betina; Olsen, Katharina E P; Struve, Carsten

    2014-01-01

    Enteroaggregative Escherichia coli (EAEC) represents a heterogeneous group of E. coli strains. The pathogenicity and clinical relevance of these bacteria are still controversial. In this review, we describe the clinical significance of EAEC regarding patterns of infection in humans, transmission...

  4. Yeast Derived LysA2 Can Control Bacterial Contamination in Ethanol Fermentation

    Directory of Open Access Journals (Sweden)

    Jun-Seob Kim

    2018-05-01

    Full Text Available Contamination of fuel-ethanol fermentations continues to be a significant problem for the corn and sugarcane-based ethanol industries. In particular, members of the Lactobacillaceae family are the primary bacteria of concern. Currently, antibiotics and acid washing are two major means of controlling contaminants. However, antibiotic use could lead to increased antibiotic resistance, and the acid wash step stresses the fermenting yeast and has limited effectiveness. Bacteriophage endolysins such as LysA2 are lytic enzymes with the potential to contribute as antimicrobials to the fuel ethanol industries. Our goal was to evaluate the potential of yeast-derived LysA2 as a means of controlling Lactobacillaceae contamination. LysA2 intracellularly produced by Pichia pastoris showed activity comparable to Escherichia coli produced LysA2. Lactic Acid Bacteria (LAB with the A4α peptidoglycan chemotype (L-Lys-D-Asp crosslinkage were the most sensitive to LysA2, though a few from that chemotype were insensitive. Pichia-expressed LysA2, both secreted and intracellularly produced, successfully improved ethanol productivity and yields in glucose (YPD60 and sucrose-based (sugarcane juice ethanol fermentations in the presence of a LysA2 susceptible LAB contaminant. LysA2 secreting Sacharomyces cerevisiae did not notably improve production in sugarcane juice, but it did control bacterial contamination during fermentation in YPD60. Secretion of LysA2 by the fermenting yeast, or adding it in purified form, are promising alternative tools to control LAB contamination during ethanol fermentation. Endolysins with much broader lytic spectrums than LysA2 could supplement or replace the currently used antibiotics or the acidic wash.

  5. Yeasts and yeast-like organisms associated with fruits and blossoms of different fruit trees.

    Science.gov (United States)

    Vadkertiová, Renáta; Molnárová, Jana; Vránová, Dana; Sláviková, Elena

    2012-12-01

    Yeasts are common inhabitants of the phyllosphere, but our knowledge of their diversity in various plant organs is still limited. This study focused on the diversity of yeasts and yeast-like organisms associated with matured fruits and fully open blossoms of apple, plum, and pear trees, during 2 consecutive years at 3 localities in southwest Slovakia. The occurrence of yeasts and yeast-like organisms in fruit samples was 2½ times higher and the yeast community more diverse than that in blossom samples. Only 2 species (Aureobasidium pullulans and Metschnikowia pulcherrima) occurred regularly in the blossom samples, whereas Galactomyces candidus, Hanseniaspora guilliermondii, Hanseniaspora uvarum, M. pulcherrima, Pichia kluyveri, Pichia kudriavzevii, and Saccharomyces cerevisiae were the most frequently isolated species from the fruit samples. The ratio of the number of samples where only individual species were present to the number of samples where 2 or more species were found (consortium) was counted. The occurrence of individual species in comparison with consortia was much higher in blossom samples than in fruit samples. In the latter, consortia predominated. Aureobasidium pullulans, M. pulcherrima, and S. cerevisiae, isolated from both the fruits and blossoms, can be considered as resident yeast species of various fruit tree species cultivated in southwest Slovakia localities.

  6. Simplified Method to Produce Human Bioactive Leukemia Inhibitory Factor in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Houman Kahroba

    2016-07-01

    Full Text Available Background Human leukemia inhibitory factor (hLIF is a poly functional cytokine with numerous regulatory effects on different cells. Main application of hLIF is maintaining pluripotency of embryonic stem cells. hLIF indicated effective work in implantation rate of fertilized eggs and multiple sclerosis (MS treatment. Low production of hLIF in eukaryotic cells and prokaryotic host’s problems for human protein production convinced us to develop a simple way to reach high amount of this widely used clinical and research factor. Objectives In this study we want to purify recombinant human leukemia inhibitory factor in single simple method. Materials and Methods This is an experimental study, gene expression: human LIF gene was codon optimized for expression in Escherichia coli and attached his-tag tail to make it extractable. After construction and transformation of vector to E. coli, isopropyl β-D-1-thiogalactopyranoside (IPTG used for induction. Single step immobilized metal affinity chromatography (IMAC used for purification confirmed by Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE and western blotting. Bioactivity of the hLIF were tested by MTT assay with TF-1 cells and CISH gene stimulation in monocyte and TF-1 by real-time PCR. Induction by 0.4 mM of IPTG in 25°C for 3 hours indicated best result for soluble expression. SPSS indicated P ˂ 0.05 that is significant for our work. Results Cloning, expression, and extraction of bio active rhLIF was successfully achieved according MTT assay and real time PCR after treatment of TF-1 and monocyte cell lines. Conclusions We developed an effective single step purification method to produce bioactive recombinant hLIF in E. coli. For the first time we used CISH gene stimulating for bioactivity test for qualifying of recombinant hLIF for application.

  7. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium

    DEFF Research Database (Denmark)

    Smith, Ida Mosbech; Baker, A; Arneborg, Nils

    2015-01-01

    distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase....... In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability......). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. SIGNIFICANCE AND IMPACT...

  8. Life History Responses and Gene Expression Profiles of the Nematode Pristionchus pacificus Cultured on Cryptococcus Yeasts.

    Directory of Open Access Journals (Sweden)

    Gaurav V Sanghvi

    Full Text Available Nematodes, the earth's most abundant metazoa are found in all ecosystems. In order to survive in diverse environments, they have evolved distinct feeding strategies and they can use different food sources. While some nematodes are specialists, including parasites of plants and animals, others such as Pristionchus pacificus are omnivorous feeders, which can live on a diet of bacteria, protozoans, fungi or yeast. In the wild, P. pacificus is often found in a necromenic association with beetles and is known to be able to feed on a variety of microbes as well as on nematode prey. However, in laboratory studies Escherichia coli OP50 has been used as standard food source, similar to investigations in Caenorhabditis elegans and it is unclear to what extent this biases the obtained results and how relevant findings are in real nature. To gain first insight into the variation in traits induced by a non-bacterial food source, we study Pristionchus-fungi interactions under laboratory conditions. After screening different yeast strains, we were able to maintain P. pacificus for at least 50-60 generations on Cryptococcus albidus and Cryptococcus curvatus. We describe life history traits of P. pacificus on both yeast strains, including developmental timing, survival and brood size. Despite a slight developmental delay and problems to digest yeast cells, which are both reflected at a transcriptomic level, all analyses support the potential of Cryptococcus strains as food source for P. pacificus. In summary, our work establishes two Cryptococcus strains as alternative food source for P. pacificus and shows change in various developmental, physiological and morphological traits, including the transcriptomic profiles.

  9. Food-Associated Lactobacillus plantarum and Yeasts Inhibit the Genotoxic Effect of 4-Nitroquinoline-1-Oxide

    Directory of Open Access Journals (Sweden)

    Roberta Prete

    2017-11-01

    Full Text Available Lactic acid bacteria and yeasts, representing the prevailing microbiota associated with different foods generally consumed without any cooking, were identified and characterized in vitro for some functional properties, such as acid-bile tolerance and antigenotoxic activity. In particular, 22 Lactobacillus plantarum strains and 14 yeasts were studied. The gastro-intestinal tract tolerance of all the strains was determined by exposing washed cell suspensions at 37°C to a simulated gastric juice (pH 2.0, containing pepsin (0.3% w/v and to a simulated small intestinal juice (pH 8.0, containing pancreatin (1 mg mL-1 and bile extract (0.5%, thus monitoring changes in total viable count. In general, following a strain-dependent behavior, all the tested strains persisted alive after combined acid-bile challenge. Moreover, many strains showed high in vitro inhibitory activity against a model genotoxin, 4-nitroquinoline-1-oxide (4-NQO, as determined by the short-term method, SOS-Chromotest. Interestingly, the supernatants from bacteria- or yeasts-genotoxin co-incubations exhibited a suppression on SOS-induction produced by 4-NQO on the tester strain Escherichia coli PQ37 (sfiA::lacZ exceeding, in general, the value of 75%. The results highlight that food associated microorganisms may reach the gut in viable form and prevent genotoxin DNA damage in situ. Our experiments can contribute to elucidate the functional role of food-associated microorganisms general recognized as safe ingested with foods as a part of the diet.

  10. Interaction Between Yeasts and Zinc

    Science.gov (United States)

    Nicola, Raffaele De; Walker, Graeme

    Zinc is an essential trace element in biological systems. For example, it acts as a cellular membrane stabiliser, plays a critical role in gene expression and genome modification and activates nearly 300 enzymes, including alcohol dehydrogenase. The present chapter will be focused on the influence of zinc on cell physiology of industrial yeast strains of Saccharomyces cerevisiae, with special regard to the uptake and subsequent utilisation of this metal. Zinc uptake by yeast is metabolism-dependent, with most of the available zinc translocated very quickly into the vacuole. At cell division, zinc is distributed from mother to daughter cells and this effectively lowers the individual cellular zinc concentration, which may become zinc depleted at the onset of the fermentation. Zinc influences yeast fermentative performance and examples will be provided relating to brewing and wine fermentations. Industrial yeasts are subjected to several stresses that may impair fermentation performance. Such stresses may also impact on yeast cell zinc homeostasis. This chapter will discuss the practical implications for the correct management of zinc bioavailability for yeast-based biotechnologies aimed at improving yeast growth, viability, fermentation performance and resistance to environmental stresses

  11. Yeast and Fungal Prions: Amyloid-Handling Systems, Amyloid Structure, and Prion Biology.

    Science.gov (United States)

    Wickner, R B; Edskes, H K; Gorkovskiy, A; Bezsonov, E E; Stroobant, E E

    2016-01-01

    Yeast prions (infectious proteins) were discovered by their outré genetic properties and have become important models for an array of human prion and amyloid diseases. A single prion protein can become any of many distinct amyloid forms (called prion variants or strains), each of which is self-propagating, but with different biological properties (eg, lethal vs mild). The folded in-register parallel β sheet architecture of the yeast prion amyloids naturally suggests a mechanism by which prion variant information can be faithfully transmitted for many generations. The yeast prions rely on cellular chaperones for their propagation, but can be cured by various chaperone imbalances. The Btn2/Cur1 system normally cures most variants of the [URE3] prion that arise. Although most variants of the [PSI+] and [URE3] prions are toxic or lethal, some are mild in their effects. Even the most mild forms of these prions are rare in the wild, indicating that they too are detrimental to yeast. The beneficial [Het-s] prion of Podospora anserina poses an important contrast in its structure, biology, and evolution to the yeast prions characterized thus far. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Expression of the human blood coagulation protein factor XIIIa in Saccharomyces cerevisiae: dependence of the expression levels from host-vector systems and medium conditions.

    Science.gov (United States)

    Bröker, M; Bäuml, O; Göttig, A; Ochs, J; Bodenbenner, M; Amann, E

    1991-03-01

    The human blood coagulation protein Factor XIIIa (FXIIIa) was expressed in Saccharomyces cerevisiae employing Escherichia coli-yeast shuttle vectors based on a 2-mu plasmid. Several factors affecting high production yield of recombinant FXIIIa were analysed. The use of the regulatable GAL-CYC1 hybrid promoter resulted in higher FXIIIa expression when compared with the constitutive ADCI promoter. Screening for suitable yeast strains for expression of FXIIIa under the transcriptional control of the GAL-CYC1 hybrid promoter revealed a broad spectrum of productivity. No obvious correlation between the expression rate and the genetic markers of the strains could be identified. The medium composition markedly influenced the FXIIIa expression rates. The expression of FXIIIa was strictly regulated by the carbon source. Glucose as the only sugar and energy source repressed the synthesis of FXIIIa, whereas addition of galactose induced FXIIIa expression. Special feeding schemes resulted in a productivity of up to 100 mg FXIIIa/l in shake flasks.

  13. Roles of C-Terminal Region of Yeast and Human Rad52 in Rad51-Nucleoprotein Filament Formation and ssDNA Annealing.

    Directory of Open Access Journals (Sweden)

    Nilesh V Khade

    Full Text Available Yeast Rad52 (yRad52 has two important functions at homologous DNA recombination (HR; annealing complementary single-strand DNA (ssDNA molecules and recruiting Rad51 recombinase onto ssDNA (recombination mediator activity. Its human homolog (hRAD52 has a lesser role in HR, and apparently lacks mediator activity. Here we show that yRad52 can load human Rad51 (hRAD51 onto ssDNA complexed with yeast RPA in vitro. This is biochemically equivalent to mediator activity because it depends on the C-terminal Rad51-binding region of yRad52 and on functional Rad52-RPA interaction. It has been reported that the N-terminal two thirds of both yRad52 and hRAD52 is essential for binding to and annealing ssDNA. Although a second DNA binding region has been found in the C-terminal region of yRad52, its role in ssDNA annealing is not clear. In this paper, we also show that the C-terminal region of yRad52, but not of hRAD52, is involved in ssDNA annealing. This suggests that the second DNA binding site is required for the efficient ssDNA annealing by yRad52. We propose an updated model of Rad52-mediated ssDNA annealing.

  14. Lactobacillus reuteri Inhibition of Enteropathogenic Escherichia coli Adherence to Human Intestinal Epithelium.

    Science.gov (United States)

    Walsham, Alistair D S; MacKenzie, Donald A; Cook, Vivienne; Wemyss-Holden, Simon; Hews, Claire L; Juge, Nathalie; Schüller, Stephanie

    2016-01-01

    Enteropathogenic Escherichia coli (EPEC) is a major cause of diarrheal infant death in developing countries, and probiotic bacteria have been shown to provide health benefits in gastrointestinal infections. In this study, we have investigated the influence of the gut symbiont Lactobacillus reuteri on EPEC adherence to the human intestinal epithelium. Different host cell model systems including non-mucus-producing HT-29 and mucus-producing LS174T intestinal epithelial cell lines as well as human small intestinal biopsies were used. Adherence of L. reuteri to HT-29 cells was strain-specific, and the mucus-binding proteins CmbA and MUB increased binding to both HT-29 and LS174T cells. L. reuteri ATCC PTA 6475 and ATCC 53608 significantly inhibited EPEC binding to HT-29 but not LS174T cells. While pre-incubation of LS174T cells with ATCC PTA 6475 did not affect EPEC attaching/effacing (A/E) lesion formation, it increased the size of EPEC microcolonies. ATCC PTA 6475 and ATCC 53608 binding to the mucus layer resulted in decreased EPEC adherence to small intestinal biopsy epithelium. Our findings show that L. reuteri reduction of EPEC adhesion is strain-specific and has the potential to target either the epithelium or the mucus layer, providing further rationale for the selection of probiotic strains.

  15. High diversity of plasmids harbouring blaCMY-2 among clinical Escherichia coli isolates from humans and companion animals in the upper Midwestern USA

    DEFF Research Database (Denmark)

    Bortolaia, Valeria; Hansen, Katrine Hartung; Nielsen, Christine

    2014-01-01

    To determine the population structure and genetic relatedness of plasmids encoding CMY-2 β-lactamase in clinical Escherichia coli from humans and companion animals within a defined geographical area....

  16. Entropy analysis in yeast DNA

    International Nuclear Information System (INIS)

    Kim, Jongkwang; Kim, Sowun; Lee, Kunsang; Kwon, Younghun

    2009-01-01

    In this article, we investigate the language structure in yeast 16 chromosomes. In order to find it, we use the entropy analysis for codons (or amino acids) of yeast 16 chromosomes, developed in analysis of natural language by Montemurro et al. From the analysis, we can see that there exists a language structure in codons (or amino acids) of yeast 16 chromosomes. Also we find that the grammar structure of amino acids of yeast 16 chromosomes has a deep relationship with secondary structure of protein.

  17. Characterization of the response chemiluminescence of neutrophils human beings to the hemolysin Escherichia coli alpha

    International Nuclear Information System (INIS)

    Garcia, J.

    2000-01-01

    Escherichia coli alpha hemolysin (AH) evoked a luminol-amplified chemiluminescence (CL) response from human polymorphonuclear leukocytes (PMN). Analysis of kinetic parameters of the PMN CL response to AH established similarities with that of PMN to the calcium ionophore A23187. PMN CL responses to both AH and A23187 were equally decreased by preincubating PMN with A63612, a hidroxamic acid derivative and lipooxigenase inhibitor, showing that the CL response to both hemolysin and ionophore share a common mechanism, probably activation of leukotriene synthesis, due to calcium entry into the cells brought about by AH and A23187. In addition, the CL response of PMN to AH was lowered by the hydroxyl radical scavenger dimethyl sulfoxide, further suggesting arachidonate metabolism is involved in CL response. (Author) [es

  18. Yeast flocculation: New story in fuel ethanol production.

    Science.gov (United States)

    Zhao, X Q; Bai, F W

    2009-01-01

    Yeast flocculation has been used in the brewing industry to facilitate biomass recovery for a long time, and thus its mechanism of yeast flocculation has been intensively studied. However, the application of flocculating yeast in ethanol production garnered attention mainly in the 1980s and 1990s. In this article, updated research progress in the molecular mechanism of yeast flocculation and the impact of environmental conditions on yeast flocculation are reviewed. Construction of flocculating yeast strains by genetic approach and utilization of yeast flocculation for ethanol production from various feedstocks were presented. The concept of self-immobilized yeast cells through their flocculation is revisited through a case study of continuous ethanol fermentation with the flocculating yeast SPSC01, and their technical and economic advantages are highlighted by comparing with yeast cells immobilized with supporting materials and regular free yeast cells as well. Taking the flocculating yeast SPSC01 as an example, the ethanol tolerance of the flocculating yeast was also discussed.

  19. Effects of gamma radiation on Sporothrix schenckii yeast cells

    Energy Technology Data Exchange (ETDEWEB)

    Lacerda, Camila M. de Sousa; Martins, Estefania Mara Nascimento; Andrade, Antero S.R. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil)], e-mail: cmsl@cdtn.br, e-mail: estefaniabio@yahoo.com.br, e-mail: antero@cdtn.br; Resende, Maria Aparecida de [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Microbiologia], e-mail: maressend@mono.icb.ufmg.br

    2009-07-01

    Sporotrichosis is a subacute or chronic infection caused by the fungus Sporothrix schenckii. Zoonotic transmission can occur after scratches or bites of animals, mainly cats, rodents, and armadillos. Up to the moment, no approved vaccine was reported for S. schenckii or to any important pathogenic fungi infection in humans, indicating the need to expand the research in this field and to explore new alternatives. The aim of this study was to evaluate the effects of gamma radiation in the viability, metabolic activity and reproductive ability of S. schenckii yeast cells for further studies on the development of a vaccine for immunization of cats and dogs. The culture of S. schenckii, in solid medium, was irradiated at doses ranging from 1.0 to 9.0 kGy. After each dose the reproductive capacity, viability and protein synthesis were estimated. The results showed that a reduction of 6 log{sub 10} cycles in the number of colonies was achieved at 6.0 kGy and after 8.0 kGy no colonies could be recovered. The viability analysis indicated that yeast cells remained viable up to 9.0 kGy. The results of protein synthesis analysis showed that the yeast cells, irradiated up to 9.0 kGy, were able to synthesize proteins. Our preliminary results indicated that for the yeast cells of S. schenckii, it is possible to find an absorbed dose in which the pathogen loses its reproductive ability, while retaining its viability, a necessary condition for the development of a radioattenuated yeast vaccine. (author)

  20. Effects of gamma radiation on Sporothrix schenckii yeast cells

    International Nuclear Information System (INIS)

    Lacerda, Camila M. de Sousa; Martins, Estefania Mara Nascimento; Andrade, Antero S.R.; Resende, Maria Aparecida de

    2009-01-01

    Sporotrichosis is a subacute or chronic infection caused by the fungus Sporothrix schenckii. Zoonotic transmission can occur after scratches or bites of animals, mainly cats, rodents, and armadillos. Up to the moment, no approved vaccine was reported for S. schenckii or to any important pathogenic fungi infection in humans, indicating the need to expand the research in this field and to explore new alternatives. The aim of this study was to evaluate the effects of gamma radiation in the viability, metabolic activity and reproductive ability of S. schenckii yeast cells for further studies on the development of a vaccine for immunization of cats and dogs. The culture of S. schenckii, in solid medium, was irradiated at doses ranging from 1.0 to 9.0 kGy. After each dose the reproductive capacity, viability and protein synthesis were estimated. The results showed that a reduction of 6 log 10 cycles in the number of colonies was achieved at 6.0 kGy and after 8.0 kGy no colonies could be recovered. The viability analysis indicated that yeast cells remained viable up to 9.0 kGy. The results of protein synthesis analysis showed that the yeast cells, irradiated up to 9.0 kGy, were able to synthesize proteins. Our preliminary results indicated that for the yeast cells of S. schenckii, it is possible to find an absorbed dose in which the pathogen loses its reproductive ability, while retaining its viability, a necessary condition for the development of a radioattenuated yeast vaccine. (author)

  1. Unraveling lipid metabolism in lipid-dependent pathogenic Malassezia yeasts

    NARCIS (Netherlands)

    Celis Ramirez, A.M.

    2017-01-01

    Malassezia yeasts are lipid-dependent fungal species that are common members of the human and animal skin microbiota. The lipid-dependency is a crucial trait in the adaptation process to grow on the skin but also plays a role in their pathogenic life style. Malassezia species can cause several skin

  2. Brewing characteristics of piezosensitive sake yeasts

    Science.gov (United States)

    Nomura, Kazuki; Hoshino, Hirofumi; Igoshi, Kazuaki; Onozuka, Haruka; Tanaka, Erika; Hayashi, Mayumi; Yamazaki, Harutake; Takaku, Hiroaki; Iguchi, Akinori; Shigematsu, Toru

    2018-04-01

    Application of high hydrostatic pressure (HHP) treatment to food processing is expected as a non-thermal fermentation regulation technology that supresses over fermentation. However, the yeast Saccharomyces cerevisiae used for Japanese rice wine (sake) brewing shows high tolerance to HHP. Therefore, we aimed to generate pressure-sensitive (piezosensitive) sake yeast strains by mating sake with piezosensitive yeast strains to establish an HHP fermentation regulation technology and extend the shelf life of fermented foods. The results of phenotypic analyses showed that the generated yeast strains were piezosensitive and exhibited similar fermentation ability compared with the original sake yeast strain. In addition, primary properties of sake brewed using these strains, such as ethanol concentration, sake meter value and sake flavor compounds, were almost equivalent to those obtained using the sake yeast strain. These results suggest that the piezosensitive strains exhibit brewing characteristics essentially equivalent to those of the sake yeast strain.

  3. Impacts of Verotoxigenic and Enterohaemorrhagic Escherichia Coli (Vtec and Ehec on Animal, Human and Food

    Directory of Open Access Journals (Sweden)

    Widodo Suwito

    2009-06-01

    Full Text Available Verotoxigenic and enterohaemorrhagic Escherichia coli, VTEC and EHEC are strains that produce a toxin on the Vero cells in vitro, therefore the toxin called verotoxin. Strain VTEC and EHEC have been isolated from human and various animal species, mainly ruminants and pigs. The prevalence of VTEC in cattle is 35%, milk 10%, cheese 1.5%. In beef meat, pork, poultry, goat and sheep is 3.7, 1.5, 1.5, 2 and 2.5%, respectively. In contrast, the human prevalence in United States, Spain and France 5 – 10, 2.5 and 3.4%, respectively. In Indonesia, nine cases of VTEC were reported by Ciptomangunkusumo Hospital and four out of nine cases were died. The verotoxin also named Shiga toxins (Stx, is active in vivo as a pathogen on the endothelial cells of the blood vessels of the gastro intestinal mucosa, kidneys, brain and other tissues of human and piglets. Verotoxin in human, ruminants and piglets causes bloody diarrhea so with ruminants. In the adult ruminants, verotoxin results in clinical signs and they act as asymptomatic carriers of VTEC and EHEC strains but in human, it produced haemorrhagic colitis (HC, haemolytic uremic syndrome (HUS and thrombocytopenia purpura (TPP. Infection in human by the most famous EHEC strain belongs to the O157:H7 serotype, through faecal contamination or environment of either food of animal origin, or other foodstuffs (fruits and vegetables. Infection from EHEC strains could be prevented by avoiding food of animal origin and unpasteurized milk, and by taking care of food hygiene for comsumption.

  4. YMDB: the Yeast Metabolome Database

    Science.gov (United States)

    Jewison, Timothy; Knox, Craig; Neveu, Vanessa; Djoumbou, Yannick; Guo, An Chi; Lee, Jacqueline; Liu, Philip; Mandal, Rupasri; Krishnamurthy, Ram; Sinelnikov, Igor; Wilson, Michael; Wishart, David S.

    2012-01-01

    The Yeast Metabolome Database (YMDB, http://www.ymdb.ca) is a richly annotated ‘metabolomic’ database containing detailed information about the metabolome of Saccharomyces cerevisiae. Modeled closely after the Human Metabolome Database, the YMDB contains >2000 metabolites with links to 995 different genes/proteins, including enzymes and transporters. The information in YMDB has been gathered from hundreds of books, journal articles and electronic databases. In addition to its comprehensive literature-derived data, the YMDB also contains an extensive collection of experimental intracellular and extracellular metabolite concentration data compiled from detailed Mass Spectrometry (MS) and Nuclear Magnetic Resonance (NMR) metabolomic analyses performed in our lab. This is further supplemented with thousands of NMR and MS spectra collected on pure, reference yeast metabolites. Each metabolite entry in the YMDB contains an average of 80 separate data fields including comprehensive compound description, names and synonyms, structural information, physico-chemical data, reference NMR and MS spectra, intracellular/extracellular concentrations, growth conditions and substrates, pathway information, enzyme data, gene/protein sequence data, as well as numerous hyperlinks to images, references and other public databases. Extensive searching, relational querying and data browsing tools are also provided that support text, chemical structure, spectral, molecular weight and gene/protein sequence queries. Because of S. cervesiae's importance as a model organism for biologists and as a biofactory for industry, we believe this kind of database could have considerable appeal not only to metabolomics researchers, but also to yeast biologists, systems biologists, the industrial fermentation industry, as well as the beer, wine and spirit industry. PMID:22064855

  5. Saccharomyces boulardii Preserves the Barrier Function and Modulates the Signal Transduction Pathway Induced in Enteropathogenic Escherichia coli-Infected T84 Cells

    Science.gov (United States)

    Czerucka, Dorota; Dahan, Stephanie; Mograbi, Baharia; Rossi, Bernard; Rampal, Patrick

    2000-01-01

    Use of the nonpathogenic yeast Saccharomyces boulardii in the treatment of infectious diarrhea has attracted growing interest. The present study designed to investigate the effect of this yeast on enteropathogenic Escherichia coli (EPEC)-associated disease demonstrates that S. boulardii abrogated the alterations induced by an EPEC strain on transepithelial resistance, [3H]inulin flux, and ZO-1 distribution in T84 cells. Moreover, EPEC-mediated apoptosis of epithelial cells was delayed in the presence of S. boulardii. The yeast did not modify the number of adherent bacteria but lowered by 50% the number of intracellular bacteria. Infection by EPEC induced tyrosine phosphorylation of several proteins in T84 cells, including p46 and p52 SHC isoforms, that was attenuated in the presence of S. boulardii. Similarly, EPEC-induced activation of the ERK1/2 mitogen-activated protein (MAP) kinase pathway was diminished in the presence of the yeast. Interestingly, inhibition of the ERK1/2 pathway with the specific inhibitor PD 98059 decreased EPEC internalization, suggesting that modulation of the ERK1/2 MAP pathway might account for the lowering of the number of intracellular bacteria observed in the presence of S. boulardii. Altogether, this study demonstrated that S. boulardii exerts a protective effect on epithelial cells after EPEC adhesion by modulating the signaling pathway induced by bacterial infection. PMID:10992512

  6. Role of wild birds as carriers of multi-drug resistant Escherichia coli and Escherichia vulneris

    Directory of Open Access Journals (Sweden)

    Mohammed Y. Shobrak

    2014-12-01

    Full Text Available Emergence and distribution of multi-drug resistant (MDR bacteria in environments pose a risk to human and animal health. A total of 82 isolates of Escherichia spp. were recovered from cloacal swabs of migrating and non-migrating wild birds. All bacterial isolates were identified and characterized morphologically and biochemically. 72% and 50% of isolates recovered from non-migrating and migrating birds, respectively, showed positive congo red dye binding (a virulence factor. Also, hemolysin production (a virulence factor was showed in 8% of isolates recovered from non-migrating birds and 75% of isolates recovered from migrating birds. All isolates recovered from non-migrating birds were found resistant to Oxacillin while all isolates recovered from migrating birds demonstrated resistance to Oxacillin, Chloramphenicol, Oxytetracycline and Lincomycin. Some bacterial isolates recovered from non-migrating birds and migrating birds exhibited MDR phenotype. The MDR isolates were further characterized by API 20E and 16S rRNA as E. coli and E. vulneris. MDR Escherichia isolates contain ~1-5 plasmids of high-molecular weights. Accordingly, wild birds could create a potential threat to human and animal health by transmitting MDR bacteria to water streams and other environmental sources through their faecal residues, and to remote regions by migration.

  7. Role of wild birds as carriers of multi-drug resistant Escherichia coli and Escherichia vulneris

    Science.gov (United States)

    Shobrak, Mohammed Y.; Abo-Amer, Aly E.

    2014-01-01

    Emergence and distribution of multi-drug resistant (MDR) bacteria in environments pose a risk to human and animal health. A total of 82 isolates of Escherichia spp. were recovered from cloacal swabs of migrating and non-migrating wild birds. All bacterial isolates were identified and characterized morphologically and biochemically. 72% and 50% of isolates recovered from non-migrating and migrating birds, respectively, showed positive congo red dye binding (a virulence factor). Also, hemolysin production (a virulence factor) was showed in 8% of isolates recovered from non-migrating birds and 75% of isolates recovered from migrating birds. All isolates recovered from non-migrating birds were found resistant to Oxacillin while all isolates recovered from migrating birds demonstrated resistance to Oxacillin, Chloramphenicol, Oxytetracycline and Lincomycin. Some bacterial isolates recovered from non-migrating birds and migrating birds exhibited MDR phenotype. The MDR isolates were further characterized by API 20E and 16S rRNA as E. coli and E. vulneris. MDR Escherichia isolates contain ~1–5 plasmids of high-molecular weights. Accordingly, wild birds could create a potential threat to human and animal health by transmitting MDR bacteria to water streams and other environmental sources through their faecal residues, and to remote regions by migration. PMID:25763023

  8. Characterization of yeast mutants lacking alkaline ceramidases YPC1 and YDC1

    DEFF Research Database (Denmark)

    Voynova, Natalia S; Mallela, Shamroop K; Vazquez, Hector M

    2014-01-01

    Humans and yeast possess alkaline ceramidases located in the early secretory pathway. Single deletions of the highly homologous yeast alkaline ceramidases YPC1 and YDC1 have very little genetic interactions or phenotypes. Here, we performed chemical-genetic screens to find deletions...... reduces chronological life span. A novel finding is that, when working backwards as a ceramide synthase in vivo, Ypc1p prefers C24 and C26 fatty acids as substrates, whereas it prefers C16:0, when solubilized in detergent and working in vitro. Therefore, its physiological activity may not only concern...... the minor ceramides containing C14 and C16. Intriguingly, so far the sole discernable benefit of conserving YPC1 for yeast resides with its ability to convey relative resistance toward H2 O2 ....

  9. Lipid Metabolic Versatility in Malassezia spp. Yeasts Studied through Metabolic Modeling

    NARCIS (Netherlands)

    Triana, Sergio; de Cock, Hans; Ohm, Robin A; Danies, Giovanna; Wösten, Han A B; Restrepo, Silvia; González Barrios, Andrés F; Celis Ramirez, Adriana

    2017-01-01

    Malassezia species are lipophilic and lipid-dependent yeasts belonging to the human and animal microbiota. Typically, they are isolated from regions rich in sebaceous glands. They have been associated with dermatological diseases such as seborrheic dermatitis, pityriasis versicolor, atopic

  10. Not your ordinary yeast: non-Saccharomyces yeasts in wine production uncovered.

    Science.gov (United States)

    Jolly, Neil P; Varela, Cristian; Pretorius, Isak S

    2014-03-01

    Saccharomyces cerevisiae and grape juice are 'natural companions' and make a happy wine marriage. However, this relationship can be enriched by allowing 'wild' non-Saccharomyces yeast to participate in a sequential manner in the early phases of grape must fermentation. However, such a triangular relationship is complex and can only be taken to 'the next level' if there are no spoilage yeast present and if the 'wine yeast' - S. cerevisiae - is able to exert its dominance in time to successfully complete the alcoholic fermentation. Winemakers apply various 'matchmaking' strategies (e.g. cellar hygiene, pH, SO2 , temperature and nutrient management) to keep 'spoilers' (e.g. Dekkera bruxellensis) at bay, and allow 'compatible' wild yeast (e.g. Torulaspora delbrueckii, Pichia kluyveri, Lachancea thermotolerans and Candida/Metschnikowia pulcherrima) to harmonize with potent S. cerevisiae wine yeast and bring the best out in wine. Mismatching can lead to a 'two is company, three is a crowd' scenario. More than 40 of the 1500 known yeast species have been isolated from grape must. In this article, we review the specific flavour-active characteristics of those non-Saccharomyces species that might play a positive role in both spontaneous and inoculated wine ferments. We seek to present 'single-species' and 'multi-species' ferments in a new light and a new context, and we raise important questions about the direction of mixed-fermentation research to address market trends regarding so-called 'natural' wines. This review also highlights that, despite the fact that most frontier research and technological developments are often focussed primarily on S. cerevisiae, non-Saccharomyces research can benefit from the techniques and knowledge developed by research on the former. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  11. Hybrids of Shigatoxigenic and Enterotoxigenic Escherichia coli (STEC/ETEC) Among Human and Animal Isolates in Finland.

    Science.gov (United States)

    Nyholm, O; Heinikainen, S; Pelkonen, S; Hallanvuo, S; Haukka, K; Siitonen, A

    2015-11-01

    Diarrhoeagenic Escherichia coli (DEC) cause serious foodborne infections in humans. Total of 450 Shigatoxigenic E. coli (STEC) strains isolated from humans, animals and environment in Finland were examined by multiplex PCR targeting the virulence genes of various DEC pathogroups simultaneously. One per cent (3/291) of the human STEC and 14% (22/159) of the animal and environmental STEC had genes typically present in enterotoxigenic E. coli (ETEC). The strains possessed genes encoding both Shiga toxin 1 and/or 2 (stx1 and/or stx2 ) and ETEC-specific heat-stable (ST) enterotoxin Ia (estIa). The identified stx subtypes were stx1a, stx1c, stx2a, stx2d and stx2g. The three human STEC/ETEC strains were isolated from the patients with haemolytic uraemic syndrome and diarrhoea and from an asymptomatic carrier. The animal STEC/ETEC strains were isolated from cattle and moose. The human and animal STEC/ETEC strains belonged to 11 serotypes, of which O2:H27, O15:H16, O101:H-, O128:H8 and O141:H8 have previously been described to be associated with human disease. Identification of multiple virulence genes offers further information for assessing the virulence potential of STEC and other DEC. The emergence of novel hybrid pathogens should be taken into account in the patient care and epidemiological surveillance. © 2015 Blackwell Verlag GmbH.

  12. Effect of yeast storage temperature and flour composition on fermentative activities of baker's yeast

    Directory of Open Access Journals (Sweden)

    Pejin Dušanka J.

    2009-01-01

    Full Text Available Baker's yeast is a set of living cells of Saccharomyces cerevisiae. It contains around 70-72% of water, 42-45% of proteins, around 40% of carbohydrates, around 7.5% of lipids (based on dry matter, and vitamin B-complex. On the basis of yeast cell analysis it can be concluded that yeast is a complex biological system which changes in time. The intensity of the changes depends on temperature. Yeast sample was stored at 4°C i 24°C for 12 days. During storage at 4°C, the content of total carbohydrates decreased from 48.81% to 37.50% (dry matter, whereas carbohydrate loss ranged from 40.81% to 29.28% at 24°C. The content of trehalose was 12.33% in the yeast sample stored at 4°C and 0.24% at 24°C. Loss of fermentative activity was 81.76% in the sample stored at 24°C for 12 days. The composition of five samples of 1st category flour was investigated. It was found that flours containing more reducing sugars and maltose enable higher fermentation activities. The flours with higher ash content (in the range 0.5-0.94% had higher contents of phytic acid. Higher ash and phytic contents in flour increased the yeast fermentative efficiency. In bakery industry, a range of ingredients has been applied to improve the product's quality such as surface active substances (emulsifiers, enzymes, sugars and fats. In the paper, the effect of some ingredients added to dough (margarine, saccharose, sodium chloride and malted barley on the yeast fermentative activity was studied. The mentioned ingredients were added to dough at different doses: 0.5, 1.0, 1.5 and 2.0%, flour basis. It was found that the investigated ingredients affected the fermentative activity of yeast and improved the bread quality.

  13. Antibiotic resistance of Verotoxigenic Escherichia coli isolated from vegetables

    Directory of Open Access Journals (Sweden)

    mojtaba boniadian

    2017-01-01

    Full Text Available Introduction: Human gastrointestinal disease caused by verotoxigenic Escherichia coli has been diagnosed for recent decades. Escherichia coli O157:H7 is the most important serotype of verotoxigenic Escherichia coli that cause hemolytic uremic syndrome and hemorrhagic colitis in humans. This study was conducted to determine the occurrence of verotoxigenic E. coli and antibiotic resistance of the isolates from vegetables. Materials and methods: A total of 500 fresh vegetable samples were collected randomly from retail shops in Shahrekord, Iran. E. coli was isolated and identified using bacteriological and biochemical tests. PCR method was used to identify the rbfE, stx1, stx2 and eae genes. Also, antibiotic resistance of the isolates was determined by disk diffusion method. Results: The results represented that among 25 isolates possess virulence genes, 40, 12 and 4% of the isolates contained eaeA, STx2, and both genes, respectively. But none of them contained H7, STx1, and rfbE genes. The antibiotic resistance pattern demonstrated that the isolates were highly resistant to Gentamycin and cefotoxime. Discussion and conclusion: The results of this study showed that the presence of verotoxigenic E.coli in vegetables; and high resistance of the isolates to antibiotics could be hazardous for public health.

  14. Characterization of epidemic IncI1-Iγ plasmids harboring ambler class A and C genes in Escherichia coli and Salmonella enterica from animals and humans

    NARCIS (Netherlands)

    Smith, Hilde; Bossers, Alex; Harders, Frank; Wu, Guanghui; Woodford, Neil; Schwarz, Stefan; Guerra, Beatriz; Rodríguez, Irene; van Essen-Zandbergen, Alieda; Brouwer, Michael; Mevius, Dik

    The aim of the study was to identify the plasmid-encoded factors contributing to the emergence and spread of epidemic IncI1-Iγ plasmids obtained from Escherichia coli and Salmonella enterica isolates from animal and human reservoirs. For this, 251 IncI1-Iγ plasmids carrying various extended-spectrum

  15. Isolation and genomic characterization of Escherichia coli O157:NM ...

    African Journals Online (AJOL)

    Human diseases caused by Escherichia coli O157:NM and E. coli O157:H7 strains have been reported throughout the world. In developed countries, serotype O157:H7 represents the major cause of human diseases; however, there have been increasing reports of non-O157 Shiga toxin (Stx)-producing E. coli strains ...

  16. Yeast strains and methods of use thereof

    OpenAIRE

    Goddard, Matthew Robert; Gardner, Richard Clague; Anfang, Nicole

    2013-01-01

    The present invention relates to yeast strains and, in particular, to yeast stains for use in fermentation processes. The invention also relates to methods of fermentation using the yeast strains of the invention either alone or in combination with other yeast strains. The invention thither relates to methods for the selection of yeast strains suitable for fermentation cultures by screening for various metabolic products and the use of specific nutrient sources.

  17. Bioactive Compounds Derived from the Yeast Metabolism of Aromatic Amino Acids during Alcoholic Fermentation

    Directory of Open Access Journals (Sweden)

    Albert Mas

    2014-01-01

    Full Text Available Metabolites resulting from nitrogen metabolism in yeast are currently found in some fermented beverages such as wine and beer. Their study has recently attracted the attention of researchers. Some metabolites derived from aromatic amino acids are bioactive compounds that can behave as hormones or even mimic their role in humans and may also act as regulators in yeast. Although the metabolic pathways for their formation are well known, the physiological significance is still far from being understood. The understanding of this relevance will be a key element in managing the production of these compounds under controlled conditions, to offer fermented food with specific enrichment in these compounds or even to use the yeast as nutritional complements.

  18. Immobilization of yeast cells by radiation-induced polymerization

    International Nuclear Information System (INIS)

    Fujimura, T.; Kaetsu, I.

    1982-01-01

    Radiation-induced polymerization method was applied to the immobilization of yeast cells. The effects of irradiation, cooling and monomer, which are neccessary for polymerization, were recovered completely by subsequent aerobical incubation of yeast cells. The ethanol productive in immobilized yeast cells increased with the increase of aerobical incubation period. The growth of yeast cells in immobilized yeast cells was indicated. The maximum ethanol productivity in immobilized yeast cell system was around three times as much as that in free yeast cell system. (orig.)

  19. Evolutionary History of Ascomyceteous Yeasts

    Energy Technology Data Exchange (ETDEWEB)

    Haridas, Sajeet; Riley, Robert; Salamov, Asaf; Goker, Markus; Klenk, Hans-Peter; Kurtzman, Cletus P.; Blackwell, Meredith; Grigoriev, Igor; Jeffries, Thomas W.

    2014-06-06

    Yeasts are important for many industrial and biotechnological processes and show remarkable diversity despite morphological similarities. We have sequenced the genomes of 16 ascomycete yeasts of taxonomic and industrial importance including members of Saccharomycotina and Taphrinomycotina. A comparison of these with several other previously published yeast genomes have added increased confidence to the phylogenetic positions of previously poorly placed species including Saitoella complicata, Babjeviella inositovora and Metschnikowia bicuspidata. Phylogenetic analysis also showed that yeasts with alternative nuclear codon usage where CUG encodes serine instead of leucine are monophyletic within the Saccharomycotina. Most of the yeasts have compact genomes with a large fraction of single exon genes with Lipomyces starkeyi and the previously published Pneumocystis jirovecii being notable exceptions. Intron analysis suggests that early diverging species have more introns. We also observed a large number of unclassified lineage specific non-simple repeats in these genomes.

  20. Cloning and characterization of a functional human homolog of Escherichia coli endonuclease III

    Science.gov (United States)

    Aspinwall, Richard; Rothwell, Dominic G.; Roldan-Arjona, Teresa; Anselmino, Catherine; Ward, Christopher J.; Cheadle, Jeremy P.; Sampson, Julian R.; Lindahl, Tomas; Harris, Peter C.; Hickson, Ian D.

    1997-01-01

    Repair of oxidative damage to DNA bases is essential to prevent mutations and cell death. Endonuclease III is the major DNA glycosylase activity in Escherichia coli that catalyzes the excision of pyrimidines damaged by ring opening or ring saturation, and it also possesses an associated lyase activity that incises the DNA backbone adjacent to apurinic/apyrimidinic sites. During analysis of the area adjacent to the human tuberous sclerosis gene (TSC2) in chromosome region 16p13.3, we identified a gene, OCTS3, that encodes a 1-kb transcript. Analysis of OCTS3 cDNA clones revealed an open reading frame encoding a predicted protein of 34.3 kDa that shares extensive sequence similarity with E. coli endonuclease III and a related enzyme from Schizosaccharomyces pombe, including a conserved active site region and an iron/sulfur domain. The product of the OCTS3 gene was therefore designated hNTH1 (human endonuclease III homolog 1). The hNTH1 protein was overexpressed in E. coli and purified to apparent homogeneity. The recombinant protein had spectral properties indicative of the presence of an iron/sulfur cluster, and exhibited DNA glycosylase activity on double-stranded polydeoxyribonucleotides containing urea and thymine glycol residues, as well as an apurinic/apyrimidinic lyase activity. Our data indicate that hNTH1 is a structural and functional homolog of E. coli endonuclease III, and that this class of enzymes, for repair of oxidatively damaged pyrimidines in DNA, is highly conserved in evolution from microorganisms to human cells. PMID:8990169

  1. Association Between Antimicrobial Resistance in Escherichia coli Isolates from Food Animals and Blood Stream Isolates from Humans in Europe: An Ecological Study

    DEFF Research Database (Denmark)

    Vieira, Antonio; Collignon, Peter; Aarestrup, Frank Møller

    2011-01-01

    Background: In addition to medical antimicrobial usage, the use of antimicrobials in food animals contributes to the occurrence of resistance among some bacterial species isolated from infections in humans. Recently, several studies have indicated that a large proportion of Escherichia coli causing...... infections in humans, especially those resistant to antimicrobials, have an animal origin.Methods: We analyzed the correlation between the prevalence of antimicrobial resistance in E. coli isolates from blood stream infections in humans and in E. coli isolates from poultry, pigs, and cattle between 2005...... and 2008 for 11 countries, using available surveillance data. We also assessed the correlation between human antimicrobial usage and the occurrence of resistance in E. coli isolates from blood stream infections.Results: Strong and significant correlations between prevalences of resistance to ampicillin (r...

  2. Characterization of recombinant human lysosomal beta-hexosaminidases produced in the methylotrophic yeast Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Angela Johana Espejo Mojica

    2016-08-01

    Full Text Available β-hexosaminidases (Hex are dimeric enzymes involved in the lysosomal degradation of glycolipids and glycans. They are formed by α- and/or β-subunits encoded byHEXA and HEXB genes, respectively. Mutations in these genes lead to Tay Sachs or Sandhoff diseases, which are neurodegenerative disorders caused by the accumulation of non-degraded glycolipids. Although tissue-derived Hex have been widely characterized, limited information is available for recombinant β-hexosaminidases. In this study, human lysosomal recombinant Hex (rhHex-A, rhHex-B, and rhHex-S were produced in the methylotrophic yeast Pichia pastoris GS115. The highest specific enzyme activities were 13,124 for rhHexA; 12,779 for rhHex-B; and 14,606 U .mg-1 for rhHex-S. These results were 25- to 50-fold higher than those obtained from normal human leukocytes. Proteins were purified and characterized at different pH and temperature conditions. All proteins were stable at acidic pH, and at 4 °C and 37 °C. At 45 °C rhHex-S was completely inactivated, while rhHex-A and rhHex-B showed high stability. This study demonstrates P. pastoris GS115 potential for polymeric lysosomal enzyme production, and describes the characterization of recombinant β-hexosaminidases produced within the same host.

  3. Sexual differentiation in fission yeast

    DEFF Research Database (Denmark)

    Egel, R; Nielsen, O; Weilguny, D

    1990-01-01

    The regulation of sexual reproduction in yeast constitutes the highest level of differentiation observed in these unicellular organisms. The various ramifications of this system involve DNA rearrangement, transcriptional control, post-translational modification (such as protein phosphorylation) a......) and receptor/signal processing. A few basic similarities are common to both fission and budding yeasts. The wiring of the regulatory circuitry, however, varies considerably between these divergent yeast groups....

  4. Type 1 diabetes in children is not a predisposing factor for oral yeast colonization.

    Science.gov (United States)

    Costa, Ana L; Silva, Branca M A; Soares, Rui; Mota, Diana; Alves, Vera; Mirante, Alice; Ramos, João C; Maló de Abreu, João; Santos-Rosa, Manuel; Caramelo, Francisco; Gonçalves, Teresa

    2017-06-01

    Type 1 diabetes mellitus (T1D) is considered a risk factor associated with oral yeast infections. The aim of this study was to evaluate the yeast oral carriage (in saliva and mucosal surface) of children with T1D and potential relation with host factors, particularly the subset of CD4+ T cells. Yeasts were quantified and identified in stimulated saliva and in cheek mucosal swabs of 133 diabetic T1D and 72 healthy control subjects. Salivary lymphocytes were quantified using flow cytometry. The presence of yeasts in the oral cavity (60% of total patients) was not affected by diabetes, metabolic control, duration of the disease, salivary flow rate or saliva buffer capacity, by age, sex, place of residence, number of daily meals, consumption of sweets or frequency of tooth brushing. Candida albicans was the most prevalent yeast species, but a higher number of yeast species was isolated in nondiabetics. T1D children with HbA1c ≤ 7.5 (metabolically controlled) presented higher number of CD4+ T salivary subsets when compared with the other groups of children (non-diabetic and nonmetabolically controlled) and also presented the highest number of individuals without oral yeast colonization. In conclusion, T1D does not predisposes for increased oral yeast colonization and a higher number of salivary CD4+T cells seems to result in the absence of oral colonization by yeasts. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Occurrence of Killer Yeast Strains in Fruit and Berry Wine Yeast Populations

    Directory of Open Access Journals (Sweden)

    Gintare Gulbiniene

    2004-01-01

    Full Text Available Apple, cranberry, chokeberry and Lithuanian red grape wine yeast populations were used for the determination of killer yeast occurrence. According to the tests of the killer characteristics and immunity the isolated strains were divided into seven groups. In this work the activity of killer toxins purified from some typical strains was evaluated. The analysed strains produced different amounts of active killer toxin and some of them possessed new industrially significant killer properties. Total dsRNA extractions in 11 killer strains of yeast isolated from spontaneous fermentations revealed that the molecular basis of the killer phenomenon was not only dsRNAs, but also unidentified genetic determinants.

  6. Mechanisms of uv mutagenesis in yeast

    International Nuclear Information System (INIS)

    Lawrence, C.W.; Christensen, R.; Schwartz, A.

    1982-01-01

    The uv mutagenesis in yeast depends on the function of the RAD6 locus, a gene that is also responsible for a substantial fraction of wild-type resistance, suggesting that this eukaryote may possess a misrepair mechanism analogous to that proposed for Escherichia coli. The molecular mechanism responsible for RAD6 repair or recovery is not yet known, but it is different from either excision or recombination-dependent repair, processes carried out by the other two main repair pathways in yeast. RAD6-dependent mutagenesis has been found to have the following characteristics. It is associated at best with only a small fraction of RAD6-dependent repair, the majority of the sensitivity of rad6 mutants being due to their lack of nonmutagenic repair. SRS2 metabolic suppressors restore a substantial fraction of uv resistance to rad6 mutants but do not restore their uv mutability. Strains containing mutations at loci (rev, umr) that are probably more directly involved in mutagenesis are only mildly sensitive, and there is a poor correlation between their sensitivity and mutational deficiency. The uv mutagenesis appears to require a large number of gene functions, perhaps ten or more. Where examined in detail, these genes have been found to be concerned in the production of only a specific range of mutational events, not all of them. Mating experiments have shown that a substantial fraction, probably 40% or more, of uv-induced mutations are untargeted, that is, occur in lesion-free regions of DNA. The uv irradiation, therefore, produces a general reduction in the normally high fidelity with which DNA is replicated on undamaged templates. It does not appear to be necessary for the causal lesion to be present in the same chromosome as the mutation it induces. The reduction in fidelity may be the consequence of the production of a diffusible factor in uv-irradiated cells, but definite evidence supporting this proposal has not yet been obtained

  7. Molecular comparisons for identification of food spoilage yeasts and prediction of species that may develop in different food products

    Science.gov (United States)

    Spoilage of foods and beverages by yeasts is often characterized by objectionable odors, appearance, taste, texture or build-up of gas in packaging containers, resulting in loss of the product. Seldom is human health compromised by products spoiled by yeasts even though some spoilage is caused by sp...

  8. Virgin olive oil yeasts: A review.

    Science.gov (United States)

    Ciafardini, Gino; Zullo, Biagi Angelo

    2018-04-01

    This review summarizes current knowledge on virgin olive oil yeasts. Newly produced olive oil contains solid particles and micro drops of vegetation water in which yeasts reproduce to become the typical microbiota of olive oil. To date, about seventeen yeast species have been isolated from different types of olive oils and their by-products, of which six species have been identified as new species. Certain yeast species contribute greatly to improving the sensorial characteristics of the newly produced olive oil, whereas other species are considered harmful as they can damage the oil quality through the production of unpleasant flavors and triacylglycerol hydrolysis. Studies carried out in certain yeast strains have demonstrated the presence of defects in olive oil treated with Candida adriatica, Nakazawaea wickerhamii and Candida diddensiae specific strains, while other olive oil samples treated with other Candida diddensiae strains were defect-free after four months of storage and categorized as extra virgin. A new acetic acid producing yeast species, namely, Brettanomyces acidodurans sp. nov., which was recently isolated from olive oil, could be implicated in the wine-vinegary defect of the product. Other aspects related to the activity of the lipase-producing yeasts and the survival of the yeast species in the flavored olive oils are also discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Growth inhibition of pathogenic bacteria and some yeasts by selected essential oils and survival of L. monocytogenes and C. albicans in apple-carrot juice.

    Science.gov (United States)

    Irkin, Reyhan; Korukluoglu, Mihriban

    2009-04-01

    Food safety is a fundamental concern of both consumers and the food industry. The increasing incidence of foodborne diseases increases the demand of using antimicrobials in foods. Spices and plants are rich in essential oils and show inhibition activity against microorganisms, which are composed of many compounds. In this research, effects of garlic, bay, black pepper, origanum, orange, thyme, tea tree, mint, clove, and cumin essential oils on Listeria monocytogenes AUFE 39237, Escherichia coli ATCC 25922, Salmonella enteritidis ATCC 13076, Proteus mirabilis AUFE 43566, Bacillus cereus AUFE 81154, Saccharomyces uvarum UUFE 16732, Kloeckera apiculata UUFE 10628, Candida albicans ATCC 10231, Candida oleophila UUPP 94365, and Metschnikowia fructicola UUPP 23067 and effects of thyme oil at a concentration of 0.5% on L. monocytogenes and C. albicans in apple-carrot juice during +4 degrees C storage (first to fifth day) were investigated. Strong antibacterial and antifungal activities of some essential oils were found. Thyme, origanum, clove, and orange essential oils were the most inhibitory against bacteria and yeasts. Cumin, tea tree, and mint oils inhibited the yeasts actively. It is concluded that some essential oils could be used as potential biopreservatives capable of controlling foodborne pathogens and food spoilage yeasts.

  10. Terroir of yeasts? – Application of FTIR spectroscopy and molecular methods for strain typing of yeasts

    Directory of Open Access Journals (Sweden)

    Gerhards Daniel

    2015-01-01

    Full Text Available The site specific influence on wine (Terroir is an often by wine producers, consumers and scientists discussed topic in the world of wine. A study on grapes and (spontaneous fermentations from six different vineyards was done to investigate the biodiversity of yeasts and to answer the question if there is a terroir of yeast and how it could be influenced. Randomly isolated yeasts were identified by FTIR-spectroscopy and molecular methods on species and strain level. Vineyard specific yeast floras would be observed but they are not such important as expected. Only a few overlapping strain patterns would be identified during both vintages. The yeast flora of the winery had a huge impact on the spontaneous fermentations, but is not really constant and influenced by different factors from outside.

  11. 21 CFR 184.1983 - Bakers yeast extract.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Bakers yeast extract. 184.1983 Section 184.1983... Listing of Specific Substances Affirmed as GRAS § 184.1983 Bakers yeast extract. (a) Bakers yeast extract... a selected strain of yeast, Saccharomyces cerevisiae. It may be concentrated or dried. (b) The...

  12. Endoplasmic reticulum involvement in yeast cell death

    International Nuclear Information System (INIS)

    Nicanor Austriaco, O.

    2012-01-01

    Yeast cells undergo programed cell death (PCD) with characteristic markers associated with apoptosis in mammalian cells including chromatin breakage, nuclear fragmentation, reactive oxygen species generation, and metacaspase activation. Though significant research has focused on mitochondrial involvement in this phenomenon, more recent work with both Saccharomyces cerevisiae and Schizosaccharomyces pombe has also implicated the endoplasmic reticulum (ER) in yeast PCD. This minireview provides an overview of ER stress-associated cell death (ER-SAD) in yeast. It begins with a description of ER structure and function in yeast before moving to a discussion of ER-SAD in both mammalian and yeast cells. Three examples of yeast cell death associated with the ER will be highlighted here including inositol starvation, lipid toxicity, and the inhibition of N-glycosylation. It closes by suggesting ways to further examine the involvement of the ER in yeast cell death.

  13. The Yeast Retrograde Response as a Model of Intracellular Signaling of Mitochondrial Dysfunction

    Directory of Open Access Journals (Sweden)

    S. Michal eJazwinski

    2012-05-01

    Full Text Available Mitochondrial dysfunction activates intracellular signaling pathways that impact yeast longevity, and the best known of these pathways is the retrograde response. More recently, similar responses have been discerned in other systems, from invertebrates to human cells. However, the identity of the signal transducers is either unknown or apparently diverse, contrasting with the well-established signaling module of the yeast retrograde response. On the other hand, it has become equally clear that several other pathways and processes interact with the retrograde response, embedding it in a network responsive to a variety of cellular states. An examination of this network supports the notion that the master regulator NFkB aggregated a variety of mitochondria-related cellular responses at some point in evolution and has become the retrograde transcription factor. This has significant consequences for how we view some of the deficits associated with aging, such as inflammation. The support for NFkB as the retrograde response transcription factor is not only based on functional analyses. It is bolstered by the fact that NFkB can regulate Myc-Max, which is activated in human cells with dysfunctional mitochondria and impacts cellular metabolism. Myc-Max is homologous to the yeast retrograde response transcription factor Rtg1-Rtg3. Further research will be needed to disentangle the pro-aging from the anti-aging effects of NFkB. Interestingly, this is also a challenge for the complete understanding of the yeast retrograde response.

  14. Screening of intact yeasts and cell extracts to reduce Scrapie prions during biotransformation of food waste.

    Science.gov (United States)

    Huyben, David; Boqvist, Sofia; Passoth, Volkmar; Renström, Lena; Allard Bengtsson, Ulrika; Andréoletti, Olivier; Kiessling, Anders; Lundh, Torbjörn; Vågsholm, Ivar

    2018-02-08

    Yeasts can be used to convert organic food wastes to protein-rich animal feed in order to recapture nutrients. However, the reuse of animal-derived waste poses a risk for the transmission of infectious prions that can cause neurodegeneration and fatality in humans and animals. The aim of this study was to investigate the ability of yeasts to reduce prion activity during the biotransformation of waste substrates-thereby becoming a biosafety hurdle in such a circular food system. During pre-screening, 30 yeast isolates were spiked with Classical Scrapie prions and incubated for 72 h in casein substrate, as a waste substitute. Based on reduced Scrapie seeding activity, waste biotransformation and protease activities, intact cells and cell extracts of 10 yeasts were further tested. Prion analysis showed that five yeast species reduced Scrapie seeding activity by approximately 1 log10 or 90%. Cryptococcus laurentii showed the most potential to reduce prion activity since both intact and extracted cells reduced Scrapie by 1 log10 and achieved the highest protease activity. These results show that select forms of yeast can act as a prion hurdle during the biotransformation of waste. However, the limited ability of yeasts to reduce prion activity warrants caution as a sole barrier to transmission as higher log reductions are needed before using waste-cultured yeast in circular food systems.

  15. Integron, Plasmid and Host Strain Characteristics of Escherichia coli from Humans and Food Included in the Norwegian Antimicrobial Resistance Monitoring Programs.

    Science.gov (United States)

    Sunde, Marianne; Simonsen, Gunnar Skov; Slettemeås, Jannice Schau; Böckerman, Inger; Norström, Madelaine

    2015-01-01

    Antimicrobial resistant Escherichia coli (n=331) isolates from humans with bloodstream infections were investigated for the presence of class 1 and class 2 integrons. The integron cassettes arrays were characterized and the findings were compared with data from similar investigations on resistant E. coli from meat and meat products (n=241) produced during the same time period. All isolates were obtained from the Norwegian monitoring programs for antimicrobial resistance in human pathogens and in the veterinary sector. Methods used included PCR, sequencing, conjugation experiments, plasmid replicon typing and subtyping, pulsed-field-gel-electrophoresis and serotyping. Integrons of class 1 and 2 occurred significantly more frequently among human isolates; 45.4% (95% CI: 39.9-50.9) than among isolates from meat; 18% (95% CI: 13.2 -23.3), (pfood source and from a human clinical sample highlights the possible role of meat as a source of resistance elements for pathogenic bacteria.

  16. 21 CFR 172.898 - Bakers yeast glycan.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Bakers yeast glycan. 172.898 Section 172.898 Food... Multipurpose Additives § 172.898 Bakers yeast glycan. Bakers yeast glycan may be safely used in food in accordance with the following conditions: (a) Bakers yeast glycan is the comminuted, washed, pasteurized, and...

  17. Transcriptional activation of a geranylgeranyl diphosphate synthase gene, GGPPS2, isolated from Scoparia dulcis by treatment with methyl jasmonate and yeast extract.

    Science.gov (United States)

    Yamamura, Y; Mizuguchi, Y; Taura, F; Kurosaki, F

    2014-10-01

    A cDNA clone, designated SdGGPPS2, was isolated from young seedlings of Scoparia dulcis. The putative amino acid sequence of the translate of the gene showed high homology with geranylgeranyl diphosphate synthase (GGPPS) from various plant sources, and the N-terminal residues exhibited the characteristics of chloroplast targeting sequence. An appreciable increase in the transcriptional level of SdGGPPS2 was observed by exposure of the leaf tissues of S. dulcis to methyl jasmonate, yeast extract or Ca(2+) ionophore A23187. In contrast, SdGGPPS1, a homologous GGPPS gene of the plant, showed no or only negligible change in the expression level upon treatment with these stimuli. The truncated protein heterologously expressed in Escherichia coli in which the putative targeting domain was deleted catalyzed the condensation of farnesyl diphosphate and isopentenyl diphosphate to liberate geranylgeranyl diphosphate. These results suggested that SdGGPPS2 plays physiological roles in methyl jasmonate and yeast extract-induced metabolism in the chloroplast of S. dulcis cells.

  18. The wine and beer yeast Dekkera bruxellensis.

    Science.gov (United States)

    Schifferdecker, Anna Judith; Dashko, Sofia; Ishchuk, Olena P; Piškur, Jure

    2014-09-01

    Recently, the non-conventional yeast Dekkera bruxellensis has been gaining more and more attention in the food industry and academic research. This yeast species is a distant relative of Saccharomyces cerevisiae and is especially known for two important characteristics: on the one hand, it is considered to be one of the main spoilage organisms in the wine and bioethanol industry; on the other hand, it is 'indispensable' as a contributor to the flavour profile of Belgium lambic and gueuze beers. Additionally, it adds to the characteristic aromatic properties of some red wines. Recently this yeast has also become a model for the study of yeast evolution. In this review we focus on the recently developed molecular and genetic tools, such as complete genome sequencing and transformation, to study and manipulate this yeast. We also focus on the areas that are particularly well explored in this yeast, such as the synthesis of off-flavours, yeast detection methods, carbon metabolism and evolutionary history. © 2014 The Authors. Yeast published by John Wiley & Sons, Ltd.

  19. Mitochondrion-mediated cell death: dissecting yeast apoptosis for a better understanding of neurodegeneration

    Energy Technology Data Exchange (ETDEWEB)

    Braun, Ralf J., E-mail: ralf.braun@uni-bayreuth.de [Institut für Zellbiologie, Universität Bayreuth, Bayreuth (Germany)

    2012-11-28

    Mitochondrial damage and dysfunction are common hallmarks for neurodegenerative disorders, including Alzheimer, Parkinson, Huntington diseases, and the motor neuron disorder amyotrophic lateral sclerosis. Damaged mitochondria pivotally contribute to neurotoxicity and neuronal cell death in these disorders, e.g., due to their inability to provide the high energy requirements for neurons, their generation of reactive oxygen species (ROS), and their induction of mitochondrion-mediated cell death pathways. Therefore, in-depth analyses of the underlying molecular pathways, including cellular mechanisms controlling the maintenance of mitochondrial function, is a prerequisite for a better understanding of neurodegenerative disorders. The yeast Saccharomyces cerevisiae is an established model for deciphering mitochondrial quality control mechanisms and the distinct mitochondrial roles during apoptosis and programmed cell death. Cell death upon expression of various human neurotoxic proteins has been characterized in yeast, revealing neurotoxic protein-specific differences. This review summarizes how mitochondria are affected in these neurotoxic yeast models, and how they are involved in the execution and prevention of cell death. I will discuss to which extent this mimics the situation in other neurotoxic model systems, and how this may contribute to a better understanding of the mitochondrial roles in the human disorders.

  20. Mitochondrion-mediated cell death: dissecting yeast apoptosis for a better understanding of neurodegeneration

    International Nuclear Information System (INIS)

    Braun, Ralf J.

    2012-01-01

    Mitochondrial damage and dysfunction are common hallmarks for neurodegenerative disorders, including Alzheimer, Parkinson, Huntington diseases, and the motor neuron disorder amyotrophic lateral sclerosis. Damaged mitochondria pivotally contribute to neurotoxicity and neuronal cell death in these disorders, e.g., due to their inability to provide the high energy requirements for neurons, their generation of reactive oxygen species (ROS), and their induction of mitochondrion-mediated cell death pathways. Therefore, in-depth analyses of the underlying molecular pathways, including cellular mechanisms controlling the maintenance of mitochondrial function, is a prerequisite for a better understanding of neurodegenerative disorders. The yeast Saccharomyces cerevisiae is an established model for deciphering mitochondrial quality control mechanisms and the distinct mitochondrial roles during apoptosis and programmed cell death. Cell death upon expression of various human neurotoxic proteins has been characterized in yeast, revealing neurotoxic protein-specific differences. This review summarizes how mitochondria are affected in these neurotoxic yeast models, and how they are involved in the execution and prevention of cell death. I will discuss to which extent this mimics the situation in other neurotoxic model systems, and how this may contribute to a better understanding of the mitochondrial roles in the human disorders.

  1. Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption.

    Science.gov (United States)

    Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

    2015-01-01

    The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry.

  2. Implication of Ca2+ in the Regulation of Replicative Life Span of Budding Yeast*

    OpenAIRE

    Tsubakiyama, Ryohei; Mizunuma, Masaki; Gengyo, Anri; Yamamoto, Josuke; Kume, Kazunori; Miyakawa, Tokichi; Hirata, Dai

    2011-01-01

    In eukaryotic cells, Ca2+-triggered signaling pathways are used to regulate a wide variety of cellular processes. Calcineurin, a highly conserved Ca2+/calmodulin-dependent protein phosphatase, plays key roles in the regulation of diverse biological processes in organisms ranging from yeast to humans. We isolated a mutant of the SIR3 gene, implicated in the regulation of life span, as a suppressor of the Ca2+ sensitivity of zds1Δ cells in the budding yeast Saccharomyces cerevisiae. Therefore, ...

  3. Between science and industry-applied yeast research.

    Science.gov (United States)

    Korhola, Matti

    2018-03-01

    I was fortunate to enter yeast research at the Alko Research Laboratories with a strong tradition in yeast biochemistry and physiology studies. At the same time in the 1980s there was a fundamental or paradigm change in molecular biology research with discoveries in DNA sequencing and other analytical and physical techniques for studying macromolecules and cells. Since that time biotechnological research has expanded the traditional fermentation industries to efficient production of industrial and other enzymes and specialty chemicals. Our efforts were directed towards improving the industrial production organisms: minerals enriched yeasts (Se, Cr, Zn) and high glutathione content yeast, baker´s, distiller´s, sour dough and wine yeasts, and the fungal Trichoderma reesei platform for enzyme production. I am grateful for the trust of my colleagues in several leadership positions at the Alko Research Laboratories, Yeast Industry Platform and at the international yeast community.

  4. Teaching microbial physiology using glucose repression phenomenon in baker's yeast as an examplele

    DEFF Research Database (Denmark)

    Vijayendran, Raghavendran; Nielsen, Jens; Olsson, Lisbeth

    2005-01-01

    The yeast Saccharomyces cerevisiae has been used by human beings since ancient times for its ability to convert sugar to alcohol. Continual exposure to glucose in the natural environment for innumerable generations has probably enabled S. cerevisiae to grow in fermentative mode on sugars by switc......The yeast Saccharomyces cerevisiae has been used by human beings since ancient times for its ability to convert sugar to alcohol. Continual exposure to glucose in the natural environment for innumerable generations has probably enabled S. cerevisiae to grow in fermentative mode on sugars...... by switching off the genes responsible for respiration even under aerobic conditions. This phenomenon is referred to as the Crabtree effect. The present review focuses on glucose repression in S. cerevisiae from a physiological perspective. Physiological studies presented involve batch and chemostat...

  5. Functional properties of the two redox-active sites in yeast protein disulphide isomerase in vitro and in vivo

    DEFF Research Database (Denmark)

    Westphal, V; Darby, N J; Winther, Jakob R.

    1999-01-01

    to that of human PDI, both in rearrangement and oxidation reactions. However, while the a domain active site of the human enzyme is more active than the a'-site, the reverse is the case for yPDI. This prompted us to set up an assay to investigate whether the situation would be different with a native yeast......-site to be most important. We furthermore show that the apparent difference between in vivo and in vitro activities is not due to catalytic contributions from the other PDI homologues found in yeast....

  6. Formation of new chromosomes as a virulence mechanism in yeast Candida glabrata

    DEFF Research Database (Denmark)

    Poláková, S.; Blume, C.; Zárate, J. A.

    2009-01-01

    , Candida glabrata, for their genome structure and stability. This organism has recently become the second most prevalent yeast pathogen in humans. Although the gene sequences were well conserved among different strains, their chromosome structures differed drastically. The most frequent events reshaping...

  7. Yeast-based biosensors: design and applications.

    Science.gov (United States)

    Adeniran, Adebola; Sherer, Michael; Tyo, Keith E J

    2015-02-01

    Yeast-based biosensing (YBB) is an exciting research area, as many studies have demonstrated the use of yeasts to accurately detect specific molecules. Biosensors incorporating various yeasts have been reported to detect an incredibly large range of molecules including but not limited to odorants, metals, intracellular metabolites, carcinogens, lactate, alcohols, and sugars. We review the detection strategies available for different types of analytes, as well as the wide range of output methods that have been incorporated with yeast biosensors. We group biosensors into two categories: those that are dependent upon transcription of a gene to report the detection of a desired molecule and those that are independent of this reporting mechanism. Transcription-dependent biosensors frequently depend on heterologous expression of sensing elements from non-yeast organisms, a strategy that has greatly expanded the range of molecules available for detection by YBBs. Transcription-independent biosensors circumvent the problem of sensing difficult-to-detect analytes by instead relying on yeast metabolism to generate easily detected molecules when the analyte is present. The use of yeast as the sensing element in biosensors has proven to be successful and continues to hold great promise for a variety of applications. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  8. Detection of Escherichia coli Shiga toxin-producing in viscera of animals bovine and chicken intended for human consumption

    Directory of Open Access Journals (Sweden)

    Zotta, Claudio Marcelo

    2016-05-01

    Full Text Available Escherichia coli producing-Shiga toxin (STEC is associated with foodborne illness (ETA. It can cause bloody diarrhea, hemorrhagic colitis, hemolytic uremic syndrome and thrombotic thrombocytopenic purpura. The aim of the study was to detect the presence of STEC in samples of organs (offal of bovine animals and chicken intended for human consumption. Between 2008-2009, 76 samples bovine entrails and 22 chicken viscera samples, were processed and underwent, as screening technique, the polymerase chain reaction (PCR for detection of multiple genes coding for the factors virulence: Shiga toxin (stx1, stx2 and rfbO157 gene coding for capsular O157 lipopolysaccharide LPS. Samples from bovine offal development showed 84.2% for coliform bacteria. These isolates showed no virulence factor that characterized as STEC or Escherichia coli O157. The chicken offal samples showed 95.5% of development for coliform bacteria, being negative for the presence of genes encoding the Shiga toxins 1 and 2 (stx1, stx2 and rfbO157 gene. While this work does not STEC was detected, the presence of coliform bacteria in the samples studied makes these foods should be considered as potentially hazardous to consume undercooked with the consequent possibility of filing ETA.

  9. Functional expression of a human GDP-L-fucose transporter in Escherichia coli.

    Science.gov (United States)

    Förster-Fromme, Karin; Schneider, Sarah; Sprenger, Georg A; Albermann, Christoph

    2017-02-01

    To investigate the translocation of nucleotide-activated sugars from the cytosol across a membrane into the endoplasmatic reticulum or the Golgi apparatus which is an important step in the synthesis of glycoproteins and glycolipids in eukaryotes. The heterologous expression of the recombinant and codon-adapted human GDP-L-fucose antiporter gene SLC35C1 (encoding an N-terminal OmpA-signal sequence) led to a functional transporter protein located in the cytoplasmic membrane of Escherichia coli. The in vitro transport was investigated using inverted membrane vesicles. SLC35C1 is an antiporter specific for GDP-L-fucose and depending on the concomitant reverse transport of GMP. The recombinant transporter FucT1 exhibited an activity for the transport of 3 H-GDP-L-fucose with a V max of 8 pmol/min mg with a K m of 4 µM. The functional expression of SLC35C1 in GDP-L-fucose overproducing E. coli led to the export of GDP-L-fucose to the culture supernatant. The export of GDP-L-fucose by E. coli provides the opportunity for the engineering of a periplasmatic fucosylation reaction in recombinant bacterial cells.

  10. Transport of Escherichia coli phage through saturated porous media considering managed aquifer recharge.

    Science.gov (United States)

    Zhang, Wenjing; Li, Shuo; Wang, Shuang; Lei, Liancheng; Yu, Xipeng; Ma, Tianyi

    2018-03-01

    Virus is one of the most potentially harmful microorganisms in groundwater. In this paper, the effects of hydrodynamic and hydrogeochemical conditions on the transportation of the colloidal virus considering managed aquifer recharge were systematically investigated. Escherichia coli phage, vB_EcoM-ep3, has a broad host range and was able to lyse pathogenic Escherichia coli. Bacteriophage with low risk to infect human has been found extensively in the groundwater environment, so it is considered as a representative model of groundwater viruses. Laboratory studies were carried out to analyze the transport of the Escherichia coli phage under varying conditions of pH, ionic strength, cation valence, flow rate, porous media, and phosphate buffer concentration. The results indicated that decreasing the pH will increase the adsorption of Escherichia coli phage. Increasing the ionic strength, either Na + or Ca 2+ , will form negative condition for the migration of Escherichia coli phage. A comparison of different cation valence tests indicated that changes in transport and deposition were more pronounced with divalent Ca 2+ than monovalent Na + . As the flow rate increases, the release of Escherichia coli phage increases and the retention of Escherichia coli phage in the aquifer medium reduces. Changes in porous media had a significant effect on Escherichia coli phage migration. With increase of phosphate buffer concentration, the suspension stability and migration ability of Escherichia coli phage are both increased. Based on laboratory-scale column experiments, a one-dimensional transport model was established to quantitatively describe the virus transport in saturated porous medium.

  11. Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E-coli (STEC) infections in the Netherlands, January 2008 to December 2011

    NARCIS (Netherlands)

    Friesema, I.; van der Zwaluw, K.; Schuurman, T.; Kooistra-Smid, M.; Franz, E.; van Duynhoven, Y.; van Pelt, W.

    2014-01-01

    The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx(2f) is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC

  12. Prevalence of candida and non-candida yeasts isolated from patients with yeast fungal infections in Tehran labs

    Directory of Open Access Journals (Sweden)

    Hashemi SJ

    2011-04-01

    Full Text Available "n 800x600 Normal 0 false false false EN-US X-NONE AR-SA MicrosoftInternetExplorer4 /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman","serif";} Background: Infections caused by opportunistic yeasts such as Candida species, Trichosporon, Rhodotorula and Saccharomyces have increased in immunocompromis-ed patients and their identification is crucial as intrinsic and acquired resistance of some yeast species to antifungal agents are on the rise. The aim of this study was to identify the organisms to the species level in order to suggest accurate and effective antifungal therapies."n"nMethods: In this study that carried out in Tehran, Iran in 2009, 200 patients with yeast infection were medically examined and clinical specimens were prepared for direct examination and culture on Sabouraud dextrose agar. Subsequently, the isolated yeast colonies were identified using various tests including culture on Corn Meal agar with Tween 80, CHROMagar Candida and casein agar. For the definite identification of organisms some biochemical tests were done based on carbohydrate assimilation by RapID Yeast Plus System kit, and, finally, a molecular method, PCR-RFLP, using Hpa II enzyme, was performed for the remaining unknown yeast species."n"nResults: A total of 211 yeast isolates were identified in 200 patients with yeast infections. The most frequent isolated yeasts were Candida albicans, 124 (58.77%, followed by Candida parapsilosis, 36 (17.06%, Candida tropicalis, 17 (8.06%, Candida glabrata, 13 (6.16%, Candida krusei, 8 (3.79%, Candida guilliermondii, 2 (0.96%, Trichosporon, 3 (1.14%, Rhodotorula, 1 (0.47%, Saccaromyces cerevisiae, 1 (0.47% and other

  13. Oral yeast colonization throughout pregnancy.

    Science.gov (United States)

    Rio, R; Simões-Silva, L; Garro, S; Silva, M-J; Azevedo, Á; Sampaio-Maia, B

    2017-03-01

    Recent studies suggest that placenta may harbour a unique microbiome that may have origin in maternal oral microbiome. Although the major physiological and hormonal adjustments observed in pregnant women lead to biochemical and microbiological modifications of the oral environment, very few studies evaluated the changes suffered by the oral microbiota throughout pregnancy. So, the aim of our study was to evaluate oral yeast colonization throughout pregnancy and to compare it with non-pregnant women. The oral yeast colonization was assessed in saliva of 30 pregnant and non-pregnant women longitudinally over a 6-months period. Demographic information was collected, a non-invasive intra-oral examination was performed and saliva flow and pH were determined. Pregnant and non-pregnant groups were similar regarding age and level of education. Saliva flow rate did not differ, but saliva pH was lower in pregnant than in non-pregnant women. Oral yeast prevalence was higher in pregnant than in non-pregnant women, either in the first or in the third trimester, but did not attain statistical significance. In individuals colonized with yeast, the total yeast quantification (Log10CFU/mL) increase from the 1st to the 3rd trimester in pregnant women, but not in non-pregnant women. Pregnancy may favour oral yeast growth that may be associated with an acidic oral environment.

  14. Biotechnological Applications of Dimorphic Yeasts

    Science.gov (United States)

    Doiphode, N.; Joshi, C.; Ghormade, V.; Deshpande, M. V.

    The dimorphic yeasts have the equilibrium between spherical growth (budding) and polarized (hyphal or pseudohyphal tip elongation) which can be triggered by change in the environmental conditions. The reversible growth phenomenon has made dimorphic yeasts as an useful model to understand fungal evolution and fungal differentiation, in general. In nature dimorphism is clearly evident in plant and animal fungal pathogens, which survive and most importantly proliferate in the respective hosts. However, number of organisms with no known pathogenic behaviour also show such a transition, which can be exploited for the technological applications due to their different biochemical make up under different morphologies. For instance, chitin and chitosan production using dimorphic Saccharomyces, Mucor, Rhizopus and Benjaminiella, oil degradation and biotransformation with yeast-form of Yarrowia species, bioremediation of organic pollutants, exopolysac-charide production by yeast-phase of Aureobasidium pullulans, to name a few. Myrothecium verrucaria can be used for seed dressing in its yeast form and it produces a mycolytic enzyme complex in its hyphal-form for the biocontrol of fungal pathogens, while Beauveria bassiana and other entomopathogens kill the insect pest by producing yeast- like cells in the insect body. The form-specific expression of protease, chitinase, lipase, ornithine decarboxylase, glutamate dehydrogenases, etc. make Benjaminiella poitrasii, Basidiobolus sp., and Mucor rouxii strains important in bioremediation, nanobiotechnology, fungal evolution and other areas.

  15. Identification of salivary components that induce transition of hyphae to yeast in Candida albicans.

    Science.gov (United States)

    Leito, Jelani T D; Ligtenberg, Antoon J M; Nazmi, Kamran; Veerman, Enno C I

    2009-10-01

    Candida albicans, the major human fungal pathogen, undergoes a reversible morphological transition from single yeast cells to pseudohyphae and hyphae filaments. The hyphae form is considered the most invasive form of the fungus. The purpose of this study is to investigate the effect of saliva on hyphae growth of C. albicans. Candida albicans hyphae were inoculated in Roswell Park Memorial Institute medium with whole saliva, parotid saliva or buffer mimicking the saliva ion composition, and cultured for 18 h at 37 degrees C under aerobic conditions with 5% CO(2). Whole saliva and parotid saliva induced transition to yeast growth, whereas the culture with buffer remained in the hyphae form. Parotid saliva was fractionated on a reverse-phase C8 column and each fraction was tested for inducing transition to yeast growth. By immunoblotting, the salivary component in the active fraction was identified as statherin, a phosphoprotein of 43 amino acids that has been implicated in remineralization of the teeth. Synthetically made statherin induced transition of hyphae to yeast. By deletion of five amino acids at the negatively charged N-terminal site (DpSpSEE), yeast-inducing activity and binding to C. albicans were increased. In conclusion, statherin induces transition to yeast of C. albicans hyphae and may thus contribute to the oral defense against candidiasis.

  16. The global transcriptional response of fission yeast to hydrogen sulfide.

    Directory of Open Access Journals (Sweden)

    Xu Jia

    Full Text Available BACKGROUND: Hydrogen sulfide (H(2S is a newly identified member of the small family of gasotransmitters that are endogenous gaseous signaling molecules that have a fundamental role in human biology and disease. Although it is a relatively recent discovery and the mechanism of H(2S activity is not completely understood, it is known to be involved in a number of cellular processes; H(2S can affect ion channels, transcription factors and protein kinases in mammals. METHODOLOGY/PRINCIPAL FINDINGS: In this paper, we have used fission yeast as a model organism to study the global gene expression profile in response to H(2S by microarray. We initially measured the genome-wide transcriptional response of fission yeast to H(2S. Through the functional classification of genes whose expression profile changed in response to H(2S, we found that H(2S mainly influences genes that encode putative or known stress proteins, membrane transporters, cell cycle/meiotic proteins, transcription factors and respiration protein in the mitochondrion. Our analysis showed that there was a significant overlap between the genes affected by H(2S and the stress response. We identified that the target genes of the MAPK pathway respond to H(2S; we also identified that a number of transporters respond to H(2S, these include sugar/carbohydrate transporters, ion transporters, and amino acid transporters. We found many mitochondrial genes to be down regulated upon H(2S treatment and that H(2S can reduce mitochondrial oxygen consumption. CONCLUSION/SIGNIFICANCE: This study identifies potential molecular targets of the signaling molecule H(2S in fission yeast and provides clues about the identity of homologues human proteins and will further the understanding of the cellular role of H(2S in human diseases.

  17. Electron transport chain in a thermotolerant yeast.

    Science.gov (United States)

    Mejía-Barajas, Jorge A; Martínez-Mora, José A; Salgado-Garciglia, Rafael; Noriega-Cisneros, Ruth; Ortiz-Avila, Omar; Cortés-Rojo, Christian; Saavedra-Molina, Alfredo

    2017-04-01

    Yeasts capable of growing and surviving at high temperatures are regarded as thermotolerant. For appropriate functioning of cellular processes and cell survival, the maintenance of an optimal redox state is critical of reducing and oxidizing species. We studied mitochondrial functions of the thermotolerant Kluyveromyces marxianus SLP1 and the mesophilic OFF1 yeasts, through the evaluation of its mitochondrial membrane potential (ΔΨ m ), ATPase activity, electron transport chain (ETC) activities, alternative oxidase activity, lipid peroxidation. Mitochondrial membrane potential and the cytoplasmic free Ca 2+ ions (Ca 2+ cyt) increased in the SLP1 yeast when exposed to high temperature, compared with the mesophilic yeast OFF1. ATPase activity in the mesophilic yeast diminished 80% when exposed to 40° while the thermotolerant SLP1 showed no change, despite an increase in the mitochondrial lipid peroxidation. The SLP1 thermotolerant yeast exposed to high temperature showed a diminution of 33% of the oxygen consumption in state 4. The uncoupled state 3 of oxygen consumption did not change in the mesophilic yeast when it had an increase of temperature, whereas in the thermotolerant SLP1 yeast resulted in an increase of 2.5 times when yeast were grown at 30 o , while a decrease of 51% was observed when it was exposed to high temperature. The activities of the ETC complexes were diminished in the SLP1 when exposed to high temperature, but also it was distinguished an alternative oxidase activity. Our results suggest that the mitochondria state, particularly ETC state, is an important characteristic of the thermotolerance of the SLP1 yeast strain.

  18. Purification and characterization of the d-xylose isomerase gene from Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Ho, N W.Y.; Rosenfeld, S; Stevis, P; Tsao, G T

    1983-11-01

    A DNA fragment containing both the Escherichia coli D-xylose isomerase (D-xylose ketol-isomerase, EC 5.3.1.5) gene and the D-xylulokinase (ATP: D-xylulose 5-phosphotransferase, EC 2.7.1.17) gene has been cloned on an E. coli plasmid. The D-xylose isomerase gene was separated from the D-xylulokinase gene by the construction of a new deletion plasmid, pLX7. The D-xylose isomerase gene cloned on pLX7 was found still to be an intact gene. The precise location of the D-xylose isomerase gene on the plasmid pLX7 was further determined by the construction of two more plasmids, pLX8 and pLX9. This is believed to be the first D-xylose isomerase gene that has been isolated and extensively purified from any organism. D-Xylose isomerase, the enzyme product of the D-xylose isomerase gene, is responsible for the conversion of D-xylose to D-xylulose, as well as D-glucose to D-fructose. It is widely believed that yeast cannot ferment D-xylose to ethanol primarily because of the lack of D-xylose isomerase in yeast. D-Xylose isomerase (also known as D-glucose isomerase) is also used for the commercial production of high-fructose syrups. The purification of the D-xylose isomerase gene may lead to the following industrial applications: (1) cloning and expression of the gene in yeast to make the latter organism capable of directly fermenting D-xylose to ethanol, and (2) cloning of the gene on a high-copy-number plasmid in a proper host to overproduce the enzyme, which should have a profound impact on the high-fructose syrup technology. 14 references.

  19. Transmission of antibiotic-resistant Escherichia coli between cattle, humans and the environment in peri-urban livestock keeping communities in Morogoro, Tanzania

    DEFF Research Database (Denmark)

    Lupindu, Athumani M; Dalsgaard, Anders; Msoffe, Peter L. M.

    2015-01-01

    Urban and peri-urban livestock farming is expanding world-widely because of increased urbanization and demands for food of animal origin. Such farming practices pose a public health risk as livestock are reservoirs of several zoonotic pathogens. In an attempt to determine the fecal transmission...... infrastructures (Odd Ratio=11.2, 95% CI=1.1-119.3) were associated with E. coli showing identical PFGE types within and between clusters. There is a need to improve animal husbandry and manure management practices to reduce risks of transmission of enteropathogens between livestock and humans in urban and peri-urban...... between livestock and people, 100 household clusters keeping cattle in close proximity of humans were selected in urban and peri-urban areas of Morogoro in Tanzania. One hundred eighteen ampicillin and tetracycline resistant Escherichia coli (40 from human stool, 50 from cattle feces, 21 from soil...

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

    Science.gov (United States)

    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.

  1. Mutational analysis of Escherichia coli elongation factor Tu in search of a role for the N-terminal region

    DEFF Research Database (Denmark)

    Mansilla, Francisco; Knudsen, Charlotte Rohde; Laurberg, M

    1998-01-01

    We have mutated lysine 2 and arginine 7 in elongation factor Tu from Escherichia coli separately either to alanine or glutamic acid. The aim of the work was to reveal the possible interactions between the conserved N-terminal part of the molecule, which is rich in basic residues and aminoacyl...... this activity. Furthermore, arginine 7 seems to play a role in regulating the binding of GTP. The three-dimensional structure of the ternary complex, EF-Tu:GTP:Phe-tRNAPhe, involving Thermus aquaticus EF-Tu and yeast Phe-tRNA(Phe), shows that Arg7 is in a position which permits salt bridge formation with Asp284...

  2. Interactions between Drosophila and its natural yeast symbionts-Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

    Science.gov (United States)

    Hoang, Don; Kopp, Artyom; Chandler, James Angus

    2015-01-01

    Yeasts play an important role in the biology of the fruit fly, Drosophila melanogaster. In addition to being a valuable source of nutrition, yeasts affect D. melanogaster behavior and interact with the host immune system. Most experiments investigating the role of yeasts in D. melanogaster biology use the baker's yeast, Saccharomyces cerevisiae. However, S. cerevisiae is rarely found with natural populations of D. melanogaster or other Drosophila species. Moreover, the strain of S. cerevisiae used most often in D. melanogaster experiments is a commercially and industrially important strain that, to the best of our knowledge, was not isolated from flies. Since disrupting natural host-microbe interactions can have profound effects on host biology, the results from D. melanogaster-S. cerevisiae laboratory experiments may not be fully representative of host-microbe interactions in nature. In this study, we explore the D. melanogaster-yeast relationship using five different strains of yeast that were isolated from wild Drosophila populations. Ingested live yeasts have variable persistence in the D. melanogaster gastrointestinal tract. For example, Hanseniaspora occidentalis persists relative to S. cerevisiae, while Brettanomyces naardenensis is removed. Despite these differences in persistence relative to S. cerevisiae, we find that all yeasts decrease in total abundance over time. Reactive oxygen species (ROS) are an important component of the D. melanogaster anti-microbial response and can inhibit S. cerevisiae growth in the intestine. To determine if sensitivity to ROS explains the differences in yeast persistence, we measured yeast growth in the presence and absence of hydrogen peroxide. We find that B. naardenesis is completely inhibited by hydrogen peroxide, while H. occidentalis is not, which is consistent with yeast sensitivity to ROS affecting persistence within the D. melanogaster gastrointestinal tract. We also compared the feeding preference of D

  3. Source and role of diacylglycerol formed during phagocytosis of opsonized yeast particles and associated respiratory burst in human neutrophils

    International Nuclear Information System (INIS)

    Della Bianca, V.; Grzeskowiak, M.; Lissandrini, D.; Rossi, F.

    1991-01-01

    The results presented in this paper demonstrate that in human neutrophils phagocytosis of C3b/bi and IgG-opsonized yeast particles is associated with activation of phospholipase D and that this reaction is the main source of diglycerides. The demonstration is based upon the following findings: (1) the challenge of neutrophils with these opsonized particles was followed by a rapid formation of [3H]alkyl-phosphatidic acid [( 3H]alkyl-PA) and [3H]alkyl-diglyceride [( 3H]alkyl-DG) in cells labeled with [3H]alkyl-lyso-phosphatidylcholine; (2) in the presence of ethanol [3H]alkyl-phosphatidylethanol was formed, and accumulation of [3H]alkyl-PA and [3H]alkyl-DG was depressed; (3) propranolol, by inhibiting the dephosphorylation of [3H]alkyl-PA, completely inhibited the accumulation of [3H]alkyl-DG and depressed by about 75% the formation of diglyceride mass. Evidence is also presented that phagocytosis of C3b/bi and IgG-opsonized yeast particles and associated respiratory burst can take place independently of diglyceride formation and of the activity of this second messenger on protein kinase C. In fact: (a) propranolol while completely inhibited the formation of diglyceride mass did not modify either the phagocytosis or respiratory burst; (b) these two processes were insensitive to staurosporine

  4. Rapid customised operon assembly by yeast recombinational cloning.

    Science.gov (United States)

    Liu, Michael A; Kenyon, Johanna J; Lee, Jason; Reeves, Peter R

    2017-06-01

    We have developed a system called the Operon Assembly Protocol (OAP), which takes advantage of the homologous recombination DNA repair pathway in Saccharomyces cerevisiae to assemble full-length operons from a series of overlapping PCR products into a specially engineered yeast-Escherichia coli shuttle vector. This flexible, streamlined system can be used to assemble several operon clones simultaneously, and each clone can be expressed in the same E. coli tester strain to facilitate direct functional comparisons. We demonstrated the utility of the OAP by assembling and expressing a series of E. coli O1A O-antigen gene cluster clones containing various gene deletions or replacements. We then used these constructs to assess the substrate preferences of several Wzx flippases, which are responsible for translocation of oligosaccharide repeat units (O units) across the inner membrane during O-antigen biosynthesis. We were able to identify several O unit structural features that appear to be important determinants of Wzx substrate preference. The OAP system should be broadly applicable for the genetic manipulation of any bacterial operon and can be modified for use in other host species. It could also have potential uses in fields such as glycoengineering.

  5. The Budding Yeast “Saccharomyces cerevisiae” as a Drug Discovery Tool to Identify Plant-Derived Natural Products with Anti-Proliferative Properties

    Directory of Open Access Journals (Sweden)

    Bouchra Qaddouri

    2011-01-01

    Full Text Available The budding yeast Saccharomyces cerevisiae is a valuable system to study cell-cycle regulation, which is defective in cancer cells. Due to the highly conserved nature of the cell-cycle machinery between yeast and humans, yeast studies are directly relevant to anticancer-drug discovery. The budding yeast is also an excellent model system for identifying and studying antifungal compounds because of the functional conservation of fungal genes. Moreover, yeast studies have also contributed greatly to our understanding of the biological targets and modes of action of bioactive compounds. Understanding the mechanism of action of clinically relevant compounds is essential for the design of improved second-generation molecules. Here we describe our methodology for screening a library of plant-derived natural products in yeast in order to identify and characterize new compounds with anti-proliferative properties.

  6. Full Data of Yeast Interacting Proteins Database (Original Version) - Yeast Interacting Proteins Database | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us Yeast Interacting Proteins Database Full Data of Yeast Interacting Proteins Database (Origin...al Version) Data detail Data name Full Data of Yeast Interacting Proteins Database (Original Version) DOI 10....18908/lsdba.nbdc00742-004 Description of data contents The entire data in the Yeast Interacting Proteins Database...eir interactions are required. Several sources including YPD (Yeast Proteome Database, Costanzo, M. C., Hoga...ematic name in the SGD (Saccharomyces Genome Database; http://www.yeastgenome.org /). Bait gene name The gen

  7. Genomics and the making of yeast biodiversity

    NARCIS (Netherlands)

    Hittinger, Chris Todd; Rokas, Antonis; Bai, Feng-Yan; Boekhout, Teun; Gonçalves, Paula; Jeffries, Thomas W; Kominek, Jacek; Lachance, Marc-André; Libkind, Diego; Rosa, Carlos A; Sampaio, José Paulo; Kurtzman, Cletus P

    2015-01-01

    Yeasts are unicellular fungi that do not form fruiting bodies. Although the yeast lifestyle has evolved multiple times, most known species belong to the subphylum Saccharomycotina (syn. Hemiascomycota, hereafter yeasts). This diverse group includes the premier eukaryotic model system, Saccharomyces

  8. A novel yeast cell-based screen identifies flavone as a tankyrase inhibitor

    International Nuclear Information System (INIS)

    Yashiroda, Yoko; Okamoto, Reika; Hatsugai, Kaori; Takemoto, Yasushi; Goshima, Naoki; Saito, Tamio; Hamamoto, Makiko; Sugimoto, Yoshikazu; Osada, Hiroyuki; Seimiya, Hiroyuki; Yoshida, Minoru

    2010-01-01

    The telomere-associated protein tankyrase 1 is a poly(ADP-ribose) polymerase and is considered to be a promising target for cancer therapy, especially for BRCA-associated cancers. However, an efficient assay system for inhibitor screening has not been established, mainly due to the difficulty of efficient preparation of the enzyme and its substrate. Here, we report a cell-based assay system for detecting inhibitory activity against tankyrase 1. We found that overexpression of the human tankyrase 1 gene causes a growth defect in the fission yeast Schizosaccharomyces pombe. Chemicals that restore the growth defect phenotype can be identified as potential tankyrase 1 inhibitors. We performed a high-throughput screen using this system, and identified flavone as a compound that restores the growth of yeast cells overexpressing tankyrase 1. Indeed, flavone inhibited poly(ADP-ribosyl)ation of proteins caused by overexpression of tankyrase 1 in yeast cells. This system allows rapid identification of inhibitory activity against tankyrase 1 and is amenable to high-throughput screening using robotics.

  9. Yeasts preservation: alternatives for lyophilisation

    OpenAIRE

    Nyanga, Loveness K.; Nout, Martinus J. R.; Smid, Eddy J.; Boekhout, Teun; Zwietering, Marcel H.

    2012-01-01

    The aim of the study was to compare the effect of two low-cost, low technology traditional methods for drying starter cultures with standard lyophilisation. Lyophilised yeast cultures and yeast cultures preserved in dry rice cakes and dry plant fibre strands were examined for viable cell counts during 6 months storage at 4 and 25 °C. None of the yeast cultures showed a significant loss in viable cell count during 6 months of storage at 4 °C upon lyophilisation and preservation in dry rice cak...

  10. Review of the recombinant human interferon gamma as an immunotherapeutic: Impacts of production platforms and glycosylation.

    Science.gov (United States)

    Razaghi, Ali; Owens, Leigh; Heimann, Kirsten

    2016-12-20

    Human interferon gamma is a cytokine belonging to a diverse group of interferons which have a crucial immunological function against mycobacteria and a wide variety of viral infections. To date, it has been approved for treatment of chronic granulomatous disease and malignant osteopetrosis, and its application as an immunotherapeutic agent against cancer is an increasing prospect. Recombinant human interferon gamma, as a lucrative biopharmaceutical, has been engineered in different expression systems including prokaryotic, protozoan, fungal (yeasts), plant, insect and mammalian cells. Human interferon gamma is commonly expressed in Escherichia coli, marketed as ACTIMMUNE ® , however, the resulting product of the prokaryotic expression system is unglycosylated with a short half-life in the bloodstream; the purification process is tedious and makes the product costlier. Other expression systems also did not show satisfactory results in terms of yields, the biological activity of the protein or economic viability. Thus, the review aims to synthesise available information from previous studies on the production of human interferon gamma and its glycosylation patterns in different expression systems, to provide direction to future research in this field. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption.

    Directory of Open Access Journals (Sweden)

    Ying Luo

    Full Text Available The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry.

  12. The in vitro synergistic inhibitory effect of human amniotic fluid and gentamicin on growth of Escherichia coli.

    Science.gov (United States)

    Miglioli, P A; Schoffel, U; Gianfranceschi, L

    1996-01-01

    The activity of serum and its synergistic effect with many antibiotics against bacteria are well known. Few reports are available on similar phenomena produced by human amniotic fluid (HAF). Thus we investigated the antibacterial activity of HAF and the presence of a synergistic effect with gentamicin (GM) against Escherichia coli strains. Antimicrobial activity was evaluated as a delay of the growth curve, using a turbidimetric method. E. coli ATCC 10798 and E. coli SC 12155 were employed as test micro-organisms in nutrient broth, and GM was used at a subinhibitory concentration. HAF exerted antibacterial activity and, cooperating with GM at subinhibitory concentration, enhanced its antibiotic activity against E. coli. The presence of Schlievert's glycoprotein in HAF could explain these results.

  13. Immunological Effects of Probiotics and their Significance to Human Health

    Science.gov (United States)

    Gill, Harsharn S.; Grover, Sunita; Batish, Virender K.; Gill, Preet

    Probiotics are defined as live microorganisms which when administered in adequate amounts confer a health benefit upon the host (FAO/WHO, 2001). Lactic acid bacteria, particularly Lactobacillus and Bifidobacterium species are commonly used as probiotics. Other less commonly used probiotics include the yeast Sacchromyces cerevisiae and some non-pathogenic Escherichia coli and Bacillus species. Studies over the past 20 years have demonstrated that probiotic intake is able to confer a range of health benefits including modulation of the immune system, protection against gastrointestinal and respiratory tract infections, lowering of blood cholesterol levels, attenuation of overt immuno-inflammatory disorders (such as inflammatory bowel disease, allergies) and anti-cancer effects. However, the strongest clinical evidence for probiotics relates to their effectiveness in improving gut health and modulating (via stimulation or regulation) the host immune system. This chapter provides an overview of the current status of our knowledge regarding the immunostimulatory and immunoregulatory effects of probiotics on the immune system and their significance to human health.

  14. Yeasts Diversity in Fermented Foods and Beverages

    Science.gov (United States)

    Tamang, Jyoti Prakash; Fleet, Graham H.

    People across the world have learnt to culture and use the essential microorganisms for production of fermented foods and alcoholic beverages. A fermented food is produced either spontaneously or by adding mixed/pure starter culture(s). Yeasts are among the essential functional microorganisms encountered in many fermented foods, and are commercially used in production of baker's yeast, breads, wine, beer, cheese, etc. In Asia, moulds are predominant followed by amylolytic and alcohol-producing yeasts in the fermentation processes, whereas in Africa, Europe, Australia and America, fermented products are prepared exclusively using bacteria or bacteria-yeasts mixed cultures. This chapter would focus on the varieties of fermented foods and alcoholic beverages produced by yeasts, their microbiology and role in food fermentation, widely used commercial starters (pilot production, molecular aspects), production technology of some common commercial fermented foods and alcoholic beverages, toxicity and food safety using yeasts cultures and socio-economy

  15. The ferric yersiniabactin uptake receptor FyuA is required for efficient biofilm formation by urinary tract infectious Escherichia coli in human urine

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Ferrieres, Lionel; Klemm, Per

    2008-01-01

    Urinary tract infection (UTI) is the most common infection in patients with indwelling urinary catheters, and bacterial biofilm formation is a major problem in this type of infection. Escherichia coli is responsible for the large majority of UTIs. Free iron is strictly limited in the human urinary...... of the most upregulated genes in biofilm; it was upregulated 63-fold in the E coli UTI strain VR50. FyuA was found to be highly important for biofilm formation in iron-poor environments such as human urine. Mutants in fyuA show aberrant biofilm formation and the cells become filamentous; a VR50fyuA mutant...... of iron greatly influences UTI strains' ability to form biofilm....

  16. Implication of Ca2+ in the regulation of replicative life span of budding yeast.

    Science.gov (United States)

    Tsubakiyama, Ryohei; Mizunuma, Masaki; Gengyo, Anri; Yamamoto, Josuke; Kume, Kazunori; Miyakawa, Tokichi; Hirata, Dai

    2011-08-19

    In eukaryotic cells, Ca(2+)-triggered signaling pathways are used to regulate a wide variety of cellular processes. Calcineurin, a highly conserved Ca(2+)/calmodulin-dependent protein phosphatase, plays key roles in the regulation of diverse biological processes in organisms ranging from yeast to humans. We isolated a mutant of the SIR3 gene, implicated in the regulation of life span, as a suppressor of the Ca(2+) sensitivity of zds1Δ cells in the budding yeast Saccharomyces cerevisiae. Therefore, we investigated a relationship between Ca(2+) signaling and life span in yeast. Here we show that Ca(2+) affected the replicative life span (RLS) of yeast. Increased external and intracellular Ca(2+) levels caused a reduction in their RLS. Consistently, the increase in calcineurin activity by either the zds1 deletion or the constitutively activated calcineurin reduced RLS. Indeed, the shortened RLS of zds1Δ cells was suppressed by the calcineurin deletion. Further, the calcineurin deletion per se promoted aging without impairing the gene silencing typically observed in short-lived sir mutants, indicating that calcineurin plays an important role in a regulation of RLS even under normal growth condition. Thus, our results indicate that Ca(2+) homeostasis/Ca(2+) signaling are required to regulate longevity in budding yeast.

  17. Expression of death receptor 4 induces caspase-independent cell death in MMS-treated yeast.

    Science.gov (United States)

    Kang, Mi-Sun; Lee, Sung-Keun; Park, Chang-Shin; Kang, Ju-Hee; Bae, Sung-Ho; Yu, Sung-Lim

    2008-11-14

    DR4, a tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, is a key element in the extrinsic pathway of TRAIL/TRAIL receptor-related apoptosis that exerts a preferential toxic effect against tumor cells. However, TRAIL and DR4 are expressed in various normal cells, and recent studies indicate that DR4 has a number of non-apoptotic functions. In this study, we evaluated the effects of human DR4 expression in yeast to determine the function of DR4 in normal cells. The expression of DR4 in yeast caused G1 arrest, which resulted in transient growth inhibition. Moreover, treatment of DR4-expressing yeast with a DNA damaging agent, MMS, elicited drastic, and sustained cell growth inhibition accompanied with massive apoptotic cell death. Further analysis revealed that cell death in the presence of DNA damage and DR4 expression was not dependent on the yeast caspase, YCA1. Taken together, these results indicate that DR4 triggers caspase-independent programmed cell death during the response of normal cells to DNA damage.

  18. Unscheduled DNA synthesis in xeroderma pigmentosum cells after microinjection of yeast photoreactivating enzyme.

    NARCIS (Netherlands)

    J.C.M. Zwetsloot; J.H.J. Hoeijmakers (Jan); W. Vermeulen (Wim); A.P.M. Eker (André); D. Bootsma (Dirk)

    1986-01-01

    textabstractPhotoreactivating enzyme (PRE) from yeast causes a light-dependent reduction of UV-induced unscheduled DNA synthesis (UDS) when injected into the cytoplasm of repair-proficieint human fibroblasts (Zwetsloot et al., 1985). This result indicates that the exogenous PRE monomerizers

  19. Genetics of Yeasts

    Science.gov (United States)

    Querol, Amparo; Fernández-Espinar, M. Teresa; Belloch, Carmela

    The use of yeasts in biotechnology processes dates back to ancient days. Before 7000 BC, beer was produced in Sumeria. Wine was made in Assyria in 3500 BC, and ancient Rome had over 250 bakeries, which were making leavened bread by 100 BC. And milk has been made into Kefyr and Koumiss in Asia for many centuries (Demain, Phaff, & Kurtzman, 1999). However, the importance of yeast in the food and beverage industries was only realized about 1860, when their role in food manufacturing became evident.

  20. Inheritance of the yeast mitochondrial genome

    DEFF Research Database (Denmark)

    Piskur, Jure

    1994-01-01

    Mitochondrion, extrachromosomal genetics, intergenic sequences, genome size, mitochondrial DNA, petite mutation, yeast......Mitochondrion, extrachromosomal genetics, intergenic sequences, genome size, mitochondrial DNA, petite mutation, yeast...

  1. 21 CFR 172.590 - Yeast-malt sprout extract.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Yeast-malt sprout extract. 172.590 Section 172.590... CONSUMPTION Flavoring Agents and Related Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout... prescribed conditions: (a) The additive is produced by partial hydrolysis of yeast extract (derived from...

  2. Yeasts in sustainable bioethanol production: A review.

    Science.gov (United States)

    Mohd Azhar, Siti Hajar; Abdulla, Rahmath; Jambo, Siti Azmah; Marbawi, Hartinie; Gansau, Jualang Azlan; Mohd Faik, Ainol Azifa; Rodrigues, Kenneth Francis

    2017-07-01

    Bioethanol has been identified as the mostly used biofuel worldwide since it significantly contributes to the reduction of crude oil consumption and environmental pollution. It can be produced from various types of feedstocks such as sucrose, starch, lignocellulosic and algal biomass through fermentation process by microorganisms. Compared to other types of microoganisms, yeasts especially Saccharomyces cerevisiae is the common microbes employed in ethanol production due to its high ethanol productivity, high ethanol tolerance and ability of fermenting wide range of sugars. However, there are some challenges in yeast fermentation which inhibit ethanol production such as high temperature, high ethanol concentration and the ability to ferment pentose sugars. Various types of yeast strains have been used in fermentation for ethanol production including hybrid, recombinant and wild-type yeasts. Yeasts can directly ferment simple sugars into ethanol while other type of feedstocks must be converted to fermentable sugars before it can be fermented to ethanol. The common processes involves in ethanol production are pretreatment, hydrolysis and fermentation. Production of bioethanol during fermentation depends on several factors such as temperature, sugar concentration, pH, fermentation time, agitation rate, and inoculum size. The efficiency and productivity of ethanol can be enhanced by immobilizing the yeast cells. This review highlights the different types of yeast strains, fermentation process, factors affecting bioethanol production and immobilization of yeasts for better bioethanol production.

  3. Yeasts in sustainable bioethanol production: A review

    Directory of Open Access Journals (Sweden)

    Siti Hajar Mohd Azhar

    2017-07-01

    Full Text Available Bioethanol has been identified as the mostly used biofuel worldwide since it significantly contributes to the reduction of crude oil consumption and environmental pollution. It can be produced from various types of feedstocks such as sucrose, starch, lignocellulosic and algal biomass through fermentation process by microorganisms. Compared to other types of microoganisms, yeasts especially Saccharomyces cerevisiae is the common microbes employed in ethanol production due to its high ethanol productivity, high ethanol tolerance and ability of fermenting wide range of sugars. However, there are some challenges in yeast fermentation which inhibit ethanol production such as high temperature, high ethanol concentration and the ability to ferment pentose sugars. Various types of yeast strains have been used in fermentation for ethanol production including hybrid, recombinant and wild-type yeasts. Yeasts can directly ferment simple sugars into ethanol while other type of feedstocks must be converted to fermentable sugars before it can be fermented to ethanol. The common processes involves in ethanol production are pretreatment, hydrolysis and fermentation. Production of bioethanol during fermentation depends on several factors such as temperature, sugar concentration, pH, fermentation time, agitation rate, and inoculum size. The efficiency and productivity of ethanol can be enhanced by immobilizing the yeast cells. This review highlights the different types of yeast strains, fermentation process, factors affecting bioethanol production and immobilization of yeasts for better bioethanol production.

  4. Comparison of extended-spectrum-β-lactamase (ESBL) carrying Escherichia coli from sewage sludge and human urinary tract infection

    International Nuclear Information System (INIS)

    Zarfel, G.; Galler, H.; Feierl, G.; Haas, D.; Kittinger, C.; Leitner, E.; Grisold, A.J.; Mascher, F.; Posch, J.; Pertschy, B.; Marth, E.; Reinthaler, F.F.

    2013-01-01

    For many years, extended-spectrum-beta-lactamase (ESBL) producing bacteria were a problem mainly located in medical facilities. Within the last decade however, ESBL-producing bacteria have started spreading into the community and the environment. In this study, ESBL-producing Escherichia coli from sewage sludge were collected, analysed and compared to ESBL-E. coli from human urinary tract infections (UTIs). The dominant ESBL-gene-family in both sample groups was bla CTX-M , which is the most prevalent ESBL-gene-family in human infection. Still, the distribution of ESBL genes and the frequency of additional antibiotic resistances differed in the two sample sets. Nevertheless, phenotyping did not divide isolates of the two sources into separate groups, suggesting similar strains in both sample sets. We speculate that an exchange is taking place between the ESBL E. coli populations in infected humans and sewage sludge, most likely by the entry of ESBL E. coli from UTIs into the sewage system. - Highlights: ► ESBL E. coli strains from sewage sludge harbour the same dominant ESBL enzymes as human isolates. ► High resistance rates for important antibiotics can be found in isolated ESBL strains. ► High phenotypic diversity of ESBL E. coli isolates from sewage sludge and from human sources. - The distribution of ESBL resistance genes in isolates from patients and environmental samples.

  5. YNK1, the yeast homolog of human metastasis suppressor NM23, is required for repair of UV radiation- and etoposide-induced DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Yang Mengmeng; Jarrett, Stuart G.; Craven, Rolf [Department of Molecular and Biomedical Pharmacology, College of Medicine, University of Kentucky, Lexington, KY 40536-0298 (United States); Kaetzel, David M. [Department of Molecular and Biomedical Pharmacology, College of Medicine, University of Kentucky, Lexington, KY 40536-0298 (United States)], E-mail: dmkaetz@uky.edu

    2009-01-15

    In humans, NM23-H1 is a metastasis suppressor whose expression is reduced in metastatic melanoma and breast carcinoma cells, and which possesses the ability to inhibit metastatic growth without significant impact on the transformed phenotype. NM23-H1 exhibits three enzymatic activities in vitro, each with potential to maintain genomic stability, a 3'-5' exonuclease and two kinases, nucleoside diphosphate kinase (NDPK), and protein histidine kinase. Herein we have investigated the potential contributions of NM23 proteins to DNA repair in the yeast, Saccharomyces cerevisiae, which contains a single NM23 homolog, YNK1. Ablation of YNK1 delayed repair of UV- and etoposide-induced nuclear DNA damage by 3-6 h. However, YNK1 had no impact upon the kinetics of MMS-induced DNA repair. Furthermore, YNK1 was not required for repair of mitochondrial DNA damage. To determine whether the nuclear DNA repair deficit manifested as an increase in mutation frequency, the CAN1 forward assay was employed. An YNK1 deletion was associated with increased mutation rates following treatment with either UV (2.6x) or MMS (1.6x). Mutation spectral analysis further revealed significantly increased rates of base substitution and frameshift mutations following UV treatment in the ynk1{delta} strain. This study indicates a novel role for YNK1 in DNA repair in yeast, and suggests an anti-mutator function that may contribute to the metastasis suppressor function of NM23-H1 in humans.

  6. YNK1, the yeast homolog of human metastasis suppressor NM23, is required for repair of UV radiation- and etoposide-induced DNA damage

    International Nuclear Information System (INIS)

    Yang Mengmeng; Jarrett, Stuart G.; Craven, Rolf; Kaetzel, David M.

    2009-01-01

    In humans, NM23-H1 is a metastasis suppressor whose expression is reduced in metastatic melanoma and breast carcinoma cells, and which possesses the ability to inhibit metastatic growth without significant impact on the transformed phenotype. NM23-H1 exhibits three enzymatic activities in vitro, each with potential to maintain genomic stability, a 3'-5' exonuclease and two kinases, nucleoside diphosphate kinase (NDPK), and protein histidine kinase. Herein we have investigated the potential contributions of NM23 proteins to DNA repair in the yeast, Saccharomyces cerevisiae, which contains a single NM23 homolog, YNK1. Ablation of YNK1 delayed repair of UV- and etoposide-induced nuclear DNA damage by 3-6 h. However, YNK1 had no impact upon the kinetics of MMS-induced DNA repair. Furthermore, YNK1 was not required for repair of mitochondrial DNA damage. To determine whether the nuclear DNA repair deficit manifested as an increase in mutation frequency, the CAN1 forward assay was employed. An YNK1 deletion was associated with increased mutation rates following treatment with either UV (2.6x) or MMS (1.6x). Mutation spectral analysis further revealed significantly increased rates of base substitution and frameshift mutations following UV treatment in the ynk1Δ strain. This study indicates a novel role for YNK1 in DNA repair in yeast, and suggests an anti-mutator function that may contribute to the metastasis suppressor function of NM23-H1 in humans

  7. Gamma irradiation induced ultrastructural changes in Paracoccidioides brasiliensis yeast cells

    International Nuclear Information System (INIS)

    Demicheli, Marina C.; Andrade, Antero S.R.; Goes, Alfredo Miranda

    2007-01-01

    Paracoccidioides brasiliensis is a thermally dimorphic fungus agent of paracoccidioidomycosis, a deep-seated systemic infection of humans with high prevalence in Latin America. Up to the moment no vaccine has still been reported. Ionizing radiation can be used to attenuate pathogens for vaccine development and we have successfully attenuated yeast cells of P. brasiliensis by gamma irradiation. The aim of the present study was to examine at ultrastructural level the effects of gamma irradiation attenuation on the morphology of P. brasiliensis yeast cells. P. brasiliensis (strain Pb-18) cultures were irradiated with a dose of 6.5 kGy. The irradiated cells were examined by scanning and also transmission electron microscopy. When examined two hours after the irradiation by scanning electron microscopy the 6.5 kGy irradiated cells presented deep folds or were collapsed. These lesions were reversible since examined 48 hours after irradiation the yeast have recovered the usual morphology. The transmission electron microscopy showed that the irradiated cells plasma membrane and cell wall were intact and preserved. Remarkable changes were found in the nucleus that was frequently in a very electrodense form. A extensive DNA fragmentation was produced by the gamma irradiation treatment. (author)

  8. Mutant power: using mutant allele collections for yeast functional genomics.

    Science.gov (United States)

    Norman, Kaitlyn L; Kumar, Anuj

    2016-03-01

    The budding yeast has long served as a model eukaryote for the functional genomic analysis of highly conserved signaling pathways, cellular processes and mechanisms underlying human disease. The collection of reagents available for genomics in yeast is extensive, encompassing a growing diversity of mutant collections beyond gene deletion sets in the standard wild-type S288C genetic background. We review here three main types of mutant allele collections: transposon mutagen collections, essential gene collections and overexpression libraries. Each collection provides unique and identifiable alleles that can be utilized in genome-wide, high-throughput studies. These genomic reagents are particularly informative in identifying synthetic phenotypes and functions associated with essential genes, including those modeled most effectively in complex genetic backgrounds. Several examples of genomic studies in filamentous/pseudohyphal backgrounds are provided here to illustrate this point. Additionally, the limitations of each approach are examined. Collectively, these mutant allele collections in Saccharomyces cerevisiae and the related pathogenic yeast Candida albicans promise insights toward an advanced understanding of eukaryotic molecular and cellular biology. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  9. Optimizing the feeding operation of recombinant Escherichia coli ...

    African Journals Online (AJOL)

    Recombinant Escherichia coli BL21 was used to produce human-like collagen in fed-batch culture. After building and analyzing the kinetic models of fed-batch cultures, the maximum specific growth rate, Yx/s and Yp/s were 0.411 h-1 , 0.428 g·g-1 and 0.0716 g/g, respectively. The square error of cell growth models, glucose ...

  10. Comet assay on tetraploid yeast cells

    DEFF Research Database (Denmark)

    Rank, Jette; Syberg, Kristian; Jensen, Klara

    2009-01-01

    Tetraploid yeast cells (Saccharomyces cerevisiae) were used in the comet assay with the intention of developing a new, fast and easy assay for detecting environmental genotoxic agents without using higher organisms. Two DNA-damaging chemicals, H2O2 and acrylamide, together with wastewater from...... three municipal treatment plants were tested for their effect on the yeast-cell DNA. The main problem with using yeast in the comet assay is the necessity to degrade the cell wall. This was achieved by using Zymolase 100 T twice during the procedure, since Zymolase 20 T did not open the cell wall....... Analytical problems that arose due to the small amount of DNA in the yeast nuclei in haploid and diploid cells, which contain 13 Mbp and 26 Mbp DNA per cell, respectively, were solved by using tetraploid yeast cells (52 Mbp) instead. DNA damage was shown after exposure to H2O2 and acrylamide. The lowest dose...

  11. Molecular characterization of the Escherichia coli asymptomatic bacteriuria strain 83972

    DEFF Research Database (Denmark)

    Klemm, Per; Hancock, Viktoria; Ulett, G.C.

    2006-01-01

    Escherichia coli 83972 is a clinical asymptomatia bacteriuric isolate that is able to colonize the human urinary bladder without inducing an immune response. Here we demonstrate that one of the mechanisms by which this strain has become attenuated is through the mutation of its genes encoding type...

  12. Microbial Quality and Antimicrobial Resistance of Staphylococcus aureus and Escherichia coli Isolated from Traditional Ice Cream in Hamadan City, West of Iran

    Directory of Open Access Journals (Sweden)

    Ghadimi

    2016-10-01

    Full Text Available Background Foodborne diseases are one of the most major public health concerns in the world. Ice cream flavors, especially the traditional ones, have a high potential for the transmission of the pathogenic bacteria. Objectives The aim of the current study is to investigate the microbiological status and antibiotic resistance of Escherichia coli and Staphylococcus aureus isolated from traditional ice cream. Methods A total of 114 traditional ice creams were randomly collected from retail stores in Hamadan, Iran. Samples were investigated for the total bacteria count (TBC and contamination with the coliform, Enterobacteriaceae and Salmonella as well as the prevalence and antibiotic resistance of Staphylococcus aureus and Escherichia coli. Results The count of Enterobacteriaceae (89.47%, mold and yeast (50%, coliform (40.35% and TBC (28.07% of samples was higher than Iran’s standard. Salmonella was not found in all samples. The prevalence of Staphylococcus aureus and Escherichia coli was confirmed in 50% and 37.72% of samples, respectively. Collected Escherichia coli had the highest antibiotic resistance to ampicillin 67.44%, nalidixic acid 39.53% and co-amoxyclav 37.21%. Staphylococcus aureus showed a higher antibiotic resistance to penicillin (82.46% of isolates and oxacillin (38% of isolates. Conclusions The results showed high contamination levels of traditional ice cream with spoilage and pathogenic microorganisms as well as considerable resistance of isolated Staphylococcus aureus and Escherichia coli to common antibiotics. Therefore, good hygienic practice during processing and personal hygiene should be considered to improve the quality of ice cream. In addition, it is necessary that the regulatory authorities carry out more control on the production centers of traditional ice cream.

  13. Revaluation of Waste Yeast from Beer Production

    Directory of Open Access Journals (Sweden)

    Nicoleta Suruceanu

    2013-11-01

    Full Text Available Brewing yeast is an important waste product from beer production. The valorification of slurry yeast mainly consists of separation of vitamins and important nitrogen compounds. The hops compounds, one of the most important raw materials in beer technology are removed beforehand valorification. The prenylflavonoids compounds from hops are important bioactive compounds that can be revaluation with proper technology. Revaluation of prenylflavonoids from waste yeast into dietary supplement, identification and quantification of xanthohumol by HPLC method. Waste yeast from brewery pilot plant of USAMV Cluj Napoca it was dried by atomization and the powder was analyzed on xanthohumol content by HPLC method. For quantification a calibration curve it was used. The process of drying by atomisation lead to a powder product. It was used malt dextrin powder for stabilisation. The final product it was encapsulated. The xanthohumol content of powdered yeast it was 1.94 µg/ml. In conclusion the slurry yeast from beer production it is an important source of prenylflavonoids compounds.

  14. Aboveground Deadwood Deposition Supports Development of Soil Yeasts

    Directory of Open Access Journals (Sweden)

    Thorsten Wehde

    2012-12-01

    Full Text Available Unicellular saprobic fungi (yeasts inhabit soils worldwide. Although yeast species typically occupy defined areas on the biome scale, their distribution patterns within a single type of vegetation, such as forests, are more complex. In order to understand factors that shape soil yeast communities, soils collected underneath decaying wood logs and under forest litter were analyzed. We isolated and identified molecularly a total of 25 yeast species, including three new species. Occurrence and distribution of yeasts isolated from these soils provide new insights into ecology and niche specialization of several soil-borne species. Although abundance of typical soil yeast species varied among experimental plots, the analysis of species abundance and community composition revealed a strong influence of wood log deposition and leakage of organic carbon. Unlike soils underneath logs, yeast communities in adjacent areas harbored a considerable number of transient (phylloplane-related yeasts reaching 30% of the total yeast quantity. We showed that distinguishing autochthonous community members and species transient in soils is essential to estimate appropriate effects of environmental factors on soil fungi. Furthermore, a better understanding of species niches is crucial for analyses of culture-independent data, and may hint to the discovery of unifying patterns of microbial species distribution.

  15. Ru(CO)3Cl(Glycinate) (CORM-3): A Carbon Monoxide–Releasing Molecule with Broad-Spectrum Antimicrobial and Photosensitive Activities Against Respiration and Cation Transport in Escherichia coli

    OpenAIRE

    Wilson, Jayne Louise; Jesse, Helen E.; Hughes, Bethan; Lund, Victoria; Naylor, Kathryn; Davidge, Kelly S.; Cook, Gregory M.; Mann, Brian E.; Poole, Robert K.

    2013-01-01

    Aims: Carbon monoxide (CO) delivered to cells and tissues by CO-releasing molecules (CO-RMs) has beneficial\\ud and toxic effects not mimicked by CO gas. The metal carbonyl Ru(CO)3Cl(glycinate) (CORM-3) is a novel, potent\\ud antimicrobial agent. Here, we established its mode of action. Results: CORM-3 inhibits respiration in several\\ud bacterial and yeast pathogens. In anoxic Escherichia coli suspensions, CORM-3 first stimulates, then inhibits\\ud respiration, but much higher concentrations of ...

  16. Development of a cultivation process for the enhancement of human interferon alpha 2b production in the oleaginous yeast, Yarrowia lipolytica

    Directory of Open Access Journals (Sweden)

    Gasmi Najla

    2011-11-01

    Full Text Available Abstract Background As an oleaginous yeast, Yarrowia lipolytica is able to assimilate hydrophobic substrates. This led to the isolation of several promoters of key enzymes of this catabolic pathway. Less is known about the behavior of Y. lipolytica in large bioreactors using these substrates. There is therefore a lack of established know-how concerning high cell density culture protocols of this yeast. Consequently, the establishment of suitable induction conditions is required, to maximize recombinant protein production under the control of these promoters. Results Human interferon α2b (huIFN α2b production in Yarrowia lipolytica was used as a model for the enhancement of recombinant protein production under the control of the oleic acid (OA-inducible promoter POX2. Cell viability and heterologous protein production were enhanced by exponential glucose feeding, to generate biomass before OA induction. The optimal biomass level before induction was determined (73 g L-1, and glucose was added with oleic acid during the induction phase. Several oleic acid feeding strategies were assessed. Continuous feeding with OA at a ratio of 0.02 g OA per g dry cell weight increased huIFNα2b production by a factor of 1.88 (425 mg L-1 and decreased the induction time (by a factor of 2.6, 21 h. huIFN α2b degradation by an aspartic protease secreted by Y. lipolytica was prevented by adding pepstatin (10 μM, leading to produce a 19-fold more active huIFN α2b (26.2 × 107 IU mg-1. Conclusion Y. lipolytica, a generally regarded as safe (GRAS microorganism is one of the most promising non conventional yeasts for the production of biologically active therapeutic proteins under the control of hydrophobic substrate-inducible promoter.

  17. [Distiller Yeasts Producing Antibacterial Peptides].

    Science.gov (United States)

    Klyachko, E V; Morozkina, E V; Zaitchik, B Ts; Benevolensky, S V

    2015-01-01

    A new method of controlling lactic acid bacteria contamination was developed with the use of recombinant Saccharomyces cerevisiae strains producing antibacterial peptides. Genes encoding the antibacterial peptides pediocin and plantaricin with codons preferable for S. cerevisiae were synthesized, and a system was constructed for their secretory expression. Recombinant S. cerevisiae strains producing antibacterial peptides effectively inhibit the growth of Lactobacillus sakei, Pediacoccus pentasaceus, Pediacoccus acidilactici, etc. The application of distiller yeasts producing antibacterial peptides enhances the ethanol yield in cases of bacterial contamination. Recombinant yeasts producing the antibacterial peptides pediocin and plantaricin can successfully substitute the available industrial yeast strains upon ethanol production.

  18. Made for Each Other: Ascomycete Yeasts and Insects.

    Science.gov (United States)

    Blackwell, Meredith

    2017-06-01

    Fungi and insects live together in the same habitats, and many species of both groups rely on each other for success. Insects, the most successful animals on Earth, cannot produce sterols, essential vitamins, and many enzymes; fungi, often yeast-like in growth form, make up for these deficits. Fungi, however, require constantly replenished substrates because they consume the previous ones, and insects, sometimes lured by volatile fungal compounds, carry fungi directly to a similar, but fresh, habitat. Yeasts associated with insects include Ascomycota (Saccharomycotina, Pezizomycotina) and a few Basidiomycota. Beetles, homopterans, and flies are important associates of fungi, and in turn the insects carry yeasts in pits, specialized external pouches, and modified gut pockets. Some yeasts undergo sexual reproduction within the insect gut, where the genetic diversity of the population is increased, while others, well suited to their stable environment, may never mate. The range of interactions extends from dispersal of yeasts on the surface of insects (e.g., cactus- Drosophila -yeast and ephemeral flower communities, ambrosia beetles, yeasts with holdfasts) to extremely specialized associations of organisms that can no longer exist independently, as in the case of yeast-like symbionts of planthoppers. In a few cases yeast-like fungus-insect associations threaten butterflies and other species with extinction. Technical advances improve discovery and identification of the fungi but also inform our understanding of the evolution of yeast-insect symbioses, although there is much more to learn.

  19. The growth of solar radiated yeast

    Energy Technology Data Exchange (ETDEWEB)

    Kraft, T.

    1995-09-01

    This researcher plans to determine if solar radiation affects the growth of yeast. The irradiated yeast was obtained from a sample exposed in space during a Space Shuttle flight of September 9-20, 1994. Further, the control groups were held at: (1) Goddard Space Flight Center (GSFC) in Greenbelt, Maryland; and (2) South Dakota School of Mines and Technology. The procedure used was based on the fact that yeast is most often used in consumable baked goods. Therefore, the yeast was incorporated into a basic Betty Crocker bread recipe. Data was collected by placing measured amounts of dough into sample containers with fifteen minute growth in height measurements collected and recorded. This researcher assumed the viability of yeast to be relative to its ability to produce carbon dioxide gas and cause the dough to rise. As all ingredients and surroundings were equal, this researcher assumed the yeast will produce the only significant difference in data collected. This researcher noted the approximate use date on all sample packages to be prior to arrival and experiment date. All dates equal, it was then assumed each would act in a similar manner of response. This assumption will allow for equally correct data collection.

  20. The growth of solar radiated yeast

    Science.gov (United States)

    Kraft, Tyrone

    1995-01-01

    This researcher plans to determine if solar radiation affects the growth of yeast. The irradiated yeast was obtained from a sample exposed in space during a Space Shuttle flight of September 9-20, 1994. Further, the control groups were held at: (1) Goddard Space Flight Center (GSFC) in Greenbelt, Maryland; and (2) South Dakota School of Mines and Technology. The procedure used was based on the fact that yeast is most often used in consumable baked goods. Therefore, the yeast was incorporated into a basic Betty Crocker bread recipe. Data was collected by placing measured amounts of dough into sample containers with fifteen minute growth in height measurements collected and recorded. This researcher assumed the viability of yeast to be relative to its ability to produce carbon dioxide gas and cause the dough to rise. As all ingredients and surroundings were equal, this researcher assumed the yeast will produce the only significant difference in data collected. This researcher noted the approximate use date on all sample packages to be prior to arrival and experiment date. All dates equal, it was then assumed each would act in a similar manner of response. This assumption will allow for equally correct data collection.

  1. Cell dualism: presence of cells with alternative membrane potentials in growing populations of bacteria and yeasts.

    Science.gov (United States)

    Ivanov, Volodymyr; Rezaeinejad, Saeid; Chu, Jian

    2013-10-01

    It is considered that all growing cells, for exception of acidophilic bacteria, have negatively charged inside cytoplasmic membrane (Δψ⁻-cells). Here we show that growing populations of microbial cells contain a small portion of cells with positively charged inside cytoplasmic membrane (Δψ⁺-cells). These cells were detected after simultaneous application of the fluorescent probes for positive membrane potential (anionic dye DIBAC⁻) and membrane integrity (propidium iodide, PI). We found in exponentially growing cell populations of Escherichia coli and Saccharomyces cerevisiae that the content of live Δψ⁻-cells was 93.6 ± 1.8 % for bacteria and 90.4 ± 4.0 % for yeasts and the content of live Δψ⁺-cells was 0.9 ± 0.3 % for bacteria and 2.4 ± 0.7 % for yeasts. Hypothetically, existence of Δψ⁺-cells could be due to short-term, about 1 min for bacteria and 5 min for yeasts, change of membrane potential from negative to positive value during the cell cycle. This change has been shown by the reversions of K⁺, Na⁺, and Ca²⁺ ions fluxes across the cell membrane during synchronous yeast culture. The transformation of Δψ(⁻-cells to Δψ⁺-cells can be explained by slow influx of K⁺ ions into Δψ⁻-cell to the trigger level of K⁺ concentration ("compression of potassium spring"), which is forming "alternative" Δψ⁺-cell for a short period, following with fast efflux of K⁺ ions out of Δψ⁺-cell ("release of potassium spring") returning cell to normal Δψ⁻ state. We anticipate our results to be a starting point to reveal the biological role of cell dualism in form of Δψ⁻- and Δψ⁺- cells.

  2. Scheffersomyces cryptocercus: a new xylose-fermenting yeast associated with the gut of wood roaches and new combinations in the Sugiyamaella yeast clade.

    Science.gov (United States)

    Urbina, Hector; Frank, Robert; Blackwell, Meredith

    2013-01-01

    The gut of wood-feeding insects is a microhabitat for a specialized community of microbes, including bacteria and several groups of eukaryotes such as nematodes, parabasalids and fungi. The characterization of gut yeast communities from a variety of insects has shown that certain yeasts often are associated with the insects. The gut of wood-feeding insects is rich in ascomycete yeasts and in particular xylose-fermenting (X-F) and assimilating yeasts have been consistently present in the gut of lignicolous insects. The objective of this study was the characterization of the yeast flora from the gut of the wood roach Cryptocercus sp. (Blattodea: Cryptocercidae). Five wood roaches were collected along the Appalachian Trail near the border between Tennessee and North Carolina, USA. We isolated 18 yeast strains from the wood roaches identified as Sugiyamaella paludigena and Sugiyamaella lignohabitans, xylose-assimilating yeasts, and Scheffersomyces cryptocercus (NRRL Y-48824(T) = CBS 12658) a new species of X-F yeast. The presence of X-F and certain non X-F yeasts in the gut of the subsocial wood roach Cryptocercus sp. extends the previous findings of associations between certain ascomycete yeasts and lignicolous insects. New combinations were made for 13 asexual members of the Sugiyamaella clade.

  3. Inactivation of Escherichia coli in soil by solarization

    International Nuclear Information System (INIS)

    Wu, S.; Nishihara, M.; Kawasaki, Y.; Yokoyama, A.; Matsuura, K.; Koga, T.; Ueno, D.; Inoue, K.; Someya, T.

    2009-01-01

    Contamination of agricultural soil by fecal pathogenic bacteria poses a potential risk of infection to humans. For the biosafety control of field soil, soil solarization in an upland field was examined to determine the efficiency of solarization on the inactivation of Escherichia coli inoculated into soil as a model microorganism for human pathogenic bacteria. Soil solarization, carried out by sprinkling water and covering the soil surface with thin plastic sheets, greatly increased the soil temperature. The daily average temperature of the solarized soil was 4–10°C higher than that of the non-solarized soil and fluctuated between 31 and 38°C. The daily highest temperature reached more than 40°C for 8 days in total in the solarized soil during the second and third weeks of the experiment. Escherichia coli in the solarized soil became undetectable (< 0.08 c.f.u. g −1 dry soil) within 4 weeks as a result, whereas E. coli survived for more than 6 weeks in the non-solarized soil. Soil solarization, however, had little influence on the total direct count and total viable count of bacteria in the soil. These results indicate that soil solarization would be useful for the biosafety control of soil contaminated by human pathogens via immature compost or animal feces. (author)

  4. Effect of increasing growth temperature on yeast fermentation ...

    African Journals Online (AJOL)

    The effect of increasing growth temperature on yeast fermentation was studied at approximately 5 oC intervals over a range of 18 – 37 oC, using one strain each of ale, lager and wine yeast. The ale and wine yeasts grew at all the temperatures tested, but lager yeast failed to grow at 37 oC. All these strains gave lower ...

  5. Yeasts preservation: alternatives for lyophilisation

    NARCIS (Netherlands)

    Nyanga, L.K.; Nout, M.J.R.; Smid, E.J.; Boekhout, T.; Zwietering, M.H.

    2012-01-01

    The aim of the study was to compare the effect of two low-cost, low technology traditional methods for drying starter cultures with standard lyophilisation. Lyophilised yeast cultures and yeast cultures preserved in dry rice cakes and dry plant fibre strands were examined for viable cell counts

  6. Biotechnology of non-Saccharomyces yeasts--the ascomycetes.

    Science.gov (United States)

    Johnson, Eric A

    2013-01-01

    Saccharomyces cerevisiae and several other yeast species are among the most important groups of biotechnological organisms. S. cerevisiae and closely related ascomycetous yeasts are the major producer of biotechnology products worldwide, exceeding other groups of industrial microorganisms in productivity and economic revenues. Traditional industrial attributes of the S. cerevisiae group include their primary roles in food fermentations such as beers, cider, wines, sake, distilled spirits, bakery products, cheese, sausages, and other fermented foods. Other long-standing industrial processes involving S. cerevisae yeasts are production of fuel ethanol, single-cell protein (SCP), feeds and fodder, industrial enzymes, and small molecular weight metabolites. More recently, non-Saccharomyces yeasts (non-conventional yeasts) have been utilized as industrial organisms for a variety of biotechnological roles. Non-Saccharomyces yeasts are increasingly being used as hosts for expression of proteins, biocatalysts and multi-enzyme pathways for the synthesis of fine chemicals and small molecular weight compounds of medicinal and nutritional importance. Non-Saccharomyces yeasts also have important roles in agriculture as agents of biocontrol, bioremediation, and as indicators of environmental quality. Several of these products and processes have reached commercial utility, while others are in advanced development. The objective of this mini-review is to describe processes currently used by industry and those in developmental stages and close to commercialization primarily from non-Saccharomyces yeasts with an emphasis on new opportunities. The utility of S. cerevisiae in heterologous production of selected products is also described.

  7. Taming wild yeast: potential of conventional and nonconventional yeasts in industrial fermentations.

    Science.gov (United States)

    Steensels, Jan; Verstrepen, Kevin J

    2014-01-01

    Yeasts are the main driving force behind several industrial food fermentation processes, including the production of beer, wine, sake, bread, and chocolate. Historically, these processes developed from uncontrolled, spontaneous fermentation reactions that rely on a complex mixture of microbes present in the environment. Because such spontaneous processes are generally inconsistent and inefficient and often lead to the formation of off-flavors, most of today's industrial production utilizes defined starter cultures, often consisting of a specific domesticated strain of Saccharomyces cerevisiae, S. bayanus, or S. pastorianus. Although this practice greatly improved process consistency, efficiency, and overall quality, it also limited the sensorial complexity of the end product. In this review, we discuss how Saccharomyces yeasts were domesticated to become the main workhorse of food fermentations, and we investigate the potential and selection of nonconventional yeasts that are often found in spontaneous fermentations, such as Brettanomyces, Hanseniaspora, and Pichia spp.

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

    Science.gov (United States)

    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.

  9. Yeast Interacting Proteins Database: YFR015C, YFR015C [Yeast Interacting Proteins Database

    Lifescience Database Archive (English)

    Full Text Available yeast homolog; expression induced by glucose limitation, nitrogen starvation, environmental stress, and entr...ression induced by glucose limitation, nitrogen starvation, environmental stress, and entry into stationary ...tion, nitrogen starvation, environmental stress, and entry into stationary phase Rows with this bait as bait..., the more highly expressed yeast homolog; expression induced by glucose limitation, nitrogen starvation, environmental

  10. [Yeast-like fungi in the gastrointestinal tract in children and adolescents with diabetes type 1].

    Science.gov (United States)

    Kowalewska, Beata; Kawko, Małgorzata; Zorena, Katarzyna; Myśliwiec, Małgorzata

    2015-01-01

    In recent years the frequency of fungal infections in human populations has increased considerably. The most common type offungus attacking the human organism is Candida albicans. Yeast-like fungi occur naturally in the oral cavity, intestines, vagina, or skin, however in amounts not dangerous to human health. The studies so far have shown that patients with diabetes type 1 (T1DM) to a large degree are exposed to complications related to fungal infections. A substantial growth of fungi observed in diabetic patients may unfavorably affect metabolic compensation, and lead to increased demand for insulin, as well as to the difficult to cure symptom infections. The weaker the immune resistance in patients with diabetes, the greater the risk of ailments related to candidiasis. The article contains a review of recent literature regarding the problems related to occurrence of yeast-like fungi in digestive tract of children with diabetes type 1. © Polish Society for Pediatric Endocrinology and Diabetology.

  11. Radiodiagnosis of yeast alveolits (a clinicoexperimental study)

    International Nuclear Information System (INIS)

    Amosov, I.S.; Smirnov, V.A.

    1984-01-01

    A clinicoroetgenological study was made of 115 workers engaged in the yeast production for different periods of time. Disorders of the respiration biomechanics were revealed depending on the period of service. These data were obtained as a result of the use of roentgenopneumopolygraphy. An experimental study was conducted to establish the nature of lesions in the bronchopulmonary system in allergic alveolitis. The effect of finely divided yeast dust on the bronchopulmonary system was studied on 132 guinea-pigs usinq microbronchography and morphological examination. As a result of the study it has been established that during the inhalation of yeast dust, notnceable dystrophy of the bronchi develops, the sizes of alveoli enlarge and part of them undergo emphysematous distension with the rupture of the interalveolar septa. In the course of the study, it has been shown that yeast dust is little agreessive, yeast alveolitis develops after many years of work. The clinical symptoms are non-specific and insignificant. X-ray and morphological changes are followed by the physical manifestations of yeast alveolitis

  12. Assessment of Shiga toxin-producing Escherichia coli isolates from wildlife meat as potential pathogens for humans.

    Science.gov (United States)

    Miko, Angelika; Pries, Karin; Haby, Sabine; Steege, Katja; Albrecht, Nadine; Krause, Gladys; Beutin, Lothar

    2009-10-01

    A total of 140 Shiga toxin-producing Escherichia coli (STEC) strains from wildlife meat (deer, wild boar, and hare) isolated in Germany between 1998 and 2006 were characterized with respect to their serotypes and virulence markers associated with human pathogenicity. The strains grouped into 38 serotypes, but eight O groups (21, 146, 128, 113, 22, 88, 6, and 91) and four H types (21, 28, 2, and 8) accounted for 71.4% and 75.7% of all STEC strains from game, respectively. Eighteen of the serotypes, including enterohemorrhagic E. coli (EHEC) O26:[H11] and O103:H2, were previously found to be associated with human illness. Genes linked to high-level virulence for humans (stx(2), stx(2d), and eae) were present in 46 (32.8%) STEC strains from game. Fifty-four STEC isolates from game belonged to serotypes which are frequently found in human patients (O103:H2, O26:H11, O113:H21, O91:H21, O128:H2, O146:H21, and O146:H28). These 54 STEC isolates were compared with 101 STEC isolates belonging to the same serotypes isolated from farm animals, from their food products, and from human patients. Within a given serotype, most STEC strains were similar with respect to their stx genotypes and other virulence attributes, regardless of origin. The 155 STEC strains were analyzed for genetic similarity by XbaI pulsed-field gel electrophoresis. O103:H2, O26:H11, O113:H21, O128:H2, and O146:H28 STEC isolates from game were 85 to 100% similar to STEC isolates of the same strains from human patients. By multilocus sequence typing, game EHEC O103:H2 strains were attributed to a clonal lineage associated with hemorrhagic diseases in humans. The results from our study indicate that game animals represent a reservoir for and a potential source of human pathogenic STEC and EHEC strains.

  13. MALDI-TOF MS as a tool to identify foodborne yeasts and yeast-like fungi.

    Science.gov (United States)

    Quintilla, Raquel; Kolecka, Anna; Casaregola, Serge; Daniel, Heide M; Houbraken, Jos; Kostrzewa, Markus; Boekhout, Teun; Groenewald, Marizeth

    2018-02-02

    Since food spoilage by yeasts causes high economic losses, fast and accurate identifications of yeasts associated with food and food-related products are important for the food industry. In this study the efficiency of the matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) to identify food related yeasts was evaluated. A CBS in-house MALDI-TOF MS database was created and later challenged with a blinded test set of 146 yeast strains obtained from food and food related products. Ninety eight percent of the strains were correctly identified with log score values>1.7. One strain, Mrakia frigida, gained a correct identification with a score value1.7. Ambiguous identifications were observed due to two incorrect reference mass spectra's found in the commercial database BDAL v.4.0, namely Candida sake DSM 70763 which was re-identified as Candida oleophila, and Candida inconspicua DSM 70631 which was re-identified as Pichia membranifaciens. MALDI-TOF MS can distinguish between most of the species, but for some species complexes, such as the Kazachstania telluris and Mrakia frigida complexes, MALDI-TOF MS showed limited resolution and identification of sibling species was sometimes problematic. Despite this, we showed that the MALDI-TOF MS is applicable for routine identification and validation of foodborne yeasts, but a further update of the commercial reference databases is needed. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Shiga toxin-producing Escherichia coli in humans and the food chain in Bangladesh

    NARCIS (Netherlands)

    Islam, M.A.

    2009-01-01

    Shiga toxin-producing Escherichia coli (STEC) are significant pathogenic bacteria that can cause severe gastrointestinal diseases and also the hemolytic-uremic syndrome. Domestic ruminants appear to be the main reservoirs of these organisms. Although Bangladesh is an endemic zone for diarrhea caused

  15. A central role for TOR signalling in a yeast model for juvenile CLN3 disease

    Directory of Open Access Journals (Sweden)

    Michael E. Bond

    2015-11-01

    Full Text Available Yeasts provide an excellent genetically tractable eukaryotic system for investigating the function of genes in their biological context, and are especially relevant for those conserved genes that cause disease. We study the role of btn1, the orthologue of a human gene that underlies an early onset neurodegenerative disease (juvenile CLN3 disease, neuronal ceroid lipofuscinosis (NCLs or Batten disease in the fission yeast Schizosaccharomyces pombe. A global screen for genetic interactions with btn1 highlighted a conserved key signalling hub in which multiple components functionally relate to this conserved disease gene. This signalling hub includes two major mitogen-activated protein kinase (MAPK cascades, and centers on the Tor kinase complexes TORC1 and TORC2. We confirmed that yeast cells modelling CLN3 disease exhibit features consistent with dysfunction in the TORC pathways, and showed that modulating TORC function leads to a comprehensive rescue of defects in this yeast disease model. The same pathways may be novel targets in the development of therapies for the NCLs and related diseases.

  16. Yeasts are essential for cocoa bean fermentation.

    Science.gov (United States)

    Ho, Van Thi Thuy; Zhao, Jian; Fleet, Graham

    2014-03-17

    Cocoa beans (Theobroma cacao) are the major raw material for chocolate production and fermentation of the beans is essential for the development of chocolate flavor precursors. In this study, a novel approach was used to determine the role of yeasts in cocoa fermentation and their contribution to chocolate quality. Cocoa bean fermentations were conducted with the addition of 200ppm Natamycin to inhibit the growth of yeasts, and the resultant microbial ecology and metabolism, bean chemistry and chocolate quality were compared with those of normal (control) fermentations. The yeasts Hanseniaspora guilliermondii, Pichia kudriavzevii and Kluyveromyces marxianus, the lactic acid bacteria Lactobacillus plantarum and Lactobacillus fermentum and the acetic acid bacteria Acetobacter pasteurianus and Gluconobacter frateurii were the major species found in the control fermentation. In fermentations with the presence of Natamycin, the same bacterial species grew but yeast growth was inhibited. Physical and chemical analyses showed that beans fermented without yeasts had increased shell content, lower production of ethanol, higher alcohols and esters throughout fermentation and lesser presence of pyrazines in the roasted product. Quality tests revealed that beans fermented without yeasts were purplish-violet in color and not fully brown, and chocolate prepared from these beans tasted more acid and lacked characteristic chocolate flavor. Beans fermented with yeast growth were fully brown in color and gave chocolate with typical characters which were clearly preferred by sensory panels. Our findings demonstrate that yeast growth and activity were essential for cocoa bean fermentation and the development of chocolate characteristics. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.

  17. Off-target effects of psychoactive drugs revealed by genome-wide assays in yeast.

    Directory of Open Access Journals (Sweden)

    Elke Ericson

    2008-08-01

    Full Text Available To better understand off-target effects of widely prescribed psychoactive drugs, we performed a comprehensive series of chemogenomic screens using the budding yeast Saccharomyces cerevisiae as a model system. Because the known human targets of these drugs do not exist in yeast, we could employ the yeast gene deletion collections and parallel fitness profiling to explore potential off-target effects in a genome-wide manner. Among 214 tested, documented psychoactive drugs, we identified 81 compounds that inhibited wild-type yeast growth and were thus selected for genome-wide fitness profiling. Many of these drugs had a propensity to affect multiple cellular functions. The sensitivity profiles of half of the analyzed drugs were enriched for core cellular processes such as secretion, protein folding, RNA processing, and chromatin structure. Interestingly, fluoxetine (Prozac interfered with establishment of cell polarity, cyproheptadine (Periactin targeted essential genes with chromatin-remodeling roles, while paroxetine (Paxil interfered with essential RNA metabolism genes, suggesting potential secondary drug targets. We also found that the more recently developed atypical antipsychotic clozapine (Clozaril had no fewer off-target effects in yeast than the typical antipsychotics haloperidol (Haldol and pimozide (Orap. Our results suggest that model organism pharmacogenetic studies provide a rational foundation for understanding the off-target effects of clinically important psychoactive agents and suggest a rational means both for devising compound derivatives with fewer side effects and for tailoring drug treatment to individual patient genotypes.

  18. Mitochondrial fission proteins regulate programmed cell death in yeast

    OpenAIRE

    Fannjiang, Yihru; Cheng, Wen-Chih; Lee, Sarah J.; Qi, Bing; Pevsner, Jonathan; McCaffery, J. Michael; Hill, R. Blake; Basañez, Gorka; Hardwick, J. Marie

    2004-01-01

    The possibility that single-cell organisms undergo programmed cell death has been questioned in part because they lack several key components of the mammalian cell death machinery. However, yeast encode a homolog of human Drp1, a mitochondrial fission protein that was shown previously to promote mammalian cell death and the excessive mitochondrial fragmentation characteristic of apoptotic mammalian cells. In support of a primordial origin of programmed cell death involving mitochondria, we fo...

  19. Genomic Evolution of the Ascomycete Yeasts

    Energy Technology Data Exchange (ETDEWEB)

    Riley, Robert; Haridas, Sajeet; Salamov, Asaf; Boundy-Mills, Kyria; Goker, Markus; Hittinger, Chris; Klenk, Hans-Peter; Lopes, Mariana; Meir-Kolthoff, Jan P.; Rokas, Antonis; Rosa, Carlos; Scheuner, Carmen; Soares, Marco; Stielow, Benjamin; Wisecaver, Jennifer H.; Wolfe, Ken; Blackwell, Meredith; Kurtzman, Cletus; Grigoriev, Igor; Jeffries, Thomas

    2015-03-16

    Yeasts are important for industrial and biotechnological processes and show remarkable metabolic and phylogenetic diversity despite morphological similarities. We have sequenced the genomes of 16 ascomycete yeasts of taxonomic and industrial importance including members of Saccharomycotina and Taphrinomycotina. Phylogenetic analysis of these and previously published yeast genomes helped resolve the placement of species including Saitoella complicata, Babjeviella inositovora, Hyphopichia burtonii, and Metschnikowia bicuspidata. Moreover, we find that alternative nuclear codon usage, where CUG encodes serine instead of leucine, are monophyletic within the Saccharomycotina. Most of the yeasts have compact genomes with a large fraction of single exon genes, and a tendency towards more introns in early-diverging species. Analysis of enzyme phylogeny gives insights into the evolution of metabolic capabilities such as methanol utilization and assimilation of alternative carbon sources.

  20. Specialist nectar-yeasts decline with urbanization in Berlin

    Science.gov (United States)

    Wehner, Jeannine; Mittelbach, Moritz; Rillig, Matthias C.; Verbruggen, Erik

    2017-03-01

    Nectar yeasts are common inhabitants of insect-pollinated flowers but factors determining their distribution are not well understood. We studied the influence of host identity, environmental factors related to pollution/urbanization, and the distance to a target beehive on local distribution of nectar yeasts within Robinia pseudoacacia L. and Tilia tomentosa Moench in Berlin, Germany. Nectar samples of six individuals per species were collected at seven sites in a 2 km radius from each target beehive and plated on YM-Agar to visualise the different morphotypes, which were then identified by sequencing a section of the 26S rDNA gene. Multivariate linear models were used to analyze the effects of all investigated factors on yeast occurrence per tree. Yeast distribution was mainly driven by host identity. The influence of the environmental factors (NO2, height of construction, soil sealing) strongly depended on the radius around the tree, similar to the distance of the sampled beehive. Incidence of specialist nectar-borne yeast species decreased with increasing pollution/urbanization index. Given that specialist yeast species gave way to generalist yeasts that have a reduced dependency on pollinators for between-flower dispersal, our results indicate that increased urbanization may restrict the movement of nectar-specialized yeasts, via limitations of pollinator foraging behavior.

  1. Isolation, genotyping, and antimicrobial resistance of zoonotic shiga toxin-producing escherichia coli

    Science.gov (United States)

    Shiga toxin-producing Escherichia coli (STEC) is an enteric pathogen linked to outbreaks of human gastroenteritis with diverse clinical spectra. Traditional culture and isolation methods, including selective enrichment and differential plating, have enabled the effective recovery of STEC. Ruminants ...

  2. Black yeast-like fungi in skin and nail

    DEFF Research Database (Denmark)

    Saunte, D M; Tarazooie, B; Arendrup, M C

    2011-01-01

    Black yeast-like fungi are rarely reported from superficial infections. We noticed a consistent prevalence of these organisms as single isolations from mycological routine specimens. To investigate the prevalence of black yeast-like fungi in skin, hair and nail specimens and to discuss...... the probability of these species to be involved in disease. Slow-growing black yeast-like fungi in routine specimens were prospectively collected and identified. A questionnaire regarding patient information was sent to physicians regarding black yeast-like fungus positive patients. A total of 20 746...... dermatological specimens were examined by culture. Black yeast-like fungi accounted for 2.2% (n = 108) of the positive cultures. Only 31.0% of the samples, culture positive for black yeast-like fungi were direct microscopy positive when compared with overall 68.8% of the culture positive specimens. The most...

  3. The unfolded protein response has a protective role in yeast models of classic galactosemia

    Directory of Open Access Journals (Sweden)

    Evandro A. De-Souza

    2014-01-01

    Full Text Available Classic galactosemia is a human autosomal recessive disorder caused by mutations in the GALT gene (GAL7 in yeast, which encodes the enzyme galactose-1-phosphate uridyltransferase. Here we show that the unfolded protein response pathway is triggered by galactose in two yeast models of galactosemia: lithium-treated cells and the gal7Δ mutant. The synthesis of galactose-1-phosphate is essential to trigger the unfolded protein response under these conditions because the deletion of the galactokinase-encoding gene GAL1 completely abolishes unfolded protein response activation and galactose toxicity. Impairment of the unfolded protein response in both yeast models makes cells even more sensitive to galactose, unmasking its cytotoxic effect. These results indicate that endoplasmic reticulum stress is induced under galactosemic conditions and underscores the importance of the unfolded protein response pathway to cellular adaptation in these models of classic galactosemia.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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

  6. Biochemical characterization of recombinant dihydroorotate dehydrogenase from the opportunistic pathogenic yeast Candida albicans

    DEFF Research Database (Denmark)

    Zameitat, E.; Gojkovic, Zoran; Knecht, Wolfgang

    2006-01-01

    Candida albicans is the most prevalent yeast pathogen in humans, and recently it has become increasingly resistant to the current antifungal agents. In this study we investigated C. albicans dihydroorotate dehydrogenase (DHODH, EC 1.3.99.11), which catalyzes the fourth step of de novo pyrimidine...

  7. Chemical signaling and insect attraction is a conserved trait in yeasts.

    Science.gov (United States)

    Becher, Paul G; Hagman, Arne; Verschut, Vasiliki; Chakraborty, Amrita; Rozpędowska, Elżbieta; Lebreton, Sébastien; Bengtsson, Marie; Flick, Gerhard; Witzgall, Peter; Piškur, Jure

    2018-03-01

    Yeast volatiles attract insects, which apparently is of mutual benefit, for both yeasts and insects. However, it is unknown whether biosynthesis of metabolites that attract insects is a basic and general trait, or if it is specific for yeasts that live in close association with insects. Our goal was to study chemical insect attractants produced by yeasts that span more than 250 million years of evolutionary history and vastly differ in their metabolism and lifestyle. We bioassayed attraction of the vinegar fly Drosophila melanogaster to odors of phylogenetically and ecologically distinct yeasts grown under controlled conditions. Baker's yeast Saccharomyces cerevisiae , the insect-associated species Candida californica , Pichia kluyveri and Metschnikowia andauensis , wine yeast Dekkera bruxellensis , milk yeast Kluyveromyces lactis , the vertebrate pathogens Candida albicans and Candida glabrata , and oleophilic Yarrowia lipolytica were screened for fly attraction in a wind tunnel. Yeast headspace was chemically analyzed, and co-occurrence of insect attractants in yeasts and flowering plants was investigated through a database search. In yeasts with known genomes, we investigated the occurrence of genes involved in the synthesis of key aroma compounds. Flies were attracted to all nine yeasts studied. The behavioral response to baker's yeast was independent of its growth stage. In addition to Drosophila , we tested the basal hexapod Folsomia candida (Collembola) in a Y-tube assay to the most ancient yeast, Y. lipolytica, which proved that early yeast signals also function on clades older than neopteran insects. Behavioral and chemical data and a search for selected genes of volatile metabolites underline that biosynthesis of chemical signals is found throughout the yeast clade and has been conserved during the evolution of yeast lifestyles. Literature and database reviews corroborate that yeast signals mediate mutualistic interactions between insects and yeasts

  8. Molecular characterization and antimicrobial resistance of faecal and urinary Escherichia coli isolated from dogs and humans in Italy

    Directory of Open Access Journals (Sweden)

    Clara Tramuta

    2014-03-01

    Full Text Available During this study, 109 faecal Escherichia coli samples isolated from 61 dogs and 48 humans were characterised according to phylogenetic group, extraintestinal virulence factors and antibiotic resistance. The isolates from dogs were predominantly distributed within phylogroup B1 (36%, while the majority of human strains belonged to phylogroup B2 (54%. The prevalence of cnf1, hlyA, papC and sfa virulence genes was significantly associated with the group B2. Canine isolates showed multidrug resistance (MDR more frequently than human strains. Since group B2 contains most of the strains that cause extraintestinal infections, all 46 B2 faecal strains were confronted against an addition population of 57 urinary E. coli strains belonging to the same phylogroup. The comparison shows that there was no significant difference in the occurrence of virulence factors or in the distribution of antibiotic resistance between faecal and urinary E. coli isolates fromd dogs. At the same time, a highly significant association was detected between multiple resistence and the source of the strains and between MDR and E. coli isolated from urine in human. This study highlighted similar features of E. coli isolated across sources and hosts. The data suggest a high prevalence of antibiotic resistance in faecal strains, which may represent a serious health risk since these strains can function as a reservoir for uropathogenic E. coli.

  9. Effect of fungicides on epiphytic yeasts associated with strawberry

    Science.gov (United States)

    Debode, Jane; Van Hemelrijck, Wendy; Creemers, Piet; Maes, Martine

    2013-01-01

    We studied the effect of two commonly used fungicides on the epiphytic yeast community of strawberry. Greenhouse and field experiments were conducted applying Switch (cyprodinil plus fludioxonil) or Signum (boscalid plus pyraclostrobin) to strawberry plants. Yeasts on leaves and fruits were assessed on treated and untreated plants at several time points via plating and denaturing gradient gel electrophoresis (DGGE) analysis. The yeast counts on plates of the treated plants were similar to the control plants. Unripe fruits had 10 times larger yeast concentrations than ripe fruits or leaves. Some dominant yeast types were isolated and in vitro tests showed that they were at least 10 times less sensitive to Switch and Signum as compared with two important fungal strawberry pathogens Botrytis cinerea and Colletotrichum acutatum, which are the targets for the fungicide control. DGGE analysis showed that the applied fungicides had no effect on the composition of the yeast communities, while the growing system, strawberry tissue, and sampling time did affect the yeast communities. The yeast species most commonly identified were Cryptococcus, Rhodotorula, and Sporobolomyces. These results point toward the potential applicability of natural occurring yeast antagonists into an integrated disease control strategy for strawberry diseases.

  10. mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli

    International Nuclear Information System (INIS)

    Zhang Weici; Xiao Weihua; Wei Haiming; Zhang Jian; Tian Zhigang

    2006-01-01

    Codon usage and thermodynamic optimization of the 5'-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5'-end of mRNA may be used as a general strategy for human protein production in E. coli

  11. Specific binding of lactoferrin to Escherichia coli isolated from human intestinal infections

    International Nuclear Information System (INIS)

    Naidu, S.S.; Erdei, J.; Forsgren, A.; Naidu, A.S.; Czirok, E.; Gado, I.; Kalfas, S.; Thoren, A.

    1991-01-01

    The degrees of human lactoferrin (HLf) and bovine lactoferrin (BLf) binding in 169 Escherichia coli strains isolated from human intestinal infections, and in an additional 68 strains isolated from healthy individuals, were examined in a 125 I-labelled protein binding assay. The binding was expressed as a percentage calculated from the total labelled ligand added to bacteria. The HLf and BLf binding to E. coli was in the range 3.7 to 73.4% and 4.8 to 61.6%, respectively. Enterotoxigenic strains demonstrated a significantly higher HLf binding (median = 19%) than enteropathogenic, enteroinvasive, enterohaemorrhagic strains or normal intestinal E. coli isolates (medians 6 to 9). Enteropathogenic strains belonging to serotypes O44 and O127 demonstrated significantly higher HLf binding compared to O26, O55, O111, O119 and O126. No significant differences in the degree of HLf or BLf binding were found between aerobactin-producing and non-producing strains. The interaction was further characterized in a high Lf-binging EPEC strain, E34663 (serotype O127). The binding was stable in the pH range 4.0 to 7.5, did not dissociate in the presence of 2M NaCl or 2M urea, and reached saturation within two h. Unlabelled HLf and BLf displaced the 125 I-HLf binding to E34663 in a dose-dependent manner. Apo- and iron-saturated forms of Lf demonstrated similar binding to E34663. Among various unlabelled subephithelial matrix proteins and carbohydrates tested (in 10 4 -fold excess) only fibronectin and fibrinogen caused a moderate inhibition of 125 I-HLf binding. According to Scatchard plot analysis, 5,400 HLf-binding sites/cell, with an affinity constant (K a ) of 1.4 x 10 -7 M, were estimated in strain E34663. These data establish the presence of a specific Lf-binding mechanism in E. coli. (au)

  12. Yeast as a Model System to Study Tau Biology

    Directory of Open Access Journals (Sweden)

    Ann De Vos

    2011-01-01

    Full Text Available Hyperphosphorylated and aggregated human protein tau constitutes a hallmark of a multitude of neurodegenerative diseases called tauopathies, exemplified by Alzheimer's disease. In spite of an enormous amount of research performed on tau biology, several crucial questions concerning the mechanisms of tau toxicity remain unanswered. In this paper we will highlight some of the processes involved in tau biology and pathology, focusing on tau phosphorylation and the interplay with oxidative stress. In addition, we will introduce the development of a human tau-expressing yeast model, and discuss some crucial results obtained in this model, highlighting its potential in the elucidation of cellular processes leading to tau toxicity.

  13. Replication dynamics of the yeast genome.

    Science.gov (United States)

    Raghuraman, M K; Winzeler, E A; Collingwood, D; Hunt, S; Wodicka, L; Conway, A; Lockhart, D J; Davis, R W; Brewer, B J; Fangman, W L

    2001-10-05

    Oligonucleotide microarrays were used to map the detailed topography of chromosome replication in the budding yeast Saccharomyces cerevisiae. The times of replication of thousands of sites across the genome were determined by hybridizing replicated and unreplicated DNAs, isolated at different times in S phase, to the microarrays. Origin activations take place continuously throughout S phase but with most firings near mid-S phase. Rates of replication fork movement vary greatly from region to region in the genome. The two ends of each of the 16 chromosomes are highly correlated in their times of replication. This microarray approach is readily applicable to other organisms, including humans.

  14. Protein functional analysis data in support of comparative proteomics of the pathogenic black yeast Exophiala dermatitidis under different temperature conditions

    Directory of Open Access Journals (Sweden)

    Donatella Tesei

    2015-12-01

    Full Text Available In the current study a comparative proteomic approach was used to investigate the response of the human pathogen black yeast Exophiala dermatitidis toward temperature treatment. Protein functional analysis – based on cellular process GO terms – was performed on the 32 temperature-responsive identified proteins. The bioinformatics analyses and data presented here provided novel insights into the cellular pathways at the base of the fungus temperature tolerance. A detailed analysis and interpretation of the data can be found in “Proteome of tolerance fine-tuning in the human pathogen black yeast Exophiala dermatitidis” by Tesei et al. (2015 [1].

  15. Flor Yeast: New Perspectives Beyond Wine Aging

    Science.gov (United States)

    Legras, Jean-Luc; Moreno-Garcia, Jaime; Zara, Severino; Zara, Giacomo; Garcia-Martinez, Teresa; Mauricio, Juan C.; Mannazzu, Ilaria; Coi, Anna L.; Bou Zeidan, Marc; Dequin, Sylvie; Moreno, Juan; Budroni, Marilena

    2016-01-01

    The most important dogma in white-wine production is the preservation of the wine aroma and the limitation of the oxidative action of oxygen. In contrast, the aging of Sherry and Sherry-like wines is an aerobic process that depends on the oxidative activity of flor strains of Saccharomyces cerevisiae. Under depletion of nitrogen and fermentable carbon sources, these yeast produce aggregates of floating cells and form an air–liquid biofilm on the wine surface, which is also known as velum or flor. This behavior is due to genetic and metabolic peculiarities that differentiate flor yeast from other wine yeast. This review will focus first on the most updated data obtained through the analysis of flor yeast with -omic tools. Comparative genomics, proteomics, and metabolomics of flor and wine yeast strains are shedding new light on several features of these special yeast, and in particular, they have revealed the extent of proteome remodeling imposed by the biofilm life-style. Finally, new insights in terms of promotion and inhibition of biofilm formation through small molecules, amino acids, and di/tri-peptides, and novel possibilities for the exploitation of biofilm immobilization within a fungal hyphae framework, will be discussed. PMID:27148192

  16. Production of Odd-Carbon Dicarboxylic Acids in Escherichia coli Using an Engineered Biotin-Fatty Acid Biosynthetic Pathway.

    Science.gov (United States)

    Haushalter, Robert W; Phelan, Ryan M; Hoh, Kristina M; Su, Cindy; Wang, George; Baidoo, Edward E K; Keasling, Jay D

    2017-04-05

    Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids.

  17. Construction of gateway-compatible yeast two-hybrid vectors for ...

    African Journals Online (AJOL)

    Yeast two-hybrid system combined with the gateway technology will greatly facilitate the cloning of interested DNA fragment into yeast two-hybrid vectors and therefore increase the efficiency of yeast two-hybrid analysis. In this study, we constructed a pair of Gateway-compatible yeast two-hybrid vectors pBTM116GW and ...

  18. Yeast species associated with wine grapes in China.

    Science.gov (United States)

    Li, Shuang-Shi; Cheng, Chao; Li, Zheng; Chen, Jing-Yu; Yan, Bin; Han, Bei-Zhong; Reeves, Malcolm

    2010-03-31

    Having more information on the yeast ecology of grapes is important for wine-makers to produce wine with high quality and typical attributes. China is a significant wine-consuming country and is becoming a serious wine-producer, but little has been reported about the yeast ecology of local ecosystems. This study provides the first step towards the exploitation of the yeast wealth in China's vine-growing regions. The aim of this study was to investigate the yeast population density and diversity on three grape varieties cultivated in four representative vine-growing regions of China. Yeast species diversity was evaluated by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequence analysis of the 5.8S internal transcribed spacer (ITS) ribosomal DNA (rDNA) region of cultivable yeasts. The grapes harbored yeast populations at 10(2)-10(6)CFU/mL, consisting mostly of non-Saccharomyces species. Seventeen different yeast species belonging to eight genera were detected on the grape samples tested, including Hanseniaspora uvarum, Cryptococcus flavescens, Pichia fermentans, Candida zemplinina, Cryptococcus carnescens, Candida inconpicua, Zygosaccharomyces fermentati, Issatchenkia terricola, Candida quercitrusa, Hanseniaspora guilliermondii, Candida bombi, Zygosaccharomyces bailii, Sporidiobolus pararoseus, Cryptococcus magnus, Metschnikowia pulcherrima, Issatchenkia orientalis and Pichia guilliermondii. H. uvarum and C. flavescens were the dominant species present on the grapes. For the first time Sporidiobolus pararoseus was discovered as an inhabitant of the grape ecosystem. The yeast community on grape berries was influenced by the grape chemical composition, vine-variety and vine-growing region. This study is the first to identify the yeast communities associated with grapes in China using molecular methods. The results enrich our knowledge of wine-related microorganisms, and can be used to promote the development of the local wine

  19. Human Bile Reduces Antimicrobial Activity of Selected Antibiotics against Enterococcus faecalis and Escherichia coli In Vitro.

    Science.gov (United States)

    Wulkersdorfer, Beatrix; Jaros, David; Eberl, Sabine; Poschner, Stefan; Jäger, Walter; Cosentini, Enrico; Zeitlinger, Markus; Schwameis, Richard

    2017-08-01

    It has been known from previous studies that body fluids, such as cerebrospinal fluid, lung surfactant, and urine, have a strong impact on the bacterial killing of many anti-infective agents. However, the influence of human bile on the antimicrobial activity of antibiotics is widely unknown. Human bile was obtained and pooled from 11 patients undergoing cholecystectomy. After sterilization of the bile fluid by gamma irradiation, its effect on bacterial killing was investigated for linezolid (LZD) and tigecycline (TGC) against Enterococcus faecalis ATCC 29212. Further, ciprofloxacin (CIP), meropenem (MEM), and TGC were tested against Escherichia coli ATCC 25922. Time-kill curves were performed in pooled human bile and Mueller-Hinton broth (MHB) over 24 h. Bacterial counts (in CFU per milliliter after 24 h) of bile growth controls were approximately equal to MHB growth controls for E. coli and approximately 2-fold greater for E. faecalis , indicating a promotion of bacterial growth by bile for the latter strain. Bile reduced the antimicrobial activity of CIP, MEM, and TGC against E. coli as well as the activity of LZD and TGC against E. faecalis This effect was strongest for TGC against the two strains. Degradation of TGC in bile was identified as the most likely explanation. These findings may have important implications for the treatment of bacterial infections of the gallbladder and biliary tract and should be explored in more detail. Copyright © 2017 American Society for Microbiology.

  20. Functional Complementation Studies Reveal Different Interaction Partners of Escherichia coli IscS and Human NFS1.

    Science.gov (United States)

    Bühning, Martin; Friemel, Martin; Leimkühler, Silke

    2017-08-29

    The trafficking and delivery of sulfur to cofactors and nucleosides is a highly regulated and conserved process among all organisms. All sulfur transfer pathways generally have an l-cysteine desulfurase as an initial sulfur-mobilizing enzyme in common, which serves as a sulfur donor for the biosynthesis of sulfur-containing biomolecules like iron-sulfur (Fe-S) clusters, thiamine, biotin, lipoic acid, the molybdenum cofactor (Moco), and thiolated nucleosides in tRNA. The human l-cysteine desulfurase NFS1 and the Escherichia coli homologue IscS share a level of amino acid sequence identity of ∼60%. While E. coli IscS has a versatile role in the cell and was shown to have numerous interaction partners, NFS1 is mainly localized in mitochondria with a crucial role in the biosynthesis of Fe-S clusters. Additionally, NFS1 is also located in smaller amounts in the cytosol with a role in Moco biosynthesis and mcm 5 s 2 U34 thio modifications of nucleosides in tRNA. NFS1 and IscS were conclusively shown to have different interaction partners in their respective organisms. Here, we used functional complementation studies of an E. coli iscS deletion strain with human NFS1 to dissect their conserved roles in the transfer of sulfur to a specific target protein. Our results show that human NFS1 and E. coli IscS share conserved binding sites for proteins involved in Fe-S cluster assembly like IscU, but not with proteins for tRNA thio modifications or Moco biosynthesis. In addition, we show that human NFS1 was almost fully able to complement the role of IscS in Moco biosynthesis when its specific interaction partner protein MOCS3 from humans was also present.

  1. Utilization of spent brewer’s yeast Saccharomyces cerevisiae for the production of yeast enzymatic hydrolysate

    Directory of Open Access Journals (Sweden)

    M Bayarjargal

    2014-09-01

    Full Text Available Spent brewer’s yeast (Saccharomyces cerevisiae is a rich source of protein, vitamins and widely used as a raw material for production of food supplements. The autolysis and enzymatic treatment of spent brewer’s yeast using Pancreatin (2.5% and Flavourzyme (2.5% were performed at 45 °C and 50 °C, respectively. The autolysis and hydrolysis processes were evaluated by determining a soluble solids, soluble protein concentration and α-amino nitrogen content in a reaction mixture. The yield of pancreatic digest and α-amino nitrogen content was high in comparison with autolysis and Flavourzyme treatment. The total solids recovery in dry Yeast hydrolysate was about 50%, a protein and α-amino nitrogen content was 55.9 and 4.8%, respectively. These results show the possibility of utilizing the spent brewer’s yeast as hydrolysate using hydrolytic enzymes and use it as a food supplement after biological experiments.DOI: http://dx.doi.org/10.5564/mjc.v12i0.179 Mongolian Journal of Chemistry Vol.12 2011: 88-91

  2. Stopped-flow studies of the reaction of D-tartronate semialdehyde-2-phosphate with human neuronal enolase and yeast enolase 1.

    Science.gov (United States)

    Brewer, John M; McKinnon, Jared S; Phillips, Robert S

    2010-03-05

    We determined the kinetics of the reaction of human neuronal enolase and yeast enolase 1 with the slowly-reacting chromophoric substrate D-tartronate semialdehyde phosphate (TSP), each in tris (tris (hydroxymethyl) aminomethane) and another buffer at several Mg2+ concentrations, 50 or 100 microM, 1 mM and 30 mM. All data were biphasic, and could be satisfactorily fit, assuming either two successive first-order reactions or two independent first-order reactions. Higher Mg2+ concentrations reduce the relative magnitude of the slower reaction. The results are interpreted in terms of a catalytically significant interaction between the two subunits of these enzymes. Copyright (c) 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  3. Substrate overlap and functional competition between human nucleotide excision repair and Escherichia coli photolyase and (A)BC excision nuclease

    International Nuclear Information System (INIS)

    Sibghat-Ullah; Sancar, Z.

    1990-01-01

    Human cell free extract prepared by the method of Manley et al. carries out repair synthesis on UV-irradiated DNA. Removal of pyrimidine dimers by photoreactivation with DNA photolyase reduces repair synthesis by about 50%. With excess enzyme in the reaction mixture photolyase reduced the repair signal by the same amount even in the absence of photoreactivating light, presumably by binding to pyrimidine dimers and interfering with the binding of human damage recognition protein. Similarly, the UvrB subunit of Escherichia coli (A)BC excinuclease when loaded onto UV-irradiated or psoralen-adducted DNA inhibited repair synthesis by cell-free extract by 75-80%. The opposite was true also as HeLa cell free extract specifically inhibited the photorepair of a thymine dimer by DNA photolyase and its removal by (A)BC excinuclease. Cell-free extracts from xeroderma pigmentosum (XP) complementation groups A and C were equally effective in blocking the E. coli repair proteins, while extracts from complementation groups D and E were ineffective in blocking the E. coli enzyme. These results suggest that XP-D and XP-E cells are defective in the damage recognition subunits(s) of human excision nuclease

  4. Selection of oleaginous yeasts for fatty acid production

    NARCIS (Netherlands)

    Lamers, Dennis; Biezen, van Nick; Martens, Dirk; Peters, Linda; Zilver, van de Eric; Jacobs-van Dreumel, Nicole; Wijffels, René H.; Lokman, Christien

    2016-01-01

    Background: Oleaginous yeast species are an alternative for the production of lipids or triacylglycerides (TAGs). These yeasts are usually non-pathogenic and able to store TAGs ranging from 20 % to 70 % of their cell mass depending on culture conditions. TAGs originating from oleaginous yeasts

  5. The tumor suppressor homolog in fission yeast, myh1+, displays a strong interaction with the checkpoint gene rad1+

    International Nuclear Information System (INIS)

    Jansson, Kristina; Warringer, Jonas; Farewell, Anne; Park, Han-Oh; Hoe, Kwang-Lae; Kim, Dong-Uk; Hayles, Jacqueline; Sunnerhagen, Per

    2008-01-01

    The DNA glycosylase MutY is strongly conserved in evolution, and homologs are found in most eukaryotes and prokaryotes examined. This protein is implicated in repair of oxidative DNA damage, in particular adenine mispaired opposite 7,8-dihydro-8-oxoguanine. Previous investigations in Escherichia coli, fission yeast, and mammalian cells show an association of mutations in MutY homologs with a mutator phenotype and carcinogenesis. Eukaryotic MutY homologs physically associate with several proteins with a role in replication, DNA repair, and checkpoint signaling, specifically the trimeric 9-1-1 complex. In a genetic investigation of the fission yeast MutY homolog, myh1 + , we show that the myh1 mutation confers a moderately increased UV sensitivity alone and in combination with mutations in several DNA repair genes. The myh1 rad1, and to a lesser degree myh1 rad9, double mutants display a synthetic interaction resulting in enhanced sensitivity to DNA damaging agents and hydroxyurea. UV irradiation of myh1 rad1 double mutants results in severe chromosome segregation defects and visible DNA fragmentation, and a failure to activate the checkpoint. Additionally, myh1 rad1 double mutants exhibit morphological defects in the absence of DNA damaging agents. We also found a moderate suppression of the slow growth and UV sensitivity of rhp51 mutants by the myh1 mutation. Our results implicate fission yeast Myh1 in repair of a wider range of DNA damage than previously thought, and functionally link it to the checkpoint pathway

  6. Genetically engineered yeast

    DEFF Research Database (Denmark)

    2014-01-01

    A genetically modified Saccharomyces cerevisiae comprising an active fermentation pathway producing 3-HP expresses an exogenous gene expressing the aminotransferase YhxA from Bacillus cereus AH1272 catalysing a transamination reaction between beta-alanine and pyruvate to produce malonate semialde......A genetically modified Saccharomyces cerevisiae comprising an active fermentation pathway producing 3-HP expresses an exogenous gene expressing the aminotransferase YhxA from Bacillus cereus AH1272 catalysing a transamination reaction between beta-alanine and pyruvate to produce malonate...... semialdehyde. The yeast may also express a 3-hydroxyisobutyrate dehydrogenase (HIBADH) and a 3-hydroxypropanoate dehydrogenase (3-HPDH) and aspartate 1-decarboxylase. Additionally the yeast may express pyruvate carboxylase and aspartate aminotransferase....

  7. The Use Of Local Product Yeast For Substitution Torula Yeast In The Formulation Of Artificial Diet Fruit Fly Larvae Bactrocera Carambolae Drew and Hancock

    International Nuclear Information System (INIS)

    Sikumbang, I.; Nasution, A.I.; Indarwatmi, M.; Kuswandi, A.N.

    2000-01-01

    The use of local product yeast I.e brewer yeast, yeast of tapai (fermented cassava), yeast of tempe (fermented soy beam), and brem(intoxicating beverage made of fermented rice) after cooked and uncooked were used to substitute torula yeast to reduce cost production for mass-rearing of fruit fly had been carried out. Artificial diet formulation consisted of torula yeast, wheat bran, nipagin, sodium benzoate, cane sugar, water and HCI ti make pH of 4. One kilogram of diet was inoculated with 1 ml of fruit fly eggs. Parameters of the experiment were, the number of pupae, weight of pupae, percentage of pupae and the percentage of viable fly. The results showed that the number of pupae were 6356 for brewers yeast with cooked and 0.942 gram/100 pupae for brem. Percentage viable emergence fly were 70%, 18.25% and 15.25% for brewers yeast with cooked and uncooked respectively. Cost production for 1.000.000 using cooked brewer yeast was reduced about Rp.179,200 or cost efficiency were 55.56%

  8. Guidelines and recommendations on yeast cell death nomenclature

    OpenAIRE

    Carmona-Gutierrez, Didac; Bauer, Maria Anna; Zimmermann, Andreas; Aguilera, Andres; Austriaco, Nicanor; Sigrist, Stephan J.

    2018-01-01

    Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cellular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the definition of accidental, regulated, and programmed forms of cell death in yeast based on a series of mor...

  9. Thymineless death is inhibited by CsrA in Escherichia coli lacking the SOS response.

    Science.gov (United States)

    Hamilton, Holly M; Wilson, Ray; Blythe, Martin; Nehring, Ralf B; Fonville, Natalie C; Louis, Edward J; Rosenberg, Susan M

    2013-11-01

    Thymineless death (TLD) is the rapid loss of colony-forming ability in bacterial, yeast and human cells starved for thymine, and is the mechanism of action of common chemotherapeutic drugs. In Escherichia coli, significant loss of viability during TLD requires the SOS replication-stress/DNA-damage response, specifically its role in inducing the inhibitor of cell division, SulA. An independent RecQ- and RecJ-dependent TLD pathway accounts for a similarly large additional component of TLD, and a third SOS- and RecQ/J-independent TLD pathway has also been observed. Although two groups have implicated the SOS-response in TLD, an SOS-deficient mutant strain from an earlier study was found to be sensitive to thymine deprivation. We performed whole-genome resequencing on that SOS-deficient strain and find that, compared with the SOS-proficient control strain, it contains five mutations in addition to the SOS-blocking lexA(Ind(-)) mutation. One of the additional mutations, csrA, confers TLD sensitivity specifically in SOS-defective strains. We find that CsrA, a carbon storage regulator, reduces TLD in SOS- or SulA-defective cells, and that the increased TLD that occurs in csrA(-) SOS-defective cells is dependent on RecQ. We consider a hypothesis in which the modulation of nucleotide pools by CsrA might inhibit TLD specifically in SOS-deficient (SulA-deficient) cells. Copyright © 2013 Elsevier B.V. All rights reserved.

  10. Guidelines and recommendations on yeast cell death nomenclature

    NARCIS (Netherlands)

    Carmona-Gutierrez, Didac; Bauer, Maria Anna; Zimmermann, Andreas; Aguilera, Andrés; Austriaco, Nicanor; Ayscough, Kathryn; Balzan, Rena; Bar-Nun, Shoshana; Barrientos, Antonio; Belenky, Peter; Blondel, Marc; Braun, Ralf J; Breitenbach, Michael; Burhans, William C; Büttner, Sabrina; Cavalieri, Duccio; Chang, Michael; Cooper, Katrina F; Côrte-Real, Manuela; Costa, Vítor; Cullin, Christophe; Dawes, Ian; Dengjel, Jörn; Dickman, Martin B; Eisenberg, Tobias; Fahrenkrog, Birthe; Fasel, Nicolas; Fröhlich, Kai-Uwe; Gargouri, Ali; Giannattasio, Sergio; Goffrini, Paola; Gourlay, Campbell W; Grant, Chris M; Greenwood, Michael T; Guaragnella, Nicoletta; Heger, Thomas; Heinisch, Jürgen; Herker, Eva; Herrmann, Johannes M; Hofer, Sebastian; Jiménez-Ruiz, Antonio; Jungwirth, Helmut; Kainz, Katharina; Kontoyiannis, Dimitrios P; Ludovico, Paula; Manon, Stéphen; Martegani, Enzo; Mazzoni, Cristina; Megeney, Lynn A; Meisinger, Chris; Nielsen, Jens; Nyström, Thomas; Osiewacz, Heinz D; Outeiro, Tiago F; Park, Hay-Oak; Pendl, Tobias; Petranovic, Dina; Picot, Stephane; Polčic, Peter; Powers, Ted; Ramsdale, Mark; Rinnerthaler, Mark; Rockenfeller, Patrick; Ruckenstuhl, Christoph; Schaffrath, Raffael; Segovia, Maria; Severin, Fedor F; Sharon, Amir; Sigrist, Stephan J; Sommer-Ruck, Cornelia; Sousa, Maria João; Thevelein, Johan M; Thevissen, Karin; Titorenko, Vladimir; Toledano, Michel B; Tuite, Mick; Vögtle, F-Nora; Westermann, Benedikt; Winderickx, Joris; Wissing, Silke; Wölfl, Stefan; Zhang, Zhaojie J; Zhao, Richard Y; Zhou, Bing; Galluzzi, Lorenzo; Kroemer, Guido; Madeo, Frank

    2018-01-01

    Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cel-lular demise. However, the investigation of yeast cell death is a relatively young field, and a widely

  11. Unraveling lipid metabolism in lipid-dependent pathogenic Malassezia yeasts

    OpenAIRE

    Celis Ramirez, A.M.

    2017-01-01

    Malassezia yeasts are lipid-dependent fungal species that are common members of the human and animal skin microbiota. The lipid-dependency is a crucial trait in the adaptation process to grow on the skin but also plays a role in their pathogenic life style. Malassezia species can cause several skin infections like dandruff or seborrheic dermatitis but also bloodstream infections. Understanding the lipid metabolism in Malassezia is essential to understand its life style as skin commensal and p...

  12. Cheese whey-induced high-cell-density production of recombinant proteins in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Neubauer Peter

    2003-04-01

    Full Text Available Abstract Background Use of lactose-rich concentrates from dairy processes for the induction of recombinant gene's expression has not received much attention although they are interesting low cost substrates for production of recombinant enzymes. Applicability of dairy waste for induction of recombinant genes in Escherichia coli was studied. Clones expressing Lactobacillus phage muramidase and Lactobacillus alcohol dehydrogenase were used for the experiments. Results Shake flask cultivations in mineral salt medium showed that cheese whey or deproteinised whey induced gene expression as efficiently as IPTG (isopropyl-β-D-thiogalactopyranoside or pure lactose. Addition of yeast extract or proteolytically degraded whey proteins did not improve the recombinant protein yield. In contrast, addition of yeast extract to the well-balanced mineral salt medium decreased the product yield. Feeding with glycerol provided sufficient amount of easily assimilable carbon source during the induction period without preventing lactose intake and induction by lactose. High-cell-density fed-batch cultivations showed that product yields comparable to IPTG-induction can be achieved by feeding bacteria with a mixture of glycerol and concentrated whey permeate during the induction. Conclusion Whey and concentrated whey permeate can be applied as an alternative inducer in recombinant high-cell-density fed-batch fermentations. The yield of the recombinant product was comparable to fermentations induced by IPTG. In low-cell-density shake flask experiments the yield was higher with whey or whey permeate than with IPTG.

  13. [Comparison of methods for the identification of the most common yeasts in the clinical microbiology laboratory].

    Science.gov (United States)

    Guelfand, L; Grisolía, P; Bozzano, C; Kaufman, S

    2003-01-01

    We evaluated different methods for the routine identification of medically important yeasts. A total of 150 clinical isolates: 25 C. albicans, 25 C. tropicalis, 25 C. glabrata, 25 C. parapsilosis, 8 C. guilliermondii, 11 C. krusei and 31 Cryptococcus neoformans were tested by Yeast Biochemical Card bioMerieux Vitek (YBC), CHROMagar Candida (CHR). The addition of yeast morphology in Corn Meal agar-Tween 80 (AM) to YBC and CHR was also evaluated. The reference methods used were: API 20C, germ tube formation, AM, Christensen urea and Birdseed agar. YBC identified 135 yeasts with an overall accuracy of 90%. Sensitivity (S) and specificity (E) were: 92-98% for C. albicans and C. tropicalis; 84-99% for C. papapsilosis; 100-99% for C. glabrata; 91-100% for C. krusei; 63-98% for C. guilliermondii and 90-99% for Cryptococcus neoformans, respectively. CHR identified correctly 100% for C. albicans, 92% for C. tropicalis and 91% for C. krusei. Both methods combined with AM provided 100% S and E. We found that YBC system was appropriate for identification of yeasts isolated from human sources. CHR was effective and easy to use for identification of C. albicans, C. tropicalis and C. krusei. The routine use of AM with both methods is recommended.

  14. Heat-treated mineral-yeast as a potent post-irradiation radioprotector

    International Nuclear Information System (INIS)

    Anzai, Kazunori; Ueno, Megumi; Nyui, Minako; Ikota, Nobuo; Kagiya, Tsutomu V.

    2008-01-01

    In vivo radioprotection of C3H mice by i.p. administration of Zn-, Mn-, Cu-, or Se-containing heat-treated Saccharomyces serevisiae yeast sample was examined. The 30-day survival of the group treated 30 min before 7.5 Gy whole-body X-irradiation with mineral-containing yeast powders suspended in 0.5% methylcellulose was significantly higher than that of control group. When mineral-yeast was administered immediately after irradiation, the survival rate was even higher and Zn- or Cu-yeast showed the highest rate (more than 90%). Although treatment with simple yeast showed a high survival rate (73%), it was significantly lower than that obtained by the Zn-yeast treatment. The effects of Zn-yeast were studied further. When the interval between irradiation and administration was varied, the protective activity of Zn-yeast decreased gradually by increasing the interval but was still significantly high for the administration at 10 h post-irradiation. The dose reduction factor of Zn-yeast (100 mg/kg, i.p. administration immediately after irradiation) was about 1.2. When the suspension of Zn-yeast was fractionated by centrifugation, the insoluble fraction showed a potent effect, while the soluble fraction had only a moderate effect. In conclusion, mineral-yeast, especially Zn-yeast, provides remarkable post-irradiation protection against lethal whole body X-irradiation. The activity is mainly attributable to the insoluble fraction, whereas some soluble components might contribute to the additional protective activity. (author)

  15. Yeast diversity and native vigor for flavor phenotypes.

    Science.gov (United States)

    Carrau, Francisco; Gaggero, Carina; Aguilar, Pablo S

    2015-03-01

    Saccharomyces cerevisiae, the yeast used widely for beer, bread, cider, and wine production, is the most resourceful eukaryotic model used for genetic engineering. A typical concern about using engineered yeasts for food production might be negative consumer perception of genetically modified organisms. However, we believe the true pitfall of using genetically modified yeasts is their limited capacity to either refine or improve the sensory properties of fermented foods under real production conditions. Alternatively, yeast diversity screening to improve the aroma and flavors could offer groundbreaking opportunities in food biotechnology. We propose a 'Yeast Flavor Diversity Screening' strategy which integrates knowledge from sensory analysis and natural whole-genome evolution with information about flavor metabolic networks and their regulation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Oral yeast carriage in patients with advanced cancer.

    Science.gov (United States)

    Davies, A N; Brailsford, S; Broadley, K; Beighton, D

    2002-04-01

    The aim of this study was to investigate oral yeast carriage amongst patients with advanced cancer. Oral rinse samples were obtained from 120 subjects. Yeasts were isolated using Sabouraud's dextrose agar and CHROMagar Candida, and were identified using a combination of the API 20 C AUX yeast identification system, species-specific PCR and 26S rDNA gene sequencing. Oral yeast carriage was present in 66% of subjects. The frequency of isolation of individual species was: Candida albicans, 46%; Candida glabrata, 18%; Candida dubliniensis, 5%; others, yeast carriage was associated with denture wearing (P = 0.006), and low stimulated whole salivary flow rate (P = 0.009). Identification of these risk factors offers new strategies for the prevention of oral candidosis in this group of patients.

  17. Influence of various scavengers of •OH radicals on the radiation sensitivity of yeast and bacteria.

    Science.gov (United States)

    Múčka, Viliam; Bláha, Pavel; Čuba, Václav; Červenák, Jaroslav

    2013-12-01

    To quantitatively investigate the influence of various •OH (hydroxyl radical) scavengers on the radiation sensitivity of yeast and bacteria, particularly to define the relationship between the protective effect of a scavenger and its •OH scavenging efficiency. In order to study the protective effect of •OH scavengers we used various concentrations of four scavengers (methanol, potassium formate, ethanol and ascorbic acid) in isotonic salt solutions. These solutions containing live yeast (Saccharomyces cerevisiae) or bacteria (Escherichia coli) were irradiated with (60)Co isotope γ -radiation using two different doses and dose rates. The number of surviving cells was determined prior to and after irradiation both in suspension with and without scavengers. The surviving fractions after irradiation with and without the scavenger were evaluated. The main results of the paper were: The surviving fraction increased approximately linearly within the measured interval with increasing concentration of the scavenger. The same dependences were found for the protecting effect depending on the scavenging efficiency. The slopes of these dependences (k) were found to be characteristic for each scavenger. The k value determined the degree in which the scavenging of •OH radicals participated in the protection of living cells. The protective effects of scavengers at the same scavenging efficiency were different and unique for each scavenger. No simple relation was found between the efficiency of scavenger k and the rate constant kOH of the reactions between scavengers and •OH radicals. Our results suggest that the studied scavengers effectively protected yeast and bacteria against ionizing radiation. Although the scavenging of •OH radicals seems to be important for protection of living cells, it is clearly not the only process on which the protection is based.

  18. Molecular cloning of the gene for the human placental GTP-binding protein Gp (G25K): Identification of this GTP-binding protein as the human homolog of the yeast cell-division-cycle protein CDC42

    International Nuclear Information System (INIS)

    Shinjo, K.; Koland, J.G.; Hart, M.J.; Narasimhan, V.; Cerione, R.A.; Johnson, D.I.; Evans, T.

    1990-01-01

    The authors have isolated cDNA clones from a human placental library that code for a low molecular weight GTP-binding protein originally designated G p (also called G25K). This identification is based on comparisons with the available peptide sequences for the purified human G p protein and the use of two highly specific anti-peptide antibodies. The predicted amino acid sequence of the protein is very similar to those of various members of the ras superfamily of low molecular weight GTP-binding proteins, including the N-, Ki-, and Ha-ras proteins (30-35% identical), the rho proteins and the rac proteins. The highest degree of sequence identity (80%) is found with the Saccharomyces cerevisiae cell division-cycle protein CDC42. The human placental gene, which they designate CDC42Hs, complements the cdc42-1 mutation in S. cerevisiae, which suggests that this GTP-binding protein is the human homolog of the yeast protein

  19. Novel brewing yeast hybrids: creation and application.

    Science.gov (United States)

    Krogerus, Kristoffer; Magalhães, Frederico; Vidgren, Virve; Gibson, Brian

    2017-01-01

    The natural interspecies Saccharomyces cerevisiae × Saccharomyces eubayanus hybrid yeast is responsible for global lager beer production and is one of the most important industrial microorganisms. Its success in the lager brewing environment is due to a combination of traits not commonly found in pure yeast species, principally low-temperature tolerance, and maltotriose utilization. Parental transgression is typical of hybrid organisms and has been exploited previously for, e.g., the production of wine yeast with beneficial properties. The parental strain S. eubayanus has only been discovered recently and newly created lager yeast strains have not yet been applied industrially. A number of reports attest to the feasibility of this approach and artificially created hybrids are likely to have a significant impact on the future of lager brewing. De novo S. cerevisiae × S. eubayanus hybrids outperform their parent strains in a number of respects, including, but not restricted to, fermentation rate, sugar utilization, stress tolerance, and aroma formation. Hybrid genome function and stability, as well as different techniques for generating hybrids and their relative merits are discussed. Hybridization not only offers the possibility of generating novel non-GM brewing yeast strains with unique properties, but is expected to aid in unraveling the complex evolutionary history of industrial lager yeast.

  20. Making Sense of the Yeast Sphingolipid Pathway.

    Science.gov (United States)

    Megyeri, Márton; Riezman, Howard; Schuldiner, Maya; Futerman, Anthony H

    2016-12-04

    Sphingolipids (SL) and their metabolites play key roles both as structural components of membranes and as signaling molecules. Many of the key enzymes and regulators of SL metabolism were discovered using the yeast Saccharomyces cerevisiae, and based on the high degree of conservation, a number of mammalian homologs were identified. Although yeast continues to be an important tool for SL research, the complexity of SL structure and nomenclature often hampers the ability of new researchers to grasp the subtleties of yeast SL biology and discover new modulators of this intricate pathway. Moreover, the emergence of lipidomics by mass spectrometry has enabled the rapid identification of SL species in yeast and rendered the analysis of SL composition under various physiological and pathophysiological conditions readily amenable. However, the complex nomenclature of the identified species renders much of the data inaccessible to non-specialists. In this review, we focus on parsing both the classical SL nomenclature and the nomenclature normally used during mass spectrometry analysis, which should facilitate the understanding of yeast SL data and might shed light on biological processes in which SLs are involved. Finally, we discuss a number of putative roles of various yeast SL species. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Improving industrial yeast strains: exploiting natural and artificial diversity

    Science.gov (United States)

    Steensels, Jan; Snoek, Tim; Meersman, Esther; Nicolino, Martina Picca; Voordeckers, Karin; Verstrepen, Kevin J

    2014-01-01

    Yeasts have been used for thousands of years to make fermented foods and beverages, such as beer, wine, sake, and bread. However, the choice for a particular yeast strain or species for a specific industrial application is often based on historical, rather than scientific grounds. Moreover, new biotechnological yeast applications, such as the production of second-generation biofuels, confront yeast with environments and challenges that differ from those encountered in traditional food fermentations. Together, this implies that there are interesting opportunities to isolate or generate yeast variants that perform better than the currently used strains. Here, we discuss the different strategies of strain selection and improvement available for both conventional and nonconventional yeasts. Exploiting the existing natural diversity and using techniques such as mutagenesis, protoplast fusion, breeding, genome shuffling and directed evolution to generate artificial diversity, or the use of genetic modification strategies to alter traits in a more targeted way, have led to the selection of superior industrial yeasts. Furthermore, recent technological advances allowed the development of high-throughput techniques, such as ‘global transcription machinery engineering’ (gTME), to induce genetic variation, providing a new source of yeast genetic diversity. PMID:24724938

  2. WdStuAp, an APSES transcription factor, is a regulator of yeast-hyphal transitions in Wangiella (Exophiala) dermatitidis.

    Science.gov (United States)

    Wang, Qin; Szaniszlo, Paul J

    2007-09-01

    APSES transcription factors are well-known regulators of fungal cellular development and differentiation. To study the function of an APSES protein in the fungus Wangiella dermatitidis, a conidiogenous and polymorphic agent of human phaeohyphomycosis with yeast predominance, the APSES transcription factor gene WdSTUA was cloned, sequenced, disrupted, and overexpressed. Analysis showed that its derived protein was most similar to the APSES proteins of other conidiogenous molds and had its APSES DNA-binding domain located in the amino-terminal half. Deletion of WdSTUA in W. dermatitidis induced convoluted instead of normal smooth colony surface growth on the rich yeast maintenance agar medium yeast extract-peptone-dextrose agar (YPDA) at 37 degrees C. Additionally, deletion of WdSTUA repressed aerial hyphal growth, conidiation, and invasive hyphal growth on the nitrogen-poor, hypha-inducing agar medium potato dextrose agar (PDA) at 25 degrees C. Ectopic overexpression of WdSTUA repressed the convoluted colony surface growth on YPDA at 37 degrees C, and also strongly repressed hyphal growth on PDA at 25 degrees C and 37 degrees C. These new results provide additional insights into the diverse roles played by APSES factors in fungi. They also suggest that the transcription factor encoded by WdSTUA is both a positive and negative morphotype regulator in W. dermatitidis and possibly other of the numerous human pathogenic, conidiogenous fungi capable of yeast growth.

  3. Protein patterns of yeast during sporulation

    International Nuclear Information System (INIS)

    Litske Petersen, J.G.; Kielland-Brandt, M.C.; Nilsson-Tillgren, T.

    1979-01-01

    High resolution two-dimensional gel electrophoresis was used to study protein synthesis during synchronous meiosis and ascospore formation of Saccharomyces cerevisiae. The stained protein patterns of samples harvested at any stage between meiotic prophase and the four-spore stage in two sporulating strains showed the same approximately 250 polypeptides. Of these only a few seemed to increase or decrease in concentration during sporulation. The characteristic pattern of sporulating yeast was identical to the pattern of glucose-grown staitonary yeast cells adapted to respiration. The latter type of cells readily initiates meiosis when transferred to sporulation medium. This pattern differed from the protein patterns of exponentially growing cells in glucose or acetate presporulation medium. Five major proteins in stationary and sporulating yeast cells were not detected in either type of exponential culture. Two-dimensional autoradiograms of [ 35 S]methionine-labelled yeast proteins revealed that some proteins were preferentially labelled during sporulation, while other proteins were labelled at later stages. These patterns differed from the auroradiograms of exponentially growing yeast cells in glucose presporulation medium in a number of spots. No differences were observed when stained gels or autoradiograms of sporulating cultures and non-sporulating strains in sporulation medium were compared. (author)

  4. Isolation and identification of radiation resistant yeasts from sea water

    International Nuclear Information System (INIS)

    Park, Jong Cheon; Jeong, Yong Uk; Kim, Du Hong; Jo, Eun A

    2011-12-01

    This study was conducted to isolate radiation-resistant yeasts from sea water for development of application technology of radiation-resistant microorganism. · Isolation of 656 yeasts from sea water and selection of 2 radiation-resistant yeasts (D 10 value >3) · Identification of isolated yeasts as Filobasidium elegans sharing 99% sequence similarity · Characterization of isolated yeast with ability to repair of the DNA damage and membrane integrity to irradiation

  5. Malassezia vespertilionis sp. nov.: a new cold-tolerant species of yeast isolated from bats

    NARCIS (Netherlands)

    Lorch, J.M.; Palmer, J.M.; Vanderwolf, K.J.; Schmidt, K.Z.; Verant, M.L.; Weller, T.J.; Blehert, D.S.

    2018-01-01

    Malassezia is a genus of medically-important, lipid-dependent yeasts that live on the skin of warmblooded animals. The 17 described species have been documented primarily on humans and domestic animals, but few studies have examined Malassezia species associated with more diverse host groups such as

  6. Identification of salivary components that induce transition of hyphae to yeast in Candida albicans

    NARCIS (Netherlands)

    Leito, J.T.D.; Ligtenberg, A.J.M.; Nazmi, K.; Veerman, E.C.I.

    2009-01-01

    Candida albicans, the major human fungal pathogen, undergoes a reversible morphological transition from single yeast cells to pseudohyphae and hyphae filaments. The hyphae form is considered the most invasive form of the fungus. The purpose of this study is to investigate the effect of saliva on

  7. Evolutionary conservation of nuclear and nucleolar targeting sequences in yeast ribosomal protein S6A

    International Nuclear Information System (INIS)

    Lipsius, Edgar; Walter, Korden; Leicher, Torsten; Phlippen, Wolfgang; Bisotti, Marc-Angelo; Kruppa, Joachim

    2005-01-01

    Over 1 billion years ago, the animal kingdom diverged from the fungi. Nevertheless, a high sequence homology of 62% exists between human ribosomal protein S6 and S6A of Saccharomyces cerevisiae. To investigate whether this similarity in primary structure is mirrored in corresponding functional protein domains, the nuclear and nucleolar targeting signals were delineated in yeast S6A and compared to the known human S6 signals. The complete sequence of S6A and cDNA fragments was fused to the 5'-end of the LacZ gene, the constructs were transiently expressed in COS cells, and the subcellular localization of the fusion proteins was detected by indirect immunofluorescence. One bipartite and two monopartite nuclear localization signals as well as two nucleolar binding domains were identified in yeast S6A, which are located at homologous regions in human S6 protein. Remarkably, the number, nature, and position of these targeting signals have been conserved, albeit their amino acid sequences have presumably undergone a process of co-evolution with their corresponding rRNAs

  8. Potentially pathogenic yeasts from soil of children’s recreational areas in the city of Łódź (Poland

    Directory of Open Access Journals (Sweden)

    Anna Wójcik

    2013-06-01

    Full Text Available Objectives: Yeasts may become potential human and animal pathogens, particularly for individuals with a depressed immune system. Their presence in the environment, especially in soil, may favour their spread into human ontocenoses. Materials and Methods: Eighty-four soil samples obtained from 21 children's recreational sites in Łódź in autumn 2010 and spring 2011 were evaluated. The yeasts were isolated by classical microbiological methods and identified on the basis of morphological and biochemical features. Results: The fungi were found in 73.8% and in 69.0% of the examined samples collected in autumn and spring, respectively. Among 97 isolates of yeasts, the species potentially pathogenic to humans and animals were Candida colliculosa, C. guilliermondii, C. humicola, C. inconspicua, C. lambica, C. lusitaniae, C. pelliculosa, C. tropicalis, Cryptococcus albidus, C. laurentii, C. neoformans, C. terreus, Kloeckera japonica, Geotrichum candidum, G. penicillatum, Rhodotorula mucilaginosa, R. glutinis, Saccharomyces cerevisiae, Sporobolomyces salmonicolor and Trichosporon cutaneum. The most frequently isolated fungi included the genus Cryptococcus (38 isolates and two species: Rhodotorula glutinis (15, Trichosporon cutaneum (14. C. neoformans, an etiological factor of cryptococcal meningitis, was present in the sandpits of 3 kindergartens. The Candida species were identified from park playgrounds and school sports fields mainly in autumn 2010 (14 isolates, in spring 2011 - only 1 isolate. The concentration of fungal species in particular samples varied considerably, but in the majority of samples, fungi were present at concentration of up to 1×102 CFU/1 g of soil. Conclusions: Yeasts were present in the soil of parks, schools and kindergarten recreational areas; the fact may pose a health risk to humans, especially to children, and this type of biological pollution should be regarded as a potential public health concern.

  9. Expression of Human CTP Synthetase in Saccharomyces cerevisiae Reveals Phosphorylation by Protein Kinase A*

    Science.gov (United States)

    Han, Gil-Soo; Sreenivas, Avula; Choi, Mal-Gi; Chang, Yu-Fang; Martin, Shelley S.; Baldwin, Enoch P.; Carman, George M.

    2005-01-01

    CTP synthetase (EC 6.3.4.2, UTP: ammonia ligase (ADP-forming)) is an essential enzyme in all organisms; it generates the CTP required for the synthesis of nucleic acids and membrane phospholipids. In this work we showed that the human CTP synthetase genes, CTPS1 and CTPS2, were functional in Saccharomyces cerevisiae and complemented the lethal phenotype of the ura7Δ ura8Δ mutant lacking CTP synthetase activity. The expression of the CTPS1-and CTPS2-encoded human CTP synthetase enzymes in the ura7Δ ura8Δ mutant was shown by immunoblot analysis of CTP synthetase proteins, the measurement of CTP synthetase activity, and the synthesis of CTP in vivo. Phosphoamino acid and phosphopeptide mapping analyses of human CTP synthetase 1 isolated from 32Pi-labeled cells revealed that the enzyme was phosphorylated on multiple serine residues in vivo. Activation of protein kinase A activity in yeast resulted in transient increases (2-fold) in the phosphorylation of human CTP synthetase 1 and the cellular level of CTP. Human CTP synthetase 1 was also phosphorylated by mammalian protein kinase A in vitro. Using human CTP synthetase 1 purified from Escherichia coli as a substrate, protein kinase A activity was dose- and time-dependent, and dependent on the concentrations of CTP synthetase1 and ATP. These studies showed that S. cerevisiae was useful for the analysis of human CTP synthetase phosphorylation. PMID:16179339

  10. The yeast spectrum of the 'tea fungus Kombucha'.

    Science.gov (United States)

    Mayser, P; Fromme, S; Leitzmann, C; Gründer, K

    1995-01-01

    The tea fungus 'Kombucha' is a symbiosis of Acetobacter, including Acetobacter xylinum as a characteristic species, and various yeasts. A characteristic yeast species or genus has not yet been identified. Kombucha is mainly cultivated in sugared black tea to produce a slightly acidulous effervescent beverage that is said to have several curative effects. In addition to sugar, the beverage contains small amounts of alcohol and various acids, including acetic acid, gluconic acid and lactic acid, as well as some antibiotic substances. To characterize the yeast spectrum with special consideration given to facultatively pathogenic yeasts, two commercially available specimens of tea fungus and 32 from private households in Germany were analysed by micromorphological and biochemical methods. Yeasts of the genera Brettanomyces, Zygosaccharomyces and Saccharomyces were identified in 56%, 29% and 26% respectively. The species Saccharomycodes ludwigii and Candida kefyr were only demonstrated in isolated cases. Furthermore, the tests revealed pellicle-forming yeasts such as Candida krusei or Issatchenkia orientalis/occidentalis as well as species of the apiculatus yeasts (Kloeckera, Hanseniaspora). Thus, the genus Brettanomyces may be a typical group of yeasts that are especially adapted to the environment of the tea fungus. However, to investigate further the beneficial effects of tea fungus, a spectrum of the other typical genera must be defined. Only three specimens showed definite contaminations. In one case, no yeasts could be isolated because of massive contamination with Penicillium spp. In the remaining two samples (from one household), Candida albicans was demonstrated. The low rate of contamination might be explained by protective mechanisms, such as formation of organic acids and antibiotic substances. Thus, subjects with a healthy metabolism do not need to be advised against cultivating Kombucha. However, those suffering from immunosuppression should preferably

  11. 21 CFR 573.750 - Pichia pastoris dried yeast.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Pichia pastoris dried yeast. 573.750 Section 573... Food Additive Listing § 573.750 Pichia pastoris dried yeast. (a) Identity. The food additive Pichia pastoris dried yeast may be used in feed formulations of broiler chickens as a source of protein not to...

  12. Teaching Microbial Physiology Using Glucose Repression Phenomenon in Baker's Yeast as an Example

    Science.gov (United States)

    Raghevendran, Vijayendran; Nielsen, Jens; Olsson, Lisbeth

    2005-01-01

    The yeast "Saccharomyces cerevisiae" has been used by human beings since ancient times for its ability to convert sugar to alcohol. Continual exposure to glucose in the natural environment for innumerable generations has probably enabled "S. cerevisiae" to grow in fermentative mode on sugars by switching off the genes responsible for respiration…

  13. Yeast signaling pathways in the oxidative stress response

    Energy Technology Data Exchange (ETDEWEB)

    Ikner, Aminah [Section of Microbiology, Division of Biological Sciences, University of California, Davis, CA 95616 (United States); Shiozaki, Kazuhiro [Section of Microbiology, Division of Biological Sciences, University of California, Davis, CA 95616 (United States)]. E-mail: kshiozaki@ucdavis.edu

    2005-01-06

    Oxidative stress that generates the reactive oxygen species (ROS) is one of the major causes of DNA damage and mutations. The 'DNA damage checkpoint' that arrests cell cycle and repairs damaged DNA has been a focus of recent studies, and the genetically amenable model systems provided by yeasts have been playing a leading role in the eukaryotic checkpoint research. However, means to eliminate ROS are likely to be as important as the DNA repair mechanisms in order to suppress mutations in the chromosomal DNA, and yeasts also serve as excellent models to understand how eukaryotes combat oxidative stress. In this article, we present an overview of the signaling pathways that sense oxidative stress and induce expression of various anti-oxidant genes in the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe and the pathogenic yeast Candida albicans. Three conserved signaling modules have been identified in the oxidative stress response of these diverse yeast species: the stress-responsive MAP kinase cascade, the multistep phosphorelay and the AP-1-like transcription factor. The structure and function of these signaling modules are discussed.

  14. Yeast signaling pathways in the oxidative stress response

    International Nuclear Information System (INIS)

    Ikner, Aminah; Shiozaki, Kazuhiro

    2005-01-01

    Oxidative stress that generates the reactive oxygen species (ROS) is one of the major causes of DNA damage and mutations. The 'DNA damage checkpoint' that arrests cell cycle and repairs damaged DNA has been a focus of recent studies, and the genetically amenable model systems provided by yeasts have been playing a leading role in the eukaryotic checkpoint research. However, means to eliminate ROS are likely to be as important as the DNA repair mechanisms in order to suppress mutations in the chromosomal DNA, and yeasts also serve as excellent models to understand how eukaryotes combat oxidative stress. In this article, we present an overview of the signaling pathways that sense oxidative stress and induce expression of various anti-oxidant genes in the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe and the pathogenic yeast Candida albicans. Three conserved signaling modules have been identified in the oxidative stress response of these diverse yeast species: the stress-responsive MAP kinase cascade, the multistep phosphorelay and the AP-1-like transcription factor. The structure and function of these signaling modules are discussed

  15. [Yeast species in vulvovaginitis candidosa].

    Science.gov (United States)

    Nemes-Nikodém, Éva; Tamási, Béla; Mihalik, Noémi; Ostorházi, Eszter

    2015-01-04

    Vulvovaginal candidiasis is the most common mycosis, however, the available information about antifungal susceptibilities of these yeasts is limited. To compare the gold standard fungal culture with a new molecular identification method and report the incidence of yeast species in vulvovaginitis candidosa. The authors studied 370 yeasts isolated from vulvovaginal candidiasis and identified them by phenotypic and molecular methods. The most common species was Candida albicans (85%), followed by Candida glabrata, and other Candida species. At present there are no recommendations for the evaluation of antifungal susceptibility of pathogenic fungal species occurring in vulvovaginal candidiasis and the natural antifungal resistance of the different species is known only. Matrix Assisted Laser Desorption Ionization Time of Flight identification can be used to differentiate the fluconazole resistant Candida dubliniensis and the sensitive Candida albicans strains.

  16. Escherichia coli O157:H7 - An Emerging Pathogen in foods of Animal Origin

    Directory of Open Access Journals (Sweden)

    Ch. Bindu Kiranmayi

    Full Text Available Escherichia coli O157:H7 is an emerging public health concern in most countries of the world. E. coli O157:H7 was known to be a human pathogen for nearly 24 years. EHEC O157 infection is estimated to be the fourth most costly food borne disease in Canada and USA, not counting the cost of possible litigation. E. coli O157:H7 and Salmonella are the leading causes of produce related outbreaks, accounting for 20 and 30% respectively. The authority of the Federal Meat Inspection Act, FSIS (Food Safety and Inspection Service declared Escherichia coli O157:H7, an adulterant in raw ground beef and enforced “zero tolerance” (USDA-FSIS, 17 December 1998. Because of the severity of these illnesses and the apparent low infective dose (less than 10 cells, Escherichia coli O157:H7 is considered one of the most serious of known food borne pathogens. Escherichia coli O157:H7 is mainly pathogenic to human but in cattle and other animals, it did not induce any clinical disease except diarrhea. So, these animals act as carriers to Escherichia coli O157:H7. The majority transmission is through eating of undercooked contaminated ground meat and consumption of raw milk, raw vegetables, fruits contaminated by water, cheese, curd and also through consumption of sprouts, lettuce and juice. The conventional isolation procedure includes growth in enrichment broth like modified EC (E. coli broth or modified tryptic soy broth (mTSB Since the infection primarily occurs via faeco-oral route, the preventive measures include food hygiene measures like proper cooking of meat, consumption of pasteurized milk, washing fruits and vegetables especially those to be eaten raw and drinking chlorine treated water and personnel hygiene measures like washing hands after toilet visits. [Veterinary World 2010; 3(8.000: 382-389

  17. Antimicrobial Activity of Endemic Herbs from Tangkahan Conservation Forest North Sumatera to Bacteria and Yeast

    Directory of Open Access Journals (Sweden)

    KIKI NURTJAHJA

    2013-12-01

    Full Text Available Tangkahan Conservation Forest in Karo County, North Sumatera has high biodiversity of endemic herbs. Many species of the wild herbs are well known used as traditional medicine not only by local people but also by people out of the area. The methanol extract of the medicinal wild herbs in Tangkahan Conservation Forest, Karo County to relief skin diseases caused by bacteria and fungi never been studied medically. The antimicrobial activity leave extract of the medicinal herbs to pathogenic microorganisms are studied. The leaves extract of kembu-kembu (Callicarpa candicans, rintih bulung (Piper muricatum, cep-cepan (Castanopsis costata, and sereh kayu (Eugenia grandis, has antimicrobial to bacteria (Bacillus sp., Escherichia coli, Serratia marcescens, Staphylococcus aureus and yeast (Candida albicans. Toxicity assay of these plants by brine shrimp method using Artemia salina indicates that cep-cepan dan sereh kayu have lethal concentration higher than kembu-kembu and rintih bulung.

  18. Transglycosylation properties of maltodextrin glucosidase (MalZ) from Escherichia coli and its application for synthesis of a nigerose-containing oligosaccharide

    Energy Technology Data Exchange (ETDEWEB)

    Song, Kyung-Mo [Center for Agricultural Biomaterials and Department of Food Science and Biotechnology, Seoul National University, Seoul 151-921 (Korea, Republic of); Shim, Jae-Hoon [Department of Biology, University of Incheon, Incheon 406-772 (Korea, Republic of); Park, Jong-Tae; Kim, Sung-Hee [Center for Agricultural Biomaterials and Department of Food Science and Biotechnology, Seoul National University, Seoul 151-921 (Korea, Republic of); Kim, Young-Wan [Department of Food and Biotechnology, Korea University, Jochiwon 339-700 (Korea, Republic of); Boos, Winfried [Department of Biology, University of Konstanz, Konstanz 78457 (Germany); Park, Kwan-Hwa, E-mail: parkkh@incheon.ac.kr [Department of Biology, University of Incheon, Incheon 406-772 (Korea, Republic of)

    2010-06-18

    The transglycosylation reaction of maltodextrin glucosidase (MalZ) cloned and purified from Escherichia coli K12 was characterized and applied to the synthesis of branched oligosaccharides. Purified MalZ preferentially catalyzed the hydrolysis of maltodextrin, {gamma}-cyclodextrin (CD), and cycloamylose (CA). In addition, when the enzyme was incubated with 5% maltotriose (G3), a series of transfer products were produced. The resulting major transfer products, annotated as T1, T2, and T3, were purified and their structures were determined by TLC, MALDI-TOF/MS, {sup 13}C NMR, and enzymatic analysis. T1 was identified as a novel compound, maltosyl {alpha}-1,3-maltose, whereas T2 and T3 were determined to be isopanose and maltosyl-{alpha}-1,6-maltose, respectively. These results indicated that MalZ transferred sugar moiety mainly to C-3 or C-6-OH of glucose of the acceptor molecule. To obtain highly concentrated transfer products, the enzyme was reacted with 10% liquefied cornstarch, and then glucose and maltose were removed by immobilized yeast. The T1 content of the resulting reaction mixture reached 9.0%. The mixture of T1 containing a nigerose moiety can have an immunopotentiating effect on the human body and may be a potential functional sugar stuff.

  19. OCCURRENCE OF ANTIBIOTIC-RESISTANT UROPATHOGENIC ESCHERICHIA COLI CLONAL GROUP A IN WASTEWATER EFFLUENTS

    Science.gov (United States)

    Isolates of Escherichia coli belonging to clonal group A (CGA), a recently described disseminated cause of drug-resistant urinary tract infections in humans, were present in four of seven sewage effluents collected from geographically dispersed areas of the United States. ...

  20. Detection of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Market-Ready Chickens in Zambia.

    Science.gov (United States)

    Chishimba, K; Hang'ombe, B M; Muzandu, K; Mshana, S E; Matee, M I; Nakajima, C; Suzuki, Y

    2016-01-01

    The frequent administering of antibiotics in the treatment of poultry diseases may contribute to emergence of antimicrobial-resistant strains. The objective of this study was to detect the presence of extended-spectrum β-lactamase- (ESBL-) producing Escherichia coli in poultry in Zambia. A total of 384 poultry samples were collected and analyzed for ESBL-producing Escherichia coli. The cultured E. coli isolates were subjected to antimicrobial susceptibility tests and the polymerase chain reaction for detection of bla CTX-M, bla SHV, and bla TEM genes. Overall 20.1%, 77/384, (95% CI; 43.2-65.5%) of total samples analyzed contained ESBL-producing Escherichia coli. The antimicrobial sensitivity test revealed that 85.7% (66/77; CI: 75.7-92) of ESBL-producing E. coli isolates conferred resistance to beta-lactam and other antimicrobial agents. These results indicate that poultry is a potential reservoir for ESBL-producing Escherichia coli. The presence of ESBL-producing Escherichia coli in poultry destined for human consumption requires strengthening of the antibiotic administering policy. This is important as antibiotic administration in food animals is gaining momentum for improved animal productivity in developing countries such as Zambia.

  1. Detection of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Market-Ready Chickens in Zambia

    Directory of Open Access Journals (Sweden)

    K. Chishimba

    2016-01-01

    Full Text Available The frequent administering of antibiotics in the treatment of poultry diseases may contribute to emergence of antimicrobial-resistant strains. The objective of this study was to detect the presence of extended-spectrum β-lactamase- (ESBL- producing Escherichia coli in poultry in Zambia. A total of 384 poultry samples were collected and analyzed for ESBL-producing Escherichia coli. The cultured E. coli isolates were subjected to antimicrobial susceptibility tests and the polymerase chain reaction for detection of blaCTX-M, blaSHV, and blaTEM genes. Overall 20.1%, 77/384, (95% CI; 43.2–65.5% of total samples analyzed contained ESBL-producing Escherichia coli. The antimicrobial sensitivity test revealed that 85.7% (66/77; CI: 75.7–92 of ESBL-producing E. coli isolates conferred resistance to beta-lactam and other antimicrobial agents. These results indicate that poultry is a potential reservoir for ESBL-producing Escherichia coli. The presence of ESBL-producing Escherichia coli in poultry destined for human consumption requires strengthening of the antibiotic administering policy. This is important as antibiotic administration in food animals is gaining momentum for improved animal productivity in developing countries such as Zambia.

  2. Functional analysis of human aromatic amino acid transporter MCT10/TAT1 using the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Uemura, Satoshi; Mochizuki, Takahiro; Kurosaka, Goyu; Hashimoto, Takanori; Masukawa, Yuki; Abe, Fumiyoshi

    2017-10-01

    Tryptophan is an essential amino acid in humans and an important serotonin and melatonin precursor. Monocarboxylate transporter MCT10 is a member of the SLC16A family proteins that mediates low-affinity tryptophan transport across basolateral membranes of kidney, small intestine, and liver epithelial cells, although the precise transport mechanism remains unclear. Here we developed a simple functional assay to analyze tryptophan transport by human MCT10 using a deletion mutant for the high-affinity tryptophan permease Tat2 in Saccharomyces cerevisiae. tat2Δtrp1 cells are defective in growth in YPD medium because tyrosine present in the medium competes for the low-affinity tryptophan permease Tat1 with tryptophan. MCT10 appeared to allow growth of tat2Δtrp1 cells in YPD medium, and accumulate in cells deficient for Rsp5 ubiquitin ligase. These results suggest that MCT10 is functional in yeast, and is subject to ubiquitin-dependent quality control. Whereas growth of Tat2-expressing cells was significantly impaired by neutral pH, that of MCT10-expressing cells was nearly unaffected. This property is consistent with the transport mechanism of MCT10 via facilitated diffusion without a need for pH gradient across the plasma membrane. Single-nucleotide polymorphisms (SNPs) are known to occur in the human MCT10 coding region. Among eight SNP amino acid changes in MCT10, the N81K mutation completely abrogated tryptophan import without any abnormalities in the expression or localization. In the MCT10 modeled structure, N81 appeared to protrude into the putative trajectory of tryptophan. Plasma membrane localization of MCT10 and the variant proteins was also verified in human embryonic kidney 293T cells. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    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.

  4. Escherichia coli O104 associated with human diarrhea, South Africa, 2004-2011.

    Science.gov (United States)

    Tau, Nomsa P; Meidany, Parastu; Smith, Anthony M; Sooka, Arvinda; Keddy, Karen H

    2012-08-01

    To determine the origin of >4,000 suspected diarrheagenic Escherichia coli strains isolated during 2004-2011 in South Africa, we identified 7 isolates as serotype O104; 5 as enteroaggregative E. coli O104:H4, and 2 as enteropathogenic E. coli O104:non-H4. Pulsed-field gel electrophoresis showed that these isolates were unrelated to the 2011 E. coli O104:H4 outbreak strain from Germany.

  5. An Evidenced-Based Scale of Disease Severity following Human Challenge with Enteroxigenic Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Chad K Porter

    Full Text Available Experimental human challenge models have played a major role in enhancing our understanding of infectious diseases. Primary outcomes have typically utilized overly simplistic outcomes that fail to entirely account for complex illness syndromes. We sought to characterize clinical outcomes associated with experimental infection with enterotoxigenic Escherichia coli (ETEC and to develop a disease score.Data were obtained from prior controlled human ETEC infection studies. Correlation and univariate regression across sign and symptom severity was performed. A multiple correspondence analysis was conducted. A 3-parameter disease score with construct validity was developed in an iterative fashion, compared to standard outcome definitions and applied to prior vaccine challenge trials.Data on 264 subjects receiving seven ETEC strains at doses from 1x105 to 1x1010 cfu were used to construct a standardized dataset. The strongest observed correlation was between vomiting and nausea (r = 0.65; however, stool output was poorly correlated with subjective activity-impacting outcomes. Multiple correspondence analyses showed covariability in multiple signs and symptoms, with severity being the strongest factor corresponding across outcomes. The developed disease score performed well compared to standard outcome definitions and differentiated disease in vaccinated and unvaccinated subjects.Frequency and volumetric definitions of diarrhea severity poorly characterize ETEC disease. These data support a disease severity score accounting for stool output and other clinical signs and symptoms. Such a score could serve as the basis for better field trial outcomes and gives an additional outcome measure to help select future vaccines that warrant expanded testing in pivotal pre-licensure trials.

  6. Kinetics of growth and sugar consumption in yeasts.

    Science.gov (United States)

    van Dijken, J P; Weusthuis, R A; Pronk, J T

    1993-01-01

    An overview is presented of the steady- and transient state kinetics of growth and formation of metabolic byproducts in yeasts. Saccharomyces cerevisiae is strongly inclined to perform alcoholic fermentation. Even under fully aerobic conditions, ethanol is produced by this yeast when sugars are present in excess. This so-called 'Crabtree effect' probably results from a multiplicity of factors, including the mode of sugar transport and the regulation of enzyme activities involved in respiration and alcoholic fermentation. The Crabtree effect in S. cerevisiae is not caused by an intrinsic inability to adjust its respiratory activity to high glycolytic fluxes. Under certain cultivation conditions, for example during growth in the presence of weak organic acids, very high respiration rates can be achieved by this yeast. S. cerevisiae is an exceptional yeast since, in contrast to most other species that are able to perform alcoholic fermentation, it can grow under strictly anaerobic conditions. 'Non-Saccharomyces' yeasts require a growth-limiting supply of oxygen (i.e. oxygen-limited growth conditions) to trigger alcoholic fermentation. However, complete absence of oxygen results in cessation of growth and therefore, ultimately, of alcoholic fermentation. Since it is very difficult to reproducibly achieve the right oxygen dosage in large-scale fermentations, non-Saccharomyces yeasts are therefore not suitable for large-scale alcoholic fermentation of sugar-containing waste streams. In these yeasts, alcoholic fermentation is also dependent on the type of sugar. For example, the facultatively fermentative yeast Candida utilis does not ferment maltose, not even under oxygen-limited growth conditions, although this disaccharide supports rapid oxidative growth.

  7. Binding properties of SUMO-interacting motifs (SIMs) in yeast.

    Science.gov (United States)

    Jardin, Christophe; Horn, Anselm H C; Sticht, Heinrich

    2015-03-01

    Small ubiquitin-like modifier (SUMO) conjugation and interaction play an essential role in many cellular processes. A large number of yeast proteins is known to interact non-covalently with SUMO via short SUMO-interacting motifs (SIMs), but the structural details of this interaction are yet poorly characterized. In the present work, sequence analysis of a large dataset of 148 yeast SIMs revealed the existence of a hydrophobic core binding motif and a preference for acidic residues either within or adjacent to the core motif. Thus the sequence properties of yeast SIMs are highly similar to those described for human. Molecular dynamics simulations were performed to investigate the binding preferences for four representative SIM peptides differing in the number and distribution of acidic residues. Furthermore, the relative stability of two previously observed alternative binding orientations (parallel, antiparallel) was assessed. For all SIMs investigated, the antiparallel binding mode remained stable in the simulations and the SIMs were tightly bound via their hydrophobic core residues supplemented by polar interactions of the acidic residues. In contrary, the stability of the parallel binding mode is more dependent on the sequence features of the SIM motif like the number and position of acidic residues or the presence of additional adjacent interaction motifs. This information should be helpful to enhance the prediction of SIMs and their binding properties in different organisms to facilitate the reconstruction of the SUMO interactome.

  8. Performance of non-conventional yeasts in co-culture with brewers’ yeast for steering ethanol and aroma production

    NARCIS (Netherlands)

    Rijswijck, van Irma M.H.; Wolkers - Rooijackers, Judith C.M.; Abee, Tjakko; Smid, Eddy J.

    2017-01-01

    Increasing interest in new beer types has stimulated the search for approaches to extend the metabolic variation of brewers’ yeast. Therefore, we tested two approaches using non-conventional yeast to create a beer with lower ethanol content and a complex aroma bouquet. First, the mono-culture

  9. Molecular cloning and characterization of genes required for nucleotide excision repair in yeast

    International Nuclear Information System (INIS)

    Friedberg, E.C.

    1987-01-01

    Nucleotide excision repair in the yeast S. cerevisiae is a complex process which involves a large number of genes. At least five of these genes (RAD1, RAD2, RAD3, RAD4 and RAD10) are absolutely required for this process and mutations in any of these genes result in no detectable excision repair in vivo. In order to understand the function of these genes in DNA repair, the authors isolated a number of them by screening a yeast genomic library for recombinant plasmids which complement the phentoype of sensitivity to ultraviolet (UV) radiation imparted to mutant strains. A plasmid containing the RAD4 gene was isolated by an alternative strategy which will be discussed. The cloned genes have been extensively characterized. It has been determined that the RAD3 gene is essential for the viability of haploid yeast cells in the absence of DNA damage. The RAD2 gene is inducible by treatment of cells with a variety of DNA-damaging agents, including UV radiation and ionizing radiation. The RAD10 gene shares considerable amino acid sequence homology with a cloned gene involved in nucleotide excision repair in human cells. Yeast is a particularly versatile organism for studying gene function by molecular and genetic approaches and emphasis is placed on many of the techniques used in the present studies

  10. The ecology of the Drosophila-yeast mutualism in wineries

    Science.gov (United States)

    2018-01-01

    The fruit fly, Drosophila melanogaster, is preferentially found on fermenting fruits. The yeasts that dominate the microbial communities of these substrates are the primary food source for developing D. melanogaster larvae, and adult flies manifest a strong olfactory system-mediated attraction for the volatile compounds produced by these yeasts during fermentation. Although most work on this interaction has focused on the standard laboratory yeast Saccharomyces cerevisiae, a wide variety of other yeasts naturally ferment fallen fruit. Here we address the open question of whether D. melanogaster preferentially associates with distinct yeasts in different, closely-related environments. We characterized the spatial and temporal dynamics of Drosophila-associated fungi in Northern California wineries that use organic grapes and natural fermentation using high-throughput, short-amplicon sequencing. We found that there is nonrandom structure in the fungal communities that are vectored by flies both between and within vineyards. Within wineries, the fungal communities associated with flies in cellars, fermentation tanks, and pomace piles are distinguished by varying abundances of a small number of yeast species. To investigate the origins of this structure, we assayed Drosophila attraction to, oviposition on, larval development in, and longevity when consuming the yeasts that distinguish vineyard microhabitats from each other. We found that wild fly lines did not respond differentially to the yeast species that distinguish winery habitats in habitat specific manner. Instead, this subset of yeast shares traits that make them attractive to and ensure their close association with Drosophila. PMID:29768432

  11. Mechanisms of antibiotic resistance to enrofloxacin in uropathogenic Escherichia coli in dog

    Science.gov (United States)

    Escherichia coli (E. coli) urinary tract infections (UTIs) are becoming a serious problem both for pets and humans (zoonosis) due to the close contact and to the increasing resistance to antibiotics. Canine E. coli represents a good experimental model useful to study this pathology. Moreover, as des...

  12. Genotypic and phenotypic characterisation of Escherichia coli strains associated with porcine pyelonephritis

    DEFF Research Database (Denmark)

    Krag, L.; Hancock, Viktoria; Aalbæk, B.

    2009-01-01

    Urinary tract infection (UTI) is a severe problem in humans as well as in many domestic animals like pigs. The most frequent infectious agent in UTI is uropathogenic Escherichia coli. Such strains have been extensively characterised with respect to virulence and fitness factors as well as clonal...

  13. Improving industrial yeast strains: exploiting natural and artificial diversity.

    Science.gov (United States)

    Steensels, Jan; Snoek, Tim; Meersman, Esther; Picca Nicolino, Martina; Voordeckers, Karin; Verstrepen, Kevin J

    2014-09-01

    Yeasts have been used for thousands of years to make fermented foods and beverages, such as beer, wine, sake, and bread. However, the choice for a particular yeast strain or species for a specific industrial application is often based on historical, rather than scientific grounds. Moreover, new biotechnological yeast applications, such as the production of second-generation biofuels, confront yeast with environments and challenges that differ from those encountered in traditional food fermentations. Together, this implies that there are interesting opportunities to isolate or generate yeast variants that perform better than the currently used strains. Here, we discuss the different strategies of strain selection and improvement available for both conventional and nonconventional yeasts. Exploiting the existing natural diversity and using techniques such as mutagenesis, protoplast fusion, breeding, genome shuffling and directed evolution to generate artificial diversity, or the use of genetic modification strategies to alter traits in a more targeted way, have led to the selection of superior industrial yeasts. Furthermore, recent technological advances allowed the development of high-throughput techniques, such as 'global transcription machinery engineering' (gTME), to induce genetic variation, providing a new source of yeast genetic diversity. © 2014 The Authors. FEMS Microbiology Reviews published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

  14. Structural investigations of yeast mannans

    International Nuclear Information System (INIS)

    Rademacher, K.H.

    1983-01-01

    Cell wall mannans were isolated from 8 different Candida species and separated in oligosaccharides by partial acetolysis. After gel chromatography specific acetolysis patterns were obtained. The 13 C NMR spectra of mannans and oligosaccharides were recorded. Signals at delta = 93.1 - 105.4 were assigned to certain chemical structures. Both the spectral patterns and the acetolysis patterns of the yeast mannans can be used for the discrimination of related yeasts. (author)

  15. Activation of waste brewer's yeast Saccharomyces cerevisiae for bread production

    OpenAIRE

    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...

  16. PMAA-stabilized ferrofluid/chitosan/yeast composite for bioapplications

    International Nuclear Information System (INIS)

    Baldikova, Eva; Prochazkova, Jitka; Stepanek, Miroslav; Hajduova, Jana; Pospiskova, Kristyna; Safarikova, Mirka; Safarik, Ivo

    2017-01-01

    A simple, one-pot process for the preparation of magnetically responsive yeast-based biocatalysts was developed. Saccharomyces cerevisiae, Candida utilis and Kluyveromyces lactis cells were successfully incorporated into chitosan gel magnetically modified with poly(methacrylic acid)-stabilized magnetic fluid (PMAA-FF) during its formation. Magnetic PMAA-FF/chitosan/yeast composites were efficiently employed for invert sugar production. The dependence of invertase activity on used yeast, amount of magnetic biocatalyst, agitation time and after reuse was studied in detail. The tested magnetic biocatalysts retained at least 69% of their initial activity after 8 reuse cycles. - Highlights: • New types of magnetically responsive yeast biocomposites were prepared. • Recently developed PMAA-stabilized magnetic fluid was used. • Three yeast species were entrapped into magnetic chitosan gel during its formation. • All biocatalysts were efficiently employed for invert sugar formation.

  17. PMAA-stabilized ferrofluid/chitosan/yeast composite for bioapplications

    Energy Technology Data Exchange (ETDEWEB)

    Baldikova, Eva, E-mail: baldie@email.cz [Global Change Research Institute, CAS, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic); Department of Applied Chemistry, Faculty of Agriculture, University of South Bohemia, Branisovska 1457, 370 05 Ceske Budejovice (Czech Republic); Prochazkova, Jitka [Global Change Research Institute, CAS, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic); Stepanek, Miroslav; Hajduova, Jana [Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University, Hlavova 2030, 128 40 Prague 2 (Czech Republic); Pospiskova, Kristyna [Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 27, 783 71 Olomouc (Czech Republic); Safarikova, Mirka [Global Change Research Institute, CAS, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic); Department of Nanobiotechnology, Biology Centre, CAS, ISB, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic); Safarik, Ivo [Global Change Research Institute, CAS, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic); Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 27, 783 71 Olomouc (Czech Republic); Department of Nanobiotechnology, Biology Centre, CAS, ISB, Na Sadkach 7, 370 05 Ceske Budejovice (Czech Republic)

    2017-04-01

    A simple, one-pot process for the preparation of magnetically responsive yeast-based biocatalysts was developed. Saccharomyces cerevisiae, Candida utilis and Kluyveromyces lactis cells were successfully incorporated into chitosan gel magnetically modified with poly(methacrylic acid)-stabilized magnetic fluid (PMAA-FF) during its formation. Magnetic PMAA-FF/chitosan/yeast composites were efficiently employed for invert sugar production. The dependence of invertase activity on used yeast, amount of magnetic biocatalyst, agitation time and after reuse was studied in detail. The tested magnetic biocatalysts retained at least 69% of their initial activity after 8 reuse cycles. - Highlights: • New types of magnetically responsive yeast biocomposites were prepared. • Recently developed PMAA-stabilized magnetic fluid was used. • Three yeast species were entrapped into magnetic chitosan gel during its formation. • All biocatalysts were efficiently employed for invert sugar formation.

  18. Detection of anabolic steroids in dietary supplements: The added value of an androgen yeast bioassay in parallel with a liquid chromatography-tandem mass spectrometry screening method

    NARCIS (Netherlands)

    Rijk, J.C.W.; Bovee, T.F.H.; Wang, S.; Poucke, C.; Peteghem, van C.; Nielen, M.W.F.

    2009-01-01

    Recently we constructed a recombinant yeast cell that expresses the human androgen receptor (hAR) and yeast enhanced green fluorescent protein (yEGFP), the latter in response to androgens. When exposed to testosterone, the concentration where half-maximal activation is reached (EC50) was 50 nM.

  19. Binding and Conversion of Selenium in Candida utilis ATCC 9950 Yeasts in Bioreactor Culture

    Directory of Open Access Journals (Sweden)

    Marek Kieliszek

    2017-02-01

    Full Text Available Selenium is considered an essential component of all living organisms. The use of yeasts as a selenium supplement in human nutrition has gained much interest over the last decade. The accumulation and biochemical transformation of selenium in yeast cells is particularly interesting to many researchers. In this article, we present the results of the determination of selenium and selenomethionine content in the biomass of feed yeast Candida utilis ATCC 9950 obtained from the culture grown in a bioreactor. The results indicated that C. utilis cells performed the biotransformation of inorganic selenium(IV to organic derivatives (e.g., selenomethionine. Selenium introduced (20–30 mg Se4+∙L−1 to the experimental media in the form of sodium(IV selenite (Na2SeO3 salt caused a significant increase in selenium content in the biomass of C. utilis,irrespective of the concentration. The highest amount of selenium (1841 μg∙gd.w.−1 was obtained after a 48-h culture in media containing 30 mg Se4+∙L−1. The highest content of selenomethionine (238.8 μg∙gd.w.−1 was found after 48-h culture from the experimental medium that was supplemented with selenium at a concentration of 20 mg Se4+∙L−1. Biomass cell in the cultures supplemented with selenium ranged from 1.5 to 14.1 g∙L−1. The results of this study indicate that yeast cell biomass of C. utilis enriched mainly with the organic forms of selenium can be a valuable source of protein. It creates the possibility of obtaining selenium biocomplexes that can be used in the production of protein-selenium dietary supplements for animals and humans

  20. Determination of the autolysis of champagne yeast by using 14C-labelled yeast

    International Nuclear Information System (INIS)

    Molnar, I.; Oura, E.; Suomalainen, H.

    1980-01-01

    The degree of autolysis of 14 C-labelled Champagne Hautvillers yeast was studied in the function of different temperatures of storage. A linear relationship was found between the length of the storage and the degree of autolysis. The rate of autolysis increased with raising the temperature of storage. The raising of the temperature by 10 deg C was followed by a 6-7% increase in the rate of autolysis. Shaking up the yeast sediment at 20-day intervals raised the rate of autolysis by 1.5-4.2%. (author)

  1. 21 CFR 73.355 - Phaffia yeast.

    Science.gov (United States)

    2010-04-01

    ... stabilized color additive mixture. Color additive mixtures for fish feed use made with phaffia yeast may... additive mixtures for coloring foods. (b) Specifications. Phaffia yeast shall conform to the following... § 501.4 of this chapter. (3) The presence of the color additive in salmonid fish that have been fed...

  2. Expression, purification, and refolding of active recombinant human E-selectin lectin and EGF domains in Escherichia coli.

    Science.gov (United States)

    Kawano, Susumu; Iyaguchi, Daisuke; Okada, Chiaki; Sasaki, Yusuke; Toyota, Eiko

    2013-06-01

    Attempts to obtain active E-selectin from Escherichia coli (E. coli) have not yet been successful. In this study, we succeeded in expressing the recombinant lectin and epidermal growth factor domain fragments of human E-selectin (rh-ESLE) in E. coli on a large-scale. The rh-ESLE protein was expressed as an inactive form in the inclusion bodies. The inactive form of rh-ESLE was denatured and solubilized by 6 M guanidine hydrochloride and then purified by Ni(2+) affinity chromatography under denaturing conditions. Denatured rh-ESLE was then refolded by a rapid-dilution method using a large amount of refolding buffer, which contained arginine and cysteine/cystine. The refolded rh-ESLE showed binding affinity for sLe(X) (K(d) = 321 nM, B(max) = 1.9 pmol/μg protein). This result suggests that the refolded rh-ESLE recovered its native and functional structure.

  3. GROWTH RATE AND IDENTIFICATION OF YEASTS IN THREE DIFFERENT MEDIA: FLOUR, SPROUT OF CEREALS AND COMMERCIAL MEDIA

    Directory of Open Access Journals (Sweden)

    P DEHGHAN

    2002-12-01

    Full Text Available Introduction. The yeasts are a large group of fungi. In addition to their different uses in food industries and pharmacology they can also cause human and animal infections under predisposing factors. For investigating the yeast"s growth phases and diagnosis of the species, they should culture in media from natural sources like the seeds on the commercial media. According to yeast"s nutritinal requierments and considering the various nutritional materials of seeds like wheat, rice, barney and better use of these seed"s compounds during sprouting, this research was done with the aim of measurment of growth rate and yeasts characterization in such media. Methods. For making seeds media, a fixed quantity of seed"s flour or sprout"s flour was added to a certain amount of agar in acidotic condition (pH=5.8. The germination of seeds was done by Hus method and for colony counting Mc, Farland tubes were used and microscopic and macroscopic characteristics were investigated and compared in both synthetic and seed media Results. The results showed that the rate of yeasts growth in all germinated seeds was more than the seeds themeselves. Statistical analysis showed no significal difference between the growth rate of yeasts in both wheat and barney sprout compared to the commercial media. In the microscopic studies, the species of Candida albicans in wheat flour has produced more chlamydoconidia than the synthetic medium of corn meal agar. Also production of the capsule in C.neoformans in the seed"s media has been better than the synthetic media. Discussion. According to the promising results obtained from the present study regard to the growth rate and differentiation of the fungal species in such media, standardization and mass production of them in our country would seem to be an productive step towards self sufficiency.

  4. Conventional and Non-Conventional Yeasts in Beer Production

    Directory of Open Access Journals (Sweden)

    Angela Capece

    2018-06-01

    Full Text Available The quality of beer relies on the activity of fermenting yeasts, not only for their good fermentation yield-efficiency, but also for their influence on beer aroma, since most of the aromatic compounds are intermediate metabolites and by-products of yeast metabolism. Beer production is a traditional process, in which Saccharomyces is the sole microbial component, and any deviation is considered a flaw. However, nowadays the brewing sector is faced with an increasing demand for innovative products, and it is diffusing the use of uncharacterized autochthonous starter cultures, spontaneous fermentation, or non-Saccharomyces starters, which leads to the production of distinctive and unusual products. Attempts to obtain products with more complex sensory characteristics have led one to prospect for non-conventional yeasts, i.e., non-Saccharomyces yeasts. These generally are characterized by low fermentation yields and are more sensitive to ethanol stress, but they provide a distinctive aroma and flavor. Furthermore, non-conventional yeasts can be used for the production of low-alcohol/non-alcoholic and light beers. This review aims to present the main findings about the role of traditional and non-conventional yeasts in brewing, demonstrating the wide choice of available yeasts, which represents a new biotechnological approach with which to target the characteristics of beer and to produce different or even totally new beer styles.

  5. Effects of paraquat on Escherichia coli: Differences between B and K-12 strains

    International Nuclear Information System (INIS)

    Kitzler, J.W.; Minakami, H.; Fridovich, I.

    1990-01-01

    Escherichia coli B and K-12 are equally susceptible to the bacteriostatic effects of aerobic paraquat, but they differed strikingly when the lethality of paraquat was evaluated. E. coli B suffered an apparent loss of viability when briefly exposed to paraquat, whereas E. coli K-12 did not. This difference depended on the ability of the B-strain, but not the K-12 strain, to retain internalized paraquat; the B strain was killed on aerobic tryptic soy-yeast extract plates during the incubation which preceded the counting of colonies. This difference in retention of paraquat between strains was demonstrated by delayed loss of viability, by growth inhibition, and by cyanide-resistant respiration after brief exposure to paraquat, washing, and testing in fresh medium. This difference was also shown by using [ 14 C]paraquat. This previously unrecognized difference between E. coli B and K-12 has been the cause of apparently contradictory reports and should lead to some reevaluation of the pertinent literature

  6. Ingestion of Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli by human peritoneal mesothelial cells

    NARCIS (Netherlands)

    Visser, C. E.; Brouwer-Steenbergen, J. J.; Schadee-Eestermans, I. L.; Meijer, S.; Krediet, R. T.; Beelen, R. H.

    1996-01-01

    In the present study we examined whether mesothelial cells can ingest and digest bacteria. The results showed that all strains were ingested. Ingested staphylococci proliferated abundantly, and only a few were digested. Escherichia coli, however, was digested during the first 8 h, whereafter the

  7. Protein N-myristoylation in Escherichia coli: Reconstitution of a eukaryotic protein modification in bacteria

    International Nuclear Information System (INIS)

    Duronio, R.J.; Jackson-Machelski, E.; Heuckeroth, R.O.; Gordon, J.I.; Olins, P.O.; Devine, C.S.; Yonemoto, W.; Slice, L.W.; Taylor, S.S.

    1990-01-01

    Protein N-myristoylation refers to the covalent attachment of a myristoyl group (C14:0), via amide linkage, to the NH 2 -terminal glycine residue of certain cellular and viral proteins. Myristoyl-CoA:protein N-myristoyltransferase (NMT) catalyzes this cotranslational modification. The authors have developed a system for studying the substrate requirements and biological effects of protein N-myristoylation as well as NMT structure-activity relationships. Expression of the yeast NMT1 gene in Escherichia coli, a bacterium that has no endogenous NMT activity, results in production of the intact 53-kDa NMT polypeptide as well as a truncated polypeptide derived from proteolytic removal of its NH 2 -terminal 39 amino acids. By using a dual plasmid system, N-myristoylation of a mammalian protein was reconstituted in E. coli by simultaneous expression of the yeast NMT1 gene and a murine cDNA encoding the catalytic (C) subunit of cAMP-dependent protein kinase (PK-A). A major advantage of the bacterial system over eukaryotic systems is the absence of endogenous NMT and substrates, providing a more straightforward way of preparing myristoylated, analog-substituted, and nonmyristoylated forms of a given protein for comparison of their structural and functional properties. The experimental system may prove useful for recapitulating other eukaryotic protein modifications in E. coli so that structure-activity relationships of modifying enzymes and their substrates can be more readily assessed

  8. Chromosomal Aneuploidy Improves the Brewing Characteristics of Sake Yeast.

    Science.gov (United States)

    Kadowaki, Masafumi; Fujimaru, Yuki; Taguchi, Seiga; Ferdouse, Jannatul; Sawada, Kazutaka; Kimura, Yuta; Terasawa, Yohei; Agrimi, Gennaro; Anai, Toyoaki; Noguchi, Hideki; Toyoda, Atsushi; Fujiyama, Asao; Akao, Takeshi; Kitagaki, Hiroshi

    2017-12-15

    The effect of chromosomal aneuploidy on the brewing characteristics of brewery yeasts has not been studied. Here we report that chromosomal aneuploidy in sake brewery yeast ( Saccharomyces cerevisiae ) leads to the development of favorable brewing characteristics. We found that pyruvate-underproducing sake yeast, which produces less off-flavor diacetyl, is aneuploid and trisomic for chromosomes XI and XIV. To confirm that this phenotype is due to aneuploidy, we obtained 45 haploids with various chromosomal additions and investigated their brewing profiles. A greater number of chromosomes correlated with a decrease in pyruvate production. Especially, sake yeast haploids with extra chromosomes in addition to chromosome XI produced less pyruvate than euploids. Mitochondrion-related metabolites and intracellular oxygen species in chromosome XI aneuploids were higher than those in euploids, and this effect was canceled in their "petite" strains, suggesting that an increase in chromosomes upregulated mitochondrial activity and decreased pyruvate levels. These findings suggested that an increase in chromosome number, including chromosome XI, in sake yeast haploids leads to pyruvate underproduction through the augmentation of mitochondrial activity. This is the first report proposing that aneuploidy in brewery yeasts improves their brewing profile. IMPORTANCE Chromosomal aneuploidy has not been evaluated in development of sake brewing yeast strains. This study shows the relationship between chromosomal aneuploidy and brewing characteristics of brewery yeast strains. High concentrations of pyruvate during sake storage give rise to α-acetolactate and, in turn, to high concentrations of diacetyl, which is considered an off-flavor. It was demonstrated that pyruvate-underproducing sake yeast is trisomic for chromosome XI and XIV. Furthermore, sake yeast haploids with extra chromosomes produced reduced levels of pyruvate and showed metabolic processes characteristic of

  9. Guidelines and recommendations on yeast cell death nomenclature

    Directory of Open Access Journals (Sweden)

    Didac Carmona-Gutierrez

    2018-01-01

    Full Text Available Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cellular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the definition of accidental, regulated, and programmed forms of cell death in yeast based on a series of morphological and biochemical criteria. Specifically, we provide consensus guidelines on the differential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death routines that are relevant for the biology of (at least some species of yeast. As this area of investigation advances rapidly, changes and extensions to this set of recommendations will be implemented in the years to come. Nonetheless, we strongly encourage the authors, reviewers and editors of scientific articles to adopt these collective standards in order to establish an accurate framework for yeast cell death research and, ultimately, to accelerate the progress of this vibrant field of research.

  10. Guidelines and recommendations on yeast cell death nomenclature

    Science.gov (United States)

    Carmona-Gutierrez, Didac; Bauer, Maria Anna; Zimmermann, Andreas; Aguilera, Andrés; Austriaco, Nicanor; Ayscough, Kathryn; Balzan, Rena; Bar-Nun, Shoshana; Barrientos, Antonio; Belenky, Peter; Blondel, Marc; Braun, Ralf J.; Breitenbach, Michael; Burhans, William C.; Büttner, Sabrina; Cavalieri, Duccio; Chang, Michael; Cooper, Katrina F.; Côrte-Real, Manuela; Costa, Vítor; Cullin, Christophe; Dawes, Ian; Dengjel, Jörn; Dickman, Martin B.; Eisenberg, Tobias; Fahrenkrog, Birthe; Fasel, Nicolas; Fröhlich, Kai-Uwe; Gargouri, Ali; Giannattasio, Sergio; Goffrini, Paola; Gourlay, Campbell W.; Grant, Chris M.; Greenwood, Michael T.; Guaragnella, Nicoletta; Heger, Thomas; Heinisch, Jürgen; Herker, Eva; Herrmann, Johannes M.; Hofer, Sebastian; Jiménez-Ruiz, Antonio; Jungwirth, Helmut; Kainz, Katharina; Kontoyiannis, Dimitrios P.; Ludovico, Paula; Manon, Stéphen; Martegani, Enzo; Mazzoni, Cristina; Megeney, Lynn A.; Meisinger, Chris; Nielsen, Jens; Nyström, Thomas; Osiewacz, Heinz D.; Outeiro, Tiago F.; Park, Hay-Oak; Pendl, Tobias; Petranovic, Dina; Picot, Stephane; Polčic, Peter; Powers, Ted; Ramsdale, Mark; Rinnerthaler, Mark; Rockenfeller, Patrick; Ruckenstuhl, Christoph; Schaffrath, Raffael; Segovia, Maria; Severin, Fedor F.; Sharon, Amir; Sigrist, Stephan J.; Sommer-Ruck, Cornelia; Sousa, Maria João; Thevelein, Johan M.; Thevissen, Karin; Titorenko, Vladimir; Toledano, Michel B.; Tuite, Mick; Vögtle, F.-Nora; Westermann, Benedikt; Winderickx, Joris; Wissing, Silke; Wölfl, Stefan; Zhang, Zhaojie J.; Zhao, Richard Y.; Zhou, Bing; Galluzzi, Lorenzo; Kroemer, Guido; Madeo, Frank

    2018-01-01

    Elucidating the biology of yeast in its full complexity has major implications for science, medicine and industry. One of the most critical processes determining yeast life and physiology is cellular demise. However, the investigation of yeast cell death is a relatively young field, and a widely accepted set of concepts and terms is still missing. Here, we propose unified criteria for the definition of accidental, regulated, and programmed forms of cell death in yeast based on a series of morphological and biochemical criteria. Specifically, we provide consensus guidelines on the differential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death routines that are relevant for the biology of (at least some species of) yeast. As this area of investigation advances rapidly, changes and extensions to this set of recommendations will be implemented in the years to come. Nonetheless, we strongly encourage the authors, reviewers and editors of scientific articles to adopt these collective standards in order to establish an accurate framework for yeast cell death research and, ultimately, to accelerate the progress of this vibrant field of research. PMID:29354647

  11. Escherichia coli O157:H7: Animal Reservoir and Sources of Human Infection

    Science.gov (United States)

    Ferens, Witold A.

    2011-01-01

    Abstract This review surveys the literature on carriage and transmission of enterohemorrhagic Escherichia coli (EHEC) O157:H7 in the context of virulence factors and sampling/culture technique. EHEC of the O157:H7 serotype are worldwide zoonotic pathogens responsible for the majority of severe cases of human EHEC disease. EHEC O157:H7 strains are carried primarily by healthy cattle and other ruminants, but most of the bovine strains are not transmitted to people, and do not exhibit virulence factors associated with human disease. Prevalence of EHEC O157:H7 is probably underestimated. Carriage of EHEC O157:H7 by individual animals is typically short-lived, but pen and farm prevalence of specific isolates may extend for months or years and some carriers, designated as supershedders, may harbor high intestinal numbers of the pathogen for extended periods. The prevalence of EHEC O157:H7 in cattle peaks in the summer and is higher in postweaned calves and heifers than in younger and older animals. Virulent strains of EHEC O157:H7 are rarely harbored by pigs or chickens, but are found in turkeys. The bacteria rarely occur in wildlife with the exception of deer and are only sporadically carried by domestic animals and synanthropic rodents and birds. EHEC O157:H7 occur in amphibian, fish, and invertebrate carriers, and can colonize plant surfaces and tissues via attachment mechanisms different from those mediating intestinal attachment. Strains of EHEC O157:H7 exhibit high genetic variability but typically a small number of genetic types predominate in groups of cattle and a farm environment. Transmission to people occurs primarily via ingestion of inadequately processed contaminated food or water and less frequently through contact with manure, animals, or infected people. PMID:21117940

  12. Analysis of UV-induced mutation spectra in Escherichia coli by DNA polymerase {eta} from Arabidopsis thaliana

    Energy Technology Data Exchange (ETDEWEB)

    Santiago, Maria Jesus [Departamento de Genetica, Facultad de Ciencias, Edificio Gregor Mendel, Campus Rabanales, Universidad de Cordoba (Spain); Alejandre-Duran, Encarna [Departamento de Genetica, Facultad de Ciencias, Edificio Gregor Mendel, Campus Rabanales, Universidad de Cordoba (Spain); Ruiz-Rubio, Manuel [Departamento de Genetica, Facultad de Ciencias, Edificio Gregor Mendel, Campus Rabanales, Universidad de Cordoba (Spain)]. E-mail: ge1rurum@uco.es

    2006-10-10

    DNA polymerase {eta} belongs to the Y-family of DNA polymerases, enzymes that are able to synthesize past template lesions that block replication fork progression. This polymerase accurately bypasses UV-associated cis-syn cyclobutane thymine dimers in vitro and therefore may contributes to resistance against sunlight in vivo, both ameliorating survival and decreasing the level of mutagenesis. We cloned and sequenced a cDNA from Arabidopsis thaliana which encodes a protein containing several sequence motifs characteristics of Pol{eta} homologues, including a highly conserved sequence reported to be present in the active site of the Y-family DNA polymerases. The gene, named AtPOLH, contains 14 exons and 13 introns and is expressed in different plant tissues. A strain from Saccharomyces cerevisiae, deficient in Pol{eta} activity, was transformed with a yeast expression plasmid containing the AtPOLH cDNA. The rate of survival to UV irradiation in the transformed mutant increased to similar values of the wild type yeast strain, showing that AtPOLH encodes a functional protein. In addition, when AtPOLH is expressed in Escherichia coli, a change in the mutational spectra is detected when bacteria are irradiated with UV light. This observation might indicate that AtPOLH could compete with DNA polymerase V and then bypass cyclobutane pyrimidine dimers incorporating two adenylates.

  13. Effect of salt hyperosmotic stress on yeast cell viability

    Directory of Open Access Journals (Sweden)

    Logothetis Stelios

    2007-01-01

    Full Text Available During fermentation for ethanol production, yeasts are subjected to different kinds of physico-chemical stresses such as: initially high sugar concentration and low temperature; and later, increased ethanol concentrations. Such conditions trigger a series of biological responses in an effort to maintain cell cycle progress and yeast cell viability. Regarding osmostress, many studies have been focused on transcriptional activation and gene expression in laboratory strains of Saccharomyces cerevisiae. The overall aim of this present work was to further our understanding of wine yeast performance during fermentations under osmotic stress conditions. Specifically, the research work focused on the evaluation of NaCl-induced stress responses of an industrial wine yeast strain S. cerevisiae (VIN 13, particularly with regard to yeast cell growth and viability. The hypothesis was that osmostress conditions energized specific genes to enable yeast cells to survive under stressful conditions. Experiments were designed by pretreating cells with different sodium chloride concentrations (NaCl: 4%, 6% and 10% w/v growing in defined media containing D-glucose and evaluating the impact of this on yeast growth and viability. Subsequent fermentation cycles took place with increasing concentrations of D-glucose (20%, 30%, 40% w/v using salt-adapted cells as inocula. We present evidence that osmostress induced by mild salt pre-treatments resulted in beneficial influences on both cell viability and fermentation performance of an industrial wine yeast strain.

  14. The primary structures of two yeast enolase genes. Homology between the 5' noncoding flanking regions of yeast enolase and glyceraldehyde-3-phosphate dehydrogenase genes.

    Science.gov (United States)

    Holland, M J; Holland, J P; Thill, G P; Jackson, K A

    1981-02-10

    Segments of yeast genomic DNA containing two enolase structural genes have been isolated by subculture cloning procedures using a cDNA hybridization probe synthesized from purified yeast enolase mRNA. Based on restriction endonuclease and transcriptional maps of these two segments of yeast DNA, each hybrid plasmid contains a region of extensive nucleotide sequence homology which forms hybrids with the cDNA probe. The DNA sequences which flank this homologous region in the two hybrid plasmids are nonhomologous indicating that these sequences are nontandemly repeated in the yeast genome. The complete nucleotide sequence of the coding as well as the flanking noncoding regions of these genes has been determined. The amino acid sequence predicted from one reading frame of both structural genes is extremely similar to that determined for yeast enolase (Chin, C. C. Q., Brewer, J. M., Eckard, E., and Wold, F. (1981) J. Biol. Chem. 256, 1370-1376), confirming that these isolated structural genes encode yeast enolase. The nucleotide sequences of the coding regions of the genes are approximately 95% homologous, and neither gene contains an intervening sequence. Codon utilization in the enolase genes follows the same biased pattern previously described for two yeast glyceraldehyde-3-phosphate dehydrogenase structural genes (Holland, J. P., and Holland, M. J. (1980) J. Biol. Chem. 255, 2596-2605). DNA blotting analysis confirmed that the isolated segments of yeast DNA are colinear with yeast genomic DNA and that there are two nontandemly repeated enolase genes per haploid yeast genome. The noncoding portions of the two enolase genes adjacent to the initiation and termination codons are approximately 70% homologous and contain sequences thought to be involved in the synthesis and processing messenger RNA. Finally there are regions of extensive homology between the two enolase structural genes and two yeast glyceraldehyde-3-phosphate dehydrogenase structural genes within the 5