Production of Lycopene in the Non-Carotenoid-Producing Yeast Yarrowia lipolytica

PubMed Central

Ketelhot, Markus; Gatter, Michael; Barth, Gerold

2014-01-01

The codon-optimized genes crtB and crtI of Pantoea ananatis were expressed in Yarrowia lipolytica under the control of the TEF1 promoter of Y. lipolytica. Additionally, the rate-limiting genes for isoprenoid biosynthesis in Y. lipolytica, GGS1 and HMG1, were overexpressed to increase the production of lycopene. All of the genes were also expressed in a Y. lipolytica strain with POX1 to POX6 and GUT2 deleted, which led to an increase in the size of lipid bodies and a further increase in lycopene production. Lycopene is located mainly within lipid bodies, and increased lipid body formation leads to an increase in the lycopene storage capacity of Y. lipolytica. Growth-limiting conditions increase the specific lycopene content. Finally, a yield of 16 mg g−1 (dry cell weight) was reached in fed-batch cultures, which is the highest value reported so far for a eukaryotic host. PMID:24375130

Yarrowia lipolytica vesicle-mediated protein transport pathways

PubMed Central

Swennen, Dominique; Beckerich, Jean-Marie

2007-01-01

Background Protein secretion is a universal cellular process involving vesicles which bud and fuse between organelles to bring proteins to their final destination. Vesicle budding is mediated by protein coats; vesicle targeting and fusion depend on Rab GTPase, tethering factors and SNARE complexes. The Génolevures II sequencing project made available entire genome sequences of four hemiascomycetous yeasts, Yarrowia lipolytica, Debaryomyces hansenii, Kluyveromyces lactis and Candida glabrata. Y. lipolytica is a dimorphic yeast and has good capacities to secrete proteins. The translocation of nascent protein through the endoplasmic reticulum membrane was well studied in Y. lipolytica and is largely co-translational as in the mammalian protein secretion pathway. Results We identified S. cerevisiae proteins involved in vesicular secretion and these protein sequences were used for the BLAST searches against Génolevures protein database (Y. lipolytica, C. glabrata, K. lactis and D. hansenii). These proteins are well conserved between these yeasts and Saccharomyces cerevisiae. We note several specificities of Y. lipolytica which may be related to its good protein secretion capacities and to its dimorphic aspect. An expansion of the Y. lipolytica Rab protein family was observed with autoBLAST and the Rab2- and Rab4-related members were identified with BLAST against NCBI protein database. An expansion of this family is also found in filamentous fungi and may reflect the greater complexity of the Y. lipolytica secretion pathway. The Rab4p-related protein may play a role in membrane recycling as rab4 deleted strain shows a modification of colony morphology, dimorphic transition and permeability. Similarly, we find three copies of the gene (SSO) encoding the plasma membrane SNARE protein. Quantification of the percentages of proteins with the greatest homology between S. cerevisiae, Y. lipolytica and animal homologues involved in vesicular transport shows that 40% of Y

Enhanced α-ketoglutaric acid production and recovery in Yarrowia lipolytica yeast by effective pH controlling.

PubMed

Morgunov, Igor G; Kamzolova, Svetlana V; Samoilenko, Vladimir A

2013-10-01

The replacement of chemical synthesis by environmentally friendly energy-efficient technologies for production of valuable metabolites is a principal strategy of developing biotechnological industry all over the world. In the present study, we develop a method for α-ketoglutaric acid (KGA) production from rapeseed oil with the use of Yarrowia lipolytica yeast. Sixty strains of Y. lipolytica yeasts were tested for their ability to produce KGA, and the strain Y. lipolytica 212 (Y. lipolytica VKM Y-2412) was selected as a promising KGA producer. Using a three-stage pH controlling, in which pH was 4.5 in the growth phase, then since 72 to 144 h, pH was maintained at 3.5 and in the later phase of acid production, the titration by KOH was switch off, selected strain produced 106.5 g l(-1) of KGA with mass yield of 0.95 g g(-1). KGA in the form of monopotassium salt was isolated from the culture broth and purified. The isolation procedure involved separation of biomass, extraction of residual triglycerides, filtrate bleaching, and acidification with mineral acid (to pH 2.8-3.4), concentration, precipitation of mineral salts, and crystallization of the product. The purity of KGA isolated from the culture filtrate reached 99.1 %.

Yarrowia lipolytica and Its Multiple Applications in the Biotechnological Industry

PubMed Central

Gonçalves, F. A. G.; Colen, G.; Takahashi, J. A.

2014-01-01

Yarrowia lipolytica is a nonpathogenic dimorphic aerobic yeast that stands out due to its ability to grow in hydrophobic environments. This property allowed this yeast to develop an ability to metabolize triglycerides and fatty acids as carbon sources. This feature enables using this species in the bioremediation of environments contaminated with oil spill. In addition, Y. lipolytica has been calling the interest of researchers due to its huge biotechnological potential, associated with the production of several types of metabolites, such as bio-surfactants, γ-decalactone, citric acid, and intracellular lipids and lipase. The production of a metabolite rather than another is influenced by the growing conditions to which Y. lipolytica is subjected. The choice of carbon and nitrogen sources to be used, as well as their concentrations in the growth medium, and the careful determination of fermentation parameters, pH, temperature, and agitation (oxygenation), are essential for efficient metabolites production. This review discusses the biotechnological potential of Y. lipolytica and the best growing conditions for production of some metabolites of biotechnological interest. PMID:24715814

A Rac Homolog Is Required for Induction of Hyphal Growth in the Dimorphic Yeast Yarrowia lipolytica

PubMed Central

Hurtado, Cleofe A. R.; Beckerich, Jean-Marie; Gaillardin, Claude; Rachubinski, Richard A.

2000-01-01

Dimorphism in fungi is believed to constitute a mechanism of response to adverse conditions and represents an important attribute for the development of virulence by a number of pathogenic fungal species. We have isolated YlRAC1, a gene encoding a 192-amino-acid protein that is essential for hyphal growth in the dimorphic yeast Yarrowia lipolytica and which represents the first Rac homolog described for fungi. YlRAC1 is not an essential gene, and its deletion does not affect the ability to mate or impair actin polarization in Y. lipolytica. However, strains lacking functional YlRAC1 show alterations in cell morphology, suggesting that the function of YlRAC1 may be related to some aspect of the polarization of cell growth. Northern blot analysis showed that transcription of YlRAC1 increases steadily during the yeast-to-hypha transition, while Southern blot analysis of genomic DNA suggested the presence of several RAC family members in Y. lipolytica. Interestingly, strains lacking functional YlRAC1 are still able to grow as the pseudohyphal form and to invade agar, thus pointing to a function for YlRAC1 downstream of MHY1, a previously isolated gene encoding a C2H2-type zinc finger protein with the ability to bind putative stress response elements and whose activity is essential for both hyphal and pseudohyphal growth in Y. lipolytica. PMID:10762235

Identification of the Transcription Factor Znc1p, which Regulates the Yeast-to-Hypha Transition in the Dimorphic Yeast Yarrowia lipolytica

PubMed Central

Martinez-Vazquez, Azul; Gonzalez-Hernandez, Angelica; Domínguez, Ángel; Rachubinski, Richard; Riquelme, Meritxell; Cuellar-Mata, Patricia; Guzman, Juan Carlos Torres

2013-01-01

The dimorphic yeast Yarrowia lipolytica is used as a model to study fungal differentiation because it grows as yeast-like cells or forms hyphal cells in response to changes in environmental conditions. Here, we report the isolation and characterization of a gene, ZNC1, involved in the dimorphic transition in Y. lipolytica. The ZNC1 gene encodes a 782 amino acid protein that contains a Zn(II)2C6 fungal-type zinc finger DNA-binding domain and a leucine zipper domain. ZNC1 transcription is elevated during yeast growth and decreases during the formation of mycelium. Cells in which ZNC1 has been deleted show increased hyphal cell formation. Znc1p-GFP localizes to the nucleus, but mutations within the leucine zipper domain of Znc1p, and to a lesser extent within the Zn(II)2C6 domain, result in a mislocalization of Znc1p to the cytoplasm. Microarrays comparing gene expression between znc1::URA3 and wild-type cells during both exponential growth and the induction of the yeast-to-hypha transition revealed 1,214 genes whose expression was changed by 2-fold or more under at least one of the conditions analyzed. Our results suggest that Znc1p acts as a transcription factor repressing hyphal cell formation and functions as part of a complex network regulating mycelial growth in Y. lipolytica. PMID:23826133

Impact of culture conditions on β-carotene encapsulation using Yarrowia lipolytica cells

NASA Astrophysics Data System (ADS)

Dang, Tran Hai; Minh, Ho Thi Thu; Van Nhi, Tran Nguyen; Ngoc, Ta Thi Minh

2017-09-01

Yeast cell was reported as an effective natural preformed material for use in encapsulation of hydrophobic compounds. The encapsulation process was normally considered as passive transfer through cellular wall and cellular membrane. Beside solubility of hydrophobic compound in phospholipid membrane or plasmolysis, membrane characteristics of yeast cell which are differed between strains and influenced by culture conditions are main factors involving the accumulation of hydrophobic compound into yeast cell. In this study, the oleaginous yeast Yarrowia lipolytica was used as micro-container shell to encapsulate a high hydrophobic compound - β-carotene. Yeast cell was cultured under different conditions and wet yeast biomass was incubated with β-carotene which was dissolved in soybean oil overnight. β-carotene accumulation was then extracted and evaluated by UV-VIS spectrometry. Optimization of culture condition was investigated using the Box-Behnken model. β-carotene encapsulation efficiency in Y. lipolytica was showed to be affected by both pH of medium and agitation conditions. The highest β-carotene encapsulation efficiency was optimized at 42.8 μg/g with Y. lipolytica cultured at pH 4.5, medium volume equal to 115 ml and agitation speed at 211 rpm.

Fatty alcohol production in Lipomyces starkeyi and Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Knoshaug, Eric

Current biological pathways to produce biofuel intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels are not cost effective. Previously, oleaginous yeasts have been investigated primarily for lipid production. However, yeasts store neutral lipids intracellularly making recovery difficult and expensive. In addition, once recovered from the cells, lipids are difficult to blend directly with the existing fuels without upgrading. We have, therefore, begun to investigate secreted fatty acid-derived products which can be easily recovered and upgraded to fuels. In this study, we successfully demonstrate the production of fatty alcohols by the oleaginous yeasts, Yarrowia lipolytica and Lipomyces starkeyi, throughmore » expression of the fatty acyl-CoA reductase gene from Marinobactor aquaeolei VT8. This strategy resulted in the production of 167 and 770 mg/L of fatty alcohols in shake flask from Y. lipolytica and L starkeyi, respectively. When using a dodecane overlay during fermentation, 92 and 99% of total fatty alcohols produced by Y. lipolytica and L. starkeyi, respectively, were extracted into the dodecane phase, which compares favorably to the 3 and 50% recovered, respectively, without the dodecane layer. In both oleaginous yeasts, long chain length, saturated fatty alcohols, i.e., hexadecanol (C16:0) and octadecanol (C18:0), were predominant and accounted for more than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Furthermore, this work demonstrates that the oleaginous yeasts, Y. lipolytica and L. starkeyi, can serve as platform organisms for the production of fatty acid-derived biofuels and bioproducts.« less

Fatty alcohol production in Lipomyces starkeyi and Yarrowia lipolytica

DOE PAGES

Wang, Wei; Wei, Hui; Knoshaug, Eric; ...

2016-10-24

Current biological pathways to produce biofuel intermediates amenable to separations and catalytic upgrading to hydrocarbon fuels are not cost effective. Previously, oleaginous yeasts have been investigated primarily for lipid production. However, yeasts store neutral lipids intracellularly making recovery difficult and expensive. In addition, once recovered from the cells, lipids are difficult to blend directly with the existing fuels without upgrading. We have, therefore, begun to investigate secreted fatty acid-derived products which can be easily recovered and upgraded to fuels. In this study, we successfully demonstrate the production of fatty alcohols by the oleaginous yeasts, Yarrowia lipolytica and Lipomyces starkeyi, throughmore » expression of the fatty acyl-CoA reductase gene from Marinobactor aquaeolei VT8. This strategy resulted in the production of 167 and 770 mg/L of fatty alcohols in shake flask from Y. lipolytica and L starkeyi, respectively. When using a dodecane overlay during fermentation, 92 and 99% of total fatty alcohols produced by Y. lipolytica and L. starkeyi, respectively, were extracted into the dodecane phase, which compares favorably to the 3 and 50% recovered, respectively, without the dodecane layer. In both oleaginous yeasts, long chain length, saturated fatty alcohols, i.e., hexadecanol (C16:0) and octadecanol (C18:0), were predominant and accounted for more than 85% of the total fatty alcohols produced. To the best of our knowledge, this is the first report of fatty alcohol production in L. starkeyi. Furthermore, this work demonstrates that the oleaginous yeasts, Y. lipolytica and L. starkeyi, can serve as platform organisms for the production of fatty acid-derived biofuels and bioproducts.« less

Multi-omics analysis reveals regulators of the response to nitrogen limitation in Yarrowia lipolytica

DOE PAGES

Pomraning, Kyle R.; Kim, Young -Mo; Nicora, Carrie D.; ...

2016-02-25

Yarrowia lipolytica is an oleaginous ascomycete yeast that stores lipids in response to limitation of nitrogen. Furthermore, while the enzymatic pathways responsible for neutral lipid accumulation in Y. lipolytica are well characterized, regulation of these pathways has received little attention. We therefore sought to characterize the response to nitrogen limitation at system-wide levels, including the proteome, phosphoproteome and metabolome, to better understand how this organism regulates and controls lipid metabolism and to identify targets that may be manipulated to improve lipid yield.

A survey of yeast from the Yarrowia clade for lipid production in dilute-acid pretreated lignocellulosic biomass hydrolysate

USDA-ARS?s Scientific Manuscript database

Yarrowia lipolytica is an oleaginous yeast species that has attracted attention as a model organism for synthesis of single cell oil. Among over 50 isolates of Y. lipolytica identified, only a few of the strains have been studied extensively. Furthermore, 12 other yeast species were recently assigne...

An ortholog of farA of Aspergillus nidulans is implicated in the transcriptional activation of genes involved in fatty acid utilization in the yeast Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Poopanitpan, Napapol; Kobayashi, Satoshi; Fukuda, Ryouichi

2010-11-26

Research highlights: {yields} POR1 is a Yarrowia lipolytica ortholog of farA involved in fatty acid response in A. nidulans. {yields} Deletion of POR1 caused growth defects on fatty acids. {yields} {Delta}por1 strain exhibited defects in the induction of genes involved in fatty acid utilization. -- Abstract: The yeast Yarrowia lipolytica effectively utilizes hydrophobic substrates such as fatty acids and n-alkanes. To identify a gene(s) regulating fatty acid utilization in Y. lipolytica, we first studied homologous genes to OAF1 and PIP2 of Saccharomyces cerevisiae, but their disruption did not change growth on oleic acid at all. We next characterized a Y.more » lipolytica gene, POR1 (primary oleate regulator 1), an ortholog of farA encoding a transcriptional activator that regulates fatty acid utilization in Aspergillus nidulans. The deletion mutant of POR1 was defective in the growth on various fatty acids, but not on glucose, glycerol, or n-hexadecane. It exhibited slight defect on n-decane. The transcriptional induction of genes involved in {beta}-oxidation and peroxisome proliferation by oleate was distinctly diminished in the {Delta}por1 strains. These data suggest that POR1 encodes a transcriptional activator widely regulating fatty acid metabolism in Y. lipolytica.« less

Expression and Characterization of Glucose Oxidase from Aspergillus niger in Yarrowia lipolytica.

PubMed

Khadivi Derakshan, Fatemeh; Darvishi, Farshad; Dezfulian, Mehrouz; Madzak, Catherine

2017-08-01

Glucose oxidase (GOX) is currently used in clinical, pharmaceutical, food and chemical industries. The aim of this study was expression and characterization of Aspergillus niger glucose oxidase gene in the yeast Yarrowia lipolytica. For the first time, the GOX gene of A. niger was successfully expressed in Y. lipolytica using a mono-integrative vector containing strong hybrid promoter and secretion signal. The highest total glucose oxidase activity was 370 U/L after 7 days of cultivation. An innovative method was used to cell wall disruption in current study, and it could be recommended to use for efficiently cell wall disruption of Y. lipolytica. Optimum pH and temperature for recombinant GOX activity were 5.5 and 37 °C, respectively. A single band with a molecular weight of 80 kDa similar to the native and pure form of A. niger GOX was observed for the recombinant GOX in SDS-PAGE analysis. Y. lipolytica is a suitable and efficient eukaryotic expression system to production of recombinant GOX in compered with other yeast expression systems and could be used to production of pure form of GOX for industrial applications.

Impacts of environmental conditions on product formation and morphology of Yarrowia lipolytica.

PubMed

Timoumi, Asma; Guillouet, Stéphane E; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2018-05-01

The yeast Yarrowia lipolytica is an industrially important microorganism with distinctive physiological and metabolic characteristics. A variety of external factors (e.g., pH, temperature, and nutrient availability) influences the behavior of the yeast and may act as stress conditions which the cells must withstand and adapt. In this mini review, the impacts of environmental factors on the morphology and metabolite production by Y. lipolytica are summarized. In this regard, detailed insights into the effectors involved in the dimorphic transition of Y. lipolytica, the cultivation conditions employed, as well as the methods applied for the morphological characterization are highlighted. Concerning the metabolism products, a special focus is addressed on lipid and citric acid metabolites which have attracted significant attention in recent years. The dependence of lipid and citric acid productivity on key process parameters, such as media composition and physico-chemical variables, is thoroughly discussed. This review attempts to provide a recent update on the topic and will serve as a meaningful resource for researchers working in the field.

Overlapping reading frames at the LYS5 locus in the yeast Yarrowia lipolytica.

PubMed Central

Xuan, J W; Fournier, P; Declerck, N; Chasles, M; Gaillardin, C

1990-01-01

Mutants affected at the LYS5 locus of Yarrowia lipolytica lack detectable dehydrogenase (SDH) activity. The LYS5 gene has previously been cloned, and we present here the sequence of the 2.5-kilobase-pair (kb) DNA fragment complementing the lys5 mutation. Two large antiparallel open reading frames (ORF1 and ORF2) were observed, flanked by potential transcription signals. Both ORFs appear to be transcribed, but several lines of evidence suggest that only ORF2 is translated and encodes SDH. (i) The global amino acid compositions of Saccharomyces cerevisiae SDH and of the putative ORF2 product are similar and that of ORF1 is dissimilar. (ii) An in-frame translational fusion of ORF2 with the Escherichia coli lacZ gene was introduced into yeast cells and resulted in a beta-galactosidase activity regulated similarly to SDH; no beta-galactosidase activity was obtained with an in-frame fusion of ORF1 with lacZ. (iii) The introduction of a stop codon at the beginning of ORF2 prevented SDH expression in yeast cells, whereas no phenotypic effect was observed when ORF1 translation was blocked. Images PMID:2388625

Engineering Yarrowia lipolytica for Use in Biotechnological Applications: A Review of Major Achievements and Recent Innovations.

PubMed

Madzak, Catherine

2018-06-25

Yarrowia lipolytica is an oleaginous saccharomycetous yeast with a long history of industrial use. It aroused interest several decades ago as host for heterologous protein production. Thanks to the development of numerous molecular and genetic tools, Y. lipolytica is now a recognized system for expressing heterologous genes and secreting the corresponding proteins of interest. As genomic and transcriptomic tools increased our basic knowledge on this yeast, we can now envision engineering its metabolic pathways for use as whole-cell factory in various bioconversion processes. Y. lipolytica is currently being developed as a workhorse for biotechnology, notably for single-cell oil production and upgrading of industrial wastes into valuable products. As it becomes more and more difficult to keep up with an ever-increasing literature on Y. lipolytica engineering technology, this article aims to provide basic and actualized knowledge on this research area. The most useful reviews on Y. lipolytica biology, use, and safety will be evoked, together with a resume of the engineering tools available in this yeast. This mini-review will then focus on recently developed tools and engineering strategies, with a particular emphasis on promoter tuning, metabolic pathways assembly, and genome editing technologies.

Engineering towards a complete heterologous cellulase secretome in Yarrowia lipolytica reveals its potential for consolidated bioprocessing

DOE PAGES

Wei, Hui; Wang, Wei; Alahuhta, Markus; ...

2014-10-16

Background: Yarrowia lipolytica is an oleaginous yeast capable of metabolizing glucose to lipids, which then accumulate intracellularly. However, it lacks the suite of cellulolytic enzymes required to break down biomass cellulose and cannot therefore utilize biomass directly as a carbon source. Toward the development of a direct microbial conversion platform for the production of hydrocarbon fuels from cellulosic biomass, the potential for Y. lipolytica to function as a consolidated bioprocessing strain was investigated by first conducting a genomic search and functional testing of its endogenous glycoside hydrolases. Once the range of endogenous enzymes was determined, the critical cellulases from Trichodermamore » reesei were cloned into Yarrowia. Results: Initially, work to express T. reesei endoglucanase II (EGII) and cellobiohydrolase (CBH) II in Y. lipolytica resulted in the successful secretion of active enzymes. However, a critical cellulase, T. reesei CBHI, while successfully expressed in and secreted from Yarrowia, showed less than expected enzymatic activity, suggesting an incompatibility (probably at the post-translational level) for its expression in Yarrowia. This result prompted us to evaluate alternative or modified CBHI enzymes. Our subsequent expression of a T. reesei-Talaromyces emersonii (Tr-Te) chimeric CBHI, Chaetomium thermophilum CBHI, and Humicola grisea CBHI demonstrated remarkably improved enzymatic activities. Specifically, the purified chimeric Tr-Te CBHI showed a specific activity on Avicel that is comparable to that of the native T. reesei CBHI. Furthermore, the chimeric Tr-Te CBHI also showed significant synergism with EGII and CBHII in degrading cellulosic substrates, using either mixed supernatants or co-cultures of the corresponding Y. lipolytica transformants. The consortia system approach also allows rational volume mixing of the transformant cultures in accordance with the optimal ratio of cellulases required for efficient

The Strictly Aerobic Yeast Yarrowia lipolytica Tolerates Loss of a Mitochondrial DNA-Packaging Protein

PubMed Central

Bakkaiova, Jana; Arata, Kosuke; Matsunobu, Miki; Ono, Bungo; Aoki, Tomoyo; Lajdova, Dana; Nebohacova, Martina; Nosek, Jozef; Miyakawa, Isamu

2014-01-01

Mitochondrial DNA (mtDNA) is highly compacted into DNA-protein structures termed mitochondrial nucleoids (mt-nucleoids). The key mt-nucleoid components responsible for mtDNA condensation are HMG box-containing proteins such as mammalian mitochondrial transcription factor A (TFAM) and Abf2p of the yeast Saccharomyces cerevisiae. To gain insight into the function and organization of mt-nucleoids in strictly aerobic organisms, we initiated studies of these DNA-protein structures in Yarrowia lipolytica. We identified a principal component of mt-nucleoids in this yeast and termed it YlMhb1p (Y. lipolytica mitochondrial HMG box-containing protein 1). YlMhb1p contains two putative HMG boxes contributing both to DNA binding and to its ability to compact mtDNA in vitro. Phenotypic analysis of a Δmhb1 strain lacking YlMhb1p resulted in three interesting findings. First, although the mutant exhibits clear differences in mt-nucleoids accompanied by a large decrease in the mtDNA copy number and the number of mtDNA-derived transcripts, its respiratory characteristics and growth under most of the conditions tested are indistinguishable from those of the wild-type strain. Second, our results indicate that a potential imbalance between subunits of the respiratory chain encoded separately by nuclear DNA and mtDNA is prevented at a (post)translational level. Third, we found that mtDNA in the Δmhb1 strain is more prone to mutations, indicating that mtHMG box-containing proteins protect the mitochondrial genome against mutagenic events. PMID:24972935

Comprehensive metabolomic, lipidomic and microscopic profiling of Yarrowia lipolytica during lipid accumulation identifies targets for increased lipogenesis

DOE PAGES

Pomraning, Kyle R.; Wei, Siwei; Karagiosis, Sue A.; ...

2015-04-23

Yarrowia lipolytica is an oleaginous ascomycete yeast that accumulates large amounts of lipids and has potential as a biofuel producing organism. Despite a growing scientific literature focused on lipid production by Y. lipolytica, there remain significant knowledge gaps regarding the key biological processes involved. We applied a combination of metabolomic and lipidomic profiling approaches as well as microscopic techniques to identify and characterize the key pathways involved in de novo lipid accumulation from glucose in batch cultured, wild-type Y. lipolytica. We found that lipids accumulated rapidly and peaked at 48 hours during the five day experiment, concurrent with a shiftmore » in amino acid metabolism. We also report that Y. lipolytica secretes disaccharides early in batch culture and reabsorbs them when extracellular glucose is depleted. Exhaustion of extracellular sugars coincided with thickening of the cell wall, suggesting that genes involved in cell wall biogenesis may be a useful target for improving the efficiency of lipid producing yeast strains.« less

A comparative study on glycerol metabolism to erythritol and citric acid in Yarrowia lipolytica yeast cells.

PubMed

Tomaszewska, Ludwika; Rakicka, Magdalena; Rymowicz, Waldemar; Rywińska, Anita

2014-09-01

Citric acid and erythritol biosynthesis from pure and crude glycerol by three acetate-negative mutants of Yarrowia lipolytica yeast was investigated in batch cultures in a wide pH range (3.0-6.5). Citric acid biosynthesis was the most effective at pH 5.0-5.5 in the case of Wratislavia 1.31 and Wratislavia AWG7. With a decreasing pH value, the direction of biosynthesis changed into erythritol synthesis accompanied by low production of citric acid. Pathways of glycerol conversion into erythritol and citric acid were investigated in Wratislavia K1 cells. Enzymatic activity was compared in cultures run at pH 3.0 and 4.5, that is, under conditions promoting the production of erythritol and citric acid, respectively. The effect of pH value (3.0 and 4.5) and NaCl presence on the extracellular production and intracellular accumulation of citric acid and erythritol was compared as well. Low pH and NaCl resulted in diminished activity of glycerol kinase, whereas such conditions stimulated the activity of glycerol-3-phosphate dehydrogenase. The presence of NaCl strongly influenced enzymes activity - the effective erythritol production was correlated with a high activity of transketolase and erythrose reductase. Therefore, presented results confirmed that transketolase and erythrose reductase are involved in the overproduction of erythritol in the cells of Y. lipolytica yeast. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Synthetic Biology Expands the Industrial Potential of Yarrowia lipolytica.

PubMed

Markham, Kelly A; Alper, Hal S

2018-06-04

The oleaginous yeast Yarrowia lipolytica is quickly emerging as the most popular non-conventional (i.e., non-model organism) yeast in the bioproduction field. With a high propensity for flux through tricarboxylic acid (TCA) cycle intermediates and biological precursors such as acetyl-CoA and malonyl-CoA, this host is especially well suited to meet our industrial chemical production needs. Recent progress in synthetic biology tool development has greatly enhanced our ability to rewire this organism, with advances in genetic component design, CRISPR technologies, and modular cloning strategies. In this review we investigate recent developments in metabolic engineering and describe how the new tools being developed help to realize the full industrial potential of this host. Finally, we conclude with our vision of the developments that will be necessary to enhance future engineering efforts. Copyright © 2018 Elsevier Ltd. All rights reserved.

Yarrowia lipolytica possesses two plasma membrane alkali metal cation/H+ antiporters with different functions in cell physiology.

PubMed

Papouskova, Klara; Sychrova, Hana

2006-04-03

The family of Nha antiporters mediating the efflux of alkali metal cations in exchange for protons across the plasma membrane is conserved in all yeast species. Yarrowia lipolytica is a dimorphic yeast, phylogenetically very distant from the model yeast Saccharomyces cerevisiae. A search in its sequenced genome revealed two genes (designated as YlNHA1 and YlNHA2) with homology to the S. cerevisiae NHA1 gene, which encodes a plasma membrane alkali metal cation/H+ antiporter. Upon heterologous expression of both YlNHA genes in S. cerevisiae, we showed that Y. lipolytica antiporters differ not only in length and sequence, but also in their affinity for individual substrates. While the YlNha1 protein mainly increased cell tolerance to potassium, YlNha2p displayed a remarkable transport capacity for sodium. Thus, Y. lipolytica is the first example of a yeast species with two plasma membrane alkali metal cation/H+ antiporters differing in their putative functions in cell physiology; cell detoxification vs. the maintenance of stable intracellular pH, potassium content and cell volume.

Overproduction of Fatty Acid Ethyl Esters by the Oleaginous Yeast Yarrowia lipolytica through Metabolic Engineering and Process Optimization.

PubMed

Gao, Qi; Cao, Xuan; Huang, Yu-Ying; Yang, Jing-Lin; Chen, Jun; Wei, Liu-Jing; Hua, Qiang

2018-05-18

Recent advances in the production of biofuels by microbes have attracted attention due to increasingly limited fossil fuels. Biodiesels, especially fatty acid ethyl esters (FAEEs), are considered a potentially fully sustainable fuel in the near future due to similarities with petrodiesels and compatibility with existing infrastructure. However, biosynthesis of FAEEs is limited by the supply of precursor lipids and acetyl-CoA. In the present study, we explored the production potential of an engineered biosynthetic pathway coupled to the addition of ethanol in the oleaginous yeast Yarrowia lipolytica. This type of yeast is able to supply a greater amount of precursor lipids than species typically used. To construct the FAEEs synthesis pathway, WS genes that encode wax ester synthases (WSs) from different species were codon-optimized and heterologously expressed in Y. lipolytica. The most productive engineered strain was found to express a WS gene from Marinobacter hydrocarbonoclasticus strain DSM 8798. To stepwisely increase FAEEs production, we optimized the promoter of WS overexpression, eliminated β-oxidation by deleting the PEX10 gene in our engineered strains, and redirected metabolic flux toward acetyl-CoA. The new engineered strain, coupled with an optimized ethanol concentration, led to an approximate 5.5-fold increase in extracellular FAEEs levels compared to the wild-type strain and a maximum FAEEs titer of 1.18 g/L in shake flask cultures. In summary, the present study demonstrated that an engineered Y. lipolytica strain possessed a high capacity for FAEEs production and may serve as a platform for more efficient biodiesel production in the future.

New finding and optimal production of a novel extracellular alkaline lipase from Yarrowia lipolytica NRRL Y-2178.

PubMed

Lee, Geon-Ho; Bae, Jae-Han; Suh, Min-Jung; Kim, In-Hwan; Hou, Ching T; Kim, Hak-Ryul

2007-06-01

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions including hydrolysis of triglycerides, transesterification, and chiral synthesis of esters under natural conditions. Although lipases from various sources have been widely used in industrial applications, such as in food, chemical, pharmaceutical, and detergent industries, there are still substantial current interests in developing new microbial lipases, specifically those functioning in abnormal conditions. We screened 17 lipase-producing yeast strains, which were prescreened for substrate specificity of lipase from more than 500 yeast strains from the Agricultural Research Service Culture Collection (Peoria, IL, U.S.A.), and selected Yarrowia lipolytica NRRL Y-2178 as a best lipase producer. This report presents new finding and optimal production of a novel extracellular alkaline lipase from Y. lipolytica NRRL Y-2178. Optimal c ulture conditions f orlipase production by Y. lipolytica NRRL Y-2178 were 72 h incubation time, 27.5 degrees C, pH 9.0. Glycerol and glucose were efficiently used as the most efficient carbon sources, and a combination of yeast extract and peptone was a good nitrogen source for lipase production by Y. lipolytica NRRL Y-2178. These results suggested that Y. lipolytica NRRL Y-2178 showsgood industrial potential as a new alkaline lipase producer.

Irradiation of Yarrowia lipolytica NRRL YB-567 creating novel strains with enhanced ammonia and oil production on protein and carbohydrate substrates

USDA-ARS?s Scientific Manuscript database

Increased interest in sustainable production of renewable diesel and other valuable bioproducts is redoubling efforts to improve economic feasibility of microbial-based oil production. The yeast Yarrowia lipolytica is capable of employing a wide variety of substrates to produce oil and valuable co-p...

Multiplex gene editing of the Yarrowia lipolytica genome using the CRISPR-Cas9 system.

PubMed

Gao, Shuliang; Tong, Yangyang; Wen, Zhiqiang; Zhu, Li; Ge, Mei; Chen, Daijie; Jiang, Yu; Yang, Sheng

2016-08-01

Yarrowia lipolytica is categorized as a generally recognized as safe (GRAS) organism and is a heavily documented, unconventional yeast that has been widely incorporated into multiple industrial fields to produce valuable biochemicals. This study describes the construction of a CRISPR-Cas9 system for genome editing in Y. lipolytica using a single plasmid (pCAS1yl or pCAS2yl) to transport Cas9 and relevant guide RNA expression cassettes, with or without donor DNA, to target genes. Two Cas9 target genes, TRP1 and PEX10, were repaired by non-homologous end-joining (NHEJ) or homologous recombination, with maximal efficiencies in Y. lipolytica of 85.6 % for the wild-type strain and 94.1 % for the ku70/ku80 double-deficient strain, within 4 days. Simultaneous double and triple multigene editing was achieved with pCAS1yl by NHEJ, with efficiencies of 36.7 or 19.3 %, respectively, and the pCASyl system was successfully expanded to different Y. lipolytica breeding strains. This timesaving method will enable and improve synthetic biology, metabolic engineering and functional genomic studies of Y. lipolytica.

Unraveling fatty acid transport and activation mechanisms in Yarrowia lipolytica.

PubMed

Dulermo, Rémi; Gamboa-Meléndez, Heber; Ledesma-Amaro, Rodrigo; Thévenieau, France; Nicaud, Jean-Marc

2015-09-01

Fatty acid (FA) transport and activation have been extensively studied in the model yeast species Saccharomyces cerevisiae but have rarely been examined in oleaginous yeasts, such as Yarrowia lipolytica. Because the latter begins to be used in biodiesel production, understanding its FA transport and activation mechanisms is essential. We found that Y. lipolytica has FA transport and activation proteins similar to those of S. cerevisiae (Faa1p, Pxa1p, Pxa2p, Ant1p) but mechanism of FA peroxisomal transport and activation differs greatly with that of S. cerevisiae. While the ScPxa1p/ScPxa2p heterodimer is essential for growth on long-chain FAs, ΔYlpxa1 ΔYlpxa2 is not impaired for growth on FAs. Meanwhile, ScAnt1p and YlAnt1p are both essential for yeast growth on medium-chain FAs, suggesting they function similarly. Interestingly, we found that the ΔYlpxa1 ΔYlpxa2 ΔYlant1 mutant was unable to grow on short-, medium-, or long-chain FAs, suggesting that YlPxa1p, YlPxa2p, and YlAnt1p belong to two different FA degradation pathways. We also found that YlFaa1p is involved in FA storage in lipid bodies and that FA remobilization largely depended on YlFat1p, YlPxa1p and YlPxa2p. This study is the first to comprehensively examine FA intracellular transport and activation in oleaginous yeast. Copyright © 2015. Published by Elsevier B.V.

Characterization of phosphatidic acid phosphatase activity in the oleaginous yeast Yarrowia lipolytica and its role in lipid biosynthesis.

PubMed

Hardman, Derell; McFalls, Daniel; Fakas, Stylianos

2017-02-01

Phosphatidic acid phosphatase (PAP) catalyses the committed step of triacylglycerol (TAG) biosynthesis and thus regulates the amounts of TAG produced by the cell. TAG is the target of biotechnological processes developed for the production of food lipids or biofuels. These processes are using oleaginous microorganisms like the yeast Yarrowia lipolytica as the TAG producers. Thus manipulating key enzymatic activities like PAP in Y. lipolytica could drive lipid biosynthesis towards TAG production and increase TAG yields. In this study, PAP activity in Y. lipolytica was characterized in detail and its role in lipid biosynthesis was addressed. PAP activity increased 2.5-fold with the addition of Mg 2+ (1 mm) in the assay mixture, which means that most of the PAP activity was due to Mg 2+ -dependent PAP enzymes (e.g. Pah1, App1). In contrast, N-ethylmaleimide (NEM) potently inhibited PAP activity, indicating the presence of NEM-sensitive PAP enzymes (e.g. App1, Lpp1). Localization studies revealed that the majority of PAP activity resides in the membrane fraction, while the cytosolic fraction harbours only a small amount of activity. PAP activity was regulated in a growth-dependent manner, being induced at the early exponential phase and declining thereafter. PAP activity did not correlate with TAG synthesis, which increased as cells progressed from the exponential phase to the early stationary phase. In stationary phase, TAG was mobilized with the concomitant synthesis of sterols and sterol esters. These results provide the first insights into the role of PAP in lipid biosynthesis by Y. lipolytica. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Engineering Yarrowia lipolytica as a platform for synthesis of drop-in transportation fuels and oleochemicals

PubMed Central

Xu, Peng; Qiao, Kangjian; Ahn, Woo Suk; Stephanopoulos, Gregory

2016-01-01

Harnessing lipogenic pathways and rewiring acyl-CoA and acyl-ACP (acyl carrier protein) metabolism in Yarrowia lipolytica hold great potential for cost-efficient production of diesel, gasoline-like fuels, and oleochemicals. Here we assessed various pathway engineering strategies in Y. lipolytica toward developing a yeast biorefinery platform for sustainable production of fuel-like molecules and oleochemicals. Specifically, acyl-CoA/acyl-ACP processing enzymes were targeted to the cytoplasm, peroxisome, or endoplasmic reticulum to generate fatty acid ethyl esters and fatty alkanes with tailored chain length. Activation of endogenous free fatty acids and the subsequent reduction of fatty acyl-CoAs enabled the efficient synthesis of fatty alcohols. Engineering a hybrid fatty acid synthase shifted the free fatty acids to a medium chain-length scale. Manipulation of alternative cytosolic acetyl-CoA pathways partially decoupled lipogenesis from nitrogen starvation and unleashed the lipogenic potential of Y. lipolytica. Taken together, the strategies reported here represent promising steps to develop a yeast biorefinery platform that potentially upgrades low-value carbons to high-value fuels and oleochemicals in a sustainable and environmentally friendly manner. PMID:27621436

Genome-scale model-driven strain design for dicarboxylic acid production in Yarrowia lipolytica.

PubMed

Mishra, Pranjul; Lee, Na-Rae; Lakshmanan, Meiyappan; Kim, Minsuk; Kim, Byung-Gee; Lee, Dong-Yup

2018-03-19

Recently, there have been several attempts to produce long-chain dicarboxylic acids (DCAs) in various microbial hosts. Of these, Yarrowia lipolytica has great potential due to its oleaginous characteristics and unique ability to utilize hydrophobic substrates. However, Y. lipolytica should be further engineered to make it more competitive: the current approaches are mostly intuitive and cumbersome, thus limiting its industrial application. In this study, we proposed model-guided metabolic engineering strategies for enhanced production of DCAs in Y. lipolytica. At the outset, we reconstructed genome-scale metabolic model (GSMM) of Y. lipolytica (iYLI647) by substantially expanding the previous models. Subsequently, the model was validated using three sets of published culture experiment data. It was finally exploited to identify genetic engineering targets for overexpression, knockout, and cofactor modification by applying several in silico strain design methods, which potentially give rise to high yield production of the industrially relevant long-chain DCAs, e.g., dodecanedioic acid (DDDA). The resultant targets include (1) malate dehydrogenase and malic enzyme genes and (2) glutamate dehydrogenase gene, in silico overexpression of which generated additional NADPH required for fatty acid synthesis, leading to the increased DDDA fluxes by 48% and 22% higher, respectively, compared to wild-type. We further investigated the effect of supplying branched-chain amino acids on the acetyl-CoA turn-over rate which is key metabolite for fatty acid synthesis, suggesting their significance for production of DDDA in Y. lipolytica. In silico model-based strain design strategies allowed us to identify several metabolic engineering targets for overproducing DCAs in lipid accumulating yeast, Y. lipolytica. Thus, the current study can provide a methodological framework that is applicable to other oleaginous yeasts for value-added biochemical production.

Mitochondria from Dipodascus (Endomyces) magnusii and Yarrowia lipolytica yeasts did not undergo a Ca²⁺-dependent permeability transition even under anaerobic conditions.

PubMed

Trendeleva, Tat'yana; Sukhanova, Evgeniya; Ural'skaya, Ludmila; Saris, Nils-Erik; Zvyagilskaya, Renata

2011-12-01

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts. The two yeast strains are good alternatives to Saccharomyces cerevisiae, being aerobes containing well structured mitochondria (thus ensuring less structural limitation to observe their appreciable swelling) and fully competent respiratory chain with three invariantly functioning energy conservation points, including Complex I, that can be involved in induction of the canonical Ca²⁺/P(i)-dependent mitochondrial permeability transition (mPTP pore) with an increased open probability when electron flux increases(Fontaine et al. J Biol Chem 273: 25734–25740, 1998; Bernardi et al. FEBS J 273:2077–2099, 2006). High amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating pore opening. Previously (Kovaleva et al. J Bioenerg Biomembr 41:239–249, 2009; Kovaleva et al. Biochemistry (Moscow) 75: 297–303, 2010) we have shown that mitochondria from Y.lipolytica and D. magnusii were very resistant to the Ca²⁺overload combined with varying concentrations of P(i),palmitic acid, SH-reagents, carboxyatractyloside (an inhibitor of ADP/ATP translocator), as well as depletion of intramitochondrial adenine nucleotide pools, deenergization of mitochondria, and shifting to acidic pH values in the presence of high [P(i)]. Here we subjected yeast mitochondria to other conditions known to induce an mPTP in animal and plant mitochondria, namely to Ca²⁺ overload under hypoxic conditions (anaerobiosis). We were unable to observe Ca²⁺-induced high permeability of the inner membrane of D. magnusii and Y. lipolytica yeast mitochondria under anaerobic conditions, thus suggesting that an mPTP-like pore, if it ever occurs in yeast mitochondria, is not coupled with the Ca²⁺ uptake. The results provide the first demonstration of ATP-dependent energization of yeast mitochondria under conditions of anaerobiosis.

Urea and urine are a viable and cost-effective nitrogen source for Yarrowia lipolytica biomass and lipid accumulation.

PubMed

Brabender, Matthew; Hussain, Murtaza Shabbir; Rodriguez, Gabriel; Blenner, Mark A

2018-03-01

Yarrowia lipolytica is an industrial yeast that has been used in the sustainable production of fatty acid-derived and lipid compounds due to its high growth capacity, genetic tractability, and oleaginous properties. This investigation examines the possibility of utilizing urea or urine as an alternative to ammonium sulfate as a nitrogen source to culture Y. lipolytica. The use of a stoichiometrically equivalent concentration of urea in lieu of ammonium sulfate significantly increased cell growth when glucose was used as the carbon source. Furthermore, Y. lipolytica growth was equally improved when grown with synthetic urine and real human urine. Equivalent or better lipid production was achieved when cells are grown on urea or urine. The successful use of urea and urine as nitrogen sources for Y. lipolytica growth highlights the potential of using cheaper media components as well as exploiting and recycling non-treated human waste streams for biotechnology processes.

Engineering ..beta..-Oxidation in Yarrowia lipolytica for Methyl Ketone Production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sanchez i Nogue, Violeta; Ramirez, Kelsey J; Singer, Christine

Medium- and long-chain methyl ketones are fatty acid-derived compounds that can be used as biofuel blending agents, flavors and fragrances. However, their large-scale production from sustainable feedstocks is currently limited due to the lack of robust microbial biocatalysts. The oleaginous yeast Yarrowia lipolytica is a promising biorefinery platform strain for the production of methyl ketones from renewable lignocellulosic biomass due to its natively high flux towards fatty acid biosynthesis. In this study, we report the metabolic engineering of Y. lipolytica to produce long- and very long-chain methyl ketones. Truncation of peroxisomal ..beta..-oxidation by chromosomal deletion of pot1 resulted in themore » biosynthesis of saturated, mono-, and diunsaturated methyl ketones in the C13-C23 range. Additional overexpression and peroxisomal targeting of a heterologous bacterial methyl ketone biosynthesis pathway yielded an initial titer of 151.5 mg/L of saturated methyl ketones. Dissolved oxygen concentrations in the cultures were found to substantially impact cell morphology and methyl ketone biosynthesis. Bioreactor cultivation under optimized conditions resulted in a titer of 314.8 mg/L of total methyl ketones, representing more than a 6000-fold increase over the parental strain. This work highlights the potential of Y. lipolytica to serve as chassis organism for the biosynthesis of acyl-thioester derived long- and very long-chain methyl ketones.« less

MHY1 Encodes a C2H2-Type Zinc Finger Protein That Promotes Dimorphic Transition in the Yeast Yarrowia lipolytica

PubMed Central

Hurtado, Cleofe A. R.; Rachubinski, Richard A.

1999-01-01

The yeast-to-hypha morphological transition (dimorphism) is typical of many pathogenic fungi. Dimorphism has been attributed to changes in temperature and nutritional status and is believed to constitute a mechanism of response to adverse conditions. We have isolated and characterized a gene, MHY1, whose transcription is dramatically increased during the yeast-to-hypha transition in Yarrowia lipolytica. Deletion of MHY1 is viable and has no effect on mating, but it does result in a complete inability of cells to undergo mycelial growth. MHY1 encodes a C2H2-type zinc finger protein, Mhy1p, which can bind putative cis-acting DNA stress response elements, suggesting that Mhy1p may act as a transcription factor. Interestingly, Mhy1p tagged with a hemagglutinin epitope was concentrated in the nuclei of actively growing cells found at the hyphal tip. PMID:10322005

A novel multigene expression construct for modification of glycerol metabolism in Yarrowia lipolytica

PubMed Central

2013-01-01

Background High supply of raw, residual glycerol from biodiesel production plants promote the search for novel biotechnological methods of its utilization. In this study we attempted modification of glycerol catabolism in a nonconventional yeast species Yarrowia lipolytica through genetic engineering approach. Results To address this, we developed a novel genetic construct which allows transferring three heterologous genes, encoding glycerol dehydratase, its reactivator and a wide-spectrum alcohol oxidoreductase under the control of glycerol-induced promoter. The three genes, tandemly arrayed in an expression cassette with a marker gene ura3, regulatory and targeting sequences (G3P dh promoter and XPR-like terminator, 28S rDNA as a target locus), were transferred into Yarrowia lipolytica cells. The obtained recombinant strain NCYC3825 was characterized at the molecular level and with respect to its biotechnological potential. Our experiments indicated that the novel recombinant strain stably borne one copy of the expression cassette and efficiently expressed heterologous alcohol oxidoreductase, while glycerol dehydratase and its reactivator were expressed at lower level. Comparative shake flask cultivations in glucose- and glycerol-based media demonstrated higher biomass production by the recombinant strain when glycerol was the main carbon source. During bioreactor (5 L) fed-batch cultivation in glycerol-based medium, the recombinant strain was characterized by relatively high biomass and lipids accumulation (up to 42 gDCW L-1, and a peak value of 38%LIPIDS of DCW, respectively), and production of high titers of citric acid (59 g L-1) and 2-phenylethanol (up to 1 g L-1 in shake flask cultivation), which are industrially attractive bioproducts. Conclusions Due to heterogeneous nature of the observed alterations, we postulate that the main driving force of the modified phenotype was faster growth in glycerol-based media, triggered by modifications in the red

Development of a novel rDNA based plasmid for enhanced cell surface display on Yarrowia lipolytica.

PubMed

Bulani, Siyavuya Ishmael; Moleleki, Lucy; Albertyn, Jacobus; Moleleki, Ntsane

2012-05-20

In this study, a novel rDNA based plasmid was developed for display of heterologous proteins on the cell surface of Yarrowia lipolytica using the C-terminal end of the glycosylphosphatidylinositol (GPI) anchored Y. lipolytica cell wall protein 1 (YlCWP1). mCherry was used as a model protein to assess the efficiency of the constructed plasmid. Y. lipolytica transformants harbouring the expression cassettes showed a purple colour phenotype on selective YNB-casamino plates as compared to control cells indicating that mCherry was displayed on the cells. Expression of mCherry on cells of Y. lipolytica was confirmed by both fluorescent microscopy and flow cytometry. Furthermore, SDS-PAGE analysis and matrix-assisted laser desorption/ionization (MALDI)-time-of (TOF)-mass spectrometry (MS) peptide mass fingerprinting (PMF) confirmed that the protein cleaved from the yeast cells using enterokinase was mCherry. Efficient cleavage of mCherry reported in this work offers an alternative purification method for displayed heterologous proteins on Y. lipolytica cells using the plasmid constructed in this study. The developed displaying system offers great potential for industrial production and purification of heterologous proteins at low cost.

Storage lipids of yeasts: a survey of nonpolar lipid metabolism in Saccharomyces cerevisiae, Pichia pastoris, and Yarrowia lipolytica.

PubMed

Koch, Barbara; Schmidt, Claudia; Daum, Günther

2014-09-01

Biosynthesis and storage of nonpolar lipids, such as triacylglycerols (TG) and steryl esters (SE), have gained much interest during the last decades because defects in these processes are related to severe human diseases. The baker's yeast Saccharomyces cerevisiae has become a valuable tool to study eukaryotic lipid metabolism because this single-cell microorganism harbors many enzymes and pathways with counterparts in mammalian cells. In this article, we will review aspects of TG and SE metabolism and turnover in the yeast that have been known for a long time and combine them with new perceptions of nonpolar lipid research. We will provide a detailed insight into the mechanisms of nonpolar lipid synthesis, storage, mobilization, and degradation in the yeast S. cerevisiae. The central role of lipid droplets (LD) in these processes will be addressed with emphasis on the prevailing view that this compartment is more than only a depot for TG and SE. Dynamic and interactive aspects of LD with other organelles will be discussed. Results obtained with S. cerevisiae will be complemented by recent investigations of nonpolar lipid research with Yarrowia lipolytica and Pichia pastoris. Altogether, this review article provides a comprehensive view of nonpolar lipid research in yeast. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Induction of a non-specific permeability transition in mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts.

PubMed

Kovaleva, Mariya V; Sukhanova, Evgeniya I; Trendeleva, Tatyana A; Zyl'kova, Marina V; Ural'skaya, Ludmila A; Popova, Kristina M; Saris, Nils-Erik L; Zvyagilskaya, Renata A

2009-06-01

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts, possessing a respiratory chain with the usual three points of energy conservation. High-amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating induction of the mitochondrial permeability transition due to opening of a pore (mPTP). Mitochondria from Y. lipolytica, lacking a natural mitochondrial Ca(2+) uptake pathway, and from D. magnusii, harboring a high-capacitive, regulated mitochondrial Ca(2+) transport system (Bazhenova et al. J Biol Chem 273:4372-4377, 1998a; Bazhenova et al. Biochim Biophys Acta 1371:96-100, 1998b; Deryabina and Zvyagilskaya Biochemistry (Moscow) 65:1352-1356, 2000; Deryabina et al. J Biol Chem 276:47801-47806, 2001) were very resistant to Ca(2+) overload. However, exposure of yeast mitochondria to 50-100 microM Ca(2+) in the presence of the Ca(2+) ionophore ETH129 induced collapse of the membrane potential, possibly due to activation of the fatty acid-dependent Ca(2+)/nH(+)-antiporter, with no classical mPTP induction. The absence of response in yeast mitochondria was not simply due to structural limitations, since large-amplitude swelling occurred in the presence of alamethicin, a hydrophobic, helical peptide, forming voltage-sensitive ion channels in lipid membranes. Ca(2+)- ETH129-induced activation of the Ca(2+)/H(+)-antiport system was inhibited and prevented by bovine serum albumin, and partially by inorganic phosphate and ATP. We subjected yeast mitochondria to other conditions known to induce the permeability transition in animal mitochondria, i.e., Ca(2+) overload (in the presence of ETH129) combined with palmitic acid (Mironova et al. J Bioenerg Biomembr 33:319-331, 2001; Sultan and Sokolove Arch Biochem Biophys 386:37-51, 2001), SH-reagents, carboxyatractyloside (an inhibitor of the ADP/ATP translocator), depletion of intramitochondrial adenine nucleotide pools

Draft Genome Sequence of the Dimorphic Yeast Yarrowia lipolytica Strain W29

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Baker, Scott E.

Here, we present the draft genome sequence of the dimorphic ascomycete yeastYarrowia lipolyticastrain W29 (ATCC 20460).Y. lipolyticais a commonly employed model for the industrial production of lipases, small molecules, and more recently for its ability to accumulate lipids.

Production of oils and fats by oleaginous microorganisms with an emphasis given to the potential of the nonconventional yeast Yarrowia lipolytica.

PubMed

Carsanba, E; Papanikolaou, S; Erten, H

2018-05-15

Recently, there has been a great upsurge of interest in studies related to several aspects of microbial lipid production, which is one of the top topics in relevant research fields due to the high demand of these fatty materials in food, medical, oleochemical and biofuel industries. Lipid accumulation by the so-called "oleaginous microorganisms" can generate more than 20% w/w of oil in dry biomass and is governed by a plethora of parameters, such as medium pH, incubation temperature, nutrient limitation and C/N (carbon/nitrogen) ratio, which drastically affect the lipid production bioprocess. Until now, considerable work has been undertaken to find the cheapest substrate to enable lipid fermentation by oleaginous microorganisms. This review principally details information regarding microbial lipids, suitable production conditions and focuses attention on using the yeast Yarrowia lipolytica to achieve these objectives. Lipid production by this yeast is discussed and the necessary conditions and suitable substrates are reviewed.

Optimization of odd chain fatty acid production by Yarrowia lipolytica.

PubMed

Park, Young-Kyoung; Dulermo, Thierry; Ledesma-Amaro, Rodrigo; Nicaud, Jean-Marc

2018-01-01

Odd chain fatty acids (odd FAs) have a wide range of applications in therapeutic and nutritional industries, as well as in chemical industries including biofuel. Yarrowia lipolytica is an oleaginous yeast considered a preferred microorganism for the production of lipid-derived biofuels and chemicals. However, it naturally produces negligible amounts of odd chain fatty acids. The possibility of producing odd FAs using Y. lipolytica was investigated. Y. lipolytica wild-type strain was shown able to grow on weak acids; acetate, lactate, and propionate. Maximal growth rate on propionate reached 0.24 ± 0.01 h -1 at 2 g/L, and growth inhibition occurred at concentration above 10 g/L. Wild-type strain accumulated lipids ranging from 7.39 to 8.14% (w/w DCW) depending on the carbon source composition, and odd FAs represented only 0.01-0.12 g/L. We here proved that the deletion of the PHD1 gene improved odd FAs production, which reached a ratio of 46.82% to total lipids. When this modification was transferred to an obese strain, engineered for improving lipid accumulation, further increase odd FAs production reaching a total of 0.57 g/L was shown. Finally, a fed-batch co-feeding strategy was optimized for further increase odd FAs production, which generated 0.75 g/L, the best production described so far in Y. lipolytica . A Y. lipolytica strain able to accumulate high level of odd chain fatty acids, mainly heptadecenoic acid, has been successfully developed. In addition, a fed-batch co-feeding strategy was optimized to further improve lipid accumulation and odd chain fatty acid content. These lipids enriched in odd chain fatty acid can (1) improve the properties of the biodiesel generated from Y. lipolytica lipids and (2) be used as renewable source of odd chain fatty acid for industrial applications. This work paves the way for further improvements in odd chain fatty acids and fatty acid-derived compound production.

Engineering β-oxidation in Yarrowia lipolytica for methyl ketone production.

PubMed

Hanko, Erik K R; Denby, Charles M; Sànchez I Nogué, Violeta; Lin, Weiyin; Ramirez, Kelsey J; Singer, Christine A; Beckham, Gregg T; Keasling, Jay D

2018-05-28

Medium- and long-chain methyl ketones are fatty acid-derived compounds that can be used as biofuel blending agents, flavors and fragrances. However, their large-scale production from sustainable feedstocks is currently limited due to the lack of robust microbial biocatalysts. The oleaginous yeast Yarrowia lipolytica is a promising biorefinery platform strain for the production of methyl ketones from renewable lignocellulosic biomass due to its natively high flux towards fatty acid biosynthesis. In this study, we report the metabolic engineering of Y. lipolytica to produce long- and very long-chain methyl ketones. Truncation of peroxisomal β-oxidation by chromosomal deletion of pot1 resulted in the biosynthesis of saturated, mono-, and diunsaturated methyl ketones in the C 13 -C 23 range. Additional overexpression and peroxisomal targeting of a heterologous bacterial methyl ketone biosynthesis pathway yielded an initial titer of 151.5 mg/L of saturated methyl ketones. Dissolved oxygen concentrations in the cultures were found to substantially impact cell morphology and methyl ketone biosynthesis. Bioreactor cultivation under optimized conditions resulted in a titer of 314.8 mg/L of total methyl ketones, representing more than a 6000-fold increase over the parental strain. This work highlights the potential of Y. lipolytica to serve as chassis organism for the biosynthesis of acyl-thioester derived long- and very long-chain methyl ketones. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Functional overexpression and characterization of lipogenesis-related genes in the oleaginous yeast Yarrowia lipolytica.

PubMed

Silverman, Andrew M; Qiao, Kangjian; Xu, Peng; Stephanopoulos, Gregory

2016-04-01

Single cell oil (SCO) is an attractive energy source due to scalability, utilization of low-cost renewable feedstocks, and type of product(s) made. Engineering strains capable of producing high lipid titers and yields is crucial to the economic viability of these processes. However, lipid synthesis in cells is a complex phenomenon subject to multiple layers of regulation, making gene target identification a challenging task. In this study, we aimed to identify genes in the oleaginous yeast Yarrowia lipolytica whose overexpression enhances lipid production by this organism. To this end, we examined the effect of the overexpression of a set of 44 native genes on lipid production in Y. lipolytica, including those involved in glycerolipid synthesis, fatty acid synthesis, central carbon metabolism, NADPH generation, regulation, and metabolite transport and characterized each resulting strain's ability to produce lipids growing on both glucose and acetate as a sole carbon source. Our results suggest that a diverse subset of genes was effective at individually influencing lipid production in Y. lipolytica, sometimes in a substrate-dependent manner. The most productive strain on glucose overexpressed the diacylglycerol acyltransferase DGA2 gene, increasing lipid titer, cellular content, and yield by 236, 165, and 246 %, respectively, over our control strain. On acetate, our most productive strain overexpressed the acylglycerol-phosphate acyltransferase SLC1 gene, with a lipid titer, cellular content, and yield increase of 99, 91, and 151 %, respectively, over the control strain. Aside from genes encoding enzymes that directly catalyze the reactions of lipid synthesis, other ways by which lipogenesis was increased in these cells include overexpressing the glycerol-3-phosphate dehydrogenase (GPD1) gene to increase production of glycerol head groups and overexpressing the 6-phosphogluconolactonase (SOL3) gene from the oxidative pentose phosphate pathway to increase NADPH

SOA genes encode proteins controlling lipase expression in response to triacylglycerol utilization in the yeast Yarrowia lipolytica.

PubMed

Desfougères, Thomas; Haddouche, Ramdane; Fudalej, Franck; Neuvéglise, Cécile; Nicaud, Jean-Marc

2010-02-01

The oleaginous yeast Yarrowia lipolytica efficiently metabolizes hydrophobic substrates such as alkanes, fatty acids or triacylglycerol. This yeast has been identified in oil-polluted water and in lipid-rich food. The enzymes involved in lipid breakdown, for use as a carbon source, are known, but the molecular mechanisms controlling the expression of the genes encoding these enzymes are still poorly understood. The study of mRNAs obtained from cells grown on oleic acid identified a new group of genes called SOA genes (specific for oleic acid). SOA1 and SOA2 are two small genes coding for proteins with no known homologs. Single- and double-disrupted strains were constructed. Wild-type and mutant strains were grown on dextrose, oleic acid and triacylglycerols. The double mutant presents a clear phenotype consisting of a growth defect on tributyrin and triolein, but not on dextrose or oleic acid media. Lipase activity was 50-fold lower in this mutant than in the wild-type strain. The impact of SOA deletion on the expression of the main extracellular lipase gene (LIP2) was monitored using a LIP2-beta-galactosidase promoter fusion protein. These data suggest that Soa proteins are components of a molecular mechanism controlling lipase gene expression in response to extracellular triacylglycerol.

Sugar versus fat: elimination of glycogen storage improves lipid accumulation in Yarrowia lipolytica.

PubMed

Bhutada, Govindprasad; Kavšcek, Martin; Ledesma-Amaro, Rodrigo; Thomas, Stéphane; Rechberger, Gerald N; Nicaud, Jean-Marc; Natter, Klaus

2017-05-01

Triacylglycerol (TAG) and glycogen are the two major metabolites for carbon storage in most eukaryotic organisms. We investigated the glycogen metabolism of the oleaginous Yarrowia lipolytica and found that this yeast accumulates up to 16% glycogen in its biomass. Assuming that elimination of glycogen synthesis would result in an improvement of lipid accumulation, we characterized and deleted the single gene coding for glycogen synthase, YlGSY1. The mutant was grown under lipogenic conditions with glucose and glycerol as substrates and we obtained up to 60% improvement in TAG accumulation compared to the wild-type strain. Additionally, YlGSY1 was deleted in a background that was already engineered for high lipid accumulation. In this obese background, TAG accumulation was also further increased. The highest lipid content of 52% was found after 3 days of cultivation in nitrogen-limited glycerol medium. Furthermore, we constructed mutants of Y. lipolytica and Saccharomyces cerevisiae that are deleted for both glycogen and TAG synthesis, demonstrating that the ability to store carbon is not essential. Overall, this work showed that glycogen synthesis is a competing pathway for TAG accumulation in oleaginous yeasts and that deletion of the glycogen synthase has beneficial effects on neutral lipid storage. © FEMS 2017.

Substrates and oxygen dependent citric acid production by Yarrowia lipolytica: insights through transcriptome and fluxome analyses.

PubMed

Sabra, Wael; Bommareddy, Rajesh Reddy; Maheshwari, Garima; Papanikolaou, Seraphim; Zeng, An-Ping

2017-05-08

Unlike the well-studied backer yeast where catabolite repression represents a burden for mixed substrate fermentation, Yarrowia lipolytica, an oleaginous yeast, is recognized for its potential to produce single cell oils and citric acid from different feedstocks. These versatilities of Y. lipolytica with regards to substrate utilization make it an attractive host for biorefinery application. However, to develop a commercial process for the production of citric acid by Y. lipolytica, it is necessary to better understand the primary metabolism and its regulation, especially for growth on mixed substrate. Controlling the dissolved oxygen concentration (pO 2 ) in Y. lipolytica cultures enhanced citric acid production significantly in cultures grown on glucose in mono- or dual substrate fermentations, whereas with glycerol as mono-substrate no significant effect of pO 2 was found on citrate production. Growth on mixed substrate with glucose and glycerol revealed a relative preference of glycerol utilization by Y. lipolytica. Under optimized conditions with pO 2 control, the citric acid titer on glucose in mono- or in dual substrate cultures was 55 and 50 g/L (with productivity of 0.6 g/L*h in both cultures), respectively, compared to a maximum of 18 g/L (0.2 g/L*h) with glycerol in monosubstrate culture. Additionally, in dual substrate fermentation, glycerol limitation was found to trigger citrate consumption despite the presence of enough glucose in pO 2 -limited culture. The metabolic behavior of this yeast on different substrates was investigated at transcriptomic and 13 C-based fluxomics levels. Upregulation of most of the genes of the pentose phosphate pathway was found in cultures with highest citrate production with glucose in mono- or in dual substrate fermentation with pO 2 control. The activation of the glyoxylate cycle in the oxygen limited cultures and the imbalance caused by glycerol limitation might be the reason for the re-consumption of citrate in

Steroid biotransformations in biphasic systems with Yarrowia lipolytica expressing human liver cytochrome P450 genes

PubMed Central

2012-01-01

Background Yarrowia lipolytica efficiently metabolizes and assimilates hydrophobic compounds such as n-alkanes and fatty acids. Efficient substrate uptake is enabled by naturally secreted emulsifiers and a modified cell surface hydrophobicity and protrusions formed by this yeast. We were examining the potential of recombinant Y. lipolytica as a biocatalyst for the oxidation of hardly soluble hydrophobic steroids. Furthermore, two-liquid biphasic culture systems were evaluated to increase substrate availability. While cells, together with water soluble nutrients, are maintained in the aqueous phase, substrates and most of the products are contained in a second water-immiscible organic solvent phase. Results For the first time we have co-expressed the human cytochromes P450 2D6 and 3A4 genes in Y. lipolytica together with human cytochrome P450 reductase (hCPR) or Y. lipolytica cytochrome P450 reductase (YlCPR). These whole-cell biocatalysts were used for the conversion of poorly soluble steroids in biphasic systems. Employing a biphasic system with the organic solvent and Y. lipolytica carbon source ethyl oleate for the whole-cell bioconversion of progesterone, the initial specific hydroxylation rate in a 1.5 L stirred tank bioreactor was further increased 2-fold. Furthermore, the product formation was significantly prolonged as compared to the aqueous system. Co-expression of the human CPR gene led to a 4-10-fold higher specific activity, compared to the co-overexpression of the native Y. lipolytica CPR gene. Multicopy transformants showed a 50-70-fold increase of activity as compared to single copy strains. Conclusions Alkane-assimilating yeast Y. lipolytica, coupled with the described expression strategies, demonstrated its high potential for biotransformations of hydrophobic substrates in two-liquid biphasic systems. Especially organic solvents which can be efficiently taken up and/or metabolized by the cell might enable more efficient bioconversion as compared

CRISPR-Cas9-Mediated Genome Editing and Transcriptional Control in Yarrowia lipolytica.

PubMed

Schwartz, Cory; Wheeldon, Ian

2018-01-01

The discovery and adaptation of RNA-guided nucleases has resulted in the rapid development of efficient, scalable, and easily accessible synthetic biology tools for targeted genome editing and transcriptional control. In these systems, for example CRISPR-Cas9 from Streptococcus pyogenes, a protein with nuclease activity is targeted to a specific nucleotide sequence by a short RNA molecule, whereupon binding it cleaves the targeted nucleotide strand. To extend this genome-editing ability to the industrially important oleaginous yeast Yarrowia lipolytica, we developed a set of easily usable and effective CRISPR-Cas9 episomal vectors. In this protocols chapter, we first present a method by which arbitrary protein-coding genes can be disrupted via indel formation after CRISPR-Cas9 targeting. A second method demonstrates how the same CRISPR-Cas9 system can be used to induce markerless gene cassette integration into the genome by inducing homologous recombination after DNA cleavage by Cas9. Finally, we describe how a catalytically inactive form of Cas9 fused to a transcriptional repressor can be used to control transcription of native genes in Y. lipolytica. The CRISPR-Cas9 tools and strategies described here greatly increase the types of genome editing and transcriptional control that can be achieved in Y. lipolytica, and promise to facilitate more advanced engineering of this important oleaginous host.

Engineering Yarrowia lipolytica to express secretory invertase with strong FBA1IN promoter.

PubMed

Hong, Seung-Pyo; Seip, John; Walters-Pollak, Dana; Rupert, Ross; Jackson, Raymond; Xue, Zhixiong; Zhu, Quinn

2012-02-01

Oleaginous yeast Yarrowia lipolytica is an important host for the production of lipid-derived compounds or heterologous proteins. Selection of strong promoters and effective expression systems is critical for heterologous protein secretion. To search for a strong promoter in Y. lipolytica, activities of FBA1, TDH1 and GPM1 promoters were compared to that of TEF1 promoter by constructing GUS reporter fusions. The FBA1 promoter activity was 2.2 and 5.5 times stronger than the TDH1 and GPM1 promoters, respectively. The FBA1IN promoter (FBA1 sequence of -826 to +169) containing an intron (+64 to +165) showed five-fold higher expression than the FBA1 promoter (-831 to -1). The transcriptional enhancement by the 5'-region within the FBA1 gene was confirmed by GPM1::FBA1 chimeric promoter construction. Using the strong FBA1IN promoter, four different S. cerevisiae SUC2 expression cassettes were tested for the SUC+ phenotype in Y. lipolytica. Functional invertase secretion was facilitated by the Xpr2 prepro-region with an additional 13 amino acids of mature Xpr2, or by the native Suc2 signal sequence. However, these two secretory signals in tandem, or the mature Suc2 with no secretory signal, did not direct secretion of functional invertase. Unlike previously reported Y. lipolytica SUC+ strains, our engineered stains secreted most of invertase into the medium. Copyright © 2011 John Wiley & Sons, Ltd.

The expression of the Cuphea palustris thioesterase CpFatB2 in Yarrowia lipolytica triggers oleic acid accumulation.

PubMed

Stefan, Alessandra; Hochkoeppler, Alejandro; Ugolini, Luisa; Lazzeri, Luca; Conte, Emanuele

2016-01-01

The conversion of industrial by-products into high-value added compounds is a challenging issue. Crude glycerol, a by-product of the biodiesel production chain, could represent an alternative carbon source for the cultivation of oleaginous yeasts. Here, we developed five minimal synthetic glycerol-based media, with different C/N ratios, and we analyzed the production of biomass and fatty acids by Yarrowia lipolytica Po1g strain. We identified two media at the expense of which Y. lipolytica was able to accumulate ∼5 g L(-1) of biomass and 0.8 g L(-1) of fatty acids (0.16 g of fatty acids per g of dry weight). These optimized media contained 0.5 g L(-1) of urea or ammonium sulfate and 20 g L(-1) of glycerol, and were devoid of yeast extract. Moreover, Y. lipolytica was engineered by inserting the FatB2 gene, coding for the CpFatB2 thioesterase from Cuphea palustris, in order to modify the fatty acid composition towards the accumulation of medium-chain fatty acids. Contrary to the expected, the expression of the heterologous gene increased the production of oleic acid, and concomitantly decreased the level of saturated fatty acids. © 2015 American Institute of Chemical Engineers.

Quantitative study of lipase secretion, extracellular lipolysis, and lipid storage in the yeast Yarrowia lipolytica grown in the presence of olive oil: analogies with lipolysis in humans.

PubMed

Najjar, Amal; Robert, Sylvie; Guérin, Clémence; Violet-Asther, Michèle; Carrière, Frédéric

2011-03-01

Lipase secretion, extracellular lipolysis, and fatty acid uptake were quantified in the yeast Yarrowia lipolytica grown in the presence of olive oil and/or glucose. Specific lipase assays, Western blot analysis, and ELISA indicated that most of the lipase activity measured in Y. lipolytica cultures resulted from the YLLIP2 lipase. Lipase production was triggered by olive oil and, during the first hours of culture, most of the lipase activity and YLLIP2 immunodetection remained associated with the yeast cells. YLLIP2 was then released in the culture medium before it was totally degraded by proteases. Olive oil triglycerides were largely degraded when the lipase was still attached to the cell wall. The fate of lipolysis products in the culture medium and inside the yeast cell, as well as lipid storage, was investigated simultaneously by quantitative TLC-FID and GC analysis. The intracellular levels of free fatty acids (FFA) and triglycerides increased transiently and were dependent on the carbon sources. A maximum fat storage of 37.8% w/w of yeast dry mass was observed with olive oil alone. A transient accumulation of saturated FFA was observed whereas intracellular triglycerides became enriched in unsaturated fatty acids. So far, yeasts have been mainly used for studying the intracellular synthesis, storage, and mobilization of neutral lipids. The present study shows that yeasts are also interesting models for studying extracellular lipolysis and fat uptake by the cell. The quantitative data obtained here allow for the first time to establish interesting analogies with gastrointestinal and vascular lipolysis in humans.

Modeling and optimization of lipid accumulation by Yarrowia lipolytica from glucose under nitrogen depletion conditions.

PubMed

Robles-Rodríguez, Carlos E; Muñoz-Tamayo, Rafael; Bideaux, Carine; Gorret, Nathalie; Guillouet, Stéphane E; Molina-Jouve, Carole; Roux, Gilles; Aceves-Lara, César A

2018-05-01

Oleaginous yeasts have been seen as a feasible alternative to produce the precursors of biodiesel due to their capacity to accumulate lipids as triacylglycerol having profiles with high content of unsaturated fatty acids. The yeast Yarrowia lipolytica is a promising microorganism that can produce lipids under nitrogen depletion conditions and excess of the carbon source. However, under these conditions, this yeast also produces citric acid (overflow metabolism) decreasing lipid productivity. This work presents two mathematical models for lipid production by Y. lipolytica from glucose. The first model is based on Monod and inhibition kinetics, and the second one is based on the Droop quota model approach, which is extended to yeast. The two models showed good agreements with the experimental data used for calibration and validation. The quota based model presented a better description of the dynamics of nitrogen and glucose dynamics leading to a good management of N/C ratio which makes this model interesting for control purposes. Then, quota model was used to evaluate, by means of simulation, a scenario for optimizing lipid productivity and lipid content. For that, a control strategy was designed by approximating the flow rates of glucose and nitrogen with piecewise linear functions. Simulation results achieved productivity of 0.95 g L -1  hr -1 and lipid content fraction of 0.23 g g -1 , which indicates that this strategy is a promising alternative for the optimization of lipid production. © 2017 Wiley Periodicals, Inc.

High-throughput fermentation screening for the yeast Yarrowia lipolytica with real-time monitoring of biomass and lipid production.

PubMed

Back, Alexandre; Rossignol, Tristan; Krier, François; Nicaud, Jean-Marc; Dhulster, Pascal

2016-08-23

Because the model yeast Yarrowia lipolytica can synthesize and store lipids in quantities up to 20 % of its dry weight, it is a promising microorganism for oil production at an industrial scale. Typically, optimization of the lipid production process is performed in the laboratory and later scaled up for industrial production. However, the scale-up process can be complicated by genetic modifications that are optimized for one set of growing conditions can confer a less-than-optimal phenotype in a different environment. To address this issue, small cultivation systems have been developed that mimic the conditions in benchtop bioreactors. In this work, we used one such microbioreactor system, the BioLector, to develop high-throughput fermentation procedures that optimize growth and lipid accumulation in Y. lipolytica. Using this system, we were able to monitor lipid and biomass production in real time throughout the culture duration. The BioLector can monitor the growth of Y. lipolytica in real time by evaluating scattered light; this produced accurate measurements until cultures reached an equivalent of OD600nm = 115 and a cell dry weight of 100 g L(-1). In addition, a lipid-specific fluorescent probe was applied which reliably monitored lipid production up to a concentration of 12 g L(-1). Through screening various growing conditions, we determined that a carbon/nitrogen ratio of 35 was the most efficient for lipid production. Further screening showed that ammonium chloride and glycerol were the most valuable nitrogen and carbon sources, respectively, for growth and lipid production. Moreover, a carbon concentration above 1 M appeared to impair growth and lipid accumulation. Finally, we used these optimized conditions to screen engineered strains of Y. lipolytica with high lipid-accumulation capability. The growth and lipid content of the strains cultivated in the BioLector were compared to those grown in benchtop bioreactors. To our knowledge, this is the

Yarrowia lipolytica NCIM 3589, a tropical marine yeast, degrades bromoalkanes by an initial hydrolytic dehalogenation step.

PubMed

Vatsal, Aakanksha; Zinjarde, Smita S; Kumar, Ameeta Ravi

2015-04-01

The widespread industrial use of organobromines which are known persistent organic pollutants has led to their accumulation in sediments and water bodies causing harm to animals and humans. While degradation of organochlorines by bacteria is well documented, information regarding degradation pathways of these recalcitrant organobromines is scarce. Hence, their fates and effects on the environment are of concern. The present study shows that a tropical marine yeast, Yarrowia lipolytica NCIM 3589 aerobically degrades bromoalkanes differing in carbon chain length and position of halogen substitution viz., 2-bromopropane (2-BP), 1-bromobutane (1-BB), 1,5 dibromopentane (1,5-DBP) and 1-bromodecane (1-BD) as seen by an increase in cell mass, release of bromide and concomitant decrease in concentration of brominated compound. The amount of bromoalkane degraded was 27.3, 21.9, 18.0 and 38.3 % with degradation rates of 0.076, 0.058, 0.046 and 0.117/day for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. The initial product formed respectively were alcohols viz., 2-propanol, 1-butanol, 1-bromo, 5-pentanol and 1-decanol as detected by GC-MS. These were further metabolized to fatty acids viz., 2-propionic, 1-butyric and 1-decanoic acid eventually leading to carbon dioxide formation. Neither higher chain nor brominated fatty acids were detected. An inducible extracellular dehalogenase responsible for removal of bromide was detected with activities of 21.07, 18.82, 18.96 and 26.67 U/ml for 2-BP, 1-BB, 1,5-DBP and 1-BD, respectively. We report here for the first time the proposed aerobic pathway of bromoalkane degradation by an eukaryotic microbe Y. lipolytica 3589, involving an initial hydrolytic dehalogenation step.

The ‘LipoYeasts’ project: using the oleaginous yeast Yarrowia lipolytica in combination with specific bacterial genes for the bioconversion of lipids, fats and oils into high‐value products

PubMed Central

Sabirova, Julia S.; Haddouche, R.; Van Bogaert, I. N.; Mulaa, F.; Verstraete, W.; Timmis, K. N.; Schmidt‐Dannert, C.; Nicaud, J. M.; Soetaert, W.

2011-01-01

Summary The oleochemical industry is currently still dominated by conventional chemistry, with biotechnology only starting to play a more prominent role, primarily with respect to the biosurfactants or lipases, e.g. as detergents, or for biofuel production. A major bottleneck for all further biotechnological applications is the problem of the initial mobilization of cheap and vastly available lipid and oil substrates, which are then to be transformed into high‐value biotechnological, nutritional or pharmacological products. Under the EU‐sponsored LipoYeasts project we are developing the oleaginous yeast Yarrowia lipolytica into a versatile and high‐throughput microbial factory that, by use of specific enzymatic pathways from hydrocarbonoclastic bacteria, efficiently mobilizes lipids by directing its versatile lipid metabolism towards the production of industrially valuable lipid‐derived compounds like wax esters (WE), isoprenoid‐derived compounds (carotenoids, polyenic carotenoid ester), polyhydroxyalkanoates (PHAs) and free hydroxylated fatty acids (HFAs). Different lipid stocks (petroleum, alkane, vegetable oil, fatty acid) and combinations thereof are being assessed as substrates in combination with different mutant and recombinant strains of Y. lipolytica, in order to modulate the composition and yields of the produced added‐value products. PMID:21255371

Citric acid production from extract of Jerusalem artichoke tubers by the genetically engineered yeast Yarrowia lipolytica strain 30 and purification of citric acid.

PubMed

Wang, Ling-Fei; Wang, Zhi-Peng; Liu, Xiao-Yan; Chi, Zhen-Ming

2013-11-01

In this study, citric acid production from extract of Jerusalem artichoke tubers by the genetically engineered yeast Yarrowia lipolytica strain 30 was investigated. After the compositions of the extract of Jerusalem artichoke tubers for citric acid production were optimized, the results showed that natural components of extract of Jerusalem artichoke tubers without addition of any other components were suitable for citric acid production by the yeast strain. During 10 L fermentation using the extract containing 84.3 g L(-1) total sugars, 68.3 g L(-1) citric acid was produced and the yield of citric acid was 0.91 g g(-1) within 336 h. At the end of the fermentation, 9.2 g L(-1) of residual total sugar and 2.1 g L(-1) of reducing sugar were left in the fermented medium. At the same time, citric acid in the supernatant of the culture was purified. It was found that 67.2 % of the citric acid in the supernatant of the culture was recovered and purity of citric acid in the crystal was 96 %.

Biomass production by novel strains of Yarrowia lipolytica using raw glycerol, derived from biodiesel production.

PubMed

Juszczyk, Piotr; Tomaszewska, Ludwika; Kita, Agnieszka; Rymowicz, Waldemar

2013-06-01

This study demonstrated the potential applicability of the isolated strains of Yarrowia lipolytica for the valorization of glycerol waste generated during biodiesel production, throughout biomass production. Twenty-one strains were isolated from different environments and identified as Y. lipolytica. Biomass production from pure glycerol (25 g L(-1)) was performed in the shake-flasks experiment. Eight strains with the best biomass production ability were chosen for studies in bioreactor (pH 3.5). The analysis of technological process parameters and biomass chemical composition demonstrated that S6 strain was the most suitable for biomass production. Its application allowed obtaining 11.7 and 12.3 g L(-1) of the biomass with 1.30 and 1.37 g L(-1) h(-1) productivity, respectively when pure and raw glycerol (25 g L(-1)) was used. In the yeast protein amino acid profile the contents of lysine, threonine and phenylalanine/tyrosine were higher than required by FAO/WHO. According to the EAAI, the nutritional value of the biomass reached up to 72.3%. Copyright © 2013 Elsevier Ltd. All rights reserved.

Study of trans-trans farnesol effect on hyphae formation by Yarrowia lipolytica.

PubMed

Nunes, Patrícia Martins Botelho; da Rocha, Silvia Maria; Amaral, Priscilla Filomena Fonseca; da Rocha-Leão, Maria Helena Miguez

2013-12-01

Dimorphism is an ability of certain fungi related to its adaptation to the environment and provides a selective advantage under stress conditions and is associated to the development of human diseases. Hyphae inducing- and inhibitory-effect of farnesol on hyphae formation by the dimorphic yeast Yarrowia lipolytica was evaluated through digital image analysis. The agitation speed of the culture was the most effective hyphae inducer in comparison to bovine calf serum and N-acetylglucosamine. In low agitation system, bovine calf serum was more effective for hyphae formation inducing 57 % of hyphae transition. Farnesol inhibited hyphae formation even in low concentration (300 μM) and this effect increased with increasing concentrations. In the presence of N-acetylglucosamine, this effect was more evident in comparison to the presence of bovine calf serum, which might have protected the cells from farnesol. Digital image analysis was an important tool to evaluate this phenomenon.

Development of recombinant Yarrowia lipolytica producing virus-like particles of a fish nervous necrosis virus.

PubMed

Luu, Van-Trinh; Moon, Hye Yun; Hwang, Jee Youn; Kang, Bo-Kyu; Kang, Hyun Ah

2017-08-01

Nervous necrosis virus (NNV) causes viral encephalopathy and retinopathy, a devastating disease of many species of cultured marine fish worldwide. In this study, we used the dimorphic non-pathogenic yeast Yarrowia lipolytica as a host to express the capsid protein of red-spotted grouper nervous necrosis virus (RGNNV-CP) and evaluated its potential as a platform for vaccine production. An initial attempt was made to express the codon-optimized synthetic genes encoding intact and N-terminal truncated forms of RGNNV-CP under the strong constitutive TEF1 promoter using autonomously replicating sequence (ARS)-based vectors. The full-length recombinant capsid proteins expressed in Y. lipolytica were detected not only as monomers and but also as trimers, which is a basic unit for formation of NNV virus-like particles (VLPs). Oral immunization of mice with whole recombinant Y. lipolytica harboring the ARS-based plasmids was shown to efficiently induce the formation of IgG against RGNNV-CP. To increase the number of integrated copies of the RGNNV-CP expression cassette, a set of 26S ribosomal DNA-based multiple integrative vectors was constructed in combination with a series of defective Ylura3 with truncated promoters as selection markers, resulting in integrants harboring up to eight copies of the RGNNV-CP cassette. Sucrose gradient centrifugation and transmission electron microscopy of this high-copy integrant were carried out to confirm the expression of RGNNV-CPs as VLPs. This is the first report on efficient expression of viral capsid proteins as VLPs in Y. lipolytica, demonstrating high potential for the Y. lipolytica expression system as a platform for recombinant vaccine production based on VLPs.

Integrated Approach To Producing High-Purity Trehalose from Maltose by the Yeast Yarrowia lipolytica Displaying Trehalose Synthase (TreS) on the Cell Surface.

PubMed

Li, Ning; Wang, Hengwei; Li, Lijuan; Cheng, Huiling; Liu, Dawen; Cheng, Hairong; Deng, Zixin

2016-08-10

An alternative strategy that integrated enzyme production, trehalose biotransformation, and bioremoval in one bioreactor was developed in this study, thus simplifying the traditional procedures used for trehalose production. The trehalose synthase gene from a thermophilic archaea, Picrophilus torridus, was first fused to the YlPir1 anchor gene and then inserted into the genome of Yarrowia lipolytica, thus yielding an engineered yeast strain. The trehalose yield reached 73% under optimal conditions. The thermal and pH stabilities of the displayed enzyme were improved compared to those of its free form purified from recombinant Escherichia coli. After biotransformation, the glucose byproduct and residual maltose were directly fermented to ethanol by a Saccharomyces cerevisiae strain. Ethanol can be separated by distillation, and high-purity trehalose can easily be obtained from the fermentation broth. The results show that this one-pot procedure is an efficient approach to the economical production of trehalose from maltose.

Comparative evaluation of 13 yeast species in the Yarrowia clade on lignocellulosic biomass hydrolysate and genetic engineering of inhibitor tolerant strains for lipid and biofuel production

USDA-ARS?s Scientific Manuscript database

Yarrowia lipolytica is an oleaginous yeast that has garnered interest for commercial production of single cell oil and other fatty acid-derived chemicals because of its GRAS status and genetic tractability. Three recent peer-reviewed studies have highlighted the possibility of lipid production by th...

Dynamic behavior of Yarrowia lipolytica in response to pH perturbations: dependence of the stress response on the culture mode.

PubMed

Timoumi, Asma; Cléret, Mégane; Bideaux, Carine; Guillouet, Stéphane E; Allouche, Yohan; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2017-01-01

Yarrowia lipolytica, a non-conventional yeast with a promising biotechnological potential, is able to undergo metabolic and morphological changes in response to environmental conditions. The effect of pH perturbations of different types (pulses, Heaviside) on the dynamic behavior of Y. lipolytica W29 strain was characterized under two modes of culture: batch and continuous. In batch cultures, different pH (4.5, 5.6 (optimal condition), and 7) were investigated in order to identify the pH inducing a stress response (metabolic and/or morphologic) in Y. lipolytica. Macroscopic behavior (kinetic parameters, yields, viability) of the yeast was slightly affected by pH. However, contrary to the culture at pH 5.6, a filamentous growth was induced in batch experiments at pH 4.5 and 7. Proportions of the filamentous subpopulation reached 84 and 93 % (v/v) under acidic and neutral conditions, respectively. Given the significant impact of neutral pH on morphology, pH perturbations from 5.6 to 7 were subsequently assayed in batch and continuous bioreactors. For both process modes, the growth dynamics remained fundamentally unaltered during exposure to stress. Nevertheless, morphological behavior of the yeast was dependent on the culture mode. Specifically, in batch bioreactors where cells proliferated at their maximum growth rate, mycelia were mainly formed. Whereas, in continuous cultures at controlled growth rates (from 0.03 to 0.20 h -1 ) even closed to the maximum growth rate of the stain (0.24 h -1 ), yeast-like forms predominated. This pointed out differences in the kinetic behavior of filamentous and yeast subpopulations, cell age distribution, and pH adaptive mechanisms between both modes of culture.

Three alcohol dehydrogenase genes and one acetyl-CoA synthetase gene are responsible for ethanol utilization in Yarrowia lipolytica.

PubMed

Gatter, Michael; Ottlik, Stephanie; Kövesi, Zsolt; Bauer, Benjamin; Matthäus, Falk; Barth, Gerold

2016-10-01

The non-conventional yeast Yarrowia lipolytica is able to utilize a wide range of different substrates like glucose, glycerol, ethanol, acetate, proteins and various hydrophobic molecules. Although most metabolic pathways for the utilization of these substrates have been clarified by now, it was not clear whether ethanol is oxidized by alcohol dehydrogenases or by an alternative oxidation system inside the cell. In order to detect the genes that are required for ethanol utilization in Y. lipolytica, eight alcohol dehydrogenase (ADH) genes and one alcohol oxidase gene (FAO1) have been identified and respective deletion strains were tested for their ability to metabolize ethanol. As a result of this, we found that the availability of ADH1, ADH2 or ADH3 is required for ethanol utilization in Y. lipolytica. A strain with deletions in all three genes is lacking the ability to utilize ethanol as sole carbon source. Although Adh2p showed by far the highest enzyme activity in an in vitro assay, the availability of any of the three genes was sufficient to enable a decent growth. In addition to ADH1, ADH2 and ADH3, an acetyl-CoA synthetase encoding gene (ACS1) was found to be essential for ethanol utilization. As Y. lipolytica is a non-fermenting yeast, it is neither able to grow under anaerobic conditions nor to produce ethanol. To investigate whether Y. lipolytica may produce ethanol, the key genes of alcoholic fermentation in S. cerevisiae, ScADH1 and ScPDC1, were overexpressed in an ADH and an ACS1 deletion strain. However, instead of producing ethanol, the respective strains regained the ability to use ethanol as single carbon source and were still not able to grow under anaerobic conditions. Copyright © 2016 Elsevier Inc. All rights reserved.

Investigating Proteome and Transcriptome Defense Response of Apples Induced by Yarrowia lipolytica.

PubMed

Zhang, Hongyin; Chen, Liangliang; Sun, Yiwen; Zhao, Lina; Zheng, Xiangfeng; Yang, Qiya; Zhang, Xiaoyun

2017-04-01

A better understanding of the mode of action of postharvest biocontrol agents on fruit surfaces is critical for the advancement of successful implementation of postharvest biocontrol products. This is due to the increasing importance of biological control of postharvest diseases over chemical and other control methods. However, most of the mechanisms involved in biological control remain unknown and need to be explored. Yarrowia lipolytica significantly inhibited blue mold decay of apples caused by Penicillium expansum. The findings also demonstrated that Y. lipolytica stimulated the activities of polyphenoloxidase, peroxidase, chitinase, l-phenylalanine ammonia lyase involved in enhancing defense responses in apple fruit tissue. Proteomic and transcriptomic analysis revealed a total of 35 proteins identified as up- and down-regulated in response to the Y. lipolytica inducement. These proteins were related to defense, biotic stimulus, and stress responses, such as pathogenesis-related proteins and dehydrin. The analysis of the transcriptome results proved that the induced resistance was mediated by a crosstalk between salicylic acid (SA) and ethylene/jasmonate (ET/JA) pathways. Y. lipolytica treatment activated the expression of isochorismate synthase gene in the SA pathway, which up-regulates the expression of PR4 in apple. The expression of 1-aminocyclopropane-1-carboxylate oxidase gene and ET-responsive transcription factors 2 and 4, which are involved in the ET pathway, were also activated. In addition, cytochrome oxidase I, which plays an important role in JA signaling for resistance acquisition, was also activated. However, not all of the genes had a positive effect on the SA and ET/JA signal pathways. As transcriptional repressors in JA signaling, TIFY3B and TIFY11B were triggered by the yeast, but the gene expression levels were relatively low. Taken together, Y. lipolytica induced the SA and ET/JA signal mediating the defense pathways by stimulating

Citric acid production in Yarrowia lipolytica SWJ-1b yeast when grown on waste cooking oil.

PubMed

Liu, Xiaoyan; Lv, Jinshun; Xu, Jiaxing; Zhang, Tong; Deng, Yuanfang; He, Jianlong

2015-03-01

In this study, citric acid was produced from waste cooking oil by Yarrowia lipolytica SWJ-1b. To get the maximal yield of citric acid, the compositions of the medium for citric acid production were optimized, and our results showed that extra nitrogen and magnesium rather than vitamin B1 and phosphate were needed for CA accumulation when using waste cooking oil. The results also indicated that the optimal initial concentration of the waste cooking oil in the medium for citric acid production was 80.0 g/l, and the ideal inoculation size was 1 × 10(7) cells/l of medium. We also reported that during 10-l fermentation, 31.7 g/l of citric acid, 6.5 g/l of isocitric acid, 5.9 g/l of biomass, and 42.1 g/100.0 g cell dry weight of lipid were attained from 80.0 g/l of waste cooking oil within 336 h. At the end of the fermentation, 94.6 % of the waste cooking oil was utilized by the cells of Y. lipolytica SWJ-1b, and the yield of citric acid was 0.4 g/g waste cooking oil, which suggested that waste cooking oil was a suitable carbon resource for citric acid production.

Influence of oxygen availability on the metabolism and morphology of Yarrowia lipolytica: insights into the impact of glucose levels on dimorphism.

PubMed

Timoumi, Asma; Bideaux, Carine; Guillouet, Stéphane E; Allouche, Yohan; Molina-Jouve, Carole; Fillaudeau, Luc; Gorret, Nathalie

2017-10-01

Dynamic behavior of Yarrowia lipolytica W29 strain under conditions of fluctuating, low, and limited oxygen supply was characterized in batch and glucose-limited chemostat cultures. In batch cultures, transient oscillations between oxygen-rich and -deprived environments induced a slight citric acid accumulation (lower than 29 mg L -1 ). By contrast, no citric acid was detected in continuous fermentations for all stress conditions: full anoxia (zero pO 2 value, 100% N 2 ), limited (zero pO 2 value, 75% of cell needs), and low (pO 2 close to 2%) dissolved oxygen (DO) levels. The macroscopic behavior (kinetic parameters, yields, viability) of Y. lipolytica was not significantly affected by the exposure to DO fluctuations under both modes of culture. Nevertheless, conditions of oxygen limitation resulted in the destabilization of the glucose-limited growth during the continuous cultivations. Morphological responses of Y. lipolytica to DO oscillations were different between batch and chemostat runs. Indeed, a yeast-to-mycelium transition was induced and progressively intensified during the batch fermentations (filamentous subpopulation reaching 74% (v/v)). While, in chemostat bioreactors, the culture consisted mainly of yeast-like cells (mean diameter not exceeding 5.7 μm) with a normal size distribution. During the continuous cultures, growth at low DO concentration did not induce any changes in Y. lipolytica morphology. Dimorphism (up to 80.5% (v/v) of filaments) was only detected under conditions of oxygen limitation in the presence of a residual glucose excess (more than 0.75 g L -1 ). These data suggest an impact of glucose levels on the signaling pathways regulating dimorphic responses in Y. lipolytica.

Integrating Cellular and Bioprocess Engineering in the Non-Conventional Yeast Yarrowia lipolytica for Biodiesel Production: A Review

PubMed Central

Xie, Dongming

2017-01-01

As one of the major biofuels to replace fossil fuel, biodiesel has now attracted more and more attention due to its advantages in higher energy density and overall less greenhouse gas generation. Biodiesel (fatty acid alkyl esters) is produced by chemically or enzymatically catalyzed transesterification of lipids from microbial cells, microalgae, oil crops, or animal fats. Currently, plant oils or waste cooking oils/fats remain the major source for biodiesel production via enzymatic route, but the production capacity is limited either by the uncertain supplement of plant oils or by the low or inconsistent quality of waste oils/fats. In the past decades, significant progresses have been made on synthesis of microalgae oils directly from CO2 via a photosynthesis process, but the production cost from any current technologies is still too high to be commercialized due to microalgae’s slow growth rate on CO2, inefficiency in photo-bioreactors, lack of efficient contamination control methods, and high cost in downstream recovery. At the same time, many oleaginous microorganisms have been studied to produce lipids via the fatty acid synthesis pathway under aerobic fermentation conditions, among them one of the most studied is the non-conventional yeast, Yarrowia lipolytica, which is able to produce fatty acids at very high titer, rate, and yield from various economical substrates. This review summarizes the recent research progresses in both cellular and bioprocess engineering in Y. lipolytica to produce lipids at a low cost that may lead to commercial-scale biodiesel production. Specific technologies include the strain engineering for using various substrates, metabolic engineering in high-yield lipid synthesis, cell morphology study for efficient substrate uptake and product formation, free fatty acid formation and secretion for improved downstream recovery, and fermentation engineering for higher productivities and less operating cost. To further improve the

Analysis of ATP-citrate lyase and malic enzyme mutants of Yarrowia lipolytica points out the importance of mannitol metabolism in fatty acid synthesis.

PubMed

Dulermo, Thierry; Lazar, Zbigniew; Dulermo, Rémi; Rakicka, Magdalena; Haddouche, Ramedane; Nicaud, Jean-Marc

2015-09-01

The role of the two key enzymes of fatty acid (FA) synthesis, ATP-citrate lyase (Acl) and malic enzyme (Mae), was analyzed in the oleaginous yeast Yarrowia lipolytica. In most oleaginous yeasts, Acl and Mae are proposed to provide, respectively, acetyl-CoA and NADPH for FA synthesis. Acl was mainly studied at the biochemical level but no strain depleted for this enzyme was analyzed in oleaginous microorganisms. On the other hand the role of Mae in FA synthesis in Y. lipolytica remains unclear since it was proposed to be a mitochondrial NAD(H)-dependent enzyme and not a cytosolic NADP(H)-dependent enzyme. In this study, we analyzed for the first time strains inactivated for corresponding genes. Inactivation of ACL1 decreases FA synthesis by 60 to 80%, confirming its essential role in FA synthesis in Y. lipolytica. Conversely, inactivation of MAE1 has no effects on FA synthesis, except in a FA overaccumulating strain where it improves FA synthesis by 35%. This result definitively excludes Mae as a major key enzyme for FA synthesis in Y. lipolytica. During the analysis of both mutants, we observed a negative correlation between FA and mannitol level. As mannitol and FA pathways may compete for carbon storage, we inactivated YlSDR, encoding a mannitol dehydrogenase converting fructose and NADPH into mannitol and NADP+. The FA content of the resulting mutant was improved by 60% during growth on fructose, demonstrating that mannitol metabolism may modulate FA synthesis in Y. lipolytica. Copyright © 2015. Published by Elsevier B.V.

Multi-omics analysis reveals regulators of the response to nitrogen limitation in Yarrowia lipolytica.

PubMed

Pomraning, Kyle R; Kim, Young-Mo; Nicora, Carrie D; Chu, Rosalie K; Bredeweg, Erin L; Purvine, Samuel O; Hu, Dehong; Metz, Thomas O; Baker, Scott E

2016-02-25

Yarrowia lipolytica is an oleaginous ascomycete yeast that stores lipids in response to limitation of nitrogen. While the enzymatic pathways responsible for neutral lipid accumulation in Y. lipolytica are well characterized, regulation of these pathways has received little attention. We therefore sought to characterize the response to nitrogen limitation at system-wide levels, including the proteome, phosphoproteome and metabolome, to better understand how this organism regulates and controls lipid metabolism and to identify targets that may be manipulated to improve lipid yield. We found that ribosome structural genes are down-regulated under nitrogen limitation, during which nitrogen containing compounds (alanine, putrescine, spermidine and urea) are depleted and sugar alcohols and TCA cycle intermediates accumulate (citrate, fumarate and malate). We identified 1219 novel phosphorylation sites in Y. lipolytica, 133 of which change in their abundance during nitrogen limitation. Regulatory proteins, including kinases and DNA binding proteins, are particularly enriched for phosphorylation. Within lipid synthesis pathways, we found that ATP-citrate lyase, acetyl-CoA carboxylase and lecithin cholesterol acyl transferase are phosphorylated during nitrogen limitation while many of the proteins involved in β-oxidation are down-regulated, suggesting that storage lipid accumulation may be regulated by phosphorylation of key enzymes. Further, we identified short DNA elements that associate specific transcription factor families with up- and down-regulated genes. Integration of metabolome, proteome and phosphoproteome data identifies lipid accumulation in response to nitrogen limitation as a two-fold result of increased production of acetyl-CoA from excess citrate and decreased capacity for β-oxidation.

High performance microbiological transformation of L-tyrosine to L-dopa by Yarrowia lipolytica NRRL-143

PubMed Central

Ali, Sikander; Shultz, Jeffry L; Ikram-ul-Haq

2007-01-01

Background The 3,4-dihydroxy phenyl L-alanine (L-dopa) is a drug of choice for Parkinson's disease, controlling changes in energy metabolism enzymes of the myocardium following neurogenic injury. Aspergillus oryzae is commonly used for L-dopa production; however, potential improvements in ease of handling, growth rate and environmental impact have led to an interest in exploiting alternative yeasts. The two important elements required for L-dopa production are intracellular tyrosinases (thus pre-grown yeast cells are required for the transformation of L-tyrosine to L-dopa) and L-ascorbate, which acts as a reducing agent. Results Pre-grown cells of Yarrowia lipolytica NRRL-143 were used for the microbiological transformation of L-tyrosine to L-dopa. Different diatomite concentrations (0.5–3.0 mg/ml) were added to the acidic (pH 3.5) reaction mixture. Maximum L-dopa biosynthesis (2.96 mg/ml L-dopa from 2.68 mg/ml L-tyrosine) was obtained when 2.0 mg/ml diatomite was added 15 min after the start of the reaction. After optimizing reaction time (30 min), and yeast cell concentration (2.5 mg/ml), an overall 12.5 fold higher L-dopa production rate was observed when compared to the control. Significant enhancements in Yp/s, Qs and qs over the control were observed. Conclusion Diatomite (2.0 mg/ml) addition 15 min after reaction commencement improved microbiological transformation of L-tyrosine to L-dopa (3.48 mg/ml; p ≤ 0.05) by Y. lipolytica NRRL-143. A 35% higher substrate conversion rate was achieved when compared to the control. PMID:17705832

Inference and interrogation of a coregulatory network in the context of lipid accumulation in Yarrowia lipolytica.

PubMed

Trébulle, Pauline; Nicaud, Jean-Marc; Leplat, Christophe; Elati, Mohamed

2017-01-01

Complex phenotypes, such as lipid accumulation, result from cooperativity between regulators and the integration of multiscale information. However, the elucidation of such regulatory programs by experimental approaches may be challenging, particularly in context-specific conditions. In particular, we know very little about the regulators of lipid accumulation in the oleaginous yeast of industrial interest Yarrowia lipolytica . This lack of knowledge limits the development of this yeast as an industrial platform, due to the time-consuming and costly laboratory efforts required to design strains with the desired phenotypes. In this study, we aimed to identify context-specific regulators and mechanisms, to guide explorations of the regulation of lipid accumulation in Y. lipolytica . Using gene regulatory network inference, and considering the expression of 6539 genes over 26 time points from GSE35447 for biolipid production and a list of 151 transcription factors, we reconstructed a gene regulatory network comprising 111 transcription factors, 4451 target genes and 17048 regulatory interactions (YL-GRN-1) supported by evidence of protein-protein interactions. This study, based on network interrogation and wet laboratory validation (a) highlights the relevance of our proposed measure, the transcription factors influence, for identifying phases corresponding to changes in physiological state without prior knowledge (b) suggests new potential regulators and drivers of lipid accumulation and (c) experimentally validates the impact of six of the nine regulators identified on lipid accumulation, with variations in lipid content from +43.2% to -31.2% on glucose or glycerol.

Extracellular expression of YlLip11 with a native signal peptide from Yarrowia lipolytica MSR80 in three different yeast hosts.

PubMed

Kumari, Arti; Baronian, Keith; Kunze, Gotthard; Gupta, Rani

2015-06-01

Lipase YlLip11 from Yarrowia lipolytica was expressed with a signal peptide encoding sequence in Arxula adeninivorans, Saccharomyces cerevisiae and Hansenula polymorpha using the Xplor®2 transformation/expression platform and an expression module with the constitutive Arxula-derived TEF1 promoter. The YlLip11 signal peptide was functional in all of the yeast hosts with 97% of the recombinant enzyme being secreted into the culture medium. However, recombinant YlLip11 with His Tag fused at C-terminal was not active. The best recombinant YlLip11 producing A. adeninivorans G1212/YRC102-YlLip11 transformant cultivated in shake flasks produced 2654 U/L lipase, followed by S. cerevisiae SEY6210/YRC103-YlLip11 (1632U/L) and H. polymorpha RB11/YRC103-YlLip11 (1144U/L). Although the biochemical parameters of YlLip11 synthesized in different hosts were similar, their glycosylation level and thermo stability differed. The protein synthesized by the H. polymorpha transformant had the highest degree of glycosylation and with a t1/2 of 60min at 70°C, exhibited the highest thermostability. Copyright © 2015 Elsevier Inc. All rights reserved.

Yarrowia lipolytica morphological mutant enables lasting in situ immobilization in bioreactor.

PubMed

Vandermies, Marie; Kar, Tambi; Carly, Frédéric; Nicaud, Jean-Marc; Delvigne, Frank; Fickers, Patrick

2018-04-26

In the present study, we have isolated and characterized a Yarrowia lipolytica morphological mutant growing exclusively in the pseudohyphal morphology. The gene responsible for this phenotype, YALI0E06519g, was identified as homologous to the mitosis regulation gene HSL1 from Saccharomyces cerevisiae. Taking advantage of its morphology, we achieved the immobilization of the Δhsl1 mutant on the metallic structured packing of immobilized-cell bioreactors. We obtained significant cell retention and growth on the support during shake flask and bioreactor experiments without an attachment step prior to the culture. The system of medium aspersion on the packing ensured oxygen availability in the absence of agitation and minimized the potential release of cells in the culture medium. Additionally, the metallic packing proved its facility of cleaning and sterilization after fermentation. This combined use of morphological mutation and bioreactor design is a promising strategy to develop continuous processes for the production of recombinant protein and metabolites using Y. lipolytica. Graphical Abstract.

Production of Medium Chain Fatty Acids by Yarrowia lipolytica: Combining Molecular Design and TALEN to Engineer the Fatty Acid Synthase.

PubMed

Rigouin, Coraline; Gueroult, Marc; Croux, Christian; Dubois, Gwendoline; Borsenberger, Vinciane; Barbe, Sophie; Marty, Alain; Daboussi, Fayza; André, Isabelle; Bordes, Florence

2017-10-20

Yarrowia lipolytica is a promising organism for the production of lipids of biotechnological interest and particularly for biofuel. In this study, we engineered the key enzyme involved in lipid biosynthesis, the giant multifunctional fatty acid synthase (FAS), to shorten chain length of the synthesized fatty acids. Taking as starting point that the ketoacyl synthase (KS) domain of Yarrowia lipolytica FAS is directly involved in chain length specificity, we used molecular modeling to investigate molecular recognition of palmitic acid (C16 fatty acid) by the KS. This enabled to point out the key role of an isoleucine residue, I1220, from the fatty acid binding site, which could be targeted by mutagenesis. To address this challenge, TALEN (transcription activator-like effector nucleases)-based genome editing technology was applied for the first time to Yarrowia lipolytica and proved to be very efficient for inducing targeted genome modifications. Among the generated FAS mutants, those having a bulky aromatic amino acid residue in place of the native isoleucine at position 1220 led to a significant increase of myristic acid (C14) production compared to parental wild-type KS. Particularly, the best performing mutant, I1220W, accumulates C14 at a level of 11.6% total fatty acids. Overall, this work illustrates how a combination of molecular modeling and genome-editing technology can offer novel opportunities to rationally engineer complex systems for synthetic biology.

Multi-responses optimization of simultaneous biosorption of cationic dyes by live yeast Yarrowia lipolytica 70562 from binary solution: Application of first order derivative spectrophotometry.

PubMed

Dil, Ebrahim Alipanahpour; Ghaedi, Mehrorang; Ghezelbash, Gholam Reza; Asfaram, Arash

2017-05-01

Present study is based on application of live yeast Yarrowia lipolytica 70562 as new biosorbent was investigated for the simultaneous biosorption of Crystal Violet (CV) and Brilliant Green (BG) from wastewater. The effect of operating parameters such as initial dye concentrations (6-14mgL -1 ), solution pH (4.0-8.0) and contact time (4-20h) was investigated by response surface methodology (RSM) for modeling and optimization of biosorption process and accordingly the best operational conditions was set as: initial CV and BG concentration of 8.0, and 10mgL -1 , pH of 7.0 and contact time of 16h. Above specified conditions lead to achievement of maximum biosorption of 98.823% and 99.927% for CV and BG dyes, respectively. The experimental equilibrium data well explained according to Langmuir isotherm model with maximum biosorption capacity of 65.359 and 56.497mgg -1 for BG and CV, respectively. The second order and intraparticle diffusion models as cooperative mechanism has high efficiency and performance for interpretation of real data. Copyright © 2017. Published by Elsevier Inc.

Biochemical characterization of Yarrowia lipolytica LIP8, a secreted lipase with a cleavable C-terminal region.

PubMed

Kamoun, Jannet; Schué, Mathieu; Messaoud, Wala; Baignol, Justine; Point, Vanessa; Mateos-Diaz, Eduardo; Mansuelle, Pascal; Gargouri, Youssef; Parsiegla, Goetz; Cavalier, Jean-François; Carrière, Frédéric; Aloulou, Ahmed

2015-02-01

Yarrowia lipolytica is a lipolytic yeast possessing 16 paralog genes coding for lipases. Little information on these lipases has been obtained and only the major secreted lipase, namely YLLIP2, had been biochemically and structurally characterized. Another secreted lipase, YLLIP8, was isolated from Y. lipolytica culture medium and compared with the recombinant enzyme produced in Pichia pastoris. N-terminal sequencing showed that YLLIP8 is produced in its active form after the cleavage of a signal peptide. Mass spectrometry analysis revealed that YLLIP8 recovered from culture medium lacks a C-terminal part of 33 amino acids which are present in the coding sequence. A 3D model of YLLIP8 built from the X-ray structure of the homologous YLLIP2 lipase shows that these truncated amino acids in YLLIP8 belong to an additional C-terminal region predicted to be mainly helical. Western blot analysis shows that YLLIP8 C-tail is rapidly cleaved upon enzyme secretion since both cell-bound and culture supernatant lipases lack this extension. Mature recombinant YLLIP8 displays a true lipase activity on short-, medium- and long-chain triacylglycerols (TAG), with an optimum activity at alkaline pH on medium chain TAG. It has no apparent regioselectivity in TAG hydrolysis, thus generating glycerol and FFAs as final lipolysis products. YLLIP8 properties are distinct from those of the 1,3-regioselective YLLIP2, acting optimally at acidic pH. These lipases are tailored for complementary roles in fatty acid uptake by Y. lipolytica. Copyright © 2014 Elsevier B.V. All rights reserved.

Deciphering how LIP2 and POX2 promoters can optimally regulate recombinant protein production in the yeast Yarrowia lipolytica.

PubMed

Sassi, Hosni; Delvigne, Frank; Kar, Tambi; Nicaud, Jean-Marc; Coq, Anne-Marie Crutz-Le; Steels, Sebastien; Fickers, Patrick

2016-09-20

In recent years, the non-conventional model yeast species Yarrowia lipolytica has received much attention because it is a useful cell factory for producing recombinant proteins. In this species, expression vectors involving LIP2 and POX2 promoters have been developed and used successfully for protein production at yields similar to or even higher than those of other cell factories, such as Pichia pastoris. However, production processes involving these promoters can be difficult to manage, especially if carried out at large scales in fed-batch bioreactors, because they require hydrophobic inducers, such as oleic acid or methyl oleate. Thus, the challenge has become to reduce loads of hydrophobic substrates while simultaneously promoting recombinant protein production. One possible solution is to replace a portion of the inducer with a co-substrate that can serve as an alternative energy source. However, implementing such an approach would require detailed knowledge of how carbon sources impact promoter regulation, which is surprisingly still lacking for the LIP2 and POX2 promoters. This study's aim was thus to better characterize promoter regulation and cell metabolism in Y. lipolytica cultures grown in media supplemented with different carbon sources. pPOX2 induction could be detected when glucose or glycerol was used as sole carbon source, which meant these carbon source could not prevent promoter induction. In addition, when a mixture of glucose and oleic acid was used in complex medium, pPOX2 induction level was lower that that of pLIP2. In contrast, pLIP2 induction was absent when glucose was present in the culture medium, which meant that cell growth could occur without any recombinant gene expression. When a 40/60 mixture of glucose and oleic acid (w/w) was used, a tenfold increase in promoter induction, as compared to when an oleic-acid-only medium was observed. It was also clear that individual cells were adapting metabolically to use both glucose and oleic

Leucine Biosynthesis Is Involved in Regulating High Lipid Accumulation in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kerkhoven, Eduard J.; Kim, Young-Mo; Wei, Siwei

ABSTRACT The yeastYarrowia lipolyticais a potent accumulator of lipids, and lipogenesis in this organism can be influenced by a variety of factors, such as genetics and environmental conditions. Using a multifactorial study, we elucidated the effects of both genetic and environmental factors on regulation of lipogenesis inY. lipolyticaand identified how two opposite regulatory states both result in lipid accumulation. This study involved comparison of a strain overexpressing diacylglycerol acyltransferase (DGA1) with a control strain grown under either nitrogen or carbon limitation conditions. A strong correlation was observed between the responses on the transcript and protein levels. Combination ofDGA1overexpression with nitrogen limitationmore » resulted in a high level of lipid accumulation accompanied by downregulation of several amino acid biosynthetic pathways, including that of leucine in particular, and these changes were further correlated with a decrease in metabolic fluxes. This downregulation was supported by the measured decrease in the level of 2-isopropylmalate, an intermediate of leucine biosynthesis. Combining the multi-omics data with putative transcription factor binding motifs uncovered a contradictory role for TORC1 in controlling lipid accumulation, likely mediated through 2-isopropylmalate and a Leu3-like transcription factor. IMPORTANCEThe ubiquitous metabolism of lipids involves refined regulation, and an enriched understanding of this regulation would have wide implications. Various factors can influence lipid metabolism, including the environment and genetics. We demonstrated, using a multi-omics and multifactorial experimental setup, that multiple factors affect lipid accumulation in the yeastYarrowia lipolytica. Using integrative analysis, we identified novel interactions between nutrient restriction and genetic factors involving regulators that are highly conserved among eukaryotes. Given that lipid metabolism is involved in many

Phenol Is the Initial Product Formed during Growth and Degradation of Bromobenzene by Tropical Marine Yeast, Yarrowia lipolytica NCIM 3589 via an Early Dehalogenation Step.

PubMed

Vatsal, Aakanksha A; Zinjarde, Smita S; RaviKumar, Ameeta

2017-01-01

Bromobenzene (BrB), a hydrophobic, recalcitrant organic compound, is listed by the environmental protection agencies as an environmental and marine pollutant having hepatotoxic, mutagenic, teratogenic, and carcinogenic effects. The tropical marine yeast Yarrowia lipolytica 3589 was seen to grow aerobically on BrB and displayed a maximum growth rate (μ max ) of 0.04 h -1 . Furthermore, we also observed an increase in cell size and sedimentation velocity for the cells grown on BrB as compared to the glucose grown cells. The cells attached to the hydrophobic bromobenzene droplets through its hydrophobic and acid-base interactions. The BrB (0.5%, 47.6 mM) was utilized by the cells with the release of a corresponding amount of bromide (12.87 mM) and yielded a cell mass of 1.86 g/L after showing 34% degradation in 96 h. Maximum dehalogenase activity of 16.16 U/mL was seen in the cell free supernatant after 24 h of growth. Identification of metabolites formed as a result of BrB degradation, namely, phenol, catechol, cis, cis muconic acid, and carbon dioxide were determined by LC-MS and GC-MS. The initial attack on bromobenzene by Y. lipolytica cells lead to the transient accumulation of phenol as an early intermediate which is being reported for the first time. Degradation of phenol led to catechol which was degraded by the ortho- cleavage pathway forming cis, cis muconic acid and then to Krebs cycle intermediates eventually leading to CO 2 production. The study shows that dehalogenation via an extracellular dehalogenase occurs prior to ring cleavage with phenol as the preliminary degradative compound being produced. The yeast was also able to grow on the degradative products, i.e., phenol and catechol, to varying degrees which would be of potential relevance in the degradation and remediation of xenobiotic environmental bromoaromatic pollutants such as bromobenzene.

Sustainable source of omega-3 eicosapentaenoic acid from metabolically engineered Yarrowia lipolytica: from fundamental research to commercial production.

PubMed

Xie, Dongming; Jackson, Ethel N; Zhu, Quinn

2015-02-01

The omega-3 fatty acids, cis-5, 8, 11, 14, and 17-eicosapentaenoic acid (C20:5; EPA) and cis-4, 7, 10, 13, 16, and 19-docosahexaenoic acid (C22:6; DHA), have wide-ranging benefits in improving heart health, immune function, mental health, and infant cognitive development. Currently, the major source for EPA and DHA is from fish oil, and a minor source of DHA is from microalgae. With the increased demand for EPA and DHA, DuPont has developed a clean and sustainable source of the omega-3 fatty acid EPA through fermentation using metabolically engineered strains of Yarrowia lipolytica. In this mini-review, we will focus on DuPont's technology for EPA production. Specifically, EPA biosynthetic and supporting pathways have been introduced into the oleaginous yeast to synthesize and accumulate EPA under fermentation conditions. This Yarrowia platform can also produce tailored omega-3 (EPA, DHA) and/or omega-6 (ARA, GLA) fatty acid mixtures in the cellular lipid profiles. Fundamental research such as metabolic engineering for strain construction, high-throughput screening for strain selection, fermentation process development, and process scale-up were all needed to achieve the high levels of EPA titer, rate, and yield required for commercial application. Here, we summarize how we have combined the fundamental bioscience and the industrial engineering skills to achieve large-scale production of Yarrowia biomass containing high amounts of EPA, which led to two commercial products, New Harvest™ EPA oil and Verlasso® salmon.

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica.

PubMed

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics.

Molecular Characterization of the Elaeis guineensis Medium-Chain Fatty Acid Diacylglycerol Acyltransferase DGAT1-1 by Heterologous Expression in Yarrowia lipolytica

PubMed Central

Aymé, Laure; Jolivet, Pascale; Nicaud, Jean-Marc; Chardot, Thierry

2015-01-01

Diacylglycerol acyltransferases (DGAT) are involved in the acylation of sn-1,2-diacylglycerol. Palm kernel oil, extracted from Elaeis guineensis (oil palm) seeds, has a high content of medium-chain fatty acids mainly lauric acid (C12:0). A putative E. guineensis diacylglycerol acyltransferase gene (EgDGAT1-1) is expressed at the onset of lauric acid accumulation in the seed endosperm suggesting that it is a determinant of medium-chain triacylglycerol storage. To test this hypothesis, we thoroughly characterized EgDGAT1-1 activity through functional complementation of a Yarrowia lipolytica mutant strain devoid of neutral lipids. EgDGAT1-1 expression is sufficient to restore triacylglycerol accumulation in neosynthesized lipid droplets. A comparative functional study with Arabidopsis thaliana DGAT1 highlighted contrasting substrate specificities when the recombinant yeast was cultured in lauric acid supplemented medium. The EgDGAT1-1 expressing strain preferentially accumulated medium-chain triacylglycerols whereas AtDGAT1 expression induced long-chain triacylglycerol storage in Y. lipolytica. EgDGAT1-1 localized to the endoplasmic reticulum where TAG biosynthesis takes place. Reestablishing neutral lipid accumulation in the Y. lipolytica mutant strain did not induce major reorganization of the yeast microsomal proteome. Overall, our findings demonstrate that EgDGAT1-1 is an endoplasmic reticulum DGAT with preference for medium-chain fatty acid substrates, in line with its physiological role in palm kernel. The characterized EgDGAT1-1 could be used to promote medium-chain triacylglycerol accumulation in microbial-produced oil for industrial chemicals and cosmetics. PMID:26581109

Prospect for Developing a Consolidated Bioprocessing (CBP) Strain Using Xylan as the Substrate: the Case Study of Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Alahuhta, Markus

2016-07-08

To achieve the goal of developing a direct microbial sugar conversion platform for the production of lipids and drop-in fuels from cellulosic biomass substrate, Yarrowia lipolytica was used to investigate its potential for being developed as CBP strain by expressing cellulase and xylanase enzymes. Y. lipolytica is known to accumulate lipids intracellularly and is capable of metabolizing glucose and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in thismore » study, we present successful expression of several xylanases in Y. lipolytica. To the best of our knowledge, this is the first study introducing heterologous hemicellulose genes into the genome of Y. lipolytica. SDS-PAGE and western blotting analysis showed that the endo-xylanase gene XynII and exo-xylosidase gene XlnD were successfully expressed and secreted, and the expressed xylanases were likely either not or sparsely glycosylated, which is advantageous for expression of heterologous proteins from any species. Enzymatic activity tests further demonstrated active expression of XynII and XlnD in Y. lipolytica. Furthermore, synergistic action on converting xylan to xylose was observed when XlnD worked in concert with XynII. XlnD was able to work on the xylo-oligomers generated by XynII, enhancing the xylan conversion to monomeric xylose. The successful expression of these xylanases in Yarrowia further advances us towards our goal to develop a direct microbial conversion process using this organism. and xylose to produce lipids; however, due to the lack of the biomass degrading enzymes, it cannot directly utilize lignocellulosic substrates as carbon sources. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful

Omega-3 production by fermentation of Yarrowia lipolytica: From fed-batch to continuous.

PubMed

Xie, Dongming; Miller, Edward; Sharpe, Pamela; Jackson, Ethel; Zhu, Quinn

2017-04-01

The omega-3 fatty acid, cis-5,8,11,14,17-eicosapentaenoic acid (C20:5; EPA) has wide-ranging benefits in improving heart health, immune function, and mental health. A sustainable source of EPA production through fermentation of metabolically engineered Yarrowia lipolytica has been developed. In this paper, key fed-batch fermentation conditions were identified to achieve 25% EPA in the yeast biomass, which is so far the highest EPA titer reported in the literature. Dynamic models of the EPA fermentation process were established for analyzing, optimizing, and scaling up the fermentation process. In addition, model simulations were used to develop a two-stage continuous process and compare to single-stage continuous and fed- batch processes. The two stage continuous process, which is equipped with a smaller growth fermentor (Stage 1) and a larger production fermentor (Stage 2), was found to be a superior process to achieve high titer, rate, and yield of EPA. A two-stage continuous fermentation experiment with Y. lipolytica strain Z7334 was designed using the model simulation and then tested in a 2 L and 5 L fermentation system for 1,008 h. Compared with the standard 2 L fed-batch process, the two-stage continuous fermentation process improved the overall EPA productivity by 80% and EPA concentration in the fermenter by 40% while achieving comparable EPA titer in biomass and similar conversion yield from glucose. During the long-term experiment it was also found that the Y. lipolytica strain evolved to reduce byproduct and increase lipid production. This is one of the few continuous fermentation examples that demonstrated improved productivity and concentration of a final product with similar conversion yield compared with a fed-batch process. This paper suggests the two-stage continuous fermentation could be an effective process to achieve improved production of omega-3 and other fermentation products where non-growth or partially growth associated kinetics

Regulation of Nitrogen Metabolism by GATA Zinc Finger Transcription Factors in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.

ABSTRACT Fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeastYarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism inY. lipolytica. Deletion of the GATA transcription factor genesgzf3andgzf2resulted in nitrogen source-specific growth defects and greater accumulation of lipids when the cells weremore » growing on a simple nitrogen source. Deletion ofgzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion ofgzf3results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, whilegzf2is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressormig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism. IMPORTANCENitrogen source is commonly used to control lipid production in industrial fungi. Here we identified regulators of nitrogen catabolite repression in the oleaginous yeast

Regulation of nitrogen metabolism by GATA zinc finger transcription factors in Yarrowia lipolytica

DOE PAGES

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.; ...

2017-02-15

Here, fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeast Yarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism in Y. lipolytica. Deletion of the GATA transcription factor genes gzf3 and gzf2 resulted in nitrogen source-specific growth defects and greatermore » accumulation of lipids when the cells were growing on a simple nitrogen source. Deletion of gzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion of gzf3 results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, while gzf2 is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressor mig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism.« less

Regulation of nitrogen metabolism by GATA zinc finger transcription factors in Yarrowia lipolytica

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pomraning, Kyle R.; Bredeweg, Erin L.; Baker, Scott E.

Here, fungi accumulate lipids in a manner dependent on the quantity and quality of the nitrogen source on which they are growing. In the oleaginous yeast Yarrowia lipolytica, growth on a complex source of nitrogen enables rapid growth and limited accumulation of neutral lipids, while growth on a simple nitrogen source promotes lipid accumulation in large lipid droplets. Here we examined the roles of nitrogen catabolite repression and its regulation by GATA zinc finger transcription factors on lipid metabolism in Y. lipolytica. Deletion of the GATA transcription factor genes gzf3 and gzf2 resulted in nitrogen source-specific growth defects and greatermore » accumulation of lipids when the cells were growing on a simple nitrogen source. Deletion of gzf1, which is most similar to activators of genes repressed by nitrogen catabolite repression in filamentous ascomycetes, did not affect growth on the nitrogen sources tested. We examined gene expression of wild-type and GATA transcription factor mutants on simple and complex nitrogen sources and found that expression of enzymes involved in malate metabolism, beta-oxidation, and ammonia utilization are strongly upregulated on a simple nitrogen source. Deletion of gzf3 results in overexpression of genes with GATAA sites in their promoters, suggesting that it acts as a repressor, while gzf2 is required for expression of ammonia utilization genes but does not grossly affect the transcription level of genes predicted to be controlled by nitrogen catabolite repression. Both GATA transcription factor mutants exhibit decreased expression of genes controlled by carbon catabolite repression via the repressor mig1, including genes for beta-oxidation, highlighting the complex interplay between regulation of carbon, nitrogen, and lipid metabolism.« less

Phenol Is the Initial Product Formed during Growth and Degradation of Bromobenzene by Tropical Marine Yeast, Yarrowia lipolytica NCIM 3589 via an Early Dehalogenation Step

PubMed Central

Vatsal, Aakanksha A.; Zinjarde, Smita S.; RaviKumar, Ameeta

2017-01-01

Bromobenzene (BrB), a hydrophobic, recalcitrant organic compound, is listed by the environmental protection agencies as an environmental and marine pollutant having hepatotoxic, mutagenic, teratogenic, and carcinogenic effects. The tropical marine yeast Yarrowia lipolytica 3589 was seen to grow aerobically on BrB and displayed a maximum growth rate (μmax) of 0.04 h-1. Furthermore, we also observed an increase in cell size and sedimentation velocity for the cells grown on BrB as compared to the glucose grown cells. The cells attached to the hydrophobic bromobenzene droplets through its hydrophobic and acid–base interactions. The BrB (0.5%, 47.6 mM) was utilized by the cells with the release of a corresponding amount of bromide (12.87 mM) and yielded a cell mass of 1.86 g/L after showing 34% degradation in 96 h. Maximum dehalogenase activity of 16.16 U/mL was seen in the cell free supernatant after 24 h of growth. Identification of metabolites formed as a result of BrB degradation, namely, phenol, catechol, cis, cis muconic acid, and carbon dioxide were determined by LC–MS and GC–MS. The initial attack on bromobenzene by Y. lipolytica cells lead to the transient accumulation of phenol as an early intermediate which is being reported for the first time. Degradation of phenol led to catechol which was degraded by the ortho- cleavage pathway forming cis, cis muconic acid and then to Krebs cycle intermediates eventually leading to CO2 production. The study shows that dehalogenation via an extracellular dehalogenase occurs prior to ring cleavage with phenol as the preliminary degradative compound being produced. The yeast was also able to grow on the degradative products, i.e., phenol and catechol, to varying degrees which would be of potential relevance in the degradation and remediation of xenobiotic environmental bromoaromatic pollutants such as bromobenzene. PMID:28690604

Physical and physiological impacts of different foam control strategies during a process involving hydrophobic substrate for the lipase production by Yarrowia lipolytica.

PubMed

Kar, Tambi; Destain, Jacqueline; Thonart, Philippe; Delvigne, Frank

2012-05-01

The potentialities for the intensification of the process of lipase production by the yeast Yarrowia lipolytica on a renewable hydrophobic substrate (methyl oleate) have to be investigated. The key factor governing the lipase yield is the intensification of the oxygen transfer rate, considering the fact that Y. lipolytica is a strict aerobe. However, considering the nature of the substrate and the capacity for protein excretion and biosurfactant production of Y. lipolytica, intensification of oxygen transfer rate is accompanied by an excessive formation of foam. Two different foam control strategies have thus been implemented: a classical chemical foam control strategy and a mechanical foam control (MFM) based on the Stirring As Foam Disruption principle. The second strategy allows foam control without any modifications of the physico-chemical properties of the broth. However, the MFM system design induced the formation of a persistent foam layer in the bioreactor. This phenomenon has led to the segregation of microbial cells between the foam phase and the liquid phase in the case of the bioreactors operated with MFM control, and induced a reduction at the level of the lipase yield. More interestingly, flow cytometry experiments have shown that the residence time of microbial cells in the foam phase tends to induce a dimorphic transition which could potentially explain the reduction of lipase excretion.

Robust signal peptides for protein secretion in Yarrowia lipolytica: identification and characterization of novel secretory tags.

PubMed

Celińska, Ewelina; Borkowska, Monika; Białas, Wojciech; Korpys, Paulina; Nicaud, Jean-Marc

2018-06-01

Upon expression of a given protein in an expression host, its secretion into the culture medium or cell-surface display is frequently advantageous in both research and industrial contexts. Hence, engineering strategies targeting folding, trafficking, and secretion of the proteins gain considerable interest. Yarrowia lipolytica has emerged as an efficient protein expression platform, repeatedly proved to be a competitive secretor of proteins. Although the key role of signal peptides (SPs) in secretory overexpression of proteins and their direct effect on the final protein titers are widely known, the number of reports on manipulation with SPs in Y. lipolytica is rather scattered. In this study, we assessed the potential of ten different SPs for secretion of two heterologous proteins in Y. lipolytica. Genomic and transcriptomic data mining allowed us to select five novel, previously undescribed SPs for recombinant protein secretion in Y. lipolytica. Their secretory potential was assessed in comparison with known, widely exploited SPs. We took advantage of Golden Gate approach, for construction of expression cassettes, and micro-volume enzymatic assays, for functional screening of large libraries of recombinant strains. Based on the adopted strategy, we identified novel secretory tags, characterized their secretory capacity, indicated the most potent SPs, and suggested a consensus sequence of a potentially robust synthetic SP to expand the molecular toolbox for engineering Y. lipolytica.

L-Phenylalanine catabolism and 2-phenylethanol synthesis in Yarrowia lipolytica--mapping molecular identities through whole-proteome quantitative mass spectrometry analysis.

PubMed

Celińska, Ewelina; Olkowicz, Mariola; Grajek, Włodzimierz

2015-08-01

A world-wide effort is now being pursued towards the development of flavors and fragrances (F&F) production independently from traditional sources, as well as autonomously from depleting fossil fuel supplies. Biotechnological production of F&F by microbes has emerged as a vivid solution to the current market limitations. Amongst a wide variety of fragrant chemicals, 2-PE is of significant interest to both scientific and industrial community. Although the general overview of the 2-PE synthesis pathway is commonly known, involvement of particular molecular identities in this pathway has not been elucidated in Yarrowia lipolytica to date. The aim of this study was mapping molecular identities involved in 2-PE synthesis in Y. lipolytica. To acquire a comprehensive landscape of the proteins that are directly and indirectly involved in L-Phe degradation and 2-PE synthesis, we took advantage of comprehensibility and sensitivity of high-throughput LC-MS/MS-quantitative analysis. Amongst a number of proteins involved in amino acid turnover and the central carbon metabolism, enzymes involved in L-Phe conversion to 2-PE have been identified. Results on yeast-to-hyphae transition in relation to the character of the provided nitrogen source have been presented. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Design of an efficient medium for heterologous protein production in Yarrowia lipolytica: case of human interferon alpha 2b.

PubMed

Gasmi, Najla; Ayed, Atef; Nicaud, Jean-Marc; Kallel, Héla

2011-05-20

The non conventional yeast Yarrowia lipolytica has aroused a strong industrial interest for heterologous protein production. However most of the studies describing recombinant protein production by this yeast rely on the use of complex media, such media are not convenient for large scale production particularly for products intended for pharmaceutical applications. In addition medium composition can also affect the production yield. Hence it is necessary to design an efficient medium for therapeutic protein expression by this host. Five different media, including four minimal media and a complex medium, were assessed in shake flasks for the production of human interferon alpha 2b (hIFN α2b) by Y. lipolytica under the control of POX2 promoter inducible with oleic acid. The chemically defined medium SM4 formulated by Invitrogen for Pichia pastoris growth was the most suitable. Using statistical experimental design this medium was further optimized. The selected minimal medium consisting in SM4 supplemented with 10 mg/l FeCl₃, 1 g/l glutamate, 5 ml/l PTM1 (Pichia Trace Metals) solution and a vitamin solution composed of myo-inositol, thiamin and biotin was called GNY medium. Compared to shake flask, bioreactor culture in GNY medium resulted in 416-fold increase of hIFN α2b production and 2-fold increase of the biological activity. Furthermore, SM4 enrichment with 5 ml/l PTM1 solution contributed to protect hIFN α2b against the degradation by the 28 kDa protease identified by zymography gel in culture supernatant. The screening of the inhibitory effect of the trace elements present in PTM1 solution on the activity of this protease was achieved using a Box-Behnken design. Statistical data analysis showed that FeCl₃ and MnSO₄ had the most inhibitory effect. We have designed an efficient medium for large scale production of heterologous proteins by Y. lipolytica. The optimized medium GNY is suitable for the production of hIFN α2b with the advantage that no

Engineering Yarrowia lipolytica to Simultaneously Produce Lipase and Single Cell Protein from Agro-industrial Wastes for Feed.

PubMed

Yan, Jinyong; Han, Bingnan; Gui, Xiaohua; Wang, Guilong; Xu, Li; Yan, Yunjun; Madzak, Catherine; Pan, Dujie; Wang, Yaofeng; Zha, Genhan; Jiao, Liangcheng

2018-01-15

Lipases are scarcely exploited as feed enzymes in hydrolysis of lipids for increasing energy supply and improving nutrient use efficiency. In this work, we performed homologous overexpression, in vitro characterization and in vivo assessment of a lipase from the yeast Yarrowia lipolytica for feed purpose. Simultaneously, a large amount of yeast cell biomass was produced, for use as single cell protein, a potential protein-rich feed resource. Three kinds of low cost agro-industrial wastes were tested as substrates for simultaneous production of lipase and single cell protein (SCP) as feed additives: sugarcane molasses, waste cooking oil and crude glycerol from biodiesel production. Sugarcane molasses appeared as the most effective cheap medium, allowing production of 16420 U/ml of lipase and 151.2 g/L of single cell protein at 10 liter fermentation scale. In vitro characterization by mimicking a gastro-intestinal environment and determination of essential amino acids of the SCP, and in vivo oral feeding test on fish all revealed that lipase, SCP and their combination were excellent feed additives. Such simultaneous production of this lipase and SCP could address two main concerns of feed industry, poor utilization of lipid and shortage of protein resource at the same time.

Evaluation of the Composition of Culture Medium for Yeast Biomass Production Using Raw Glycerol from Biodiesel Synthesis

PubMed Central

dos Santos, Elisane Odriosolla; Michelon, Mariano; Furlong, Eliana Badiale; Burkert, Janaína Fernandes de Medeiros; Kalil, Susana Juliano; Burkert, Carlos André Veiga

2012-01-01

The work herewith investigated the production of yeast biomass as a source of protein, using Yarrowia lipolytica NRRL YB-423 and raw glycerol from biodiesel synthesis as the main carbon source. A significant influence of glycerol concentration, initial pH and yeast extract concentration on biomass and protein content was observed according to the 2v5-1 fractional design. These factors were further evaluated using a central composite design and response surface methodology, and an empirical model for protein content was established and validated. The biomass of Yarrowia lipolytica NRRL YB-423 reached 19.5 ± 1.0 g/L in shaken flasks cultivation, with a protein content of 20.1 ± 0.6% (w/w). PMID:24031849

Oleaginous yeast Yarrowia lipolytica culture with synthetic and food waste-derived volatile fatty acids for lipid production.

PubMed

Gao, Ruiling; Li, Zifu; Zhou, Xiaoqin; Cheng, Shikun; Zheng, Lei

2017-01-01

The sustainability of microbial lipids production from traditional carbon sources, such as glucose or glycerol, is problematic given the high price of raw materials. Considerable efforts have been directed to minimize the cost and find new alternative carbon sources. Volatile fatty acids (VFAs) are especially attractive raw materials, because they can be produced from a variety of organic wastes fermentation. Therefore, the use of volatile fatty acids as carbon sources seems to be a feasible strategy for cost-effective microbial lipid production. Lipid accumulation in Y. lipolytica using synthetic and food waste-derived VFAs as substrates was systematically compared and evaluated in batch cultures. The highest lipid content obtained with acetic, butyric, and propionic acids reached 31.62 ± 0.91, 28.36 ± 0.74, and 28.91 ± 0.66%, respectively. High concentrations of VFA inhibited cell growth in the following order: butyric acid > propionic acid > acetic acid. Within a 30-day experimental period, Y. lipolytica could adapt up to 20 g/L acetic acid, whereas the corresponding concentration of propionic acid and butyric acid were 10 and 5 g/L, respectively. Cultures on a VFA mixture showed that the utilization of different types of VFA by Y. lipolytica was not synchronized but rather performed in a step-wise manner. Although yeast fermentation is an exothermic process, and the addition of VFA will directly affect the pH of the system by increasing environmental acidity, cultures at a cultivation temperature of 38 °C and uncontrolled pH demonstrated that Y. lipolytica had high tolerance in the high temperature and acidic environment when a low concentration (2.5 g/L) of either synthetic or food waste-derived VFA was used. However, batch cultures fed with food fermentate yielded lower lipid content (18.23 ± 1.12%) and lipid productivity (0.12 ± 0.02 g/L/day). The lipid composition obtained with synthetic and food waste-derived VFA was similar to

Use of Plackett-Burman design for rapid screening of nitrogen and carbon sources for the production of lipase in solid state fermentation by Yarrowia lipolytica from mustard oil cake (Brassica napus).

PubMed

Imandi, Sarat Babu; Karanam, Sita Kumari; Garapati, Hanumantha Rao

2013-01-01

Mustard oil cake (Brassica napus), the residue obtained after extraction of mustard oil from mustard oil seeds, was investigated for the production of lipase under solid state fermentation (SSF) using the marine yeast Yarrowia lipolytica NCIM 3589. Process parameters such as incubation time, biomass concentration, initial moisture content, carbon source concentration and nitrogen source concentration of the medium were optimized. Screening of ten nitrogen and five carbon sources has been accomplished with the help of Plackett-Burman design. The highest lipase activity of 57.89 units per gram of dry fermented substrate (U/gds) was observed with the substrate of mustard oil cake in four days of fermentation.

Optimized invertase expression and secretion cassette for improving Yarrowia lipolytica growth on sucrose for industrial applications.

PubMed

Lazar, Zbigniew; Rossignol, Tristan; Verbeke, Jonathan; Crutz-Le Coq, Anne-Marie; Nicaud, Jean-Marc; Robak, Małgorzata

2013-11-01

Yarrowia lipolytica requires the expression of a heterologous invertase to grow on a sucrose-based substrate. This work reports the construction of an optimized invertase expression cassette composed of Saccharomyces cerevisiae Suc2p secretion signal sequence followed by the SUC2 sequence and under the control of the strong Y. lipolytica pTEF promoter. This new construction allows a fast and optimal cleavage of sucrose into glucose and fructose and allows cells to reach the maximum growth rate. Contrary to pre-existing constructions, the expression of SUC2 is not sensitive to medium composition in this context. The strain JMY2593, expressing this new cassette with an optimized secretion signal sequence and a strong promoter, produces 4,519 U/l of extracellular invertase in bioreactor experiments compared to 597 U/l in a strain expressing the former invertase construction. The expression of this cassette strongly improved production of invertase and is suitable for simultaneously high production level of citric acid from sucrose-based media.

Developing cellulolytic Yarrowia lipolytica as a platform for the production of valuable products in consolidated bioprocessing of cellulose.

PubMed

Guo, Zhong-Peng; Robin, Julien; Duquesne, Sophie; O'Donohue, Michael Joseph; Marty, Alain; Bordes, Florence

2018-01-01

Both industrial biotechnology and the use of cellulosic biomass as feedstock for the manufacture of various commercial goods are prominent features of the bioeconomy. In previous work, with the aim of developing a consolidated bioprocess for cellulose bioconversion, we conferred cellulolytic activity of Yarrowia lipolytica , one of the most widely studied "nonconventional" oleaginous yeast species. However, further engineering this strain often leads to the loss of previously introduced heterologous genes due to the presence of multiple LoxP sites when using Cre -recombinase to remove previously employed selection markers. In the present study, we first optimized the strategy of expression of multiple cellulases and rescued selection makers to obtain an auxotrophic cellulolytic Y. lipolytica strain. Then we pursued the quest, exemplifying how this cellulolytic Y. lipolytica strain can be used as a CBP platform for the production of target products. Our results reveal that overexpression of SCD1 gene, encoding stearoyl-CoA desaturase, and DGA1 , encoding acyl-CoA:diacylglycerol acyltransferase, confers the obese phenotype to the cellulolytic Y. lipolytica . When grown in batch conditions and minimal medium, the resulting strain consumed 12 g/L cellulose and accumulated 14% (dry cell weight) lipids. Further enhancement of lipid production was achieved either by the addition of glucose or by enhancing cellulose consumption using a commercial cellulase cocktail. Regarding the latter option, although the addition of external cellulases is contrary to the concept of CBP, the amount of commercial cocktail used remained 50% lower than that used in a conventional process (i.e., without internalized production of cellulases). The introduction of the LIP2 gene into cellulolytic Y. lipolytica led to the production of a strain capable of producing lipase 2 while growing on cellulose. Remarkably, when the strain was grown on glucose, the expression of six cellulases did not

Citric acid production from partly deproteinized whey under non-sterile culture conditions using immobilized cells of lactose-positive and cold-adapted Yarrowia lipolytica B9.

PubMed

Arslan, Nazli Pinar; Aydogan, Mehmet Nuri; Taskin, Mesut

2016-08-10

The present study was performed to produce citric acid (CA) from partly deproteinized cheese whey (DPCW) under non-sterile culture conditions using immobilized cells of the cold-adapted and lactose-positive yeast Yarrowia lipolytica B9. DPCW was prepared using the temperature treatment of 90°C for 15min. Sodium alginate was used as entrapping agent for cell immobilization. Optimum conditions for the maximum CA production (33.3g/L) in non-sterile DPCW medium were the temperature of 20°C, pH 5.5, additional lactose concentration of 20g/L, sodium alginate concentration of 2%, number of 150 beads/100mL and incubation time of 120h. Similarly, maximum citric acid/isocitric acid (CA/ICA) ratio (6.79) could be reached under these optimal conditions. Additional nitrogen and phosphorus sources decreased CA concentration and CA/ICA ratio. Immobilized cells were reused in three continuous reaction cycles without any loss in the maximum CA concentration. The unique combination of low pH and temperature values as well as cell immobilization procedure could prevent undesired microbial contaminants during CA production. This is the first work on CA production by cold-adapted microorganisms under non-sterile culture conditions. Besides, CA production using a lactose-positive strain of the yeast Y. lipolytica was investigated for the first time in the present study. Copyright © 2016 Elsevier B.V. All rights reserved.

[Activation of the alternative oxidase of Yarrowia lipolytica by adenosine 5'-monophosphate].

PubMed

Medentsev, A G; Arinbasarova, A Iu; Smirnova, N M; Akimenko, V K

2004-01-01

The study of the effect of nucleoside phosphates on the activity of cyanide-resistant oxidase in the mitochondria and the submitochondrial particles of Yarrowia lipolytica showed that adenosine monophosphate (5'-AMP, AMP) did not stimulate the respiration of the intact mitochondria. The incubation of the mitochondria at room temperature (25 degrees C) for 3-5 h or their treatment with ultrasound, phospholipase A, and detergent Triton X-100 at a low temperature inactivated the cyanide-resistant alternative oxidase. The inactivated alternative oxidase could be reactivated by AMP. The reactivating effect of AMP was enhanced by azolectin. Some other nucleoside phosphates also showed reactivating ability in the following descending order. AMP = GMP > GDP > GTP > XMP > IMP. The apparent reaction rate constant Km for AMP upon the reactivation of the alternative oxidase of mitochondria treated with Triton X-100 or incubated at 25 degrees C was 12.5 and 20 microM, respectively. The Km for AMP upon the reactivation of the alternative oxidase of submitochondrial particles was 15 microM. During the incubation of yeast cells under conditions promoting the development of alternative oxidase, the content of adenine nucleotides (AMP, ADP, and ATP) in the cells and their respiration tended to decrease. The subsequent addition of cyanide to the cells activated their respiration, diminished the intracellular content of ATP three times, and augmented the content of AMP five times. These data suggest that the stimulation of cell respiration by cyanide may be due to the activation of alternative oxidase by AMP.

Mutants of Yarrowia lipolytica NCIM 3589 grown on waste cooking oil as a biofactory for biodiesel production.

PubMed

Katre, Gouri; Ajmera, Namasvi; Zinjarde, Smita; RaviKumar, Ameeta

2017-10-24

Oleaginous yeasts are fast emerging as a possible feedstock for biodiesel production. Yarrowia lipolytica, a model oleaginous yeast is known to utilize a variety of hydrophobic substrates for lipid accumulation including waste cooking oil (WCO). Approaches to increase lipid content in this yeast include metabolic engineering which requires manipulation of multiple genes in the lipid biosynthesis pathway. A classical and cost-effective approach, namely, random chemical mutagenesis on the yeast can lead to increased production of biodiesel as is explored here. In this study, chemical mutagenesis using the alkylating agent, N- methyl-N'-nitro-N-nitrosoguanidine (MNNG) as well as an additional treatment with cerulenin, a fatty acid synthase inhibitor generated 800 mutants of Y. lipolytica NCIM 3589 (761 MNNG treated and 39 MNNG + cerulenin treated). A three-stage screening using Sudan Black B plate technique, Nile red fluorimetry and total lipid extraction using solvent was performed, which enabled selection of ten high lipid yielding mutants. Time course studies of all the ten mutants were further undertaken in terms of biomass, lipid yield and lipid content to select three stable mutants (YlB6, YlC7 and YlE1) capable of growing and accumulating lipid on WCO, with lipid contents of 55, 60 and 67% as compared to 45% for the wild type. The mutants demonstrated increased volumetric lipid productivities (0.062, 0.044 and 0.041 g L -1  h -1 ) as compared to the wild type (0.033 g L -1  h -1 ). The fatty acid profile of the three mutants consisted of a high content of C16 and C18 saturated and monounsaturated fatty acids and was found to be suitable for biodiesel production. The fuel properties, namely, density, kinematic viscosity, total acid number, iodine value of the three mutants were evaluated and found to lie within the limits specified by internationally accepted standards. Additionally, it was noted that the mutants demonstrated better cetane numbers and

Functional analysis of recombinant human and Yarrowia lipolytica O-GlcNAc transferases expressed in Saccharomyces cerevisiae.

PubMed

Oh, Hye Ji; Moon, Hye Yun; Cheon, Seon Ah; Hahn, Yoonsoo; Kang, Hyun Ah

2016-10-01

O-linked β-N-acetylglucosamine (O-GlcNAc) glycosylation is an important post-translational modification in many cellular processes. It is mediated by O-GlcNAc transferases (OGTs), which catalyze the addition of O-GlcNAc to serine or threonine residues of the target proteins. In this study, we expressed a putative Yarrowia lipolytica OGT (YlOGT), the only homolog identified in the subphylum Saccharomycotina through bioinformatics analysis, and the human OGT (hOGT) as recombinant proteins in Saccharomyces cerevisiae, and performed their functional characterization. Immunoblotting assays using antibody against O-GlcNAc revealed that recombinant hOGT (rhOGT), but not the recombinant YlOGT (rYlOGT), undergoes auto-O-GlcNAcylation in the heterologous host S. cerevisiae. Moreover, the rhOGT expressed in S. cerevisiae showed a catalytic activity during in vitro assays using casein kinase II substrates, whereas no such activity was obtained in rYlOGT. However, the chimeric human-Y. lipolytica OGT, carrying the human tetratricopeptide repeat (TPR) domain along with the Y. lipolytica catalytic domain (CTD), mediated the transfer of O-GlcNAc moiety during the in vitro assays. Although the overexpression of full-length OGTs inhibited the growth of S. cerevisiae, no such inhibition was obtained upon overexpression of only the CTD fragment, indicating the role of TPR domain in growth inhibition. This is the first report on the functional analysis of the fungal OGT, indicating that the Y. lipolytica OGT retains its catalytic activity, although the physiological role and substrates of YlOGT remain to be elucidated.

Occurrence and diversity of marine yeasts in Antarctica environments

NASA Astrophysics Data System (ADS)

Zhang, Xue; Hua, Mingxia; Song, Chunli; Chi, Zhenming

2012-03-01

A total of 28 yeast strains were obtained from the sea sediment of Antarctica. According to the results of routine identification and molecular characterization, the strains belonged to species of Yarrowia lipolytica, Debaryomyces hansenii, Rhodotorula slooffiae, Rhodotorula mucilaginosa, Sporidiobolus salmonicolor, Aureobasidium pullulans, Mrakia frigida and Guehomyces pullulans, respectively. The Antarctica yeasts have wide potential applications in biotechnology, for some of them can produce β-galactosidase and killer toxins.

Cloning, Expression, and Biochemical Characterization of an Enantioselective Lipase, YLIP9, from Yarrowia lipolytica MSR80.

PubMed

Syal, Poonam; Gupta, Rani

2015-05-01

A novel lipase gene, ylip9, of Yarrowia lipolytica MSR80 was cloned and expressed in pEZZ18-HB101 system and was 99% identical to YLIP9 of Y. lipolytica CLIB122. It was purified using IgG-Sepharose as ZZ fused YLIP9 and had specific activity of 0.8 U/mg. ZZ-YLIP9 was most active at pH 8.0 and 70 °C. It was stable over a wide pH range of 3.0-11.0 and 100 % active at 70 °C up to 2 h and had t1/2 of 286.42 min at 80 °C. It showed high specificity toward p-nitrophenyldecanoate with kcat and catalytic efficiency of 30.17 s(-1) and 16.67 mM(-1) s(-1), respectively. It was non-regioselective, but an S-enantioselective lipase and the percentage conversion were enhanced in presence of hexane. ZZ-YLIP9 was stable in all of the organic solvents used, and its activity was enhanced by solvents having logP value less than 2.

Citric acid production from hydrolysate of pretreated straw cellulose by Yarrowia lipolytica SWJ-1b using batch and fed-batch cultivation.

PubMed

Liu, Xiaoyan; Lv, Jinshun; Zhang, Tong; Deng, Yuanfang

2015-01-01

In this study, crude cellulase produced by Trichoderma reesei Rut-30 was used to hydrolyze pretreated straw. After the compositions of the hydrolysate of pretreated straw were optimized, the study showed that natural components of pretreated straw without addition of any other components such as (NH4)2SO4, KH2PO4, or Mg(2+) were suitable for citric acid production by Yarrowia lipolytica SWJ-1b, and the optimal ventilatory capacity was 10.0 L/min/L medium. Batch and fed-batch production of citric acid from the hydrolysate of pretreated straw by Yarrowia lipolytica SWJ-1b has been investigated. In the batch cultivation, 25.4 g/L and 26.7 g/L citric acid were yields from glucose and hydrolysate of straw cellulose, respectively, while the cultivation time was 120 hr. In the three-cycle fed-batch cultivation, citric acid (CA) production was increased to 42.4 g/L and the cultivation time was extended to 240 hr. However, iso-citric acid (ICA) yield in fed-batch cultivation (4.0 g/L) was similar to that during the batch cultivation (3.9 g/L), and only 1.6 g/L of reducing sugar was left in the medium at the end of fed-batch cultivation, suggesting that most of the added carbon was used in the cultivation.

D-stat culture for studying the metabolic shifts from oxidative metabolism to lipid accumulation and citric acid production in Yarrowia lipolytica.

PubMed

Ochoa-Estopier, Abril; Guillouet, Stéphane E

2014-01-20

Lipid accumulation in oleaginous yeasts is triggered by nutrient imbalance in the culture medium between the carbon source in excess and the nitrogen source in limiting concentration. However Yarrowia lipolytica when cultivated on glucose as the sole carbon source, mainly produces citric acid upon nitrogen limitation over lipid accumulation (only 5-10% triacylglycerol). Therefore for developing bioprocess for the production of triacylglycerol from renewable carbon source as glucose it is of first importance to control this imbalance in order to avoid citric acid production during TAG accumulation. Using D-stat cultivation system, where the N/C was linearly decreased using a constant change rate we were able to identify the N/C ratio inducing TAG accumulation (0.085NmolCmol(-1)) and citric acid (0.021NmolCmol(-1)). We therefore demonstrated that it was possible to accumulate lipids without excretion citric acid as long as the N/C was within this indicated range. Moreover enzyme specific activities measurement during the D-stat indicated that ATP-citrate lyase, malic enzyme and acetyl-coA carboxylase were strongly induced at the onset of lipid accumulation and showed different patterns when citric acid was excreted. Our results give relevant information for future industrial bioprocess development concerning the production of lipids using renewable carbohydrate substrates as an alternative way to produce synthons for fuel or chemical industry. By controlling the N/C over the fermentation process on glucose Y. lipolytica can accumulate lipids without excreting citric acid. Copyright © 2013 Elsevier B.V. All rights reserved.

Dual CRISPR-Cas9 Cleavage Mediated Gene Excision and Targeted Integration in Yarrowia lipolytica.

PubMed

Gao, Difeng; Smith, Spencer; Spagnuolo, Michael; Rodriguez, Gabriel; Blenner, Mark

2018-05-29

CRISPR-Cas9 technology has been successfully applied in Yarrowia lipolytica for targeted genomic editing including gene disruption and integration; however, disruptions by existing methods typically result from small frameshift mutations caused by indels within the coding region, which usually resulted in unnatural protein. In this study, a dual cleavage strategy directed by paired sgRNAs is developed for gene knockout. This method allows fast and robust gene excision, demonstrated on six genes of interest. The targeted regions for excision vary in length from 0.3 kb up to 3.5 kb and contain both non-coding and coding regions. The majority of the gene excisions are repaired by perfect nonhomologous end-joining without indel. Based on this dual cleavage system, two targeted markerless integration methods are developed by providing repair templates. While both strategies are effective, homology mediated end joining (HMEJ) based method are twice as efficient as homology recombination (HR) based method. In both cases, dual cleavage leads to similar or improved gene integration efficiencies compared to gene excision without integration. This dual cleavage strategy will be useful for not only generating more predictable and robust gene knockout, but also for efficient targeted markerless integration, and simultaneous knockout and integration in Y. lipolytica. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The RAD52 ortholog of Yarrowia lipolytica is essential for nuclear integrity and DNA repair.

PubMed

Campos-Góngora, Eduardo; Andaluz, Encarnación; Bellido, Alberto; Ruiz-Herrera, José; Larriba, German

2013-08-01

Yarrowia lipolytica (Yl) is a dimorphic fungus that has become a well-established model for a number of biological processes, including secretion of heterologous and chimerical proteins. However, little is known on the recombination machinery responsible for the integration in the genome of the exogenous DNA encoding for those proteins. We have carried out a phenotypic analysis of rad52 deletants of Y. lipolytica. YlRad52 exhibited 20-30% identity with Rad52 homologues of other eukaryotes, including Saccharomyces cerevisiae and Candida albicans. Ylrad52-Δ strains formed colonies on YPD-agar plates which were spinier and smaller than those from wild type, whereas in YPD liquid cultures they exhibited a decreased grow rate and contained cells with aberrant morphology and fragmented chromatin, supporting a role for homologous recombination (HR) in genome stability under nondamaging conditions. In addition, Ylrad52 mutants showed moderate to high sensitivity to UV light, oxidizing agents and compounds that cause single- (SSB) and double-strand breaks (DSB), indicating an important role for Rad52 in DNA repair. These findings extend to Yl previous observations indicating that RAD52 is a crucial gene for DNA repair in other fungi, including S. cerevisiae, C. albicans and Schizosaccharomyces pombe. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

An evolutionary metabolic engineering approach for enhancing lipogenesis in Yarrowia lipolytica.

PubMed

Liu, Leqian; Pan, Anny; Spofford, Caitlin; Zhou, Nijia; Alper, Hal S

2015-05-01

Lipogenic organisms provide an ideal platform for biodiesel and oleochemical production. Through our previous rational metabolic engineering efforts, lipogenesis titers in Yarrowia lipolytica were significantly enhanced. However, the resulting strain still suffered from decreased biomass generation rates. Here, we employ a rapid evolutionary metabolic engineering approach linked with a floating cell enrichment process to improve lipogenesis rates, titers, and yields. Through this iterative process, we were able to ultimately improve yields from our prior strain by 55% to achieve production titers of 39.1g/L with upwards of 76% of the theoretical maximum yield of conversation. Isolated cells were saturated with up to 87% lipid content. An average specific productivity of 0.56g/L/h was achieved with a maximum instantaneous specific productivity of 0.89g/L/h during the lipid production phase in fermentation. Genomic sequencing of the evolved strains revealed a link between a decrease/loss of function mutation of succinate semialdehyde dehydrogenase, uga2, suggesting the importance of gamma-aminobutyric acid assimilation in lipogenesis. This linkage was validated through gene deletion experiments. This work presents an improved host strain that can serve as a platform for efficient oleochemical production. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Production of Laccase by Recombinant Yarrowia lipolytica from Molasses: Bioprocess Development Using Statistical Modeling and Increase Productivity in Shake-Flask and Bioreactor Cultures.

PubMed

Darvishi, Farshad; Moradi, Marzieh; Madzak, Catherine; Jolivalt, Claude

2017-03-01

Laccases are used in numerous applications, from green degradation of various xenobiotic compounds, waste detoxification, textile dye bleaching, and delignification of lignocellulose materials to biofuel production. In this study, the recombinant Yarrowia lipolytica YL4 strain carrying the white-rot fungus Trametes versicolor laccase IIIb gene was used for laccase production from beet molasses as an agro-industrial residue. Response surface methodology was used to statistical optimization of the production of laccase by Y. lipolytica using an industrial medium containing molasses which allows a six times increase in laccase activity compared to primary medium contains glucose after 144 h. In bioreactor cultivation after 48 h, laccase production reached to 3.7- and 22.5-fold more than optimized and primary media in shake-flask cultures, respectively. Laccase productivity in bioreactor (0.0937 U/h) was higher than shake-flask culture (0.0084 U/h). The present study provides valuable information about statistical optimization of bioprocess development for cost-effective production of laccase and other heterologous proteins in Y. lipolytica from beet molasses as sole carbon source, thus allowing the valorization and decreasing environmental pollution of this agro-industrial waste.

The growth, properties and interactions of yeasts and bacteria associated with the maturation of Camembert and blue-veined cheeses.

PubMed

Addis, E; Fleet, G H; Cox, J M; Kolak, D; Leung, T

2001-09-19

The growth of yeasts and bacteria were monitored during the maturation of Camembert and blue-veined cheese produced in Australia. Yeasts were prominent throughout maturation, growing to 10(5)-10(9)/g, depending on the manufacturer. Debaryomyces hansenii predominated, but there were lesser, inconsistent contributions from Yarrowia lipolytica. Of the non-lactic acid bacteria, Acinetobacter species were significant during the maturation of Camembert but not blue-veined cheeses, and grew to 10(6)-10(8) cfu/g. Staphylococcus and Micrococcus species were consistently isolated from the cheeses with Staphylococcus xylosus growing to 10(5)-10(9) cfu/g, depending on the product. Lactic acid bacteria (10(7)-10(9) cfu/g) were present throughout maturation but were not identified. Interactions between the various yeasts and bacterial isolates were examined. Several strains of D. hansenii exhibited killer activity but not against Y. lipolytica. None of the yeasts were antagonistic towards the bacteria but some strains of D. hansenii enhanced the growth of Y. lipolytica and S. xylosus. The yeast and bacterial isolates exhibited various degrees of extracellular proteolytic and lipolytic activities.

Development of New Genetic Manipulation Tools for Metabolic Engineering of Diatoms

DTIC Science & Technology

2008-08-28

protein L41 has been shown in a variety of yeasts to be involved in resistance to anisomycin and cycloheximide 7𔄂. A conserved mutation from proline...atipitis Yarrowia lipolytica Candida troplcalis S. cerevisiae A. nidulans Oryza sativa Homo sapiens P. tricornutum T. pseudonana...Pichia stipitis Yarrowia lipolytica Candida troplcalis S. cerevisiae A. nidulans Oryza sativa Homo sapiens P

Metabolic Engineering of Oleaginous Yeasts for Fatty Alcohol Production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Knoshaug, Eric

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

Proteomic analysis of the response of α-ketoglutarate-producer Yarrowia lipolytica WSH-Z06 to environmental pH stimuli.

PubMed

Guo, Hongwei; Wan, Hui; Chen, Hongwen; Fang, Fang; Liu, Song; Zhou, Jingwen

2016-10-01

During bioproduction of short-chain carboxylates, a shift in pH is a common strategy for enhancing the biosynthesis of target products. Based on two-dimensional gel electrophoresis, comparative proteomics analysis of general and mitochondrial protein samples was used to investigate the cellular responses to environmental pH stimuli in the α-ketoglutarate overproducer Yarrowia lipolytica WSH-Z06. The lower environmental pH stimuli tensioned intracellular acidification and increased the level of reactive oxygen species (ROS). A total of 54 differentially expressed protein spots were detected, and 11 main cellular processes were identified to be involved in the cellular response to environmental pH stimuli. Slight decrease in cytoplasmic pH enhanced the cellular acidogenicity by elevating expression level of key enzymes in tricarboxylic acid cycle (TCA cycle). Enhanced energy biosynthesis, ROS elimination, and membrane potential homeostasis processes were also employed as cellular defense strategies to compete with environmental pH stimuli. Owing to its antioxidant role of α-ketoglutarate, metabolic flux shifted to α-ketoglutarate under lower pH by Y. lipolytica in response to acidic pH stimuli. The identified differentially expressed proteins provide clues for understanding the mechanisms of the cellular responses and for enhancing short-chain carboxylate production through metabolic engineering or process optimization strategies in combination with manipulation of environmental conditions.

Ylpex5 mutation partially suppresses the defective hyphal growth of a Yarrowia lipolytica ceramide synthase mutant, Yllac1, by recovering lipid raft polarization and vacuole morphogenesis.

PubMed

Bal, Jyotiranjan; Lee, Hye-Jeong; Cheon, Seon Ah; Lee, Kyung Jin; Oh, Doo-Byoung; Kim, Jeong-Yoon

2013-01-01

Sphingolipids are involved in cell differentiation and morphogenesis in eukaryotic cells. In this study, YlLac1p, a ceramide synthase required for glucosylceramide (GlcCer) synthesis, was found to be essential for hyphal growth in Yarrowia lipolytica. Y. lipolytica GlcCer was shown to be composed of a C16:0 fatty acid, which is hydroxylated at C2, and a C18:2 long chain base, which is unsaturated at both C4 and C8 and methylated at C9. Domain swapping analysis revealed that the entire TRAM/Lag1/CLN8 (TLC) domain, not the Lag1 motif, is crucial for the function of YlLac1p. YlDes1p, the C4 desaturase of the ceramide synthesized by YlLac1p, was also required for Y. lipolytica morphogenesis. Both Yllac1Δ and Yldes1Δ mutants neither polarize lipid rafts nor form normal vacuoles. Interestingly, mutation in YlPEX5, which encode a peroxisomal targeting signal receptor, partially suppressed the defective hyphal growth of Yllac1Δ. The Yllac1ΔYlpex5Δ mutant restored the ability to polarize lipid rafts and to form normal vacuoles, although it could not synthesize GlcCer. Taken together, our results suggest that GlcCer or GlcCer derivatives may be involved in hyphal morphogenesis in Y. lipolytica, at least in part, by affecting polarization of lipid rafts and vacuole morphogenesis. Copyright © 2012 Elsevier Inc. All rights reserved.

Screening of a thiamine-auxotrophic yeast for alpha-ketoglutaric acid overproduction.

PubMed

Zhou, Jingwen; Zhou, Haiyan; Du, Guocheng; Liu, Liming; Chen, Jian

2010-09-01

To obtain a thiamine-auxotrophic yeast strain that overproduces alpha-ketoglutaric acid (alpha-KG) from glycerol and to investigate nutrient effects on alpha-KG production. Yeast strain WSH-Z06, a thiamine auxotroph that gave high yields of alpha-KG from glycerol, was obtained by screening for ampicillin/kanamycin resistance and thiamine auxotrophy. The strain was identified as Yarrowia lipolytica based on physiological, chemical, and phylogenetic analysis. The ability of the strain to convert glycerol to alpha-KG was analysed by investigating the effects of nutritional factors, including thiamine, riboflavin, nitrogen sources, and calcium ion. Thiamine and calcium ion concentration had the greatest effect on alpha-KG accumulation. Under optimal conditions, a yield of 39.2 g l(-1)alpha-KG was obtained from 100 g l(-1) glycerol, with 16.84 g l(-1) pyruvate as a by-product. The current work provides a method for screening for an alpha-KG overproducer. Nutrients have a significant impact on alpha-KG production in the yeast strain presented here. The alpha-KG-overproducing yeast strain Y. lipolytica WSH-Z06 is a promising parent strain for further metabolic engineering to lower by-product accumulation and accelerate glycerol utilization.

Optimization of a low-cost hyperosmotic medium and establishing the fermentation kinetics of erythritol production by Yarrowia lipolytica from crude glycerol.

PubMed

Yang, Li-Bo; Zhan, Xiao-Bei; Zhu, Li; Gao, Min-Jie; Lin, Chi-Chung

2016-05-18

The production of erythritol by Yarrowia lipolytica from low-cost substitutable substrates for high yield was investigated. Crude glycerol, urea, and NaCl related to osmotic pressure were the most significant factors affecting erythritol production. An artificial neural network model and genetic algorithm were used to search the optimal composition of the significant factors and locate the resulting erythritol yield. Medium with 232.39 g/L crude glycerol, 1.57 g/L urea, and 31.03 g/L NaCl led to predictive maximum erythritol concentration of 110.7 g/L. The erythritol concentration improved from 50.4 g/L to 109.2 g/L with the optimized medium, which was reproducible. Erythritol fermentation kinetics were investigated in a batch system. Multistep fermentation kinetic models with hyperosmotic inhibitory effects were developed. The resulting mathematical equations provided a good description of temporal variations such as microbial growth (X), substrate consumption (S), and product formation (P) in erythritol fermentation. The accordingly derived model is the first reported model for fermentative erythritol production from glycerol, providing useful information to optimize the growth of Y. lipolytica and contributing visual description for the erythritol fermentation process under high osmotic pressure, as well as improvement of productivity and efficiency.

Co-expression of Exo-inulinase and Endo-inulinase Genes in the Oleaginous Yeast Yarrowia lipolytica for Efficient Single Cell Oil Production from Inulin.

PubMed

Shi, Nianci; Mao, Weian; He, Xiaoxia; Chi, Zhe; Chi, Zhenming; Liu, Guanglei

2018-05-01

Yarrowia lipolytica is a promising platform for the single cell oil (SCO) production. In this study, a transformant X+N8 in which exo- and endo-inulinase genes were co-expressed could produce an inulinase activity of 124.33 U/mL within 72 h. However, the inulinase activity of a transformant X2 carrying a single exo-inulinase gene was only 47.33 U/mL within 72 h. Moreover, the transformant X+N8 could accumulate 48.13% (w/w) SCO from inulin and the cell dry weight reached 13.63 g/L within 78 h, which were significantly higher than those of the transformant X2 (41.87% (w/w) and 11.23 g/L) under the same conditions. In addition, inulin hydrolysis and utilization of the transformant X+N8 were also more efficient than those of the transformant X2 during the fermentation process. These results demonstrated that the co-expression of the exo- and endo-inulinase genes significantly enhanced the SCO production from inulin due to the improvement of the inulinase activity and the synergistic action of exo- and endo-inulinase. Besides, over 95.01% of the fatty acids from the transformant X+N8 were C16-C18, especially C18:1 (53.10%), suggesting that the fatty acids could be used as feedstock for biodiesel production.

Improvement of erythrose reductase activity, deletion of by-products and statistical media optimization for enhanced erythritol production from Yarrowia lipolytica mutant 49.

PubMed

Ghezelbash, Gholam Reza; Nahvi, Iraj; Emamzadeh, Rahman

2014-08-01

The purpose of the present investigation was to produce erythritol by Yarrowia lipolytica mutant without any by-products. Mutants of Y. lipolytica were generated by ultra-violet for enhancing erythrose reductase (ER) activity and erythritol production. The mutants showing the highest ER activity were screened by triphenyl tetrazolium chloride agar plate assay. Productivity of samples was analyzed by thin-layer chromatography and high-performance liquid chromatography equipped with the refractive index detector. One of the mutants named as mutant 49 gave maximum erythritol production without any other by-products (particularly glycerol). Erythritol production and specific ER activity in mutant 49 increased to 1.65 and 1.47 times, respectively, in comparison with wild-type strain. The ER gene of wild and mutant strains was sequenced and analyzed. A general comparison of wild and mutant gene sequences showed the replacement of Asp(270) with Glu(270) in ER protein. In order to enhance erythritol production, we used a three component-three level-one response Box-Behnken of response surface methodology model. The optimum medium composition for erythritol production was found to be (g/l) glucose 279.49, ammonium sulfate 9.28, and pH 5.41 with 39.76 erythritol production.

A comparative analysis of single cell and droplet-based FACS for improving production phenotypes: Riboflavin overproduction in Yarrowia lipolytica.

PubMed

Wagner, James M; Liu, Leqian; Yuan, Shuo-Fu; Venkataraman, Maya V; Abate, Adam R; Alper, Hal S

2018-04-23

Evolutionary approaches to strain engineering inherently require the identification of suitable selection techniques for the product and phenotype of interest. In this work, we undertake a comparative analysis of two related but functionally distinct methods of high-throughput screening: traditional single cell fluorescence activated cell sorting (single cell FACS) and microdroplet-enabled FACS (droplet FACS) using water/oil/water (w/o/w) emulsions. To do so, we first engineer and evolve the non-conventional yeast Yarrowia lipolytica for high extracellular production of riboflavin (vitamin B2), an innately fluorescent product. Following mutagenesis and adaptive evolution, a direct parity-matched comparison of these two selection strategies was conducted. Both single cell FACS and droplet FACS led to significant increases in total riboflavin titer (32 and 54 fold relative to the parental PO1f strain, respectively). However, single cell FACS favored intracellular riboflavin accumulation (with only 70% of total riboflavin secreted) compared with droplet FACS that favored extracellular product accumulation (with 90% of total riboflavin secreted). We find that for the test case of riboflavin, the extent of secretion and total production were highly correlated. The resulting differences in production modes and levels clearly demonstrate the significant impact that selection approaches can exert on final evolutionary outcomes in strain engineering. Moreover, we note that these results provide a cautionary tale when intracellular read-outs of product concentration (including signals from biosensors) are used as surrogates for total production of potentially secreted products. In this regard, these results demonstrate that extracellular production is best assayed through an encapsulation technique when performing high throughput screening. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Gene repression via multiplex gRNA strategy in Y. lipolytica.

PubMed

Zhang, Jin-Lai; Peng, Yang-Zi; Liu, Duo; Liu, Hong; Cao, Ying-Xiu; Li, Bing-Zhi; Li, Chun; Yuan, Ying-Jin

2018-04-20

The oleaginous yeast Yarrowia lipolytica is a promising microbial cell factory due to their biochemical characteristics and native capacity to accumulate lipid-based chemicals. To create heterogenous biosynthesis pathway and manipulate metabolic flux in Y. lipolytica, numerous studies have been done for developing synthetic biology tools for gene regulation. CRISPR interference (CRISPRi), as an emerging technology, has been applied for specifically repressing genes of interest. In this study, we established CRISPRi systems in Y. lipolytica based on four different repressors, that was DNase-deactivated Cpf1 (dCpf1) from Francisella novicida, deactivated Cas9 (dCas9) from Streptococcus pyogenes, and two fusion proteins (dCpf1-KRAB and dCas9-KRAB). Ten gRNAs that bound to different regions of gfp gene were designed and the results indicated that there was no clear correlation between the repression efficiency and targeting sites no matter which repressor protein was used. In order to rapidly yield strong gene repression, a multiplex gRNAs strategy based on one-step Golden-brick assembly technology was developed. High repression efficiency 85% (dCpf1) and 92% (dCas9) were achieved in a short time by making three different gRNAs towards gfp gene simultaneously, which avoided the need of screening effective gRNA loci in advance. Moreover, two genes interference including gfp and vioE and three genes repression including vioA, vioB and vioE in protodeoxy-violaceinic acid pathway were also realized. Taken together, successful CRISPRi-mediated regulation of gene expression via four different repressors dCpf1, dCas9, dCpf1-KRAB and dCas9-KRAB in Y. lipolytica is achieved. And we demonstrate a multiplexed gRNA targeting strategy can efficiently achieve transcriptional simultaneous repression of several targeted genes and different sites of one gene using the one-step Golden-brick assembly. This timesaving method promised to be a potent transformative tool valuable for

Accumulation and metabolism of selenium by yeast cells.

PubMed

Kieliszek, Marek; Błażejak, Stanisław; Gientka, Iwona; Bzducha-Wróbel, Anna

2015-07-01

This paper examines the process of selenium bioaccumulation and selenium metabolism in yeast cells. Yeast cells can bind elements in ionic from the environment and permanently integrate them into their cellular structure. Up to now, Saccharomyces cerevisiae, Candida utilis, and Yarrowia lipolytica yeasts have been used primarily in biotechnological studies to evaluate binding of minerals. Yeast cells are able to bind selenium in the form of both organic and inorganic compounds. The process of bioaccumulation of selenium by microorganisms occurs through two mechanisms: extracellular binding by ligands of membrane assembly and intracellular accumulation associated with the transport of ions across the cytoplasmic membrane into the cell interior. During intracellular metabolism of selenium, oxidation, reduction, methylation, and selenoprotein synthesis processes are involved, as exemplified by detoxification processes that allow yeasts to survive under culture conditions involving the elevated selenium concentrations which were observed. Selenium yeasts represent probably the best absorbed form of this element. In turn, in terms of wide application, the inclusion of yeast with accumulated selenium may aid in lessening selenium deficiency in a diet.

Metabolic engineering of yeast for lignocellulosic biofuel production.

PubMed

Jin, Yong-Su; Cate, Jamie Hd

2017-12-01

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

Characterization of the two intracellular lipases of Y. lipolytica encoded by TGL3 and TGL4 genes: new insights into the role of intracellular lipases and lipid body organisation.

PubMed

Dulermo, Thierry; Tréton, Brigitte; Beopoulos, Athanasios; Kabran Gnankon, Affoué Philomène; Haddouche, Ramdane; Nicaud, Jean-Marc

2013-09-01

Eukaryotes store lipids in a specialised organelle, the lipid body (LB), mainly as triglycerides (TAGs). Both the rates of synthesis and degradation contribute to the control of the accumulation of TAGs. The synthesis of TAGs in yeasts has been well documented, especially in the model yeast Saccharomyces cerevisiae and in the oleaginous yeast Yarrowia lipolytica. However, descriptions of the processes involved in TAG degradation are more scarce and mostly for S. cerevisiae. Here, we report the characterisation of two Y. lipolytica genes, YlTGL3 and YlTGL4, encoding intracellular lipases involved in TAG degradation. The two proteins are localised in lipid bodies, and YlTgl4 was mainly found at the interface between LBs. Surprisingly, the spatial organisation of YlTgl3 and YlTgl4 depends on the culture medium and on the physiological phase of the cell. Inactivation of one or both genes doubles the lipid accumulation capacity of Y. lipolytica, increasing the cell's capacity to accumulate TAGs. The amino acid sequence of YlTgl4 contains the consensus sequence motif (G/A)XSXG, typical of serine hydrolases, whereas YlTgl3 does not. Single and double mutants are unable to degrade TAGs, and higher expression of YlTgl4 correlates with TAG degradation. Therefore, we propose that YlTgl4 is the main lipase responsible for TAG degradation and that YlTgl3 may act as a positive regulator of YlTgl4 rather than a functional lipase. Thus, contrary to S. cerevisiae, Y. lipolytica possesses two intracellular lipases with distinct roles and with distinct localisations in the LB. © 2013. Published by Elsevier B.V. All rights reserved.

Harnessing the Effect of pH on Lipid Production in Batch Cultures of Yarrowia lipolytica SKY7.

PubMed

Kuttiraja, Mathiazhakan; Dhouha, Ayed; Tyagi, Rajeshwar Dayal

2018-04-01

The objective of this research was to investigate the kinetics of lipid production by Yarrowia lipolytica SKY7 in the crude glycerol-supplemented media with and without the control of pH. Lipid and citric acid production were improved with the pH control condition. There was no significant difference observed in the biomass concentration with or without the pH control. In the pH-controlled experiments, the biomass and lipid concentration reached 18 and 7.78 g/L, (45.5% w/w), respectively, with lipid yield (Yp/s) of 0.179 g/g at 60 h of fermentation. The lipid production was directly correlated with growth and the process was defined as growth associated. After 60 h of fermentation, the lipid degradation was noticed in the pH-controlled reactor whereas it occurred after 84 h in the pH-uncontrolled reactor. Apart from lipid, citric acid was produced as the major extracellular product in both fermentations but the much lower concentration in uncontrolled pH. Based on the experimental results, it is evident that controlling the pH will enhance the lipid production by 15% compared to pH-uncontrolled fermentation.

Presence and changes in populations of yeasts on raw and processed poultry products stored at refrigeration temperature.

PubMed

Ismail, S A; Deak, T; El-Rahman, H A; Yassien, M A; Beuchat, L R

2000-12-05

A study was undertaken to determine populations and profiles of yeast species on fresh and processed poultry products upon purchase from retail supermarkets and after storage at 5 degrees C until shelf life expiration, and to assess the potential role of these yeasts in product spoilage. Fifty samples representing 15 commercial raw, marinated, smoked, or roasted chicken and turkey products were analyzed. Yeast populations were determined by plating on dichloran rose bengal chloramphenicol (DRBC) agar and tryptone glucose yeast extract (TGY) agar. Proteolytic activity was determined using caseinate and gelatin agars and lipolytic activity was determined on plate count agar supplemented with tributyrin. Populations of aerobic microorganisms were also determined. Initial populations of yeasts (log10 cfu/g) ranged from less than 1 (detection limit) to 2.89, and increased by the expiration date to 0.37-5.06, indicating the presence of psychrotrophic species. Highest initial populations were detected in raw chicken breast, wings, and ground chicken, as well as in turkey necks and legs, whereas roasted chicken and turkey products contained less than 1 log10 cfu/g. During storage, yeast populations increased significantly (P < or = 0.05) in whole chicken, ground chicken, liver, heart and gizzard, and in ground turkey and turkey sausage. Isolates (152 strains) of yeasts from poultry products consisted of 12 species. Yarrowia lipolytica and Candida zeylanoides were predominant, making up 39 and 26% of the isolates, respectively. Six different species of basidiomycetous yeasts representing 24% of the isolates were identified. Most Y. lipolytica strains showed strong proteolytic and lipolytic activities, whereas C. zeylanoides was weakly lipolytic. Results suggest that yeasts, particularly Y. lipolytica, may play a more prominent role than previously recognized in the spoilage of fresh and processed poultry stored at 5 degrees C.

Yeast synthetic biology for the production of recombinant therapeutic proteins.

PubMed

Kim, Hyunah; Yoo, Su Jin; Kang, Hyun Ah

2015-02-01

The production of recombinant therapeutic proteins is one of the fast-growing areas of molecular medicine and currently plays an important role in treatment of several diseases. Yeasts are unicellular eukaryotic microbial host cells that offer unique advantages in producing biopharmaceutical proteins. Yeasts are capable of robust growth on simple media, readily accommodate genetic modifications, and incorporate typical eukaryotic post-translational modifications. Saccharomyces cerevisiae is a traditional baker's yeast that has been used as a major host for the production of biopharmaceuticals; however, several nonconventional yeast species including Hansenula polymorpha, Pichia pastoris, and Yarrowia lipolytica have gained increasing attention as alternative hosts for the industrial production of recombinant proteins. In this review, we address the established and emerging genetic tools and host strains suitable for recombinant protein production in various yeast expression systems, particularly focusing on current efforts toward synthetic biology approaches in developing yeast cell factories for the production of therapeutic recombinant proteins. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

Irradiation of Yarrowia lipolytica NRRL YB-567 creating novel strains with enhanced ammonia and oil production on protein and carbohydrate substrates.

PubMed

Lindquist, Mitch R; López-Núñez, Juan Carlos; Jones, Marjorie A; Cox, Elby J; Pinkelman, Rebecca J; Bang, Sookie S; Moser, Bryan R; Jackson, Michael A; Iten, Loren B; Kurtzman, Cletus P; Bischoff, Kenneth M; Liu, Siqing; Qureshi, Nasib; Tasaki, Kenneth; Rich, Joseph O; Cotta, Michael A; Saha, Badal C; Hughes, Stephen R

2015-11-01

Increased interest in sustainable production of renewable diesel and other valuable bioproducts is redoubling efforts to improve economic feasibility of microbial-based oil production. Yarrowia lipolytica is capable of employing a wide variety of substrates to produce oil and valuable co-products. We irradiated Y. lipolytica NRRL YB-567 with UV-C to enhance ammonia (for fertilizer) and lipid (for biodiesel) production on low-cost protein and carbohydrate substrates. The resulting strains were screened for ammonia and oil production using color intensity of indicators on plate assays. Seven mutant strains were selected (based on ammonia assay) and further evaluated for growth rate, ammonia and oil production, soluble protein content, and morphology when grown on liver infusion medium (without sugars), and for growth on various substrates. Strains were identified among these mutants that had a faster doubling time, produced higher maximum ammonia levels (enzyme assay) and more oil (Sudan Black assay), and had higher maximum soluble protein levels (Bradford assay) than wild type. When grown on plates with substrates of interest, all mutant strains showed similar results aerobically to wild-type strain. The mutant strain with the highest oil production and the fastest doubling time was evaluated on coffee waste medium. On this medium, the strain produced 0.12 g/L ammonia and 0.20 g/L 2-phenylethanol, a valuable fragrance/flavoring, in addition to acylglycerols (oil) containing predominantly C16 and C18 residues. These mutant strains will be investigated further for potential application in commercial biodiesel production.

Production of Eight Different Hydride Complexes and Nitrite Release from 2,4,6-Trinitrotoluene by Yarrowia lipolytica▿ †

PubMed Central

Ziganshin, Ayrat M.; Gerlach, Robin; Borch, Thomas; Naumov, Anatoly V.; Naumova, Rimma P.

2007-01-01

2,4,6-Trinitrotoluene (TNT) transformation by the yeast strain Yarrowia lipolytica AN-L15 was shown to occur via two different pathways. Direct aromatic ring reduction was the predominant mechanism of TNT transformation, while nitro group reduction was observed to be a minor pathway. Although growth of Y. lipolytica AN-L15 was inhibited initially in the presence of TNT, TNT transformation was observed, indicating that the enzymes necessary for TNT reduction were present initially. Aromatic ring reduction resulted in the transient accumulation of eight different TNT-hydride complexes, which were characterized using high-performance liquid chromatography, UV-visible diode array detection, and negative-mode atmospheric pressure chemical ionization mass spectrometry (APCI-MS). APCI-MS analysis revealed three different groups of TNT-hydride complexes with molecular ions at m/z 227, 228, and 230, which correspond to TNT-mono- and dihydride complexes and protonated dihydride isomers, respectively. One of the three protonated dihydride complex isomers detected appears to release nitrite in the presence of strain AN-L15. This release of nitrite is of particular interest since it can provide a pathway towards complete degradation and detoxification of TNT. PMID:17933928

High-oleate yeast oil without polyunsaturated fatty acids.

PubMed

Tsakraklides, Vasiliki; Kamineni, Annapurna; Consiglio, Andrew L; MacEwen, Kyle; Friedlander, Jonathan; Blitzblau, Hannah G; Hamilton, Maureen A; Crabtree, Donald V; Su, Austin; Afshar, Jonathan; Sullivan, John E; LaTouf, W Greg; South, Colin R; Greenhagen, Emily H; Shaw, A Joe; Brevnova, Elena E

2018-01-01

Oleate-enriched triacylglycerides are well-suited for lubricant applications that require high oxidative stability. Fatty acid carbon chain length and degree of desaturation are key determinants of triacylglyceride properties and the ability to manipulate fatty acid composition in living organisms is critical to developing a source of bio-based oil tailored to meet specific application requirements. We sought to engineer the oleaginous yeast Yarrowia lipolytica for production of high-oleate triacylglyceride oil. We studied the effect of deletions and overexpressions in the fatty acid and triacylglyceride synthesis pathways to identify modifications that increase oleate levels. Oleic acid accumulation in triacylglycerides was promoted by exchanging the native ∆9 fatty acid desaturase and glycerol-3-phosphate acyltransferase with heterologous enzymes, as well as deletion of the Δ12 fatty acid desaturase and expression of a fatty acid elongase. By combining these engineering steps, we eliminated polyunsaturated fatty acids and created a Y. lipolytica strain that accumulates triglycerides with > 90% oleate content. High-oleate content and lack of polyunsaturates distinguish this triacylglyceride oil from plant and algal derived oils. Its composition renders the oil suitable for applications that require high oxidative stability and further demonstrates the potential of Y. lipolytica as a producer of tailored lipid profiles.

Interactions between yeasts and bacteria in the smear surface-ripened cheeses.

PubMed

Corsetti, A; Rossi, J; Gobbetti, M

2001-09-19

In the initial phase of ripening, the microflora of bacterial smear surface-ripened cheeses such as Limburger, Taleggio, Brick, Münster and Saint-Paulin and that of surface mould-ripened cheeses such as Camembert and Brie may be similar, but at the end of the ripening, bacteria such as Brevibacterium spp., Arthrobacter spp., Micrococcus spp., Corynebacterium spp. and moulds such as Penicillium camemberti are, respectively, the dominant microorganisms. Yeasts such as Candida spp., Cryptococcus spp., Debaryomyces spp., Geotrichum candidum, Pichia spp., Rhodotorula spp., Saccharomyces spp. and Yarrowia lipolytica are often and variably isolated from the smear surface-ripened cheeses. Although not dominant within the microorganisms of the smear surface-ripened cheeses, yeasts establish significant interactions with moulds and especially bacteria, including surface bacteria and lactic acid bacteria. Some aspects of the interactions between yeasts and bacteria in such type of cheeses are considered in this paper.

Green and sustainable succinic acid production from crude glycerol by engineered Yarrowia lipolytica via agricultural residue based in situ fibrous bed bioreactor.

PubMed

Li, Chong; Gao, Shi; Yang, Xiaofeng; Lin, Carol Sze Ki

2018-02-01

In situ fibrous bed bioreactor (isFBB) for efficient succinic acid (SA) production by Yarrowia lipolytica was firstly developed in our former study. In this study, agricultural residues including wheat straw, corn stalk and sugarcane bagasse were investigated for the improvement of isFBB, and sugarcane bagasse was demonstrated to be the best immobilization material. With crude glycerol as the sole carbon source, optimization for isFBB batch fermentation was carried out. Under the optimal conditions of 20g sugarcane bagasse as immobilization material, 120gL -1 crude glycerol as carbon source and 4Lmin -1 of aeration rate, the resultant SA concentration was 53.6gL -1 with an average productivity of 1.45gL -1 h -1 and a SA yield of 0.45gg -1 . By feeding crude glycerol, SA titer up to 209.7gL -1 was obtained from fed batch fermentation, which was the highest value that ever reported. Copyright © 2017 Elsevier Ltd. All rights reserved.

Improved performance of Yarrowia lipolytica lipase-catalyzed kinetic resolution of (R,S)-2-octanol by an integrated strategy of interfacial activation, bioimprinting and immobilization.

PubMed

Liu, Ying; Guo, Chen; Sun, Xi-Tong; Liu, Chun-Zhao

2013-08-01

Yarrowia lipolytica lipase (YLL) demonstrated an (R)-enantiopreference for efficient resolution of (R,S)-2-octanol. The activity, enantioselectivity, the ratio of substrate to enzyme, acetaldehyde tolerance, and operational stability of YLL were improved by an integrated strategy of interfacial activation, bioimprinting, and immobilization. In comparison with the control, both the enzymatic activity and enantioselectivity increased by a factor of 8.85 and 2.75 by the integrated strategy, respectively. Fifty-one percentage of conversion with 220 of enantioselectivity was obtained using the immobilized YLL prepared by the integrated strategy at a ratio of 104 of substrate to enzyme loaded. The immobilized YLL retained 97% of its initial activity without a decrease in enantioselectivity after 10 successive reuse cycles. Together these results will result in a promising strategy with the YYL for efficient resolution of (R,S)-2-octanol in practice. Copyright © 2013 Elsevier Ltd. All rights reserved.

Synthetic biology and molecular genetics in non-conventional yeasts: Current tools and future advances.

PubMed

Wagner, James M; Alper, Hal S

2016-04-01

Coupling the tools of synthetic biology with traditional molecular genetic techniques can enable the rapid prototyping and optimization of yeast strains. While the era of yeast synthetic biology began in the well-characterized model organism Saccharomyces cerevisiae, it is swiftly expanding to include non-conventional yeast production systems such as Hansenula polymorpha, Kluyveromyces lactis, Pichia pastoris, and Yarrowia lipolytica. These yeasts already have roles in the manufacture of vaccines, therapeutic proteins, food additives, and biorenewable chemicals, but recent synthetic biology advances have the potential to greatly expand and diversify their impact on biotechnology. In this review, we summarize the development of synthetic biological tools (including promoters and terminators) and enabling molecular genetics approaches that have been applied in these four promising alternative biomanufacturing platforms. An emphasis is placed on synthetic parts and genome editing tools. Finally, we discuss examples of synthetic tools developed in other organisms that can be adapted or optimized for these hosts in the near future. Copyright © 2015 Elsevier Inc. All rights reserved.

Enhancement of methanol resistance of Yarrowia lipolytica lipase 2 using β-cyclodextrin as an additive: Insights from experiments and molecular dynamics simulation.

PubMed

Cao, Hao; Jiang, Yang; Zhang, Haiyang; Nie, Kaili; Lei, Ming; Deng, Li; Wang, Fang; Tan, Tianwei

2017-01-01

The methanol resistance of lipase is a critical parameter in enzymatic biodiesel production. In the present work, the methanol resistance of Yarrowia lipolytica Lipase 2 (YLLIP2) was significantly improved using β-cyclodextrin (β-CD) as an additive. According to the results, YLLIP2 with β-CD exhibited approximately 7000U/mg specific activity in 30wt% methanol for 60min compared with no activity without β-CD under the same conditions. Molecular dynamics (MD) simulation results indicated that the β-CD molecules weakened the conformational change of YLLIP2 and maintained a semi-open state of the lid by overcoming the interference caused by methanol molecules. Furthermore, the β-CD molecule could directly stabilize "pathway" regions (e.g., Asp61-Asp67) and indirectly stabilize "pathway" regions (e.g., Gly44-Phe50) by forming hydrogen bonds with "pathway" regions and nearby "pathway" regions, respectively. The regions stabilized by the β-CD molecule then prevented the closure of active pockets, thus retaining the enzymatic activity of YLLIP2 with β-CD in methanol solvent. Copyright © 2016. Published by Elsevier Inc.

The Gene YALI0E20207g from Yarrowia lipolytica Encodes an N-Acetylglucosamine Kinase Implicated in the Regulated Expression of the Genes from the N-Acetylglucosamine Assimilatory Pathway

PubMed Central

Flores, Carmen-Lisset; Gancedo, Carlos

2015-01-01

The non-conventional yeast Yarrowia lipolytica possesses an ORF, YALI0E20207g, which encodes a protein with an amino acid sequence similar to hexokinases from different organisms. We have cloned that gene and determined several enzymatic properties of its encoded protein showing that it is an N-acetylglucosamine (NAGA) kinase. This conclusion was supported by the lack of growth in NAGA of a strain carrying a YALI0E20207g deletion. We named this gene YlNAG5. Expression of YlNAG5 as well as that of the genes encoding the enzymes of the NAGA catabolic pathway—identified by a BLAST search—was induced by this sugar. Deletion of YlNAG5 rendered that expression independent of the presence of NAGA in the medium and reintroduction of the gene restored the inducibility, indicating that YlNag5 participates in the transcriptional regulation of the NAGA assimilatory pathway genes. Expression of YlNAG5 was increased during sporulation and homozygous Ylnag5/Ylnag5 diploid strains sporulated very poorly as compared with a wild type isogenic control strain pointing to a participation of the protein in the process. Overexpression of YlNAG5 allowed growth in glucose of an Ylhxk1glk1 double mutant and produced, in a wild type background, aberrant morphologies in different media. Expression of the gene in a Saccharomyces cerevisiae hxk1 hxk2 glk1 triple mutant restored ability to grow in glucose. PMID:25816199

The secretory pathway: exploring yeast diversity.

PubMed

Delic, Marizela; Valli, Minoska; Graf, Alexandra B; Pfeffer, Martin; Mattanovich, Diethard; Gasser, Brigitte

2013-11-01

Protein secretion is an essential process for living organisms. In eukaryotes, this encompasses numerous steps mediated by several hundred cellular proteins. The core functions of translocation through the endoplasmic reticulum membrane, primary glycosylation, folding and quality control, and vesicle-mediated secretion are similar from yeasts to higher eukaryotes. However, recent research has revealed significant functional differences between yeasts and mammalian cells, and even among diverse yeast species. This review provides a current overview of the canonical protein secretion pathway in the model yeast Saccharomyces cerevisiae, highlighting differences to mammalian cells as well as currently unresolved questions, and provides a genomic comparison of the S. cerevisiae pathway to seven other yeast species where secretion has been investigated due to their attraction as protein production platforms, or for their relevance as pathogens. The analysis of Candida albicans, Candida glabrata, Kluyveromyces lactis, Pichia pastoris, Hansenula polymorpha, Yarrowia lipolytica, and Schizosaccharomyces pombe reveals that many - but not all - secretion steps are more redundant in S. cerevisiae due to duplicated genes, while some processes are even absent in this model yeast. Recent research obviates that even where homologous genes are present, small differences in protein sequence and/or differences in the regulation of gene expression may lead to quite different protein secretion phenotypes. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

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

PubMed

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

2017-09-01

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

Chemical signaling and insect attraction is a conserved trait in yeasts.

PubMed

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

Comparative Lipidomic Profiling of S. cerevisiae and Four Other Hemiascomycetous Yeasts

PubMed Central

Hein, Eva-Maria; Hayen, Heiko

2012-01-01

Glycerophospholipids (GP) are the building blocks of cellular membranes and play essential roles in cell compartmentation, membrane fluidity or apoptosis. In addition, GPs are sources for multifunctional second messengers. Whereas the genome and proteome of the most intensively studied eukaryotic model organism, the baker’s yeast (Saccharomyces cerevisiae), are well characterized, the analysis of its lipid composition is still at the beginning. Moreover, different yeast species can be distinguished on the DNA, RNA and protein level, but it is currently unknown if they can also be differentiated by determination of their GP pattern. Therefore, the GP compositions of five different yeast strains, grown under identical environmental conditions, were elucidated using high performance liquid chromatography coupled to negative electrospray ionization-hybrid linear ion trap-Fourier transform ion cyclotron resonance mass spectrometry in single and multistage mode. Using this approach, relative quantification of more than 100 molecular species belonging to nine GP classes was achieved. The comparative lipidomic profiling of Saccharomyces cerevisiae, Saccharomyces bayanus, Kluyveromyces thermotolerans, Pichia angusta, and Yarrowia lipolytica revealed characteristic GP profiles for each strain. However, genetically related yeast strains show similarities in their GP compositions, e.g., Saccharomyces cerevisiae and Saccharomyces bayanus. PMID:24957378

Comparison of nitrogen depletion and repletion on lipid production in yeast and fungal species

DOE PAGES

Yang, Shihui; Wang, Wei; Wei, Hui; ...

2016-08-29

Although it is well known that low nitrogen stimulates lipid accumulation, especially for algae and some oleaginous yeast, few studies have been conducted in fungal species, especially on the impact of different nitrogen deficiency strategies. In this study, we use two promising consolidated bioprocessing (CBP) candidates to examine the impact of two nitrogen deficiency strategies on lipid production, which are the extensively investigated oleaginous yeast Yarrowia lipolytica, and the commercial cellulase producer Trichoderma reesei. We first utilized bioinformatics approaches to reconstruct the fatty acid metabolic pathway and demonstrated the presence of a triacylglycerol (TAG) biosynthesis pathway in Trichoderma reesei. Wemore » then examined the lipid production of Trichoderma reesei and Y. lipomyces in different media using two nitrogen deficiency strategies of nitrogen natural repletion and nitrogen depletion through centrifugation. Our results demonstrated that nitrogen depletion was better than nitrogen repletion with about 30% lipid increase for Trichoderma reesei and Y. lipomyces, and could be an option to improve lipid production in both oleaginous yeast and filamentous fungal species. The resulting distinctive lipid composition profiles indicated that the impacts of nitrogen depletion on yeast were different from those for fungal species. Under three types of C/N ratio conditions, C16 and C18 fatty acids were the predominant forms of lipids for both Trichoderma reesei and Y. lipolytica. In addition, while the overall fatty acid methyl ester (FAME) profiles of Trichoderma reesei were similar, the overall FAME profiles of Y. lipolytica observed a shift. The fatty acid metabolic pathway reconstructed in this work supports previous reports of lipid production in T. reesei, and provides a pathway for future omics studies and metabolic engineering efforts. Further investigation to identify the genetic targets responsible for the effect of nitrogen depletion

Comparison of nitrogen depletion and repletion on lipid production in yeast and fungal species

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Shihui; Wang, Wei; Wei, Hui

Although it is well known that low nitrogen stimulates lipid accumulation, especially for algae and some oleaginous yeast, few studies have been conducted in fungal species, especially on the impact of different nitrogen deficiency strategies. In this study, we use two promising consolidated bioprocessing (CBP) candidates to examine the impact of two nitrogen deficiency strategies on lipid production, which are the extensively investigated oleaginous yeast Yarrowia lipolytica, and the commercial cellulase producer Trichoderma reesei. We first utilized bioinformatics approaches to reconstruct the fatty acid metabolic pathway and demonstrated the presence of a triacylglycerol (TAG) biosynthesis pathway in Trichoderma reesei. Wemore » then examined the lipid production of Trichoderma reesei and Y. lipomyces in different media using two nitrogen deficiency strategies of nitrogen natural repletion and nitrogen depletion through centrifugation. Our results demonstrated that nitrogen depletion was better than nitrogen repletion with about 30% lipid increase for Trichoderma reesei and Y. lipomyces, and could be an option to improve lipid production in both oleaginous yeast and filamentous fungal species. The resulting distinctive lipid composition profiles indicated that the impacts of nitrogen depletion on yeast were different from those for fungal species. Under three types of C/N ratio conditions, C16 and C18 fatty acids were the predominant forms of lipids for both Trichoderma reesei and Y. lipolytica. In addition, while the overall fatty acid methyl ester (FAME) profiles of Trichoderma reesei were similar, the overall FAME profiles of Y. lipolytica observed a shift. The fatty acid metabolic pathway reconstructed in this work supports previous reports of lipid production in T. reesei, and provides a pathway for future omics studies and metabolic engineering efforts. Further investigation to identify the genetic targets responsible for the effect of nitrogen depletion

CE separation of proteins and yeasts dynamically modified by PEG pyrenebutanoate with fluorescence detection.

PubMed

Horká, Marie; Růzicka, Filip; Holá, Veronika; Slais, Karel

2007-07-01

The optimized protocols of the bioanalytes separation, proteins and yeasts, dynamically modified by the nonionogenic tenside PEG pyrenebutanoate, were applied in CZE and CIEF with the acidic gradient in pH range 2-5.5, both with fluorescence detection. PEG pyrenebutanoate was used as a buffer additive for a dynamic modification of proteins and/or yeast samples. The narrow peaks of modified analytes were detected. The values of the pI's of the labeled proteins were calculated using new fluorescent pI markers in CIEF and they were found to be comparable with pI's of the native compounds. As an example of the possible use of the suggested CIEF technique, the mixed cultures of yeasts, Candida albicans, Candida glabrata, Candida kefyr, Candida krusei, Candida lusitaniae, Candida parapsilosis, Candida tropicalis, Candida zeylanoides, Geotrichum candidum, Saccharomyces cerevisiae, Trichosporon asahii and Yarrowia lipolytica, were reproducibly focused and separated with high sensitivity. Using UV excitation for the on-column fluorometric detection, the minimum detectable amounts of analytes, femtograms of proteins and down to ten cells injected on the separation capillary, were estimated.

Origins of cell-to-cell bioprocessing diversity and implications of the extracellular environment revealed at the single-cell level

DOE PAGES

Vasdekis, A. E.; Silverman, A. M.; Stephanopoulos, G.

2015-12-14

We probed the lipid expression dynamics of the oleaginous yeast Yarrowia Lipolytica. We observed that neutral lipid expression is sporadic. By performing single-cell analysis, we found that such noise emanates from the metabolic reaction level. Our results provide an alternative insight into the regulation and phenotypic variability of lipogenesis.

Oleaginous yeasts: Promising platforms for the production of oleochemicals and biofuels.

PubMed

Adrio, José L

2017-09-01

Oleaginous yeasts have a unique physiology that makes them the best suited hosts for the production of lipids, oleochemicals, and diesel-like fuels. Their high lipogenesis, capability of growing on many different carbon sources (including lignocellulosic sugars), easy large-scale cultivation, and an increasing number of genetic tools are some of the advantages that have encouraged their use to develop sustainable processes. This mini-review summarizes the metabolic engineering strategies developed in oleaginous yeasts within the last 2 years to improve process metrics (titer, yield, and productivity) for the production of lipids, free fatty acids, fatty acid-based chemicals (e.g., fatty alcohols, fatty acid ethyl esters), and alkanes. During this short period of time, tremendous progress has been made in Yarrowia lipolytica, the model oleaginous yeast, which has been engineered to improve lipid production by different strategies including increasing lipogenic pathway flux and biosynthetic precursors, and blocking degradation pathways. Moreover, remarkable advances have also been reported in Rhodosporidium toruloides and Lipomyces starkey despite the limited genetic tools available for these two very promising hosts. Biotechnol. Bioeng. 2017;114: 1915-1920. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Comparison of Yeasts as Hosts for Recombinant Protein Production.

PubMed

Vieira Gomes, Antonio Milton; Souza Carmo, Talita; Silva Carvalho, Lucas; Mendonça Bahia, Frederico; Parachin, Nádia Skorupa

2018-04-29

Recombinant protein production emerged in the early 1980s with the development of genetic engineering tools, which represented a compelling alternative to protein extraction from natural sources. Over the years, a high level of heterologous protein was made possible in a variety of hosts ranging from the bacteria Escherichia coli to mammalian cells. Recombinant protein importance is represented by its market size, which reached $1654 million in 2016 and is expected to reach $2850.5 million by 2022. Among the available hosts, yeasts have been used for producing a great variety of proteins applied to chemicals, fuels, food, and pharmaceuticals, being one of the most used hosts for recombinant production nowadays. Historically, Saccharomyces cerevisiae was the dominant yeast host for heterologous protein production. Lately, other yeasts such as Komagataella sp., Kluyveromyces lactis , and Yarrowia lipolytica have emerged as advantageous hosts. In this review, a comparative analysis is done listing the advantages and disadvantages of using each host regarding the availability of genetic tools, strategies for cultivation in bioreactors, and the main techniques utilized for protein purification. Finally, examples of each host will be discussed regarding the total amount of protein recovered and its bioactivity due to correct folding and glycosylation patterns.

Physiological uncoupling of mitochondrial oxidative phosphorylation. Studies in different yeast species.

PubMed

Guerrero-Castillo, Sergio; Araiza-Olivera, Daniela; Cabrera-Orefice, Alfredo; Espinasa-Jaramillo, Juan; Gutiérrez-Aguilar, Manuel; Luévano-Martínez, Luís A; Zepeda-Bastida, Armando; Uribe-Carvajal, Salvador

2011-06-01

Under non-phosphorylating conditions a high proton transmembrane gradient inhibits the rate of oxygen consumption mediated by the mitochondrial respiratory chain (state IV). Slow electron transit leads to production of reactive oxygen species (ROS) capable of participating in deleterious side reactions. In order to avoid overproducing ROS, mitochondria maintain a high rate of O(2) consumption by activating different exquisitely controlled uncoupling pathways. Different yeast species possess one or more uncoupling systems that work through one of two possible mechanisms: i) Proton sinks and ii) Non-pumping redox enzymes. Proton sinks are exemplified by mitochondrial unspecific channels (MUC) and by uncoupling proteins (UCP). Saccharomyces. cerevisiae and Debaryomyces hansenii express highly regulated MUCs. Also, a UCP was described in Yarrowia lipolytica which promotes uncoupled O(2) consumption. Non-pumping alternative oxido-reductases may substitute for a pump, as in S. cerevisiae or may coexist with a complete set of pumps as in the branched respiratory chains from Y. lipolytica or D. hansenii. In addition, pumps may suffer intrinsic uncoupling (slipping). Promising models for study are unicellular parasites which can turn off their aerobic metabolism completely. The variety of energy dissipating systems in eukaryote species is probably designed to control ROS production in the different environments where each species lives.

Comparison of volatile sulphur compound production by cheese-ripening yeasts from methionine and methionine-cysteine mixtures.

PubMed

López Del Castillo-Lozano, M; Delile, A; Spinnler, H E; Bonnarme, P; Landaud, S

2007-07-01

Production of volatile sulphur compounds (VSC) was assessed in culture media supplemented with L-methionine or L-methionine/L-cysteine mixtures, using five cheese-ripening yeasts: Debaryomyces hansenii DH47(8), Kluyveromyces lactis KL640, Geotrichum candidum GC77, Yarrowia lipolytica YL200 and Saccharomyces cerevisiae SC45(3). All five yeasts produced VSC with L-methionine or L-methionine/L-cysteine, but different VSC profiles were found. GC77 and YL200 produced dimethyldisulphide and trace levels of dimethyltrisulphide while DH47(8), KL640 and SC45(3) produced mainly methionol and low levels of methional. S-methylthioacetate was produced by all the yeasts but at different concentrations. DH47(8), KL640 and SC45(3) also produced other minor VSC including 3-methylthiopropyl acetate, ethyl-3-methylthiopropanoate, a thiophenone, and an oxathiane. However, VSC production diminished in a strain-dependent behaviour when L-cysteine was supplemented, even at a low concentration (0.2 g l(-1)). This effect was due mainly to a significant decrease in L-methionine consumption in all the yeasts except YL200. Hydrogen sulphide produced by L-cysteine catabolism did not seem to contribute to VSC generation at the acid pH of yeast cultures. The significance of such results in the cheese-ripening context is discussed.

Changes in volatile profile of soybean residue (okara) upon solid-state fermentation by yeasts.

PubMed

Vong, Weng Chan; Liu, Shao-Quan

2017-01-01

Soybean residue (okara), a by-product of soymilk, is produced in large volumes by the soy food industry and is often discarded due to its undesirable flavour. As it contains a considerable amount of protein and fats, biotransformation of okara to improve its flavour presents an opportunity for alternative utilisation. This paper evaluated 10 yeasts in the solid-state fermentation of okara based on their volatile profiles as analysed with HS-SPME GC-MS/FID. Four 'dairy yeasts' (Geotrichum candidum, Yarrowia lipolytica, Debaryomyces hansenii and Kluyveromyces lactis) and six 'wine yeasts' (Saccharomyces cerevisiae, Lachancea thermotolerans, Metschnikowia pulcherrima, Pichia kluyveri, Torulaspora delbrueckii, and Williopsis saturnus) were studied. The main off-odourants in okara, hexanal and trans-2-hexenal, significantly decreased after fermentation due to their bioconversion into methyl ketones and/or esters. The okara fermented by dairy yeasts contained greater proportions of methyl ketones, while that by wine yeasts contained more ethyl and acetyl esters. Notably, the okara fermented by W. saturnus contained 13 esters and the total GC-FID peak area of esters was about 380 times that in fresh okara, leading to a perceptible fruity note. Okara can be exploited as an inexpensive substrate for bioflavour extraction and/or a more pleasant food ingredient via yeast fermentation. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Heterologous expression of lipases YLIP4, YLIP5, YLIP7, YLIP13, and YLIP15 from Yarrowia lipolytica MSR80 in Escherichia coli: Substrate specificity, kinetic comparison, and enantioselectivity.

PubMed

Syal, Poonam; Gupta, Rani

2017-11-01

Five lipase genes, ylip4, ylip5, ylip7, ylip13, and ylip15, from Yarrowia lipolytica MSR80 were cloned and expressed in the pEZZ18-HB101 system. The lipases shared maximum sequence identity with Candida galli lipase, whereas they shared structural similarity with YLIP2 of Y. lipolytica CLIB122. The enzymes, purified using IgG sepharose, had specific activities in the range of 7-25 U mg -1 . Biochemical characteristics of all the lipases varied with respect to thermostability, substrate specificity, and enantioselectivity. All the enzymes were most active at neutral or slightly alkaline pH and were stable in the pH range 3.0-8.0, except YLIP4, which showed 50% stability at pH 10.0. Temperature optima of all the lipases varied from 30 to 50 ºC. YLIP15 and YLIP13 were most thermostable with a t 1/2 of 138 and 112 Min, respectively, at 60 °C. The lipases exhibited varied substrate specificity on p-nitrophenyl esters ranging from short-chain specificity (YLIP15), mid-chain specificity (YLIP4, YLIP5, YLIP7), and long-chain specificity (YLIP13). Catalytic efficiency on p-nitrophenylcaprate was highest for YLIP13 (67 × 10 3 mM -1 min -1 ) and lowest for YLIP15 (6.7 × 10 3 mM -1 min -1 ). YLIP13 was S-enantioselective, and YLIP15 was R-enantioselective with enantiomeric excess of 53 and 36%, respectively. Of all five lipases, YLIP13 and YLIP15 could be considered as industrially important enzymes as they were thermostable and enantioselective. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

Fine Structure of Tibetan Kefir Grains and Their Yeast Distribution, Diversity, and Shift

PubMed Central

Lu, Man; Wang, Xingxing; Sun, Guowei; Qin, Bing; Xiao, Jinzhou; Yan, Shuling; Pan, Yingjie; Wang, Yongjie

2014-01-01

Tibetan kefir grains (TKGs), a kind of natural starter for fermented milk in Tibet, China, host various microorganisms of lactic acid bacteria, yeasts, and occasionally acetic acid bacteria in a polysaccharide/protein matrix. In the present study, the fine structure of TKGs was studied to shed light on this unusual symbiosis with stereomicroscopy and thin sections. The results reveal that TKGs consist of numerous small grain units, which are characterized by a hollow globular structure with a diameter between 2.0 and 9.0 mm and a wall thickness of approximately 200 µm. A polyhedron-like net structure, formed mainly by the bacteria, was observed in the wall of the grain units, which has not been reported previously to our knowledge. Towards the inside of the grain unit, the polyhedron-like net structures became gradually larger in diameter and fewer in number. Such fine structures may play a crucial role in the stability of the grains. Subsequently, the distribution, diversity, and shift of yeasts in TKGs were investigated based on thin section, scanning electron microscopy, cloning and sequencing of D1/D2 of the 26S rRNA gene, real-time quantitative PCR, and in situ hybridization with specific fluorescence-labeled oligonucleotide probes. These show that (i) yeasts appear to localize on the outer surface of the grains and grow normally together to form colonies embedded in the bacterial community; (ii) the diversity of yeasts is relatively low on genus level with three dominant species – Saccharomyces cerevisiae, Kluyveromyces marxianus, and Yarrowia lipolytica; (iii) S. cerevisiae is the stable predominant yeast species, while the composition of Kluyveromyces and Yarrowia are subject to change over time. Our results indicate that TKGs are relatively stable in structure, and culture conditions to some extent shape the microbial community and interaction in kefir grains. These findings pave the way for further study of the specific symbiotic associations between S

Detergent assisted lipid extraction from wet yeast biomass for biodiesel: A response surface methodology approach.

PubMed

Yellapu, Sravan Kumar; Bezawada, Jyothi; Kaur, Rajwinder; Kuttiraja, Mathiazhakan; Tyagi, Rajeshwar D

2016-10-01

The lipid extraction from the microbial biomass is a tedious and high cost dependent process. In the present study, detergent assisted lipids extraction from the culture of the yeast Yarrowia lipolytica SKY-7 was carried out. Response surface methodology (RSM) was used to investigate the effect of three principle parameters (N-LS concentration, time and temperature) on microbial lipid extraction efficiency % (w/w). The results obtained by statistical analysis showed that the quadratic model fits in all cases. Maximum lipid recovery of 95.3±0.3% w/w was obtained at the optimum level of process variables [N-LS concentration 24.42mg (equal to 48mgN-LS/g dry biomass), treatment time 8.8min and reaction temperature 30.2°C]. Whereas the conventional chloroform and methanol extraction to achieve total lipid recovery required 12h at 60°C. The study confirmed that oleaginous yeast biomass treatment with N-lauroyl sarcosine would be a promising approach for industrial scale microbial lipid recovery. Copyright © 2016 Elsevier Ltd. All rights reserved.

Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions.

PubMed

Wei, Yongjun; Siewers, Verena; Nielsen, Jens

2017-05-01

Cocoa butter (CB) extracted from cocoa beans is the main raw material for chocolate production. However, growing chocolate demands and limited CB production has resulted in a shortage of CB supply. CB is mainly composed of three different kinds of triacylglycerols (TAGs), POP (C16:0-C18:1-C16:0), POS (C16:0-C18:1-C18:0), and SOS (C18:0-C18:1-C18:0). The storage lipids of yeasts, mainly TAGs, also contain relative high-level of C16 and C18 fatty acids and might be used as CB-like lipids (CBL). In this study, we cultivated six different yeasts, including one non-oleaginous yeast strain, Saccharomyces cerevisiae CEN.PK113-7D, and five oleaginous yeast strains, Trichosporon oleaginosus DSM11815, Rhodotorula graminis DSM 27356, Lipomyces starkeyi DSM 70296, Rhodosporidium toruloides DSM 70398, and Yarrowia lipolytica CBS 6124, in nitrogen-limited medium and compared their CBL production ability. Under the same growth conditions, we found that TAGs were the main lipids in all six yeasts and that T. oleaginosus can produce more TAGs than the other five yeasts. Less than 3% of the total TAGs were identified as potential SOS in the six yeasts. However, T. oleaginosus produced 27.8% potential POP and POS at levels of 378 mg TAGs/g dry cell weight, hinting that this yeast may have potential as a CBL production host after further metabolic engineering in future.

Targeted mutations and MD simulations of a methanol-stable lipase YLIP9 from Yarrowia lipolytica MSR80 to develop a biodiesel enzyme.

PubMed

Syal, Poonam; Verma, Ved Vrat; Gupta, Rani

2017-11-01

Biodiesel, an environment friendly alternative for fuels, contains methyl esters of long-chain fatty acids. Our group has reported a methanol-stable YLIP9 from Yarrowia lipolytica MSR80 that shows poor catalysis of long-chain fatty acids. To shift its substrate specificity, residues within lid and binding pocket were identified for sequential mutations using YLIP2 as the template. Of the two point mutations (Glu116Leu and Ser119Val) introduced in the lid, the former mutation (YLIP9L1) increased the catalytic rate by ∼2-fold without any change in substrate specificity. In this mutant, six binding pocket residues (Bp2-Bp7) were further mutated to obtain six double mutants. YLIP9L1Bp3 showed significant shift in substrate specificity towards long-chain pNPesters with 11-fold increase in catalytic efficiency than YLIP9. Double mutations also led to increased thermostability and lowered activation energy of YLIP9L1Bp3 thereby shifting its optimum temperature from 60°C to 50°C. In silico molecular dynamics simulations revealed improved lid flexibility and increased catalytic triad volume in YLIP9L1Bp3. The enzyme YLIP9L1Bp3 was methanol-stable having selectivity for long-chain fatty acids with improved catalytic efficiency. Its application as a biodiesel enzyme was validated by transesterification of palm oil in presence of methanol, where it showed 8-fold increase in conversion of oil to methyl esters. Copyright © 2017 Elsevier B.V. All rights reserved.

The expression of glycerol facilitators from various yeast species improves growth on glycerol of Saccharomyces cerevisiae.

PubMed

Klein, Mathias; Islam, Zia-Ul; Knudsen, Peter Boldsen; Carrillo, Martina; Swinnen, Steve; Workman, Mhairi; Nevoigt, Elke

2016-12-01

Glycerol is an abundant by-product during biodiesel production and additionally has several assets compared to sugars when used as a carbon source for growing microorganisms in the context of biotechnological applications. However, most strains of the platform production organism Saccharomyces cerevisiae grow poorly in synthetic glycerol medium. It has been hypothesized that the uptake of glycerol could be a major bottleneck for the utilization of glycerol in S. cerevisiae . This species exclusively relies on an active transport system for glycerol uptake. This work demonstrates that the expression of predicted glycerol facilitators (Fps1 homologues) from superior glycerol-utilizing yeast species such as Pachysolen tannophilus , Komagataella pastoris , Yarrowia lipolytica and Cyberlindnera jadinii significantly improves the growth performance on glycerol of the previously selected glycerol-consuming S. cerevisiae wild-type strain (CBS 6412-13A). The maximum specific growth rate increased from 0.13 up to 0.18 h -1 and a biomass yield coefficient of 0.56 g DW /g glycerol was observed. These results pave the way for exploiting the assets of glycerol in the production of fuels, chemicals and pharmaceuticals based on baker's yeast.

An interfacial and comparative in vitro study of gastrointestinal lipases and Yarrowia lipolytica LIP2 lipase, a candidate for enzyme replacement therapy.

PubMed

Bénarouche, Anaïs; Point, Vanessa; Carrière, Frédéric; Cavalier, Jean-François

2014-07-01

Lipolytic activities of Yarrowia lipolytica LIP2 lipase (YLLIP2), human pancreatic (HPL) and dog gastric (DGL) lipases were first compared using lecithin-stabilized triacylglycerol (TAG) emulsions (Intralipid) at various pH and bile salt concentrations. Like DGL, YLLIP2 was able to hydrolyze TAG droplets covered by a lecithin monolayer, while HPL was not directly active on that substrate. These results were in good agreement with the respective kinetics of adsorption on phosphatidylcholine (PC) monomolecular films of the same three lipases, YLLIP2 being the most tensioactive lipase. YLLIP2 adsorption onto a PC monolayer spread at the air/water interface was influenced by pH-dependent changes in the enzyme/lipid interfacial association constant (KAds) which was optimum at pH 6.0 on long-chain egg PC monolayer, and at pH 5.0 on medium chain dilauroylphosphatidylcholine film. Using substrate monolayers (1,2-dicaprin, trioctanoin), YLLIP2 displayed the highest lipolytic activities on both substrates in the 25-35 mN m(-1) surface pressure range. YLLIP2 was active in a large pH range and displayed a pH-dependent activity profile combining DGL and HPL features at pH values found in the stomach (pH 3-5) and in the intestine (pH 6-7), respectively. The apparent maximum activity of YLLIP2 was observed at acidic pH 4-6 and was therefore well correlated with an efficient interfacial binding at these pH levels, whatever the type of interfaces (Intralipid emulsions, substrate or PC monolayers). All these findings support the use of YLLIP2 in enzyme replacement therapy for the treatment of pancreatic exocrine insufficiency, a pathological situation in which an acidification of intestinal contents occurs. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Improvement of soil characteristics and growth of Dorycnium pentaphyllum by amendment with agrowastes and inoculation with AM fungi and/or the yeast Yarowia lipolytica.

PubMed

Medina, A; Vassileva, M; Caravaca, F; Roldán, A; Azcón, R

2004-08-01

The effectiveness of two microbiologically treated agrowastes [dry olive cake (DOC) and/or sugar beet (SB)] on plant growth, soil enzymatic activities and other soil characteristics was determined in a natural soil from a desertified area. Dorycnium pentaphyllum, a legume plant adapted to stress situations, was the test plant to evaluate the effect of inoculation of native arbuscular mycorrhizal (AM) fungi and/or Yarowia lipolytica (a dry soil adapted yeast) on amended and non-amended soils. Plant growth and nutrition, symbiotic developments and soil enzymatic activities were limited in non-amended soil where microbial inoculations did not improve plant development. The lack of nodules formation and AM colonization can explain the limited plant growth in this natural soil. The effectiveness and performance of inocula applied was only evident in amended soils. AM colonization and spores number in natural soil were increased by amendments and the inoculation with Y. lipolytica promoted this value. The effect of the inoculations on plant N-acquisition was only important in AM-inoculated plants growing in SB medium. Enzymatic activities as urease and protease activities were particularly increased in DOC amended soil meanwhile dehydrogenase activity was greatest in treatments inoculated with Y. lipolytica in SB added soil. The biological activities in rhizosphere of agrowaste amended soil, used as indices of changes in soil properties and fertility, were affected not only by the nature of amendments but also by the inoculant applied. All these results show that the lignocellulosic agrowastes treated with a selected microorganism and its further interaction with beneficial microbial groups (native AM fungi and/or Y. lipolytica) is a useful tool to modify soil physico-chemical, biological and fertility properties that enhance the plant performance probably by making nutrients more available to plants.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Wei; Wei, Hui; Alahuhta, Markus

In order to develop a direct microbial sugar conversion platform for the production of lipids, drop-in fuels and chemicals from cellulosic biomass substrate, we chose Yarrowia lipolytica as a viable demonstration strain. Y. lipolytica is known to accumulate lipids intracellularly and is capable of metabolizing sugars to produce lipids; however, it lacks the lignocellulose-degrading enzymes needed to break down biomass directly. While research is continuing on the development of a Y. lipolytica strain able to degrade cellulose, in this study, we present successful expression of several xylanases in Y. lipolytica. The XynII and XlnD expressing Yarrowia strains exhibited an abilitymore » to grow on xylan mineral plates. This was shown by Congo Red staining of halo zones on xylan mineral plates. Enzymatic activity tests further demonstrated active expression of XynII and XlnD in Y. lipolytica. Furthermore, synergistic action in converting xylan to xylose was observed when XlnD acted in concert with XynII. Finally, the successful expression of these xylanases in Yarrowia further advances us toward our goal to develop a direct microbial conversion process using this organism.« less

Yeast mitochondrial HMG proteins: DNA-binding properties of the most evolutionarily divergent component of mitochondrial nucleoids.

PubMed

Bakkaiova, Jana; Marini, Victoria; Willcox, Smaranda; Nosek, Jozef; Griffith, Jack D; Krejci, Lumir; Tomaska, Lubomir

2015-12-08

Yeast mtDNA is compacted into nucleoprotein structures called mitochondrial nucleoids (mt-nucleoids). The principal mediators of nucleoid formation are mitochondrial high-mobility group (HMG)-box containing (mtHMG) proteins. Although these proteins are some of the fastest evolving components of mt-nucleoids, it is not known whether the divergence of mtHMG proteins on the level of their amino acid sequences is accompanied by diversification of their biochemical properties. In the present study we performed a comparative biochemical analysis of yeast mtHMG proteins from Saccharomyces cerevisiae (ScAbf2p), Yarrowia lipolytica (YlMhb1p) and Candida parapsilosis (CpGcf1p). We found that all three proteins exhibit relatively weak binding to intact dsDNA. In fact, ScAbf2p and YlMhb1p bind quantitatively to this substrate only at very high protein to DNA ratios and CpGcf1p shows only negligible binding to dsDNA. In contrast, the proteins exhibit much higher preference for recombination intermediates such as Holliday junctions (HJ) and replication forks (RF). Therefore, we hypothesize that the roles of the yeast mtHMG proteins in maintenance and compaction of mtDNA in vivo are in large part mediated by their binding to recombination/replication intermediates. We also speculate that the distinct biochemical properties of CpGcf1p may represent one of the prerequisites for frequent evolutionary tinkering with the form of the mitochondrial genome in the CTG-clade of hemiascomycetous yeast species. © 2016 Authors.

Efficient resource recycling from liquid digestate by microalgae-yeast mixed culture and the assessment of key gene transcription related to nitrogen assimilation in microalgae.

PubMed

Qin, Lei; Liu, Lu; Wang, Zhongming; Chen, Weining; Wei, Dong

2018-05-18

To determine the feasibility of microalgae-yeast mixed culture using the liquid digestate of dairy wastewater (LDDW) for biofuels and single cell protein (SCP) production, the cell growth, nutrient removal and outputs evaluation of the mono and mixed culture of Chlorella vulgaris and Yarrowia lipolytica in LDDW were investigated by adding glycerol as carbon source. The results showed that the mixed culture could enhance the biological utilization efficiency of nitrogen and phosphorus, and obtain higher yield of biomass (1.62 g/L), lipid (0.31 g/L), protein (0.51 g/L), and higher heating value (34.06 KJ/L). Compared with the mono culture of C. vulgaris, a decline of the transcription level in nitrate reductase and glutamine synthetase II genes in C. vulgaris was observed in the mixed culture when ammonia was sufficient. The results suggest the possibility of using the mixed culture for the efficient treatment of LDDW and resources recycling. Copyright © 2018. Published by Elsevier Ltd.

Process Analysis of Variables for Standardization of Antifungal Susceptibility Testing of Nonfermentative Yeasts ▿

PubMed Central

Zaragoza, Oscar; Mesa-Arango, Ana C.; Gómez-López, Alicia; Bernal-Martínez, Leticia; Rodríguez-Tudela, Juan Luis; Cuenca-Estrella, Manuel

2011-01-01

Nonfermentative yeasts, such as Cryptococcus spp., have emerged as fungal pathogens during the last few years. However, standard methods to measure their antifungal susceptibility (antifungal susceptibility testing [AST]) are not completely reliable due to the impaired growth of these yeasts in standard media. In this work, we have compared the growth kinetics and the antifungal susceptibilities of representative species of nonfermentative yeasts such as Cryptococcus neoformans, Cryptococcus gattii, Cryptococcus albidus, Rhodotorula spp., Yarrowia lipolytica, Geotrichum spp., and Trichosporon spp. The effect of the growth medium (RPMI medium versus yeast nitrogen base [YNB]), glucose concentration (0.2% versus 2%), nitrogen source (ammonium sulfate), temperature (30°C versus 35°C), shaking, and inoculum size (103, 104, and 105 cells) were analyzed. The growth rate, lag phase, and maximum optical density were obtained from each growth experiment, and after multivariate analysis, YNB-based media demonstrated a significant improvement in the growth of yeasts. Shaking, an inoculum size of 105 CFU/ml, and incubation at 30°C also improved the growth kinetics of organisms. Supplementation with ammonium sulfate and with 2% glucose did not have any effect on growth. We also tested the antifungal susceptibilities of all the isolates by the reference methods of the CLSI and EUCAST, the EUCAST method with shaking, YNB under static conditions, and YNB with shaking. MIC values obtained under different conditions showed high percentages of agreement and significant correlation coefficient values between them. MIC value determinations according to CLSI and EUCAST standards were rather complicated, since more than half of isolates tested showed a limited growth index, hampering endpoint determinations. We conclude that AST conditions including YNB as an assay medium, agitation of the plates, reading after 48 h of incubation, an inoculum size of 105 CFU/ml, and incubation at 30�

Safety assessment of EPA-rich triglyceride oil produced from yeast: genotoxicity and 28-day oral toxicity in rats.

PubMed

Belcher, Leigh A; MacKenzie, Susan A; Donner, Maria; Sykes, Greg P; Frame, Steven R; Gillies, Peter J

2011-02-01

The 28-day repeat-dose oral and genetic toxicity of eicosapentaenoic acid triglyceride oil (EPA oil) produced from genetically modified Yarrowia lipolytica yeast were assessed. Groups of rats received 0 (olive oil), 940, 1880, or 2820 mg EPA oil/kg/day, or fish oil (sardine/anchovy source) by oral gavage. Lower total serum cholesterol was seen in all EPA and fish oil groups. Liver weights were increased in the medium and high-dose EPA (male only), and fish oil groups but were considered non-adverse physiologically adaptive responses. Increased thyroid follicular cell hypertrophy was observed in male high-dose EPA and fish oil groups, and was considered to be an adaptive response to high levels of polyunsaturated fatty acids. No adverse test substance-related effects were observed on body weight, nutritional, or other clinical or anatomic pathology parameters. The oil was not mutagenic in the in vitro Ames or mouse lymphoma assay, and was not clastogenic in the in vivo mouse micronucleus test. In conclusion, exposure for 28 days to EPA oil derived from yeast did not produce adverse effects at doses up to 2820 mg/kg/day and was not genotoxic. The safety profile of the EPA oil in these tests was comparable to a commercial fish oil. Copyright © 2010 Elsevier Inc. All rights reserved.

Microbial Interactions within a Cheese Microbial Community▿ †

PubMed Central

Mounier, Jérôme; Monnet, Christophe; Vallaeys, Tatiana; Arditi, Roger; Sarthou, Anne-Sophie; Hélias, Arnaud; Irlinger, Françoise

2008-01-01

The interactions that occur during the ripening of smear cheeses are not well understood. Yeast-yeast interactions and yeast-bacterium interactions were investigated within a microbial community composed of three yeasts and six bacteria found in cheese. The growth dynamics of this community was precisely described during the ripening of a model cheese, and the Lotka-Volterra model was used to evaluate species interactions. Subsequently, the effects on ecosystem functioning of yeast omissions in the microbial community were evaluated. It was found both in the Lotka-Volterra model and in the omission study that negative interactions occurred between yeasts. Yarrowia lipolytica inhibited mycelial expansion of Geotrichum candidum, whereas Y. lipolytica and G. candidum inhibited Debaryomyces hansenii cell viability during the stationary phase. However, the mechanisms involved in these interactions remain unclear. It was also shown that yeast-bacterium interactions played a significant role in the establishment of this multispecies ecosystem on the cheese surface. Yeasts were key species in bacterial development, but their influences on the bacteria differed. It appeared that the growth of Arthrobacter arilaitensis or Hafnia alvei relied less on a specific yeast function because these species dominated the bacterial flora, regardless of which yeasts were present in the ecosystem. For other bacteria, such as Leucobacter sp. or Brevibacterium aurantiacum, growth relied on a specific yeast, i.e., G. candidum. Furthermore, B. aurantiacum, Corynebacterium casei, and Staphylococcus xylosus showed reduced colonization capacities in comparison with the other bacteria in this model cheese. Bacterium-bacterium interactions could not be clearly identified. PMID:17981942

The requirement of sterol glucoside for pexophagy in yeast is dependent on the species and nature of peroxisome inducers.

PubMed

Nazarko, Taras Y; Polupanov, Andriy S; Manjithaya, Ravi R; Subramani, Suresh; Sibirny, Andriy A

2007-01-01

Sterol glucosyltransferase, Ugt51/Atg26, is essential for both micropexophagy and macropexophagy of methanol-induced peroxisomes in Pichia pastoris. However, the role of this protein in pexophagy in other yeast remained unclear. We show that oleate- and amine-induced peroxisomes in Yarrowia lipolytica are degraded by Atg26-independent macropexophagy. Surprisingly, Atg26 was also not essential for macropexophagy of oleate- and amine-induced peroxisomes in P. pastoris, suggesting that the function of sterol glucoside (SG) in pexophagy is both species and peroxisome inducer specific. However, the rates of degradation of oleate- and amine-induced peroxisomes in P. pastoris were reduced in the absence of SG, indicating that P. pastoris specifically uses sterol conversion by Atg26 to enhance selective degradation of peroxisomes. However, methanol-induced peroxisomes apparently have lost the redundant ability to be degraded without SG. We also show that the P. pastoris Vac8 armadillo repeat protein is not essential for macropexophagy of methanol-, oleate-, or amine-induced peroxisomes, which makes PpVac8 the first known protein required for the micropexophagy, but not for the macropexophagy, machinery. The uniqueness of Atg26 and Vac8 functions under different pexophagy conditions demonstrates that not only pexophagy inducers, such as glucose or ethanol, but also the inducers of peroxisomes, such as methanol, oleate, or primary amines, determine the requirements for subsequent pexophagy in yeast.

Evolutionary relationships among pathogenic Candida species and relatives.

PubMed Central

Barns, S M; Lane, D J; Sogin, M L; Bibeau, C; Weisburg, W G

1991-01-01

Small subunit rRNA sequences have been determined for 10 of the most clinically important pathogenic species of the yeast genus Candida (including Torulopsis [Candida] glabrata and Yarrowia [Candida] lipolytica) and for Hansenula polymorpha. Phylogenetic analyses of these sequences and those of Saccharomyces cerevisiae, Kluyveromyces marxianus var. lactis, and Aspergillus fumigatus indicate that Candida albicans, C. tropicalis, C. parapsilosis, and C. viswanathii form a subgroup within the genus. The remaining significant pathogen, T. glabrata, falls into a second, distinct subgroup and is specifically related to S. cerevisiae and more distantly related to C. kefyr (psuedotropicalis) and K. marxianus var. lactis. The 18S rRNA sequence of Y. lipolytica has evolved rapidly in relation to the other Candida sequences examined and appears to be only distantly related to them. As anticipated, species of several other genera appear to bear specific relationships to members of the genus Candida. PMID:2007550

Influence of selected factors on browning of Camembert cheese.

PubMed

Carreira, Alexandra; Dillinger, Klaus; Eliskases-Lechner, Frieda; Loureiro, Virgílio; Ginzinger, Wolfgang; Rohm, Harald

2002-05-01

Experimental Camembert cheeses were made to investigate the effects on browning of the following factors: inoculation with Yarrowia lipolytica, the use of Penicillium candidum strains with different proteolytic activity, the addition of tyrosine, and the addition of Mn2+ thus leading to 16 different variants of cheese. Two physical colour parameters were used to describe browning, depending on the location in the cheeses: a whiteness index for the outside browning (mould mycelium), and a brownness index for the inside browning (surface of the cheese body). Mn2+ promoted a significant increase of browning at both locations, whereas Yar. lipolytica had the opposite effect. Outside browning was significantly more intense when using the Pen. candidum strain with higher proteolytic activity. A significant interaction was found between Yar. lipolytica and Pen. candidum. The yeast had no effect in combination with a low proteolytic strain of Pen. candidum, but significantly reduced proteolysis and browning in combination with a high proteolytic strain of Pen. candidum. We further confirmed that both strains of Pen. candidum were able to produce brown pigments from tyrosine and thus both are presumably responsible for the browning activity in this type of cheese.

Characterization of the newly isolated ω-oxidizing yeast Candida sorbophila DS02 and its potential applications in long-chain dicarboxylic acid production.

PubMed

Lee, Heeseok; Sugiharto, Yohanes Eko Chandra; Lee, Seunghoon; Park, Gyuyeon; Han, Changpyo; Jang, Hyeran; Jeon, Wooyoung; Park, Heejoon; Ahn, Jungoh; Kang, Kyungbo; Lee, Hongwoen

2017-08-01

α, ω-Dicarboxylic acids (DCAs) are multipurpose chemicals widely used in polymers, perfumes, plasticizers, lubricants, and adhesives. The biotransformation of DCAs from alkanes and fatty acids by microorganisms has attracted recent interest, since synthesis via chemical oxidation causes problems in terms of the environment and safety. We isolated an ω-oxidizing yeast from a wastewater disposal facility of a petrochemical factory by chemostat enrichment culture. The haploid strain identified as Candida sorbophila DS02 grew on glucose and dodecane, exhibiting greater cell shrinkage on the latter. In flask cultures with mixed alkanes (C10-16) and fatty acid methyl esters (C10-16), DS02 used mixed alkanes simultaneously unlike Candida tropicalis and Yarrowia lipolytica and showed high substrate resistance. In flask cultures with acrylic acid-a known inhibitor of β-oxidation-DS02 produced 0.28 g/l dodecanedioic acid (DDDA) from dodecane, similar to wild-type C. tropicalis ATCC 20336. In fed-batch fermentation, DS02 produced 9.87 g/l DDDA, which was 5.7-fold higher than wild-type C. tropicalis. These results suggest that C. sorbophila strain DS02 has potential applications for the large-scale production of DCA.

21 CFR 173.165 - Candida lipolytica.

Code of Federal Regulations, 2010 CFR

2010-04-01

... description for Candida lipolytica variety lipolytica listed in “The Yeasts—A Toxonomic Study,” 2d Ed. (1970... equivalent). Activate as follows: Slurry 900 grams of silica gel reagent with 2 liters of purified water in a 3-liter beaker. Cool the mixture and pour into a 80 × 900 chromatographic column with coarse fritted...

21 CFR 173.165 - Candida lipolytica.

Code of Federal Regulations, 2011 CFR

2011-04-01

... description for Candida lipolytica variety lipolytica listed in “The Yeasts—A Toxonomic Study,” 2d Ed. (1970... equivalent). Activate as follows: Slurry 900 grams of silica gel reagent with 2 liters of purified water in a 3-liter beaker. Cool the mixture and pour into a 80 × 900 chromatographic column with coarse fritted...

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi

DOE PAGES

Xu, Qi; Knoshaug, Eric P.; Wang, Wei; ...

2017-07-24

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose twomore » prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Qi; Knoshaug, Eric P.; Wang, Wei

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose twomore » prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for

Expression and secretion of fungal endoglucanase II and chimeric cellobiohydrolase I in the oleaginous yeast Lipomyces starkeyi.

PubMed

Xu, Qi; Knoshaug, Eric P; Wang, Wei; Alahuhta, Markus; Baker, John O; Yang, Shihui; Vander Wall, Todd; Decker, Stephen R; Himmel, Michael E; Zhang, Min; Wei, Hui

2017-07-24

Lipomyces starkeyi is one of the leading lipid-producing microorganisms reported to date; its genetic transformation was only recently reported. Our aim is to engineer L. starkeyi to serve in consolidated bioprocessing (CBP) to produce lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates. To evaluate L. starkeyi in this role, we first conducted a genome analysis, which revealed the absence of key endo- and exocellulases in this yeast, prompting us to select and screen four signal peptides for their suitability for the overexpression and secretion of cellulase genes. To compensate for the cellulase deficiency, we chose two prominent cellulases, Trichoderma reesei endoglucanase II (EG II) and a chimeric cellobiohydrolase I (TeTrCBH I) formed by fusion of the catalytic domain from Talaromyces emersonii CBH I with the linker peptide and cellulose-binding domain from T. reesei CBH I. The systematically tested signal peptides included three peptides from native L. starkeyi and one from Yarrowia lipolytica. We found that all four signal peptides permitted secretion of active EG II. We also determined that three of these signal peptides worked for expression of the chimeric CBH I; suggesting that our design criteria for selecting these signal peptides was effective. Encouragingly, the Y. lipolytica signal peptide was able to efficiently guide secretion of the chimeric TeTrCBH I protein from L. starkeyi. The purified chimeric TeTrCBH I showed high activity against the cellulose in pretreated corn stover and the purified EG II showed high endocellulase activity measured by the CELLG3 (Megazyme) method. Our results suggest that L. starkeyi is capable of expressing and secreting core fungal cellulases. Moreover, the purified EG II and chimeric TeTrCBH I displayed significant and potentially useful enzymatic activities, demonstrating that engineered L. starkeyi has the potential to function as an oleaginous CBP strain for biofuel

Effect of scenedesmus acuminatus green algae extracts on the development of Candida lipolytic yeast in gas condensate-containing media

NASA Technical Reports Server (NTRS)

Bilmes, B. I.; Kasymova, G. A.; Runov, V. I.; Karavayeva, N. N.

1980-01-01

Data are given of a comparative study of the growth and development as well as the characteristics of the biomass of the C. Lipolytica yeast according to the content of raw protein, protein, lipids, vitamins in the B group, and residual hydrocarbons during growth in media with de-aromatized gas-condensate FNZ as the carbon source with aqueous and alcohol extracts of S. acuminatus as the biostimulants. It is shown that the decoction and aqueous extract of green algae has the most intensive stimulating effect on the yeast growth. When a decoction of algae is added to the medium, the content of residual hydrocarbons in the biomass of C. lipolytica yeast is reduced by 4%; the quantity of protein, lipids, thamine and inositol with replacement of the yeast autolysate by the decoction of algae is altered little.

[Yeast irrigation enhances the nutritional content in hydroponic green maize fodder].

PubMed

Bedolla-Torres, Martha H; Palacios Espinosa, Alejandro; Palacios, Oskar A; Choix, Francisco J; Ascencio Valle, Felipe de Jesús; López Aguilar, David R; Espinoza Villavicencio, José Luis; de Luna de la Peña, Rafael; Guillen Trujillo, Ariel; Avila Serrano, Narciso Y; Ortega Pérez, Ricardo

2015-01-01

The objective of this study was to evaluate the effect of irrigation with yeasts (Debaryomyces hansenii var. Fabry, Yarowia lipolytica YIBCS002, Yarowia lipolytica var. BCS and Candida pseudointermedia) on the final nutritional content of hydroponic green maize fodder (Zea Zea mays L.), applied at different fodder growth stages (1. seed-seedling stage, 2. seedling-plant 20cm, 3. during all the culture). Irrespective of the fodder growth stages at which they were applied, all yeasts tested enhanced the content of raw protein, lipids, ash, moisture and energy. The percentage of electrolytes (Na, K, Cl, sulphates, Ca and Mg) showed different responses depending on the kind of yeast applied; D. hansenii exhibited the highest increment in all electrolytes, except for phosphorous. We conclude that the addition of yeasts belonging to the genera Debaryomyces, Candida and Yarowia to the irrigation solution of hydroponic systems enhances the nutrient content of green fodder. This kind of irrigation can be applied to generate high commercial value cultures in limited spaces. Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Molecular characterization and expression of microbial inulinase genes.

PubMed

Liu, Guang-Lei; Chi, Zhe; Chi, Zhen-Ming

2013-05-01

Many genes encoding exo- and endo-inulinases from bacteria, yeasts and filamentous fungi have been cloned and characterized. All the inulinases have several conserved motifs, such as WMND(E)PNGL, RDP, EC(V)P, SVEVF, Q and FS(T), which play an important role in inulinase catalysis and substrate binding. However, the exo-inulinases produced by yeasts has no conserved motif SVEVF and the yeasts do not produce any endo-inulinase. Exo- and endo-inulinases found in different microorganisms cluster separately at distant positions from each other. Most of the cloned inulinase genes have been expressed in Yarrowia lipolytica, Saccharomyces cerevisiae, Pichia pastoris, Klyuveromyces lactis and Escherichia coli, respectively. The recombinant inulinases produced and the engineered hosts using the cloned inulinase genes have many potential applications. Expression of most of the inulinase genes is repressed by glucose and fructose and induced by inulin and sucrose. However, the detailed mechanisms of the repression and induction are still unknown.

Metabolic Engineering of Oleaginous Yeasts for Production of Fuels and Chemicals

PubMed Central

Shi, Shuobo; Zhao, Huimin

2017-01-01

Oleaginous yeasts have been increasingly explored for production of chemicals and fuels via metabolic engineering. Particularly, there is a growing interest in using oleaginous yeasts for the synthesis of lipid-related products due to their high lipogenesis capability, robustness, and ability to utilize a variety of substrates. Most of the metabolic engineering studies in oleaginous yeasts focused on Yarrowia that already has plenty of genetic engineering tools. However, recent advances in systems biology and synthetic biology have provided new strategies and tools to engineer those oleaginous yeasts that have naturally high lipid accumulation but lack genetic tools, such as Rhodosporidium, Trichosporon, and Lipomyces. This review highlights recent accomplishments in metabolic engineering of oleaginous yeasts and recent advances in the development of genetic engineering tools in oleaginous yeasts within the last 3 years. PMID:29167664

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

PubMed

Shi, Shuobo; Zhao, Huimin

2017-01-01

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

Evolution of the carboxylate Jen transporters in fungi.

PubMed

Lodi, Tiziana; Diffels, Julie; Goffeau, André; Baret, Philippe V

2007-08-01

Synteny analysis is combined with sequence similarity and motif identification to trace the evolution of the putative monocarboxylate (lactate/pyruvate) transporters Jen1p and the dicarboxylate (succinate/fumarate/malate) transporters Jen2p in Hemiascomycetes yeasts and Euascomycetes fungi. It is concluded that a precursor form of Jen1p, named here preJen1p, arose by the duplication of an ancestral Jen2p, during the speciation of Yarrowia lipolytica, which was transferred into a new syntenic context. The Jen1p transporters differentiated from preJen1p in Kluyveromyces lactis, before the Whole Genome Duplication (WGD), and are conserved as a single copy in the Saccharomyces species. In contrast, the ancestral Jen2p was definitively lost just prior to the WGD and is absent in Saccharomyces.

Zinc Finger Transcription Factors Displaced SREBP Proteins as the Major Sterol Regulators during Saccharomycotina Evolution

PubMed Central

Maguire, Sarah L.; Wang, Can; Holland, Linda M.; Brunel, François; Neuvéglise, Cécile; Nicaud, Jean-Marc; Zavrel, Martin; White, Theodore C.; Wolfe, Kenneth H.; Butler, Geraldine

2014-01-01

In most eukaryotes, including the majority of fungi, expression of sterol biosynthesis genes is regulated by Sterol-Regulatory Element Binding Proteins (SREBPs), which are basic helix-loop-helix transcription activators. However, in yeasts such as Saccharomyces cerevisiae and Candida albicans sterol synthesis is instead regulated by Upc2, an unrelated transcription factor with a Gal4-type zinc finger. The SREBPs in S. cerevisiae (Hms1) and C. albicans (Cph2) have lost a domain, are not major regulators of sterol synthesis, and instead regulate filamentous growth. We report here that rewiring of the sterol regulon, with Upc2 taking over from SREBP, likely occurred in the common ancestor of all Saccharomycotina. Yarrowia lipolytica, a deep-branching species, is the only genome known to contain intact and full-length orthologs of both SREBP (Sre1) and Upc2. Deleting YlUPC2, but not YlSRE1, confers susceptibility to azole drugs. Sterol levels are significantly reduced in the YlUPC2 deletion. RNA-seq analysis shows that hypoxic regulation of sterol synthesis genes in Y. lipolytica is predominantly mediated by Upc2. However, YlSre1 still retains a role in hypoxic regulation; growth of Y. lipolytica in hypoxic conditions is reduced in a Ylupc2 deletion and is abolished in a Ylsre1/Ylupc2 double deletion, and YlSre1 regulates sterol gene expression during hypoxia adaptation. We show that YlSRE1, and to a lesser extent YlUPC2, are required for switching from yeast to filamentous growth in hypoxia. Sre1 appears to have an ancestral role in the regulation of filamentation, which became decoupled from its role in sterol gene regulation by the arrival of Upc2 in the Saccharomycotina. PMID:24453983

Biotechnological Applications of Dimorphic Yeasts

NASA Astrophysics Data System (ADS)

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.

Investigation of the effect of biologically active threo-Ds-isocitric acid on oxidative stress in Paramecium caudatum.

PubMed

Morgunov, Igor G; Karpukhina, Olga V; Kamzolova, Svetlana V; Samoilenko, Vladimir A; Inozemtsev, Anatoly N

2018-01-02

The effect of biologically active form (threo-Ds-) of isocitric acid (ICA) on oxidative stress was studied using the infusorian Paramecium caudatum stressed by hydrogen peroxide and salts of some heavy metals (Cu, Pb, Zn, and Cd). ICA at concentrations between 0.5 and 10 mM favorably influenced the infusorian cells with oxidative stress induced by the toxicants studied. The maximal antioxidant effect of ICA was observed at its concentration 10 mM irrespective of the toxicant used (either H 2 O 2 or heavy metal ions). ICA was found to be a more active antioxidant than ascorbic acid. Biologically active pharmaceutically pure threo-Ds-ICA was produced through cultivation of the yeast Yarrowia lipolytica and isolated from the culture liquid in the form of crystalline monopotassium salt with a purity of 99.9%.

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic.

PubMed

Song, Bo; Rong, Yan-Jun; Zhao, Ming-Xin; Chi, Zhen-Ming

2013-08-01

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.

Metabolic engineering of microbial competitive advantage for industrial fermentation processes.

PubMed

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

2016-08-05

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

Carbohydrate and energy-yielding metabolism in non-conventional yeasts.

PubMed

Flores, C L; Rodríguez, C; Petit, T; Gancedo, C

2000-10-01

Sugars are excellent carbon sources for all yeasts. Since a vast amount of information is available on the components of the pathways of sugar utilization in Saccharomyces cerevisiae it has been tacitly assumed that other yeasts use sugars in the same way. However, although the pathways of sugar utilization follow the same theme in all yeasts, important biochemical and genetic variations on it exist. Basically, in most non-conventional yeasts, in contrast to S. cerevisiae, respiration in the presence of oxygen is prominent for the use of sugars. This review provides comparative information on the different steps of the fundamental pathways of sugar utilization in non-conventional yeasts: glycolysis, fermentation, tricarboxylic acid cycle, pentose phosphate pathway and respiration. We consider also gluconeogenesis and, briefly, catabolite repression. We have centered our attention in the genera Kluyveromyces, Candida, Pichia, Yarrowia and Schizosaccharomyces, although occasional reference to other genera is made. The review shows that basic knowledge is missing on many components of these pathways and also that studies on regulation of critical steps are scarce. Information on these points would be important to generate genetically engineered yeast strains for certain industrial uses.

Purification and characterization of an alkaline protease from the marine yeast Aureobasidium pullulans for bioactive peptide production from different sources.

PubMed

Ma, Chunling; Ni, Xiumei; Chi, Zhenming; Ma, Liyan; Gao, Lingmei

2007-01-01

The extracellular alkaline protease in the supernatant of cell culture of the marine yeast Aureobasidium pullulans 10 was purified to homogeneity with a 2.1-fold increase in specific protease activity as compared to that in the supernatant by ammonium sulfate fractionation, gel filtration chromatography (Sephadex G-75), and anion-exchange chromatography (DEAE Sepharose Fast Flow). According to the sodium dodecyl sulfate-polyacrylamide gel electrophoresis data, the molecular mass of the purified enzyme was estimated to be 32.0 kDa. The optimal pH and temperature of the purified enzyme were 9.0 and 45 degrees C, respectively. The enzyme was activated by Cu(2+) (at a concentration of 1.0 mM) and Mn(2+) and inhibited by Hg(2+), Fe(2+), Fe(3+), Zn(2+), and Co(2+). The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride, but weakly inhibited by EDTA, 1-10-phenanthroline, and iodoacetic acid. The K(m) and V(max) values of the purified enzyme for casein were 0.25 mg/ml and 0.0286 micromol/min/mg of protein, respectively. After digestion of shrimp protein, spirulina (Arthospira platensis) protein, proteins of marine yeast strains N3C (Yarrowia lipolytica) and YA03a (Hanseniaspora uvarum), milk protein, and casein with the purified alkaline protease, angiotensin I converting enzyme (ACE) inhibitory activities of the resulting peptides reached 85.3%, 12.1%, 29.8%, 22.8%, 14.1%, and 15.5%, respectively, while the antioxidant activities of these were 52.1%. 54.6%, 25.1%, 35%, 12.5%, and 24.2%, respectively, indicating that ACE inhibitory activity of the resulting peptides from the shrimp protein and antioxidant activity of those produced from the spirulina protein were the highest, respectively. These results suggest that the bioactive peptides produced by digestion of the shrimp protein with the purified alkaline protease have potential applications in the food and pharmaceutical industries.

Complementation and Genetic Recombination in Candida lipolytica

PubMed Central

Bassel, John; Warfei, Jean; Mortimer, Robert

1971-01-01

Nutritional requirements were introduced into wild-type, heterothallic strains of Candida lipolytica by exposing the cells to X rays. Complementing hybrids were recovered from mixtures of the auxotrophic strains, and genetic recombination was observed in individually isolated ascospores from the hybrid strains. PMID:5122814

Production, characterization and gene cloning of the extracellular enzymes from the marine-derived yeasts and their potential applications.

PubMed

Chi, Zhenming; Chi, Zhe; Zhang, Tong; Liu, Guanglei; Li, Jing; Wang, Xianghong

2009-01-01

In this review article, the extracellular enzymes production, their properties and cloning of the genes encoding the enzymes from marine yeasts are overviewed. Several yeast strains which could produce different kinds of extracellular enzymes were selected from the culture collection of marine yeasts available in this laboratory. The strains selected belong to different genera such as Yarrowia, Aureobasidium, Pichia, Metschnikowia and Cryptococcus. The extracellular enzymes include cellulase, alkaline protease, aspartic protease, amylase, inulinase, lipase and phytase, as well as killer toxin. The conditions and media for the enzyme production by the marine yeasts have been optimized and the enzymes have been purified and characterized. Some genes encoding the extracellular enzymes from the marine yeast strains have been cloned, sequenced and expressed. It was found that some properties of the enzymes from the marine yeasts are unique compared to those of the homologous enzymes from terrestrial yeasts and the genes encoding the enzymes in marine yeasts are different from those in terrestrial yeasts. Therefore, it is of very importance to further study the enzymes and their genes from the marine yeasts. This is the first review on the extracellular enzymes and their genes from the marine yeasts.

Yeast diversity associated to sediments and water from two Colombian artificial lakes

PubMed Central

Silva-Bedoya, L.M.; Ramírez-Castrillón, M.; Osorio-Cadavid, E.

2014-01-01

In Colombia, knowledge of the yeast and yeast-like fungi community is limited because most studies have focused on species with clinical importance. Sediments and water represent important habitats for the study of yeast diversity, especially for yeast species with industrial, biotechnological, and bioremediation potential. The main purpose of this study was to identify and compare the diversity of yeast species associated with sediment and water samples from two artificial lakes in Universidad del Valle (Cali-Colombia). Yeast samplings were performed from fifteen sediment samples and ten water samples. Grouping of similar isolates was initially based on colony and cell morphology, which was then complemented by micro/mini satellite primed PCR banding pattern analysis by using GTG5 as single primer. A representative isolate for each group established was chosen for D1/D2 domain sequencing and identification. In general, the following yeast species were identified: Candida albicans, Candida diversa, Candida glabrata, Candida pseudolambica, Cryptococcus podzolicus, Cryptococcus rajasthanensis, Cryptococcus laurentii, Williopsis saturnus, Hanseniaspora thailandica, Hanseniaspora uvarum, Rhodotorula mucilaginosa, Saccharomyces cerevisiae, Torulaspora delbrueckii, Torulaspora pretoriensis, Tricosporon jirovecii, Trichosporon laibachii and Yarrowia lypolitica. Two possible new species were also found, belonging to the Issatchenkia sp. and Bullera sp. genera. In conclusion, the lakes at the Universidad del Valle campus have significant differences in yeast diversity and species composition between them. PMID:24948924

The occurrence and growth of yeasts in Camembert and blue-veined cheeses.

PubMed

Roostita, R; Fleet, G H

1996-01-01

Yeast populations greater than 10(6) cfu/g were found in approximately 54% and 36%, respectively in surface samples of retail Camembert (85 samples) and Blue-veined (45 samples) cheeses. The most predominant species isolated were Debaryomyces hansenii, Candida catenulata, C. lipolytica, C. kefyr, C. intermedia, Saccharomyces cerevisiae, Cryptococcus albidus and Kluyveromyces marxianus. The salt concentration of the surface samples of the cheeses varied between 2.5-5.5% (w/w) (Camembert) and 7.5-8.3 (Blue-veined), depending upon brand, and influenced the yeast ecology, especially the presence of S. cerevisiae. Yeasts grew to populations of 10(6)-10(8) cfu/g when cheeses were stored at either 25 degrees C or 10 degrees C. These populations decreased on continued storage at 25 degrees C, but such decreases were not so evident on storage at 10 degrees C. The properties of yeasts influencing their occurrence and growth in cheese were: fermentation/assimilation of lactose; production of extracellular lipolytic and proteolytic enzymes, utilisation of lactic and citric acids; and growth at 10 degrees C.

An original method for producing acetaldehyde and diacetyl by yeast fermentation.

PubMed

Rosca, Irina; Petrovici, Anca Roxana; Brebu, Mihai; Stoica, Irina; Minea, Bogdan; Marangoci, Narcisa

In this study a natural culture medium that mimics the synthetic yeast peptone glucose medium used for yeast fermentations was designed to screen and select yeasts capable of producing high levels of diacetyl and acetaldehyde. The presence of whey powder and sodium citrate in the medium along with manganese and magnesium sulfate enhanced both biomass and aroma development. A total of 52 yeasts strains were cultivated in two different culture media, namely, yeast peptone glucose medium and yeast acetaldehyde-diacetyl medium. The initial screening of the strains was based on the qualitative reaction of the acetaldehyde with Schiff's reagent (violet color) and diacetyl with Brady's reagent (yellow precipitate). The fermented culture media of 10 yeast strains were subsequently analyzed by gas chromatography to quantify the concentration of acetaldehyde and diacetyl synthesized. Total titratable acidity values indicated that a total titratable acidity of 5.5°SH, implying culture medium at basic pH, was more favorable for the acetaldehyde biosynthesis using strain D15 (Candida lipolytica; 96.05mgL -1 acetaldehyde) while a total titratable acidity value of 7°SH facilitated diacetyl flavor synthesis by strain D38 (Candida globosa; 3.58mgL -1 diacetyl). Importantly, the results presented here suggest that this can be potentially used in the baking industry. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

A critical tyrosine residue determines the uncoupling protein-like activity of the yeast mitochondrial oxaloacetate carrier.

PubMed

Luévano-Martínez, Luis A; Barba-Ostria, Carlos; Araiza-Olivera, Daniela; Chiquete-Félix, Natalia; Guerrero-Castillo, Sergio; Rial, Eduardo; Georgellis, Dimitris; Uribe-Carvajal, Salvador

2012-04-01

The mitochondrial Oac (oxaloacetate carrier) found in some fungi and plants catalyses the uptake of oxaloacetate, malonate and sulfate. Despite their sequence similarity, transport specificity varies considerably between Oacs. Indeed, whereas ScOac (Saccharomyces cerevisiae Oac) is a specific anion-proton symporter, the YlOac (Yarrowia lipolytica Oac) has the added ability to transport protons, behaving as a UCP (uncoupling protein). Significantly, we identified two amino acid changes at the matrix gate of YlOac and ScOac, tyrosine to phenylalanine and methionine to leucine. We studied the role of these amino acids by expressing both wild-type and specifically mutated Oacs in an Oac-null S. cerevisiae strain. No phenotype could be associated with the methionine to leucine substitution, whereas UCP-like activity was dependent on the presence of the tyrosine residue normally expressed in the YlOac, i.e. Tyr-ScOac mediated proton transport, whereas Phe-YlOac lost its protonophoric activity. These findings indicate that the UCP-like activity of YlOac is determined by the tyrosine residue at position 146.

Adhesion of Bacillus subtilis and Pseudoalteromonas lipolytica to steel in a seawater environment and their effects on corrosion.

PubMed

Guo, Zhangwei; Liu, Tao; Cheng, Y Frank; Guo, Na; Yin, Yansheng

2017-09-01

In a marine environment, Bacillus subtilis and Pseudoalteromonas lipolytica are commonly found in the biofilms adherent to low-alloy engineering steel, and they have distinct effects on corrosion. In the present work, this phenomenon was investigated through the study of various materials characterization methods, electrochemical techniques, and contact angle measurements. It was found that the surface film formed on the steel in the presence of B. subtilis was compact, uniform, free of cracks, and hydrophobic. However, the film formed in the presence of P. lipolytica was loose, rough, heterogeneous, and hydrophilic. The main components of the films formed in the presence of B. subtilis and P. lipolytica were polysaccharides/TasA amyloid fibers and proteins/carboxylic acid, respectively. The composition, structure, and properties of the surface films formed on the steel were associated with different effects on corrosion. The presence of B. subtilis enhances the steel's resistance to corrosion, whereas corrosion was increased by the presence of P. lipolytica. In short, the compact and hydrophobic biofilm of B. subtilis appears to inhibit the corrosion of steel, while the loose, hydrophilic film of P. lipolytica tends to induce pitting corrosion. Copyright © 2017 Elsevier B.V. All rights reserved.

Surface Microflora of Four Smear-Ripened Cheeses

PubMed Central

Mounier, Jérôme; Gelsomino, Roberto; Goerges, Stefanie; Vancanneyt, Marc; Vandemeulebroecke, Katrien; Hoste, Bart; Scherer, Siegfried; Swings, Jean; Fitzgerald, Gerald F.; Cogan, Timothy M.

2005-01-01

The microbial composition of smear-ripened cheeses is not very clear. A total of 194 bacterial isolates and 187 yeast isolates from the surfaces of four Irish farmhouse smear-ripened cheeses were identified at the midpoint of ripening using pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR, and 16S rRNA gene sequencing for identifying and typing the bacteria and Fourier transform infrared spectroscopy and mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP) analysis for identifying and typing the yeast. The yeast microflora was very uniform, and Debaryomyces hansenii was the dominant species in the four cheeses. Yarrowia lipolytica was also isolated in low numbers from one cheese. The bacteria were highly diverse, and 14 different species, Corynebacterium casei, Corynebacterium variabile, Arthrobacter arilaitensis, Arthrobacter sp., Microbacterium gubbeenense, Agrococcus sp. nov., Brevibacterium linens, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus saprophyticus, Micrococcus luteus, Halomonas venusta, Vibrio sp., and Bacillus sp., were identified on the four cheeses. Each cheese had a more or less unique microflora with four to nine species on its surface. However, two bacteria, C. casei and A. arilaitensis, were found on each cheese. Diversity at the strain level was also observed, based on the different PFGE patterns and mtDNA RFLP profiles of the dominant bacterial and yeast species. None of the ripening cultures deliberately inoculated onto the surface were reisolated from the cheeses. This study confirms the importance of the adventitious, resident microflora in the ripening of smear cheeses. PMID:16269673

Clinical, microbiological, and experimental animal studies of Candida lipolytica.

PubMed Central

Walsh, T J; Salkin, I F; Dixon, D M; Hurd, N J

1989-01-01

Candida lipolytica was recovered from six patients in three different clinical centers. The index isolate caused a persistent fungemia with catheter-associated Candida thrombophlebitis, the second isolate was from a polymicrobial sinusitis, and the remaining four isolates were involved in tissue colonization. These and 20 other isolates were consistent in their morphological and physiological characteristics. All formed true hyphae and blastoconidia on cornmeal-Tween 80 agar and all assimilated glucose, glycerol, and erythritol. In a murine model of disseminated candidiasis, the index isolate that caused clinical fungemia caused no mortality and produced only two lesions on a kidney, as determined at necropsy. The nine isolates selected for in vitro antifungal susceptibility studies had intermediate susceptibilities to amphotericin B but were susceptible to ketoconazole. We conclude that C. lipolytica is a weakly virulent pathogen which may require an intravascular foreign body to cause fungemia. Images PMID:2745702

Sustainable conversion of coffee and other crop wastes to biofuels and bioproducts using coupled biochemical and thermochemical processes in a multi-stage biorefinery concept.

PubMed

Hughes, Stephen R; López-Núñez, Juan Carlos; Jones, Marjorie A; Moser, Bryan R; Cox, Elby J; Lindquist, Mitch; Galindo-Leva, Luz Angela; Riaño-Herrera, Néstor M; Rodriguez-Valencia, Nelson; Gast, Fernando; Cedeño, David L; Tasaki, Ken; Brown, Robert C; Darzins, Al; Brunner, Lane

2014-10-01

The environmental impact of agricultural waste from the processing of food and feed crops is an increasing concern worldwide. Concerted efforts are underway to develop sustainable practices for the disposal of residues from the processing of such crops as coffee, sugarcane, or corn. Coffee is crucial to the economies of many countries because its cultivation, processing, trading, and marketing provide employment for millions of people. In coffee-producing countries, improved technology for treatment of the significant amounts of coffee waste is critical to prevent ecological damage. This mini-review discusses a multi-stage biorefinery concept with the potential to convert waste produced at crop processing operations, such as coffee pulping stations, to valuable biofuels and bioproducts using biochemical and thermochemical conversion technologies. The initial bioconversion stage uses a mutant Kluyveromyces marxianus yeast strain to produce bioethanol from sugars. The resulting sugar-depleted solids (mostly protein) can be used in a second stage by the oleaginous yeast Yarrowia lipolytica to produce bio-based ammonia for fertilizer and are further degraded by Y. lipolytica proteases to peptides and free amino acids for animal feed. The lignocellulosic fraction can be ground and treated to release sugars for fermentation in a third stage by a recombinant cellulosic Saccharomyces cerevisiae, which can also be engineered to express valuable peptide products. The residual protein and lignin solids can be jet cooked and passed to a fourth-stage fermenter where Rhodotorula glutinis converts methane into isoprenoid intermediates. The residues can be combined and transferred into pyrocracking and hydroformylation reactions to convert ammonia, protein, isoprenes, lignins, and oils into renewable gas. Any remaining waste can be thermoconverted to biochar as a humus soil enhancer. The integration of multiple technologies for treatment of coffee waste has the potential to

The RNA polymerase III-dependent family of genes in hemiascomycetes: comparative RNomics, decoding strategies, transcription and evolutionary implications

PubMed Central

Marck, Christian; Kachouri-Lafond, Rym; Lafontaine, Ingrid; Westhof, Eric; Dujon, Bernard; Grosjean, Henri

2006-01-01

We present the first comprehensive analysis of RNA polymerase III (Pol III) transcribed genes in ten yeast genomes. This set includes all tRNA genes (tDNA) and genes coding for SNR6 (U6), SNR52, SCR1 and RPR1 RNA in the nine hemiascomycetes Saccharomyces cerevisiae, Saccharomyces castellii, Candida glabrata, Kluyveromyces waltii, Kluyveromyces lactis, Eremothecium gossypii, Debaryomyces hansenii, Candida albicans, Yarrowia lipolytica and the archiascomycete Schizosaccharomyces pombe. We systematically analysed sequence specificities of tRNA genes, polymorphism, variability of introns, gene redundancy and gene clustering. Analysis of decoding strategies showed that yeasts close to S.cerevisiae use bacterial decoding rules to read the Leu CUN and Arg CGN codons, in contrast to all other known Eukaryotes. In D.hansenii and C.albicans, we identified a novel tDNA-Leu (AAG), reading the Leu CUU/CUC/CUA codons with an unusual G at position 32. A systematic ‘p-distance tree’ using the 60 variable positions of the tRNA molecule revealed that most tDNAs cluster into amino acid-specific sub-trees, suggesting that, within hemiascomycetes, orthologous tDNAs are more closely related than paralogs. We finally determined the bipartite A- and B-box sequences recognized by TFIIIC. These minimal sequences are nearly conserved throughout hemiascomycetes and were satisfactorily retrieved at appropriate locations in other Pol III genes. PMID:16600899

Yeast cell surface display for lipase whole cell catalyst and its applications

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Yun; Zhang, Rui; Lian, Zhongshuai

The cell surface display technique allows for the expression of target proteins or peptides on the microbial cell surface by fusing an appropriate protein as an anchoring motif. Yeast display systems, such as Pichia pastoris, Yarowia lipolytica and Saccharomyces cerevisiae, are ideal, alternative and extensive display systems with the advantage of simple genetic manipulation and post-translational modification of expressed heterologous proteins. Engineered yeasts show high performance characteristics and variant utilizations. Herein, we comprehensively summarize the variant factors affecting lipase whole cell catalyst activity and display efficiency, including the structure and size of target proteins, screening anchor proteins, type and chainmore » length of linkers, and the appropriate matching rules among the above-mentioned display units. Furthermore, we also address novel approaches to enhance stability and activity of recombinant lipases, such as VHb gene co-expression, multi-enzyme co-display technique, and the micro-environmental interference and self-assembly techniques. Finally, we represent the variety of applications of whole cell surface displayed lipases on yeast cells in non-aqueous phases, including synthesis of esters, PUFA enrichment, resolution of chiral drugs, organic synthesis and biofuels. We demonstrate that the lipase surface display technique is a powerful tool for functionalizing yeasts to serve as whole cell catalysts, and increasing interest is providing an impetus for broad application of this technique.« less

Metrological aspects of enzyme production

NASA Astrophysics Data System (ADS)

Kerber, T. M.; Dellamora-Ortiz, G. M.; Pereira-Meirelles, F. V.

2010-05-01

Enzymes are frequently used in biotechnology to carry out specific biological reactions, either in industrial processes or for the production of bioproducts and drugs. Microbial lipases are an important group of biotechnologically valuable enzymes that present widely diversified applications. Lipase production by microorganisms is described in several published papers; however, none of them refer to metrological evaluation and the estimation of the uncertainty in measurement. Moreover, few of them refer to process optimization through experimental design. The objectives of this work were to enhance lipase production in shaken-flasks with Yarrowia lipolytica cells employing experimental design and to evaluate the uncertainty in measurement of lipase activity. The highest lipolytic activity obtained was about three- and fivefold higher than the reported activities of CRMs BCR-693 and BCR-694, respectively. Lipase production by Y. lipolytica cells aiming the classification as certified reference material is recommended after further purification and stability studies.

Rapid fabrication of three-dimensional structures for dielectrophoretic sorting of lipid-containing organisms

NASA Astrophysics Data System (ADS)

Schor, Alisha R.; Buie, Cullen R.

2016-10-01

In this work, we demonstrate a microfluidic particle sorter consisting of three-dimensional, conducting microposts. Our sorter uses dielectrophoresis (DEP) to sort high- and low-lipid phenotypes of the yeast Yarrowia lipolytica. Y. lipolytica is one of the many microorganisms being explored as a hydrocarbon source for biodiesel, Omega-3 additives, and other products derived from fatty acids. A rapid, non-destructive, lipid-based sorting tool would accelerate the commercialization of these products. Our device consists of an array of 105, 25 μm wide gold microposts that span the height of a 15 μm channel. This array generates an electric field in a microfluidic device that is uniform through the channel height, but has a custom-shaped non-uniformity in the horizontal directions. This is crucial in order to achieve continuous sorting using DEP, as it ensures all cells are exposed to the same conditions throughout the channel height. By using very low currents (100 μA), we are able to electroplate these post arrays in fewer than 15 min. This is an order of magnitude improvement over previous reports of electroplated microstructures. With an applied signal of 250 MHz, 2.6 V pp in our device, we separate a heterogeneous population with a purity of 97.8% in the low-lipid stream and 71.4% in the high-lipid stream. The high-lipid stream purity can be improved by adjusting the spacing of the array. This unique protocol for the rapid fabrication of 3D microstructures has enabled the creation of a non-invasive sorting tool for genetically engineered, lipid-producing organisms. The ability to screen organisms based on lipid content will alleviate one of the major bottlenecks in commercialization of microbial biofuels.

Harnessing biodiesel-producing microbes: from genetic engineering of lipase to metabolic engineering of fatty acid biosynthetic pathway.

PubMed

Yan, Jinyong; Yan, Yunjun; Madzak, Catherine; Han, Bingnan

2017-02-01

Microbial production routes, notably whole-cell lipase-mediated biotransformation and fatty-acids-derived biosynthesis, offer new opportunities for synthesizing biodiesel. They compare favorably to immobilized lipase and chemically catalyzed processes. Genetically modified whole-cell lipase-mediated in vitro route, together with in vivo and ex vivo microbial biosynthesis routes, constitutes emerging and rapidly developing research areas for effective production of biodiesel. This review presents recent advances in customizing microorganisms for producing biodiesel, via genetic engineering of lipases and metabolic engineering (including system regulation) of fatty-acids-derived pathways. Microbial hosts used include Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris and Aspergillus oryzae. These microbial cells can be genetically modified to produce lipases under different forms: intracellularly expressed, secreted or surface-displayed. They can be metabolically redesigned and systematically regulated to obtain balanced biodiesel-producing cells, as highlighted in this study. Such genetically or metabolically modified microbial cells can support not only in vitro biotransformation of various common oil feedstocks to biodiesel, but also de novo biosynthesis of biodiesel from glucose, glycerol or even cellulosic biomass. We believe that the genetically tractable oleaginous yeast Yarrowia lipolytica could be developed to an effective biodiesel-producing microbial cell factory. For this purpose, we propose several engineered pathways, based on lipase and wax ester synthase, in this promising oleaginous host.

Study of Holtermanniella wattica, Leucosporidium creatinivorum, Naganishia adeliensis, Solicoccozyma aeria, and Solicoccozyma terricola for their lipogenic aptitude from different carbon sources.

PubMed

Filippucci, Sara; Tasselli, Giorgia; Scardua, Alessandro; Di Mauro, Simone; Cramarossa, Maria Rita; Perini, Davide; Turchetti, Benedetta; Onofri, Andrea; Forti, Luca; Buzzini, Pietro

2016-01-01

The ability of some microorganisms to accumulate lipids is well known; however, only recently the number of studies on microbial lipid biosynthesis for obtaining oleochemical products, namely biofuels and some building blocks for chemistry, is rapidly and spectacularly increased. Since 1990s, some oleaginous yeasts were studied for their ability to accumulate lipids up to 60-70% of their dry weight. Due to the vast array of engineering techniques currently available, the recombinant DNA technology was the main approach followed so far for obtaining lipid-overproducing yeasts, mainly belonging to the Yarrowia lipolytica . However, an alternative approach can be offered by worldwide diversity as source of novel oleaginous yeasts. Lipogenic aptitude of a number of yeast strains has been reviewed, but many of these studies utilized a limited number of species and/or different culture conditions that make impossible the comparison of different results. Accordingly, the lipogenic aptitude inside the yeast world is still far from being fully explored, and finding new oleaginous yeast species can acquire a strategic importance. Holtermanniella wattica , Leucosporidium creatinivorum , Naganishia adeliensis , Solicoccozyma aeria, and Solicoccozyma terricola strains were selected as a result of a large-scale screening on 706 yeasts (both Ascomycota and Basidiomycota). Lipid yields and fatty acid profiles of selected strains were evaluated at 20 and 25 °C on glucose, and on glycerol, xylose, galactose, sucrose, maltose, and cellobiose. A variable fatty acid profile was observed in dependence of both temperature and different carbon sources. On the whole, L. creatinivorum exhibited the highest performances: total lipid yield (Y L ) >7 g/l on glucose and glycerol, % of intracellular lipids on cell biomass (Y L /DW) >70% at 20 °C on glucose, lipid coefficient (Y L /Glu) around 20% on glucose, and daily productivity (Y L /d) on glucose and sucrose >1.6 g/(l*d). This study

Population dynamics of hydrocarbon-oxidizing yeasts introduced into oil-contaminated soils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kulichevskaya, I.S.; Panikov, N.S.; Guzev, V.S.

A pure culture of the yeastlike fungus Candida lipolytica, able to actively degrade crude oil, was isolated. In preliminary trials, an optimal dose for its introduction was adjusted (10{sup 8} cells/g soil) to ensure its predominance in contaminated soil. Laboratory incubation experiments in which the population dynamics of the introduced species and indigenous soil bacteria and the dynamics of soil respiration activity were followed showed that active proliferation of the introduced species in soil is accompanied by its elimination as a result of grazing by microfauna. The most favorable conditions for the development of introduced yeasts were found to bemore » provided in gray and gray forest soil, whereas in soddy-podzolic soil, their growth and oil degradation were retarded. The obtained results indicate that introduction of the tested culture can significantly increase the rate of oil degradation. In uncontaminated soil, the introduced species is rapidly eliminated. 9 refs., 5 figs.« less

Diversity of spoilage fungi associated with various French dairy products.

PubMed

Garnier, Lucille; Valence, Florence; Pawtowski, Audrey; Auhustsinava-Galerne, Lizaveta; Frotté, Nicolas; Baroncelli, Riccardo; Deniel, Franck; Coton, Emmanuel; Mounier, Jérôme

2017-01-16

Yeasts and molds are responsible for dairy product spoilage, resulting in significant food waste and economic losses. Yet, few studies have investigated the diversity of spoilage fungi encountered in dairy products. In the present study, 175 isolates corresponding to 105 from various spoiled dairy products and 70 originating from dairy production environments, were identified using sequencing of the ITS region, the partial β-tubulin, calmodulin and/or EFα genes, and the D1-D2 domain of the 26S rRNA gene for filamentous fungi and yeasts, respectively. Among the 41 species found in spoiled products, Penicillium commune and Penicillium bialowiezense were the most common filamentous fungi, representing around 10% each of total isolates while Meyerozyma guilliermondii and Trichosporon asahii were the most common yeasts (4.8% each of total isolates). Several species (e.g. Penicillium antarcticum, Penicillium salamii and Cladosporium phyllophilum) were identified for the first time in dairy products or their environment. In addition, numerous species were identified in both spoiled products and their corresponding dairy production environment suggesting that the latter acts as a primary source of contamination. Secondly, the resistance to chemical preservatives (sodium benzoate, calcium propionate, potassium sorbate and natamycin) of 10 fungal isolates representative of the observed biodiversity was also evaluated. Independently of the fungal species, natamycin had the lowest minimum inhibitory concentration (expressed in gram of preservative/l), followed by potassium sorbate, sodium benzoate and calcium propionate. In the tested conditions, Cladosporium halotolerans and Didymella pinodella were the most sensitive fungi while Yarrowia lipolytica and Candida parapsilosis were the most resistant towards the tested preservatives. This study provides interesting information on the occurrence of fungal contaminants in dairy products and environments that may help developing

Novel efficient promoter of the mitochondrial porin, voltage-dependent anion channel (VDAC), in the genome of the Yarrowia lipolytica yeast.

PubMed

Kulanbaewa, F F; Sekova, V Yu; Isakova, E P; Deryabina, Y I; Nikolaev, A V

2016-09-01

This article presents the characteristics of the highly inducible promoter of the gene encoding the mitochondrial porin, the voltage-dependent anion channel (VDAC). This promoter is recommended for use in new genetic constructs both in basic research for assessing the adaptive strategy of lower eukaryotes under adverse conditions and in designing new highly competitive transformants producing economically important compounds (proteins, lipids, and organic acids) on its basis.

"Bligh and Dyer" and Folch Methods for Solid-Liquid-Liquid Extraction of Lipids from Microorganisms. Comprehension of Solvatation Mechanisms and towards Substitution with Alternative Solvents.

PubMed

Breil, Cassandra; Abert Vian, Maryline; Zemb, Thomas; Kunz, Werner; Chemat, Farid

2017-03-27

Bligh and Dyer (B & D) or Folch procedures for the extraction and separation of lipids from microorganisms and biological tissues using chloroform/methanol/water have been used tens of thousands of times and are "gold standards" for the analysis of extracted lipids. Based on the Conductor-like Screening MOdel for realistic Solvatation (COSMO-RS), we select ethanol and ethyl acetate as being potentially suitable for the substitution of methanol and chloroform. We confirm this by performing solid-liquid extraction of yeast ( Yarrowia lipolytica IFP29 ) and subsequent liquid-liquid partition-the two steps of routine extraction. For this purpose, we consider similar points in the ternary phase diagrams of water/methanol/chloroform and water/ethanol/ethyl acetate, both in the monophasic mixtures and in the liquid-liquid miscibility gap. Based on high performance thin-layer chromatography (HPTLC) to obtain the distribution of lipids classes, and gas chromatography coupled with a flame ionisation detector (GC/FID) to obtain fatty acid profiles, this greener solvents pair is found to be almost as effective as the classic methanol-chloroform couple in terms of efficiency and selectivity of lipids and non-lipid material. Moreover, using these bio-sourced solvents as an alternative system is shown to be as effective as the classical system in terms of the yield of lipids extracted from microorganism tissues, independently of their apparent hydrophilicity.

“Bligh and Dyer” and Folch Methods for Solid–Liquid–Liquid Extraction of Lipids from Microorganisms. Comprehension of Solvatation Mechanisms and towards Substitution with Alternative Solvents

PubMed Central

Breil, Cassandra; Abert Vian, Maryline; Zemb, Thomas; Kunz, Werner; Chemat, Farid

2017-01-01

Bligh and Dyer (B & D) or Folch procedures for the extraction and separation of lipids from microorganisms and biological tissues using chloroform/methanol/water have been used tens of thousands of times and are “gold standards” for the analysis of extracted lipids. Based on the Conductor-like Screening MOdel for realistic Solvatation (COSMO-RS), we select ethanol and ethyl acetate as being potentially suitable for the substitution of methanol and chloroform. We confirm this by performing solid–liquid extraction of yeast (Yarrowia lipolytica IFP29) and subsequent liquid–liquid partition—the two steps of routine extraction. For this purpose, we consider similar points in the ternary phase diagrams of water/methanol/chloroform and water/ethanol/ethyl acetate, both in the monophasic mixtures and in the liquid–liquid miscibility gap. Based on high performance thin-layer chromatography (HPTLC) to obtain the distribution of lipids classes, and gas chromatography coupled with a flame ionisation detector (GC/FID) to obtain fatty acid profiles, this greener solvents pair is found to be almost as effective as the classic methanol–chloroform couple in terms of efficiency and selectivity of lipids and non-lipid material. Moreover, using these bio-sourced solvents as an alternative system is shown to be as effective as the classical system in terms of the yield of lipids extracted from microorganism tissues, independently of their apparent hydrophilicity. PMID:28346372

Application of metabolic controls for the maximization of lipid production in semicontinuous fermentation.

PubMed

Xu, Jingyang; Liu, Nian; Qiao, Kangjian; Vogg, Sebastian; Stephanopoulos, Gregory

2017-07-03

Acetic acid can be generated through syngas fermentation, lignocellulosic biomass degradation, and organic waste anaerobic digestion. Microbial conversion of acetate into triacylglycerols for biofuel production has many advantages, including low-cost or even negative-cost feedstock and environmental benefits. The main issue stems from the dilute nature of acetate produced in such systems, which is costly to be processed on an industrial scale. To tackle this problem, we established an efficient bioprocess for converting dilute acetate into lipids, using the oleaginous yeast Yarrowia lipolytica in a semicontinuous system. The implemented design used low-strength acetic acid in both salt and acid forms as carbon substrate and a cross-filtration module for cell recycling. Feed controls for acetic acid and nitrogen based on metabolic models and online measurement of the respiratory quotient were used. The optimized process was able to sustain high-density cell culture using acetic acid of only 3% and achieved a lipid titer, yield, and productivity of 115 g/L, 0.16 g/g, and 0.8 g⋅L -1 ⋅h -1 , respectively. No carbon substrate was detected in the effluent stream, indicating complete utilization of acetate. These results represent a more than twofold increase in lipid production metrics compared with the current best-performing results using concentrated acetic acid as carbon feed.

A surprisingly large RNase P RNA in Candida glabrata

PubMed Central

KACHOURI, RYM; STRIBINSKIS, VILIUS; ZHU, YANGLONG; RAMOS, KENNETH S.; WESTHOF, ERIC; LI, YONG

2005-01-01

We have found an extremely large ribonuclease P (RNase P) RNA (RPR1) in the human pathogen Candida glabrata and verified that this molecule is expressed and present in the active enzyme complex of this hemiascomycete yeast. A structural alignment of the C. glabrata sequence with 36 other hemiascomycete RNase P RNAs (abbreviated as P RNAs) allows us to characterize the types of insertions. In addition, 15 P RNA sequences were newly characterized by searching in the recently sequenced genomes Candida albicans, C. glabrata, Debaryomyces hansenii, Eremothecium gossypii, Kluyveromyces lactis, Kluyveromyces waltii, Naumovia castellii, Saccharomyces kudriavzevii, Saccharomyces mikatae, and Yarrowia lipolytica; and by PCR amplification for other Candida species (Candida guilliermondii, Candida krusei, Candida parapsilosis, Candida stellatoidea, and Candida tropicalis). The phylogenetic comparative analysis identifies a hemiascomycete secondary structure consensus that presents a conserved core in all species with variable insertions or deletions. The most significant variability is found in C. glabrata P RNA in which three insertions exceeding in total 700 nt are present in the Specificity domain. This P RNA is more than twice the length of any other homologous P RNAs known in the three domains of life and is eight times the size of the smallest. RNase P RNA, therefore, represents one of the most diversified noncoding RNAs in terms of size variation and structural diversity. PMID:15987816

Protease and lipase activities of fungal and bacterial strains derived from an artisanal raw ewe's milk cheese.

PubMed

Ozturkoglu-Budak, Sebnem; Wiebenga, Ad; Bron, Peter A; de Vries, Ronald P

2016-11-21

We previously identified the microbiota present during cheese ripening and observed high protease and lipase activity in Divle Cave cheese. To determine the contribution of individual isolates to enzyme activities, we investigated a range of species representing this microbiota for their proteolytic and lipolytic ability. In total, 17 fungal, 5 yeast and 18 bacterial strains, previously isolated from Divle Cave cheese, were assessed. Qualitative protease and lipase activities were performed on skim-milk agar and spirit-blue lipase agar, respectively, and resulted in a selection of strains for quantitative assays. For the quantitative assays, the strains were grown on minimal medium containing irradiated Divle Cave cheese, obtained from the first day of ripening. Out of 16 selected filamentous fungi, Penicillium brevicompactum, Penicillium cavernicola and Penicillium olsonii showed the highest protease activity, while Mucor racemosus was the best lipase producer. Yarrowia lipolytica was the best performing yeast with respect to protease and lipase activity. From the 18 bacterial strains, 14 and 11 strains, respectively showed protease and lipase activity in agar plates. Micrococcus luteus, Bacillus stratosphericus, Brevibacterium antiquum, Psychrobacter glacincola and Pseudomonas proteolytica displayed the highest protease and lipase activity. The proteases of yeast and filamentous fungi were identified as mainly aspartic protease by specific inhibition with Pepstatin A, whereas inhibition by PMSF (phenylmethylsulfonyl fluoride) indicated that most bacterial enzymes belong to serine type protease. Our results demonstrate that aspartic proteases, which usually have high milk clotting activity, are predominantly derived from fungal strains, and therefore fungal enzymes appear to be more suitable for use in the cheese industry. Microbial enzymes studied in this research might be alternatives for rennin (chymosin) from animal source because of their low cost and stable

Role of charge screening and delocalization for lipophilic cation permeability of model and mitochondrial membranes.

PubMed

Trendeleva, Tatiana A; Sukhanova, Evgenia I; Rogov, Anton G; Zvyagilskaya, Renata A; Seveina, Inna I; Ilyasova, Tatiana M; Cherepanov, Dmitry A; Skulachev, Vladimir P

2013-09-01

The effects of the mitochondria-targeted lipophilic cation dodecyltriphenylphosphonium (C12TPP, the charge is delocalized and screened by bulky hydrophobic residues) and those of lipophilic cations decyltriethylammonium bromide and cetyltrimethylammonium bromide (C10TEA and C16TMA, the charges are localized and screened by less bulky residues) on bilayer planar phospholipid membranes and tightly-coupled mitochondria from the yeast Yarrowia lipolytica have been compared. In planar membranes, C12TPP was found to generate a diffusion potential as if it easily penetrates these membranes. In the presence of palmitate, C12TPP induced H(+) permeability like plastoquinonyl decyltriphenilphosphonium that facilitates transfer of fatty acid anions (Severin et al., PNAS, 2010, 107, 663-668). C12TPP was shown to stimulate State 4 respiration of mitochondria and caused a mitochondrial membrane depolarization with a half-maximal effect at 6μM. Besides, C12TPP profoundly potentiated the uncoupling effect of endogenous or added fatty acids. C10TEA and C16TMA inhibited State 4 respiration and decreased the membrane potential, though at much higher concentrations than C12TPP, and they did not promote the uncoupling action of fatty acids. These relationships were modeled by molecular dynamics. They can be explained by different membrane permeabilities for studied cations, which in turn are due to different availabilities of the positive charge in these cations to water dipoles. Copyright © 2012 Elsevier B.V. All rights reserved.

Organic Acid Production by Filamentous Fungi

DOE Office of Scientific and Technical Information (OSTI.GOV)

Magnuson, Jon K.; Lasure, Linda L.

Many of the commercial production processes for organic acids are excellent examples of fungal biotechnology. However, unlike penicillin, the organic acids have had a less visible impact on human well-being. Indeed, organic acid fermentations are often not even identified as fungal bioprocesses, having been overshadowed by the successful deployment of the β-lactam processes. Yet, in terms of productivity, fungal organic acid processes may be the best examples of all. For example, commercial processes using Aspergillus niger in aerated stirred-tank-reactors can convert glucose to citric acid with greater than 80% efficiency and at final concentrations in hundreds of grams per liter.more » Surprisingly, this phenomenal productivity has been the object of relatively few research programs. Perhaps a greater understanding of this extraordinary capacity of filamentous fungi to produce organic acids in high concentrations will allow greater exploitation of these organisms via application of new knowledge in this era of genomics-based biotechnology. In this chapter, we will explore the biochemistry and modern genetic aspects of the current and potential commercial processes for making organic acids. The organisms involved, with a few exceptions, are filamentous fungi, and this review is limited to that group. Although yeasts including Saccharomyces cerevisiae, species of Rhodotorula, Pichia, and Hansenula are important organisms in fungal biotechnology, they have not been significant for commercial organic acid production, with one exception. The yeast, Yarrowia lipolytica, and related yeast species, may be in use commercially to produce citric acid (Lopez-Garcia, 2002). Furthermore, in the near future engineered yeasts may provide new commercial processes to make lactic acid (Porro, Bianchi, Ranzi, Frontali, Vai, Winkler, & Alberghina, 2002). This chapter is divided into two parts. The first contains a review of the commercial aspects of current and potential large

Biodegradation of micropollutant naproxen with a selected fungal strain and identification of metabolites.

PubMed

Aracagök, Y Doruk; Göker, Hakan; Cihangir, Nilüfer

2017-05-01

Pharmaceuticals are widely used for treating human and animal diseases. Naproxen [(S) 6-methoxy-α-methyl-2-naphthalene acetic acid] and its sodium salt are members of the α-arylpropionic acid group of nonsteroidal anti-inflammatory drugs. Due to excessive usage of naproxen, this drug has been determined even in drinking water. In this study, four fungal strains Phanerochaete chrysosporium, Funalia trogii, Aspergillus niger, and Yarrowia lipolytica were investigated in terms of naproxen removal abilities. According to LC/MS data, A. niger was found the most efficient strain with 98% removal rate. Two main by-products of fungal transformation, O-desmethylnaproxen and 7-hydroxynaproxen, were identified by using LC/MS, 1HNMR, and 13CNMR. Our results showed that O-demethylation and hydroxylation of naproxen is catalyzed by cytochrome P450 enzyme system.

Waste Soybean Oil and Corn Steep Liquor as Economic Substrates for Bioemulsifier and Biodiesel Production by Candida lipolytica UCP 0998

PubMed Central

Souza, Adriana Ferreira; Rodriguez, Dayana M.; Ribeaux, Daylin R.; Luna, Marcos A. C.; Lima e Silva, Thayse A.; Andrade, Rosileide F. Silva; Gusmão, Norma B.; Campos-Takaki, Galba M.

2016-01-01

Almost all oleaginous microorganisms are available for biodiesel production, and for the mechanism of oil accumulation, which is what makes a microbial approach economically competitive. This study investigated the potential that the yeast Candida lipolytica UCP0988, in an anamorphous state, has to produce simultaneously a bioemulsifier and to accumulate lipids using inexpensive and alternative substrates. Cultivation was carried out using waste soybean oil and corn steep liquor in accordance with 22 experimental designs with 1% inoculums (107 cells/mL). The bioemulsifier was produced in the cell-free metabolic liquid in the late exponential phase (96 h), at Assay 4 (corn steep liquor 5% and waste soybean oil 8%), with 6.704 UEA, IE24 of 96.66%, and showed an anionic profile. The emulsion formed consisted of compact small and stable droplets (size 0.2–5 µm), stable at all temperatures, at pH 2 and 4, and 2% salinity, and showed an ability to remove 93.74% of diesel oil from sand. The displacement oil (ODA) showed 45.34 cm2 of dispersion (central point of the factorial design). The biomass obtained from Assay 4 was able to accumulate lipids of 0.425 g/g biomass (corresponding to 42.5%), which consisted of Palmitic acid (28.4%), Stearic acid (7.7%), Oleic acid (42.8%), Linoleic acid (19.0%), and γ-Linolenic acid (2.1%). The results showed the ability of C. lipopytica to produce both bioemulsifier and biodiesel using the metabolic conversion of waste soybean oil and corn steep liquor, which are economic renewable sources. PMID:27669227

Unravel lipid accumulation mechanism in oleaginous yeast through single cell systems biology study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xie, Xiaoliang; Ding, Shiyou

Searching for alternative and clean energy is one of the most important tasks today. Our research aimed at finding the best living condition for certain types of oleaginous yeasts for efficient lipid production. We found that R. glutinis yeast cells has great variability in lipid production among cells while Y. lipolytica cells has similar oil production ability. We found some individual cells shows much higher level of oil production. In order to further study these cases, we employed a label-free chemical sensitive microscopy method call stimulated Raman scattering (SRS). With SRS, we could measure the lipid content in each cell.more » We combined SRS microscopy with microfluidic device so that we can isolate cells with high fat content. We also developed SRS imaging technique that has higher imaging speed, which is highly desirable for high throughput cell screening and sorting. Since these cells has similar genome, it must be the transcriptome caused their difference in oil production. We developed a single cell transcriptome sequencing method to study which genes are responsible for elevated oil production. These methods that are developed for this project can easily be applied for many other areas of research. For example, the single transcriptome can be used to study the transcriptomes of other cell types. The high-speed SRS microscopy techniques can be used to speed up chemical imaging for lablefree histology or imaging distribution of chemicals in tissues of live mice or in humans. The developed microfluidic platform can be used to sort other type of cells, e.g., white blood cells for diagnosis of cancer or other blood diseases.« less

Human recombinant lysosomal enzymes produced in microorganisms.

PubMed

Espejo-Mojica, Ángela J; Alméciga-Díaz, Carlos J; Rodríguez, Alexander; Mosquera, Ángela; Díaz, Dennis; Beltrán, Laura; Díaz, Sergio; Pimentel, Natalia; Moreno, Jefferson; Sánchez, Jhonnathan; Sánchez, Oscar F; Córdoba, Henry; Poutou-Piñales, Raúl A; Barrera, Luis A

2015-01-01

Lysosomal storage diseases (LSDs) are caused by accumulation of partially degraded substrates within the lysosome, as a result of a function loss of a lysosomal protein. Recombinant lysosomal proteins are usually produced in mammalian cells, based on their capacity to carry out post-translational modifications similar to those observed in human native proteins. However, during the last years, a growing number of studies have shown the possibility to produce active forms of lysosomal proteins in other expression systems, such as plants and microorganisms. In this paper, we review the production and characterization of human lysosomal proteins, deficient in several LSDs, which have been produced in microorganisms. For this purpose, Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, and Ogataea minuta have been used as expression systems. The recombinant lysosomal proteins expressed in these hosts have shown similar substrate specificities, and temperature and pH stability profiles to those produced in mammalian cells. In addition, pre-clinical results have shown that recombinant lysosomal enzymes produced in microorganisms can be taken-up by cells and reduce the substrate accumulated within the lysosome. Recently, metabolic engineering in yeasts has allowed the production of lysosomal enzymes with tailored N-glycosylations, while progresses in E. coli N-glycosylations offer a potential platform to improve the production of these recombinant lysosomal enzymes. In summary, microorganisms represent convenient platform for the production of recombinant lysosomal proteins for biochemical and physicochemical characterization, as well as for the development of ERT for LSD. Copyright © 2015 Elsevier Inc. All rights reserved.

Microbial diversity and dynamics during the production of May bryndza cheese.

PubMed

Pangallo, Domenico; Saková, Nikoleta; Koreňová, Janka; Puškárová, Andrea; Kraková, Lucia; Valík, Lubomír; Kuchta, Tomáš

2014-01-17

paradoxus. The diversity of yeasts and fungi encompassed Alternaria alternata, "Ascomycete sp.", Aspergillus fumigatus, Beauveria brongniartii, Candida xylopsoci, C. inconspicua, Cladosporium cladosporioides, Debaromyces hansenii, Fomes fomentarius, Galactomyces candidus, Gymnoascus reesii, Chaetomium globosum, Kluyveromyces marxianus, Metarhizium anisopliae, Penicillium aurantiogriseum, P. camemberti, P. freii, P. polonicum, P. viridicatum, Pichia kudriavzevii, Sordaria alcina, Trichosporon lactis and Yarrowia lipolytica. © 2013.

Integrated bioprocess for conversion of gaseous substrates to liquids

PubMed Central

Hu, Peng; Chakraborty, Sagar; Kumar, Amit; Woolston, Benjamin; Liu, Hongjuan; Emerson, David; Stephanopoulos, Gregory

2016-01-01

In the quest for inexpensive feedstocks for the cost-effective production of liquid fuels, we have examined gaseous substrates that could be made available at low cost and sufficiently large scale for industrial fuel production. Here we introduce a new bioconversion scheme that effectively converts syngas, generated from gasification of coal, natural gas, or biomass, into lipids that can be used for biodiesel production. We present an integrated conversion method comprising a two-stage system. In the first stage, an anaerobic bioreactor converts mixtures of gases of CO2 and CO or H2 to acetic acid, using the anaerobic acetogen Moorella thermoacetica. The acetic acid product is fed as a substrate to a second bioreactor, where it is converted aerobically into lipids by an engineered oleaginous yeast, Yarrowia lipolytica. We first describe the process carried out in each reactor and then present an integrated system that produces microbial oil, using synthesis gas as input. The integrated continuous bench-scale reactor system produced 18 g/L of C16-C18 triacylglycerides directly from synthesis gas, with an overall productivity of 0.19 g⋅L−1⋅h−1 and a lipid content of 36%. Although suboptimal relative to the performance of the individual reactor components, the presented integrated system demonstrates the feasibility of substantial net fixation of carbon dioxide and conversion of gaseous feedstocks to lipids for biodiesel production. The system can be further optimized to approach the performance of its individual units so that it can be used for the economical conversion of waste gases from steel mills to valuable liquid fuels for transportation. PMID:26951649

Yeast Based Sensors

NASA Astrophysics Data System (ADS)

Shimomura-Shimizu, Mifumi; Karube, Isao

Since the first microbial cell sensor was studied by Karube et al. in 1977, many types of yeast based sensors have been developed as analytical tools. Yeasts are known as facultative anaerobes. Facultative anaerobes can survive in both aerobic and anaerobic conditions. The yeast based sensor consisted of a DO electrode and an immobilized omnivorous yeast. In yeast based sensor development, many kinds of yeast have been employed by applying their characteristics to adapt to the analyte. For example, Trichosporon cutaneum was used to estimate organic pollution in industrial wastewater. Yeast based sensors are suitable for online control of biochemical processes and for environmental monitoring. In this review, principles and applications of yeast based sensors are summarized.

Extracellular Polysaccharides Produced by Yeasts and Yeast-Like Fungi

NASA Astrophysics Data System (ADS)

van Bogaert, Inge N. A.; de Maeseneire, Sofie L.; Vandamme, Erick J.

Several yeasts and yeast-like fungi are known to produce extracellular polysaccharides. Most of these contain D-mannose, either alone or in combination with other sugars or phosphate. A large chemical and structural variability is found between yeast species and even among different strains. The types of polymers that are synthesized can be chemically characterized as mannans, glucans, phosphoman-nans, galactomannans, glucomannans and glucuronoxylomannans. Despite these differences, almost all of the yeast exopolysaccharides display some sort of biological activity. Some of them have already applications in chemistry, pharmacy, cosmetics or as probiotic. Furthermore, some yeast exopolysaccharides, such as pullulan, exhibit specific physico-chemical and rheological properties, making them useful in a wide range of technical applications. A survey is given here of the production, the characteristics and the application potential of currently well studied yeast extracellular polysaccharides.

Yeast Infection (Vaginal)

MedlinePlus

Yeast infection (vaginal) Overview A vaginal yeast infection is a fungal infection that causes irritation, discharge and intense itchiness ... symptoms Causes The fungus candida causes a vaginal yeast infection. Your vagina naturally contains a balanced mix of yeast, including ...

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

PubMed

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.

A novel phagocytic receptor (CgNimC) from Pacific oyster Crassostrea gigas with lipopolysaccharide and gram-negative bacteria binding activity.

PubMed

Wang, Weilin; Liu, Rui; Zhang, Tao; Zhang, Ran; Song, Xuan; Wang, Lingling; Song, Linsheng

2015-03-01

Phagocytosis is an evolutionarily conserved process to ingest the invading microbes and apoptotic or necrotic corpses, playing vital roles in defensing invaders and maintenance of normal physiological conditions. In the present study, a new Nimrod family phagocytic receptor with three EGF-like domains was identified in Pacific oyster Crassostrea gigas (designated CgNimC). CgNimC shared homology with other identified multiple EGF-like domain containing proteins. The mRNA transcripts of CgNimC were mainly distributed in mantle and hemocytes. Its relative expression level in hemocytes was significantly (P < 0.01) up-regulated after the injection of bacteria Vibrio anguillarum. Different to the NimC in Drosophila and Anopheles gambiae, the recombinant protein of CgNimC (rCgNimC) could bind directly to two gram-negative bacteria V. anguillarum and Vibrio splendidus, but not to gram-positive bacteria Staphylococci aureus, Micrococcus luteus or fungi Yarrowia lipolytica and Pichia pastoris. The affinity of rCgNimC toward M. luteus and Y. lipolytica was enhanced when the microorganisms were pre-incubated with the cell free hemolymph. rCgNimC exhibited higher affinity to lipopolysaccharide (LPS) and relatively lower affinity to peptidoglycan (PGN), while no affinity to glucan (GLU). After the CgNimC receptor was blocked by anti-rCgNimC antibody in vitro, the phagocytic rate of hemocytes toward two gram-negative bacteria V. anguillarum and V. splendidus was reduced significantly (P < 0.05), but no significant change of phagocytic rate was observed toward M. luteus and Y. lipolytica. All these results implied that CgNimC, with significant binding capability to LPS and gram-negative bacteria, was a novel phagocytic receptor involved in immune response of Pacific oyster. Further, it was speculated that receptors of Nimrod family might function as a phagocytic receptor to recognize PAMPs on the invaders and its recognition could be promoted by opsonization of molecules in

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

PubMed Central

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 density and species composition. When yeast densities were compared in pieces of the same fruits assigned to different treatments, fruits that developed low yeast densities in the absence of flies developed significantly higher yeast densities when exposed to larvae. Across all of the fruits, larvae regulated yeast densities within narrow limits, as compared to a much wider range of yeast densities that developed in pieces of the same fruits not exposed to flies. Larvae also affected yeast species composition, dramatically reducing species diversity across fruits, reducing variation in yeast communities from one fruit to the next (beta diversity), and encouraging the consistent development of a yeast community composed of three species of yeast (Candida californica, C. zemplinina, and Pichia kluvyeri), all of which were palatable to larvae. Larvae excreted viable cells of these three yeast species in their fecal pools, and discouraged the growth of filamentous fungi, processes which may have contributed to their effects on the yeast communities in banana fruits. These and other findings suggest that D. melanogaster adults and their larval offspring together engage in ‘niche construction’, facilitating a predictable microbial environment in the fruit substrates in which the larvae live and develop. PMID:22860093

A novel junctional adhesion molecule A (CgJAM-A-L) from oyster (Crassostrea gigas) functions as pattern recognition receptor and opsonin.

PubMed

Liu, Conghui; Wang, Mengqiang; Jiang, Shuai; Wang, Lingling; Chen, Hao; Liu, Zhaoqun; Qiu, Limei; Song, Linsheng

2016-02-01

Junctional adhesion molecule (JAM), a subfamily of immunoglobulin superfamily (IgSF) with a couple of immunoglobulin domains, can act as regulator in homeostasis and inflammation of vertebrates. In the present study, a structural homolog of JAM-A (designated CgJAM-A-L) was screened out from oyster, Crassostrea gigas, through a search of JAM-A D1 domain (N-terminal Ig domain in JAM-A). The cDNA of CgJAM-A-L was of 1188 bp encoding a predicted polypeptide of 395 amino acids. The immunoreactive area of CgJAM-A-L mainly distributed over the plasma membrane of hemocytes. After Vibro splendidus or tumor necrosis factor (CgTNF-1) stimulation, the mRNA transcripts of CgJAM-A-L in hemocytes increased significantly by 4.46-fold and 9.00-fold (p < 0.01) of those in control group, respectively. The recombinant CgJAM-A-L protein (rCgJAM-A-L) could bind multiple PAMPs including lipopolysaccharides (LPS), peptidoglycan (PGN), lipoteichoic acid (LTA), mannose (MAN), β-glucan (GLU) and poly(I:C), and various microorganisms including Micrococcus luteus, Staphylococcus aureus, Escherichia coli, Vibro anguillarum, V. splendidus, Pastoris pastoris and Yarrowia lipolytica. The phagocytic rates of oyster hemocytes towards Gram-negative bacteria V. anguillarum and yeast P. pastoris were significantly enhanced after the incubation of rCgJAM-A-L, and even increased more significantly after the pre-incubation of rCgJAM-A-L with microbes (p < 0.01). The results collectively indicated that CgJAM-A-L functioned as an important pattern recognition receptor (PRR) and opsonin in the immune defense against invading pathogen in oyster. Moreover, as the most primitive specie with homolog of JAMs, the information of CgJAM-A-L in oyster would provide useful clues for the evolutionary study of JAMs and immunoglobulins. Copyright © 2015 Elsevier Ltd. All rights reserved.

Molecular and biochemical characterisation of two aspartic proteinases TcAP1 and TcAP2 from Theobroma cacao seeds.

PubMed

Laloi, Maryse; McCarthy, James; Morandi, Olivia; Gysler, Christof; Bucheli, Peter

2002-09-01

Aspartic proteinase (EC 3.4.23) activity plays a pivotal role in the degradation of Theobroma cacao L. seed proteins during the fermentation step of cacao bean processing. Therefore, this enzyme is believed to be critical for the formation of the peptide and amino acid cocoa flavor precursors that occurs during fermentation. Using cDNA cloning and northern blot analysis, we show here that there are at least two distinct aspartic proteinase genes ( TcAP1 and TcAP2) expressed during cacao seed development. Both genes are expressed early during seed development and their mRNA levels decrease towards the end of seed maturation. TcAP2 is expressed at a much higher level than TcAP1, although the expression of TcAP1 increases slightly during germination. The proteins encoded by TcAP1 and TcAP2 are relatively different from each other (73% identity). This, and the fact that the two corresponding genes have different expression patterns, suggests that the TcAP1 and TcAP2 proteins may have different functions in the maturing seeds and during germination. Because the TcAP2 gene is expressed at a much higher level during seed development than TcAP1, it is likely that the TcAP2 protein is primarily responsible for the majority of the industrially important protein hydrolysis that occurs during cacao bean fermentation. Finally, TcAP2 has been functionally expressed in the yeast Yarrowia lipolytica. The secreted recombinant protein is able to hydrolyse bovine haemoglobin at acidic pH and is sensitive to pepstatin A, confirming that TcAP2 encodes an aspartic proteinase, and strongly suggests that this gene encodes the well-characterized aspartic proteinase of mature cacao seeds.

Synthesis, structural characterization and photoluminescence properties of a novel La(III) complex

NASA Astrophysics Data System (ADS)

Köse, Muhammet; Ceyhan, Gökhan; Atcı, Emine; McKee, Vickie; Tümer, Mehmet

2015-05-01

In this study, a novel La(III) complex [La(H2L)2(NO3)3(MeOH)] of a Schiff base ligand was synthesized and characterized by spectroscopic and analytical methods. Single crystals of the complex suitable for X-ray diffraction study were obtained by slow diffusion of diethyl ether into a MeOH solution of the complex which was found to crystallise as [La(H2L)2(NO3)3(MeOH)]ṡ2MeOHṡH2O. The structure was solved in monoclinic crystal system, P21/n space group with unit cell parameters a = 10.5641(11), b = 12.6661(16), c = 16.0022(17) Å, α = 67.364(2), β = 83.794(2)°, γ = 70.541(2)°, V = 1862.9(4) Å3 and Z = 2 with R final value of 0.526. In the complex, the La(III) ion is ten-coordinated by O atoms, five of which come from three nitrate ions, four from the two Schiff base ligands and one from MeOH oxygen atom. The Schiff base ligands in the structure are in a zwitter ion form with the phenolic H transferred to the imine N atom. Thermal properties of the La(III) complex were examined by thermogravimetric analysis and the complex was found to be thermally stable up to 310 °C. The Schiff base ligand and its La(II) complex were screened for their in vitro antimicrobial activity against Bacillus megaterium, Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus (Gram positive bacteria), Klebsiella pneumonia, Escherichia coli, Enterobacter aerogenes, Pseudomonas aeruginosa (Gram negative bacteria), Candida albicans,Yarrowia lipolytica (fungus) and Saccharomyces cerevisiae (yeast). The complex shows more antimicrobial activity than the free ligand.

Prions in Yeast

PubMed Central

Liebman, Susan W.; Chernoff, Yury O.

2012-01-01

The concept of a prion as an infectious self-propagating protein isoform was initially proposed to explain certain mammalian diseases. It is now clear that yeast also has heritable elements transmitted via protein. Indeed, the “protein only” model of prion transmission was first proven using a yeast prion. Typically, known prions are ordered cross-β aggregates (amyloids). Recently, there has been an explosion in the number of recognized prions in yeast. Yeast continues to lead the way in understanding cellular control of prion propagation, prion structure, mechanisms of de novo prion formation, specificity of prion transmission, and the biological roles of prions. This review summarizes what has been learned from yeast prions. PMID:22879407

Yeast for virus research

PubMed Central

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

L-arabinose fermenting yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Min; Singh, Arjun; Suominen, Pirkko

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.

L-arabinose fermenting yeast

DOEpatents

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

2014-09-23

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.

Prevention of Yeast Spoilage in Feed and Food by the Yeast Mycocin HMK

PubMed Central

Lowes, K. F.; Shearman, C. A.; Payne, J.; MacKenzie, D.; Archer, D. B.; Merry, R. J.; Gasson, M. J.

2000-01-01

The yeast Williopsis mrakii produces a mycocin or yeast killer toxin designated HMK; this toxin exhibits high thermal stability, high pH stability, and a broad spectrum of activity against other yeasts. We describe construction of a synthetic gene for mycocin HMK and heterologous expression of this toxin in Aspergillus niger. Mycocin HMK was fused to a glucoamylase protein carrier, which resulted in secretion of biologically active mycocin into the culture media. A partial purification protocol was developed, and a comparison with native W. mrakii mycocin showed that the heterologously expressed mycocin had similar physiological properties and an almost identical spectrum of biological activity against a number of yeasts isolated from silage and yoghurt. Two food and feed production systems prone to yeast spoilage were used as models to assess the ability of mycocin HMK to act as a biocontrol agent. The onset of aerobic spoilage in mature maize silage was delayed by application of A. niger mycocin HMK on opening because the toxin inhibited growth of the indigenous spoilage yeasts. This helped maintain both higher lactic acid levels and a lower pH. In yoghurt spiked with dairy spoilage yeasts, A. niger mycocin HMK was active at all of the storage temperatures tested at which yeast growth occurred, and there was no resurgence of resistant yeasts. The higher the yeast growth rate, the more effective the killing action of the mycocin. Thus, mycocin HMK has potential applications in controlling both silage spoilage and yoghurt spoilage caused by yeasts. PMID:10698773

Effect of wine yeast monoculture practice on the biodiversity of non-Saccharomyces yeasts.

PubMed

Ganga, M A; Martínez, C

2004-01-01

The objective of this work was to study the effect of the use of Saccharomyces cerevisiae monocultures over the biodiversity of non-Saccharomyces yeasts in wine-producing areas in Chile. Microvinifications were carried out with grape musts of two areas. In one of them, the fermentation is carried out mainly in a spontaneous manner, whereas in the other the musts are inoculated with commercial yeasts. The isolated yeasts were identified by the internal transcribed (ITS)/restriction fragment length polymorphism technique. In the industrial production area less variability of yeast genera was observed as compared with the traditional area, an observation that is greatest at the end of the fermentation. Furthermore, a study of the production of extracellular enzymes was done. The majority of the yeasts showed at least one of the activities assayed with the exception of beta-glycosidase. The results suggest that in the industrialized area the diversity of yeasts is less in the traditional area. Likewise, the potentiality of the non-Saccharomyces yeasts as enzyme producers with industrial interest has been confirmed. This study shows the negative effect of the use of monocultures over the biodiversity of yeasts in wine-producing regions.

Thermosyntropha lipolytica gen. nov., sp. nov., a lipolytic, anaerobic, alkalitolerant, thermophilic bacterium utilizing short- and long-chain fatty acids in syntrophic coculture with a methanogenic archaeum.

PubMed

Svetlitshnyi, V; Rainey, F; Wiegel, J

1996-10-01

Three strains of an anaerobic thermophilic organoheterotrophic lipolytic alkalitolerant bacterium, Thermosyntropha lipolytica gen. nov., sp. nov. (type strain JW/VS-265T; DSM 11003), were isolated from alkaline hot springs of Lake Bogoria (Kenya). The cells were nonmotile, non-spore forming, straight or slightly curved rods. At 60 degrees C the pH range for growth determined at 25 degrees C [pH25 degrees C] was 7.15 to 9.5, with an optimum between 8.1 and 8.9 (pH60 degrees C of 7.6 and 8.1). At a pH25 degrees C of 8.5 the temperature range for growth was from 52 to 70 degrees C, with an optimum between 60 and 66 degrees C. The shortest doubling time was around 1 h. In pure culture the bacterium grew in a mineral base medium supplemented with yeast extract, tryptone, Casamino Acids, betaine, and crotonate as carbon sources, producing acetate as a major product and constitutively a lipase. During growth in the presence of olive oil, free long-chain fatty acids were accumulated in the medium but the pure culture could not utilize olive oil, triacylglycerols, short- and long-chain fatty acids, and glycerol for growth. In syntrophic coculture (Methanobacterium strain JW/VS-M29) the lipolytic bacteria grew on triacylglycerols and linear saturated and unsaturated fatty acids with 4 to 18 carbon atoms, but glycerol was not utilized. Fatty acids with even numbers of carbon atoms were degraded to acetate and methane, while from odd-numbered fatty acids 1 mol of propionate per mol of fatty acid was additionally formed. 16S rDNA sequence analysis identified Syntrophospora and Syntrophomonas spp. as closest phylogenetic neighbors.

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

PubMed

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.

L-arabinose fermenting yeast

DOEpatents

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##

Nitrile Metabolizing Yeasts

NASA Astrophysics Data System (ADS)

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

Genome dynamics and evolution in yeasts: A long-term yeast-bacteria competition experiment

PubMed Central

Katz, Michael; Knecht, Wolfgang; Compagno, Concetta; Piškur, Jure

2018-01-01

There is an enormous genetic diversity evident in modern yeasts, but our understanding of the ecological basis of such diversifications in nature remains at best fragmented so far. Here we report a long-term experiment mimicking a primordial competitive environment, in which yeast and bacteria co-exist and compete against each other. Eighteen yeasts covering a wide phylogenetic background spanning approximately 250 million years of evolutionary history were used to establish independent evolution lines for at most 130 passages. Our collection of hundreds of modified strains generated through such a rare two-species cross-kingdom competition experiment re-created the appearance of large-scale genomic rearrangements and altered phenotypes important in the diversification history of yeasts. At the same time, the methodology employed in this evolutionary study would also be a non-gene-technological method of reprogramming yeast genomes and then selecting yeast strains with desired traits. Cross-kingdom competition may therefore be a method of significant value to generate industrially useful yeast strains with new metabolic traits. PMID:29624585

Characterization of self-generated variants in Pseudoalteromonas lipolytica biofilm with increased antifouling activities.

PubMed

Zeng, Zhenshun; Guo, Xing-Pan; Li, Baiyuan; Wang, Pengxia; Cai, Xingsheng; Tian, Xinpeng; Zhang, Si; Yang, Jin-Long; Wang, Xiaoxue

2015-12-01

Pseudoalteromonas is widespread in various marine environments, and most strains can affect invertebrate larval settlement and metamorphosis by forming biofilms. However, the impact and the molecular basis of population diversification occurring in Pseudoalteromonas biofilms are poorly understood. Here, we show that morphological diversification is prevalent in Pseudoalteromonas species during biofilm formation. Two types of genetic variants, wrinkled (frequency of 12±5%) and translucent (frequency of 5±3%), were found in Pseudoalteromonas lipolytica biofilms. The inducing activities of biofilms formed by the two variants on larval settlement and metamorphosis of the mussel Mytilus coruscus were significantly decreased, suggesting strong antifouling activities. Using whole-genome re-sequencing combined with genetic manipulation, two genes were identified to be responsible for the morphology alternations. A nonsense mutation in AT00_08765 led to a wrinkled morphology due to the overproduction of cellulose, whereas a point mutation in AT00_17125 led to a translucent morphology via a reduction in capsular polysaccharide production. Taken together, the results suggest that the microbial behavior on larval settlement and metamorphosis in marine environment could be affected by the self-generated variants generated during the formation of marine biofilms, thereby rendering potential application in biocontrol of marine biofouling.

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

PubMed

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.

Vaginal yeast infection

MedlinePlus

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 in the ...

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

PubMed

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.

Marine yeast isolation and industrial application.

PubMed

Zaky, Abdelrahman Saleh; Tucker, Gregory A; Daw, Zakaria Yehia; Du, Chenyu

2014-09-01

Over the last century, terrestrial yeasts have been widely used in various industries, such as baking, brewing, wine, bioethanol and pharmaceutical protein production. However, only little attention has been given to marine yeasts. Recent research showed that marine yeasts have several unique and promising features over the terrestrial yeasts, for example higher osmosis tolerance, higher special chemical productivity and production of industrial enzymes. These indicate that marine yeasts have great potential to be applied in various industries. This review gathers the most recent techniques used for marine yeast isolation as well as the latest applications of marine yeast in bioethanol, pharmaceutical and enzyme production fields. © 2014 The Authors FEMS Yeast Research published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

Yeast ecology of Kombucha fermentation.

PubMed

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.

Marine yeast isolation and industrial application

PubMed Central

Zaky, Abdelrahman Saleh; Tucker, Gregory A; Daw, Zakaria Yehia; Du, Chenyu

2014-01-01

Over the last century, terrestrial yeasts have been widely used in various industries, such as baking, brewing, wine, bioethanol and pharmaceutical protein production. However, only little attention has been given to marine yeasts. Recent research showed that marine yeasts have several unique and promising features over the terrestrial yeasts, for example higher osmosis tolerance, higher special chemical productivity and production of industrial enzymes. These indicate that marine yeasts have great potential to be applied in various industries. This review gathers the most recent techniques used for marine yeast isolation as well as the latest applications of marine yeast in bioethanol, pharmaceutical and enzyme production fields. PMID:24738708

Wine yeasts for the future.

PubMed

Fleet, Graham H

2008-11-01

International competition within the wine market, consumer demands for newer styles of wines and increasing concerns about the environmental sustainability of wine production are providing new challenges for innovation in wine fermentation. Within the total production chain, the alcoholic fermentation of grape juice by yeasts is a key process where winemakers can creatively engineer wine character and value through better yeast management and, thereby, strategically tailor wines to a changing market. This review considers the importance of yeast ecology and yeast metabolic reactions in determining wine quality, and then discusses new directions for exploiting yeasts in wine fermentation. It covers criteria for selecting and developing new commercial strains, the possibilities of using yeasts other than those in the genus of Saccharomyces, the prospects for mixed culture fermentations and explores the possibilities for high cell density, continuous fermentations.

Discussion of teleomorphic and anamorphic Ascomycetous yeasts and yeast-like taxa

USDA-ARS?s Scientific Manuscript database

The relationship of ascomycetous yeasts with other members of the ascomycete fungi (Ascomycota) has been controversial for over 100 years. Because yeasts are morphologically simple, it was proposed that they represent primitive forms of ascomycetes (e.g., Guilliermond 1912). Alternatively, the ide...

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

PubMed

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.

Forces in yeast flocculation

NASA Astrophysics Data System (ADS)

El-Kirat-Chatel, Sofiane; Beaussart, Audrey; Vincent, Stéphane P.; Abellán Flos, Marta; Hols, Pascal; Lipke, Peter N.; Dufrêne, Yves F.

2015-01-01

In the baker's yeast Saccharomyces cerevisiae, cell-cell adhesion (``flocculation'') is conferred by a family of lectin-like proteins known as the flocculin (Flo) proteins. Knowledge of the adhesive and mechanical properties of flocculins is important for understanding the mechanisms of yeast adhesion, and may help controlling yeast behaviour in biotechnology. We use single-molecule and single-cell atomic force microscopy (AFM) to explore the nanoscale forces engaged in yeast flocculation, focusing on the role of Flo1 as a prototype of flocculins. Using AFM tips labelled with mannose, we detect single flocculins on Flo1-expressing cells, showing they are widely exposed on the cell surface. When subjected to force, individual Flo1 proteins display two distinct force responses, i.e. weak lectin binding forces and strong unfolding forces reflecting the force-induced extension of hydrophobic tandem repeats. We demonstrate that cell-cell adhesion bonds also involve multiple weak lectin interactions together with strong unfolding forces, both associated with Flo1 molecules. Single-molecule and single-cell data correlate with microscale cell adhesion behaviour, suggesting strongly that Flo1 mechanics is critical for yeast flocculation. These results favour a model in which not only weak lectin-sugar interactions are involved in yeast flocculation but also strong hydrophobic interactions resulting from protein unfolding.

The yeast actin cytoskeleton.

PubMed

Mishra, Mithilesh; Huang, Junqi; Balasubramanian, Mohan K

2014-03-01

The actin cytoskeleton is a complex network of dynamic polymers, which plays an important role in various fundamental cellular processes, including maintenance of cell shape, polarity, cell division, cell migration, endocytosis, vesicular trafficking, and mechanosensation. Precise spatiotemporal assembly and disassembly of actin structures is regulated by the coordinated activity of about 100 highly conserved accessory proteins, which nucleate, elongate, cross-link, and sever actin filaments. Both in vivo studies in a wide range of organisms from yeast to metazoans and in vitro studies of purified proteins have helped shape the current understanding of actin dynamics and function. Molecular genetics, genome-wide functional analysis, sophisticated real-time imaging, and ultrastructural studies in concert with biochemical analysis have made yeast an attractive model to understand the actin cytoskeleton, its molecular dynamics, and physiological function. Studies of the yeast actin cytoskeleton have contributed substantially in defining the universal mechanism regulating actin assembly and disassembly in eukaryotes. Here, we review some of the important insights generated by the study of actin cytoskeleton in two important yeast models the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

[Thermoresistance in Saccharomyces cerevisiae yeasts].

PubMed

Kaliuzhin, V A

2011-01-01

Under natural conditions, yeast Saccharomyces cerevisiae reproduce, as a rule, on the surface of solid or liquid medium. Thus, life cycle of yeast populations is substantially influenced by diurnal changes in ambient temperature. The pattern in the response of unrestricted yeast S. cerevisiae culture to changes in the temperature of cultivation is revealed experimentally. Yeast population, in the absence of environmental constraints on the functioning of cell chemosmotic bioenergetic system, demonstrates the ability of thermoresistance when the temperature of cultivation switches from the range of 12-36 degrees C to 37.5-40 degrees C. During the transient period that is associated with the temperature switching and lasts from 1 to 4 turnover cycles, yeast reproduction rate remains 1.5-2 times higher than under stationary conditions. This is due to evolutionary acquired adaptive activity of cell chemosmotic system. After the adaptive resources exhausting, yeast thermoresistance fully recovers at the temperature range of 12-36 degrees C within one generation time under conditions of both restricted and unrestricted nourishment. Adaptive significance of such thermoresistance seems obvious enough--it allows maintaining high reproduction rate in yeast when ambient temperature is reaching a brief maximum shortly after noon.

Interaction Between Yeasts and Zinc

NASA Astrophysics Data System (ADS)

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

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

Yeast flocculation: New story in fuel ethanol production.

PubMed

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.

Brewing characteristics of piezosensitive sake yeasts

NASA Astrophysics Data System (ADS)

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.

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Bakers yeast glycan. 172.898 Section 172.898 Food... 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 dried cell walls of the yeast...

Opportunistic Pathogenic Yeasts

NASA Astrophysics Data System (ADS)

Banerjee, Uma

Advances in medical research, made during the last few decades, have improved the prophylactic, diagnostic and therapeutic capabilities for variety of infections/diseases. However, many of the prophylactic and therapeutic procedures have been seen in many instances to exact a price of host-vulnerability to an expanding group of opportunistic pathogens and yeasts are one of the important members in it. Fortunately amongst the vast majority of yeasts present in nature only few are considered to have the capability to cause infections when certain opportunities predisposes and these are termed as ‘opportunistic pathogenic yeasts.’ However, the term ‘pathogenic’ is quite tricky, as it depends of various factors of the host, the ‘bug’ and the environment to manifest the clinical infection. The borderline is expanding. In the present century with unprecedented increase in number of immune-compromised host in various disciplines of health care settings, where any yeast, which has the capability to grow at 37 ° C (normal body temperature of human), can be pathogenic and cause infection in particular situation

Yeast Droplets

NASA Astrophysics Data System (ADS)

Nguyen, Baochi; Upadhyaya, Arpita; van Oudenaarden, Alexander; Brenner, Michael

2002-11-01

It is well known that the Young's law and surface tension govern the shape of liquid droplets on solid surfaces. Here we address through experiments and theory the shape of growing aggregates of yeast on agar substrates, and assess whether these ideas still hold. Experiments are carried out on Baker's yeast, with different levels of expressions of an adhesive protein governing cell-cell and cell-substrate adhesion. Changing either the agar concentration or the expression of this protein modifies the local contact angle of a yeast droplet. When the colony is small, the shape is a spherical cap with the contact angle obeying Young's law. However, above a critical volume this structure is unstable, and the droplet becomes nonspherical. We present a theoretical model where this instability is caused by bulk elastic effects. The model predicts that the transition depends on both volume and contact angle, in a manner quantitatively consistent with our experiments.

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

PubMed

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.

Utilization of inulin-containing waste in industrial fermentations to produce biofuels and bio-based chemicals.

PubMed

Hughes, Stephen R; Qureshi, Nasib; López-Núñez, Juan Carlos; Jones, Marjorie A; Jarodsky, Joshua M; Galindo-Leva, Luz Ángela; Lindquist, Mitchell R

2017-04-01

Inulins are polysaccharides that belong to an important class of carbohydrates known as fructans and are used by many plants as a means of storing energy. Inulins contain 20 to several thousand fructose units joined by β-2,1 glycosidic bonds, typically with a terminal glucose unit. Plants with high concentrations of inulin include: agave, asparagus, coffee, chicory, dahlia, dandelion, garlic, globe artichoke, Jerusalem artichoke, jicama, onion, wild yam, and yacón. To utilize inulin as its carbon and energy source directly, a microorganism requires an extracellular inulinase to hydrolyze the glycosidic bonds to release fermentable monosaccharides. Inulinase is produced by many microorganisms, including species of Aspergillus, Kluyveromyces, Penicillium, and Pseudomonas. We review various inulinase-producing microorganisms and inulin feedstocks with potential for industrial application as well as biotechnological efforts underway to develop sustainable practices for the disposal of residues from processing inulin-containing crops. A multi-stage biorefinery concept is proposed to convert cellulosic and inulin-containing waste produced at crop processing operations to valuable biofuels and bioproducts using Kluyveromyces marxianus, Yarrowia lipolytica, Rhodotorula glutinis, and Saccharomyces cerevisiae as well as thermochemical treatments.

Genome-scale metabolic modeling of Mucor circinelloides and comparative analysis with other oleaginous species.

PubMed

Vongsangnak, Wanwipa; Klanchui, Amornpan; Tawornsamretkit, Iyarest; Tatiyaborwornchai, Witthawin; Laoteng, Kobkul; Meechai, Asawin

2016-06-01

We present a novel genome-scale metabolic model iWV1213 of Mucor circinelloides, which is an oleaginous fungus for industrial applications. The model contains 1213 genes, 1413 metabolites and 1326 metabolic reactions across different compartments. We demonstrate that iWV1213 is able to accurately predict the growth rates of M. circinelloides on various nutrient sources and culture conditions using Flux Balance Analysis and Phenotypic Phase Plane analysis. Comparative analysis of three oleaginous genome-scale models, including M. circinelloides (iWV1213), Mortierella alpina (iCY1106) and Yarrowia lipolytica (iYL619_PCP) revealed that iWV1213 possesses a higher number of genes involved in carbohydrate, amino acid, and lipid metabolisms that might contribute to its versatility in nutrient utilization. Moreover, the identification of unique and common active reactions among the Zygomycetes oleaginous models using Flux Variability Analysis unveiled a set of gene/enzyme candidates as metabolic engineering targets for cellular improvement. Thus, iWV1213 offers a powerful metabolic engineering tool for multi-level omics analysis, enabling strain optimization as a cell factory platform of lipid-based production. Copyright © 2016 Elsevier B.V. All rights reserved.

Lager Yeast Comes of Age

PubMed Central

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

Yeast killer systems.

PubMed Central

Magliani, W; Conti, S; Gerloni, M; Bertolotti, D; Polonelli, L

1997-01-01

The killer phenomenon in yeasts has been revealed to be a multicentric model for molecular biologists, virologists, phytopathologists, epidemiologists, industrial and medical microbiologists, mycologists, and pharmacologists. The surprisingly widespread occurrence of the killer phenomenon among taxonomically unrelated microorganisms, including prokaryotic and eukaryotic pathogens, has engendered a new interest in its biological significance as well as its theoretical and practical applications. The search for therapeutic opportunities by using yeast killer systems has conceptually opened new avenues for the prevention and control of life-threatening fungal diseases through the idiotypic network that is apparently exploited by the immune system in the course of natural infections. In this review, the biology, ecology, epidemiology, therapeutics, serology, and idiotypy of yeast killer systems are discussed. PMID:9227858

Yeasts as distinct life forms of fungi

USDA-ARS?s Scientific Manuscript database

This review describes all presently recognized genera of the Ascomycete yeasts (Saccharomycotina, budding yeasts, and the Taphrinomycotina, fission yeasts and related) as well as all currently recognized genera of the Basidiomycete yeasts. This update will be the lead chapter for a book entitled “Ye...

Study of amyloids using yeast

PubMed Central

Wickner, Reed B.; Kryndushkin, Dmitry; Shewmaker, Frank; McGlinchey, Ryan; Edskes, Herman K.

2012-01-01

Summary Saccharomyces cerevisiae has been a useful model organism in such fields as the cell cycle, regulation of transcription, protein trafficking and cell biology, primarily because of its ease of genetic manipulation. This is no less so in the area of amyloid studies. The endogenous yeast amyloids described to date include prions, infectious proteins (Table 1), and some cell wall proteins (1). and amyloids of humans and a fungal prion have also been studied using the yeast system. Accordingly, the emphasis of this chapter will be on genetic, biochemical, cell biological and physical methods particularly useful in the study of yeast prions and other amyloids studied in yeast. We limit our description of these methods to those aspects which have been most useful in studying yeast prions, citing more detailed expositions in the literature. Volumes on yeast genetics methods (2–4), and on amyloids and prions (5, 6) are useful, and Masison has edited a volume of Methods on “Identification, analysis and characterization of fungal prions” which covers some of this territory (7). We also outline some useful physical methods, pointing the reader to more extensive and authoratative descriptions. PMID:22528100

Evolutionary History of Ascomyceteous Yeasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haridas, Sajeet; Riley, Robert; Salamov, Asaf

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 amore » 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.« less

Eighteen new oleaginous yeast species.

PubMed

Garay, Luis A; Sitepu, Irnayuli R; Cajka, Tomas; Chandra, Idelia; Shi, Sandy; Lin, Ting; German, J Bruce; Fiehn, Oliver; Boundy-Mills, Kyria L

2016-07-01

Of 1600 known species of yeasts, about 70 are known to be oleaginous, defined as being able to accumulate over 20 % intracellular lipids. These yeasts have value for fundamental and applied research. A survey of yeasts from the Phaff Yeast Culture Collection, University of California Davis was performed to identify additional oleaginous species within the Basidiomycota phylum. Fifty-nine strains belonging to 34 species were grown in lipid inducing media, and total cell mass, lipid yield and triacylglycerol profiles were determined. Thirty-two species accumulated at least 20 % lipid and 25 species accumulated over 40 % lipid by dry weight. Eighteen of these species were not previously reported to be oleaginous. Triacylglycerol profiles were suitable for biodiesel production. These results greatly expand the number of known oleaginous yeast species, and reveal the wealth of natural diversity of triacylglycerol profiles within wild-type oleaginous Basidiomycetes.

Oral yeast colonization throughout pregnancy

PubMed Central

Rio, Rute; Simões-Silva, Liliana; Garro, Sofia; Silva, Mário-Jorge; Azevedo, Álvaro

2017-01-01

Background 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. Material and Methods 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. Results 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. Conclusions Pregnancy may favour oral yeast growth that may be associated with an acidic oral environment. Key words:Oral yeast, fungi, pregnancy, saliva pH. PMID:28160578

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

PubMed

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'.

Nutrient supplements boost yeast transformation efficiency

PubMed Central

Yu, Sheng-Chun; Dawson, Alexander; Henderson, Alyssa C.; Lockyer, Eloise J.; Read, Emily; Sritharan, Gayathri; Ryan, Marjah; Sgroi, Mara; Ngou, Pok M.; Woodruff, Rosie; Zhang, Ruifeng; Ren Teen Chia, Travis; Liu, Yu; Xiang, Yiyu; Spanu, Pietro D.

2016-01-01

Efficiency of yeast transformation is determined by the rate of yeast endocytosis. The aim of this study was to investigate the effect of introducing amino acids and other nutrients (inositol, adenine, or p-aminobenzoic acid) in the transformation medium to develop a highly efficient yeast transformation protocol. The target of rapamycin complex 1 (TORC1) kinase signalling complex influences the rate of yeast endocytosis. TORC signaling is induced by amino acids in the media. Here, we found that increasing the concentration of amino acids and other nutrients in the growth media lead to an increase yeast transformation efficiency up to 107 CFU per μg plasmid DNA and per 108 cells with a 13.8 kb plasmid DNA. This is over 130 times that of current published methods. This improvement may facilitate more efficient experimentation in which transformation efficiency is critical, such as yeast two-hybrid screening. PMID:27760994

Virgin olive oil yeasts: A review.

PubMed

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.

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Bakers yeast extract. 184.1983 Section 184.1983... GRAS § 184.1983 Bakers yeast extract. (a) Bakers yeast extract is the food ingredient resulting from concentration of the solubles of mechanically ruptured cells of a selected strain of yeast, Saccharomyces...

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false 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...

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false 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...

History of genome editing in yeast.

PubMed

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.

Inventions on baker's yeast strains and specialty ingredients.

PubMed

Gélinas, Pierre

2009-06-01

Baker's yeast is one of the oldest food microbial starters. Between 1927 and 2008, 165 inventions on more than 337 baker's yeast strains were patented. The first generation of patented yeast strains claimed improved biomass yield at the yeast plant, higher gassing power in dough or better survival to drying to prepare active dry baker's yeast. Especially between 1980 and 1995, a major interest was given to strains for multiple bakery applications such as dough with variable sugar content and stored at refrigeration (cold) or freezing temperatures. During the same period, genetically engineered yeast strains became very popular but did not find applications in the baking industry. Since year 2000, patented baker's yeast strains claimed aroma, anti-moulding or nutritive properties to better meet the needs of the baking industry. In addition to patents on yeast strains, 47 patents were issued on baker's yeast specialty ingredients for niche markets. This review shows that patents on baker's yeast with improved characteristics such as aromatic or nutritive properties have regularly been issued since the 1920's. Overall, it also confirms recent interest for a very wide range of tailored-made yeast-based ingredients for bakery applications.

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false 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...

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false 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...

The wine and beer yeast Dekkera bruxellensis

PubMed Central

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

2014-01-01

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. PMID:24932634

The wine and beer yeast Dekkera bruxellensis.

PubMed

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.

Biotechnology of non-Saccharomyces yeasts-the basidiomycetes.

PubMed

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

Between science and industry-applied yeast research.

PubMed

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.

Yeast-based biosensors: design and applications.

PubMed

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.

Oral yeast colonization throughout pregnancy.

PubMed

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.

Yeasts of the soil – obscure but precious

PubMed Central

2018-01-01

Abstract Pioneering studies performed in the nineteenth century demonstrated that yeasts are present in below‐ground sources. Soils were regarded more as a reservoir for yeasts that reside in habitats above it. Later studies showed that yeast communities in soils are taxonomically diverse and different from those above‐ground. Soil yeasts possess extraordinary adaptations that allow them to survive in a wide range of environmental conditions. A few species are promising sources of yeast oils and have been used in agriculture as potential antagonists of soil‐borne plant pathogens or as plant growth promoters. Yeasts have been studied mainly in managed soils such as vineyards, orchards and agricultural fields, and to a lesser extent under forests and grasslands. Our knowledge of soil yeasts is further biased towards temperate and boreal forests, whereas data from Africa, the Americas and Asia are scarce. Although soil yeast communities are often species‐poor in a single sample, they are more diverse on the biotope level. Soil yeasts display pronounced endemism along with a surprisingly high proportion of currently unidentified species. However, like other soil inhabitants, yeasts are threatened by habitat alterations owing to anthropogenic activities such as agriculture, deforestation and urbanization. In view of the rapid decline of many natural habitats, the study of soil yeasts in undisturbed or low‐managed biotopes is extremely valuable. The purpose of this review is to encourage researchers, both biologists and soil scientists, to include soil yeasts in future studies. PMID:29365211

Electron transport chain in a thermotolerant yeast.

PubMed

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.

Yeasts in floral nectar: a quantitative survey

PubMed Central

Herrera, Carlos M.; de Vega, Clara; Canto, Azucena; Pozo, María I.

2009-01-01

Background and Aims One peculiarity of floral nectar that remains relatively unexplored from an ecological perspective is its role as a natural habitat for micro-organisms. This study assesses the frequency of occurrence and abundance of yeast cells in floral nectar of insect-pollinated plants from three contrasting plant communities on two continents. Possible correlations between interspecific differences in yeast incidence and pollinator composition are also explored. Methods The study was conducted at three widely separated areas, two in the Iberian Peninsula (Spain) and one in the Yucatán Peninsula (Mexico). Floral nectar samples from 130 species (37–63 species per region) in 44 families were examined microscopically for the presence of yeast cells. For one of the Spanish sites, the relationship across species between incidence of yeasts in nectar and the proportion of flowers visited by each of five major pollinator categories was also investigated. Key Results Yeasts occurred regularly in the floral nectar of many species, where they sometimes reached extraordinary densities (up to 4 × 105 cells mm−3). Depending on the region, between 32 and 44 % of all nectar samples contained yeasts. Yeast cell densities in the order of 104 cells mm−3 were commonplace, and densities >105 cells mm−3 were not rare. About one-fifth of species at each site had mean yeast cell densities >104 cells mm−3. Across species, yeast frequency and abundance were directly correlated with the proportion of floral visits by bumble-bees, and inversely with the proportion of visits by solitary bees. Conclusions Incorporating nectar yeasts into the scenario of plant–pollinator interactions opens up a number of intriguing avenues for research. In addition, with yeasts being as ubiquitous and abundant in floral nectars as revealed by this study, and given their astounding metabolic versatility, studies focusing on nectar chemical features should carefully control for the presence

Interactions between Drosophila and its natural yeast symbionts—Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

PubMed Central

Hoang, Don; Kopp, Artyom

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

Interactions between Drosophila and its natural yeast symbionts-Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

PubMed

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

Evaluation of Automated Yeast Identification System

NASA Technical Reports Server (NTRS)

McGinnis, M. R.

1996-01-01

One hundred and nine teleomorphic and anamorphic yeast isolates representing approximately 30 taxa were used to evaluate the accuracy of the Biolog yeast identification system. Isolates derived from nomenclatural types, environmental, and clinica isolates of known identity were tested in the Biolog system. Of the isolates tested, 81 were in the Biolog database. The system correctly identified 40, incorrectly identified 29, and was unable to identify 12. Of the 28 isolates not in the database, 18 were given names, whereas 10 were not. The Biolog yeast identification system is inadequate for the identification of yeasts originating from the environment during space program activities.

Cloning, expression and characterization of a lipase gene from marine bacterium Pseudoalteromonas lipolytica SCSIO 04301

NASA Astrophysics Data System (ADS)

Su, Hongfei; Mai, Zhimao; Zhang, Si

2016-12-01

A lipase gene, lip1233, isolated from Pseudoalteromonas lipolytica SCSIO 04301, was cloned and expressed in E. coli. The enzyme comprised 810 amino acid residues with a deduced molecular weight of 80 kDa. Lip1233 was grouped into the lipase family X because it contained a highly conserved motif GHSLG. The recombinant enzyme was purified with Ni-NTA affinity chromatography. The optimal temperature and pH value of Lip1233 were 45°C and 8.0, respectively. It retained more than 70% of original activity after being incubated in pH ranging from 6.0 to 9.5 for 30 min. It was stable when the temperature was below 45°C, but was unstable when the temperature was above 55°C. Most metal ions tested had no significant effect on the activity of Lip1233. Lip1233 remained more than original activity in some organic solvents at the concentration of 30% (v/v). It retained more than 30% activity after incubated in pure organic solvents for 12 h, while in hexane the activity was nearly 100%. Additionally, Lip1233 exhibited typical halotolerant characteristic as it was active under 4M NaCl. Lip1233 powder could catalyze efficiently the synthesis of fructose esters in hexane at 40°C. These characteristics demonstrated that Lip1233 is applicable to elaborate food processing and organic synthesis.

Genomics and the making of yeast biodiversity.

PubMed

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.

Accelerating Yeast Prion Biology using Droplet Microfluidics

NASA Astrophysics Data System (ADS)

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.

[Groups and sources of yeasts in house dust].

PubMed

Glushakova, A M; Zheltikova, T M; Chernov, I Iu

2004-01-01

House dust contains bacteria, mycelial fungi, microarthropods, and yeasts. The house dust samples collected in 25 apartments in Moscow and the Moscow region were found to contain yeasts belonging to the genera Candida, Cryptococcus, Debaryomyces, Rhodotorula, Sporobolomyces, and Trichosporon. The most frequently encountered microorganisms were typical epiphytic yeasts, such as Cryptococcus diffluens and Rhodotorula mucilaginosa, which are capable of long-term preservation in an inactive state. The direct source of epiphytic yeasts occurring in the house dust might be the indoor plants, which were contaminated with these yeasts, albeit to a lesser degree than outdoor plants. Along with the typical epiphytic yeasts, the house dust contained the opportunistic yeast pathogens Candida catenulata, C. guillermondii, C. haemulonii, C. rugosa, and C. tropicalis, which are known as the causal agents of candidiasis. We failed to reveal any correlation between the abundance of particular yeast species in the house dust, residential characteristics, and the atopic dermatitis of the inhabitants.

Genomics and the making of yeast biodiversity

USDA-ARS?s Scientific Manuscript database

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 ...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Yeast-malt sprout extract. 172.590 Section 172.590... Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout extract, as described in this section, may... produced by partial hydrolysis of yeast extract (derived from Saccharomyces cereviseae, Saccharomyces...

Yeasts Diversity in Fermented Foods and Beverages

NASA Astrophysics Data System (ADS)

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

Genetics of Yeasts

NASA Astrophysics Data System (ADS)

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.

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2010 CFR

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...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false 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...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false 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...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 3 2011-04-01 2011-04-01 false 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...

Yeasts in sustainable bioethanol production: A review.

PubMed

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.

Yeast and Mammalian Metallothioneins Functionally Substitute for Yeast Copper-Zinc Superoxide Dismutase

NASA Astrophysics Data System (ADS)

Tamai, Katherine T.; Gralla, Edith B.; Ellerby, Lisa M.; Valentine, Joan S.; Thiele, Dennis J.

1993-09-01

Copper-zinc superoxide dismutase catalyzes the disproportionation of superoxide anion to hydrogen peroxide and dioxygen and is thought to play an important role in protecting cells from oxygen toxicity. Saccharomyces cerevisiae strains lacking copper-zinc superoxide dismutase, which is encoded by the SOD1 gene, are sensitive to oxidative stress and exhibit a variety of growth defects including hypersensitivity to dioxygen and to superoxide-generating drugs such as paraquat. We have found that in addition to these known phenotypes, SOD1-deletion strains fail to grow on agar containing the respiratory carbon source lactate. We demonstrate here that expression of the yeast or monkey metallothionein proteins in the presence of copper suppresses the lactate growth defect and some other phenotypes associated with SOD1-deletion strains, indicating that copper metallothioneins substitute for copper-zinc superoxide dismutase in vivo to protect cells from oxygen toxicity. Consistent with these results, we show that yeast metallothionein mRNA levels are dramatically elevated under conditions of oxidative stress. Furthermore, in vitro assays demonstrate that yeast metallothionein, purified or from whole-cell extracts, exhibits copper-dependent antioxidant activity. Taken together, these data suggest that both yeast and mammalian metallothioneins may play a direct role in the cellular defense against oxidative stress by functioning as antioxidants.

Experimental Systems to Study Yeast Pexophagy.

PubMed

Yamashita, Shun-Ichi; Oku, Masahide; Sakai, Yasuyoshi; Fujiki, Yukio

2017-01-01

Peroxisome abundance is tightly regulated according to the physiological contexts, through regulations of both proliferation and degradation of the organelles. Here, we describe detailed methods to analyze processes for autophagic degradation of peroxisomes, termed pexophagy, in yeast organisms. The assay systems include a method for biochemical detection of pexophagy completion, and one for microscopic visualization of specialized membrane structures acting in pexophagy. As a model yeast organism utilized in studies of pexophagy, the methylotrophic yeast Komagataella phaffii (Pichia pastoris) is referred to in this chapter and related information on the studies with baker's yeast (Saccharomyces cerevisiae) is also included. The described techniques facilitate elucidation of molecular machineries for pexophagy and understanding of peroxisome-selective autophagic pathways.

[Distiller Yeasts Producing Antibacterial Peptides].

PubMed

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.

Differential Adsorption of Ochratoxin A and Anthocyanins by Inactivated Yeasts and Yeast Cell Walls during Simulation of Wine Aging

PubMed Central

Petruzzi, Leonardo; Baiano, Antonietta; De Gianni, Antonio; Sinigaglia, Milena; Corbo, Maria Rosaria; Bevilacqua, Antonio

2015-01-01

The adsorption of ochratoxin A (OTA) by yeasts is a promising approach for the decontamination of musts and wines, but some potential competitive or interactive phenomena between mycotoxin, yeast cells, and anthocyanins might modify the intensity of the phenomenon. The aim of this study was to examine OTA adsorption by two strains of Saccharomyces cerevisiae (the wild strain W13, and the commercial isolate BM45), previously inactivated by heat, and a yeast cell wall preparation. Experiments were conducted using Nero di Troia red wine contaminated with 2 μg/L OTA and supplemented with yeast biomass (20 g/L). The samples were analyzed periodically to assess mycotoxin concentration, chromatic characteristics, and total anthocyanins over 84 days of aging. Yeast cell walls revealed the highest OTA-adsorption in comparison to thermally-inactivated cells (50% vs. 43% toxin reduction), whilst no significant differences were found for the amount of adsorbed anthocyanins in OTA-contaminated and control wines. OTA and anthocyanins adsorption were not competitive phenomena. Unfortunately, the addition of yeast cells to wine could cause color loss; therefore, yeast selection should also focus on this trait to select the best strain. PMID:26516913

Made for Each Other: Ascomycete Yeasts and Insects.

PubMed

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.

The growth of solar radiated yeast

NASA Technical Reports Server (NTRS)

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.

The growth of solar radiated yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kraft, T.

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 containersmore » 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.« less

Freeze-drying of yeast cultures.

PubMed

Bond, Chris

2007-01-01

A method is described that allows yeast species to be stored using a variation on the standard freeze-drying method, which employs evaporative cooling in a two-stage process. Yeast cultures are placed in glass ampoules after having been mixed with a lyoprotectant. Primary drying is carried out using a centrifuge head connected to a standard freeze-dryer. Once the centrifuge head is running, air is removed and evaporated liquid is captured in the freeze-dryer. Centrifugation continues for 15 min and primary drying for a further 3 h. The ampoules are constricted using a glass blowing torch. They are then placed on the freeze-dryer manifold for secondary drying under vacuum overnight, using phosphorus pentoxide as a desiccant. The ampoules are sealed and removed from the manifold by melting the constricted section. Although the process causes an initial large drop in viability, further losses after storage are minimal. Yeast strains have remained viable for more than 30 yr when stored using this method and sufficient cells are recovered to produce new working stocks. Although survival rates are strain specific, nearly all National Collection of Yeast Cultures strains covering most yeast genera, have been successfully stored with little or no detectable change in strain characteristics.

Biotechnology of non-Saccharomyces yeasts--the ascomycetes.

PubMed

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.

Yeast proteome map (last update).

PubMed

Perrot, Michel; Moes, Suzette; Massoni, Aurélie; Jenoe, Paul; Boucherie, Hélian

2009-10-01

The identification of proteins separated on 2-D gels is essential to exploit the full potential of 2-D gel electrophoresis for proteomic investigations. For this purpose we have undertaken the systematic identification of Saccharomyces cerevisiae proteins separated on 2-D gels. We report here the identification by mass spectrometry of 100 novel yeast protein spots that have so far not been tackled due to their scarcity on our standard 2-D gels. These identifications extend the number of protein spots identified on our yeast 2-D proteome map to 716. They correspond to 485 unique proteins. Among these, 154 were resolved into several isoforms. The present data set can now be expanded to report for the first time a map of 363 protein isoforms that significantly deepens our knowledge of the yeast proteome. The reference map and a list of all identified proteins can be accessed on the Yeast Protein Map server (www.ibgc.u-bordeaux2.fr/YPM).

Septin Organization and Functions in Budding Yeast

PubMed Central

Glomb, Oliver; Gronemeyer, Thomas

2016-01-01

The septins are a conserved family of GTP-binding proteins present in all eukaryotic cells except plants. They were originally discovered in the baker's yeast Saccharomyces cerevisiae that serves until today as an important model organism for septin research. In yeast, the septins assemble into a highly ordered array of filaments at the mother bud neck. The septins are regulators of spatial compartmentalization in yeast and act as key players in cytokinesis. This minireview summarizes the recent findings about structural features and cell biology of the yeast septins. PMID:27857941

Taming wild yeast: potential of conventional and nonconventional yeasts in industrial fermentations.

PubMed

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.

Yeast: A Research Organism for Teaching Genetics.

ERIC Educational Resources Information Center

Manney, Thomas R.; Manney, Monta L.

1992-01-01

Explains why laboratory strains of bakers yeast, Saccharomyces cerevisiae, are particularly suited for classroom science activities. Describes the sexual life cycle of yeast and the genetic system with visible mutations. Presents an overview of activities that can be done with yeast and gives a source for teachers to obtain more information. (PR)

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

PubMed

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.

Yeast fuel cell: Application for desalination

NASA Astrophysics Data System (ADS)

Mardiana, Ummy; Innocent, Christophe; Cretin, Marc; Buchari, Buchari; Gandasasmita, Suryo

2016-02-01

Yeasts have been implicated in microbial fuel cells as biocatalysts because they are non-pathogenic organisms, easily handled and robust with a good tolerance in different environmental conditions. Here we investigated baker's yeast Saccharomyces cerevisiae through the oxidation of glucose. Yeast was used in the anolyte, to transfer electrons to the anode in the presence of methylene blue as mediator whereas K3Fe(CN)6 was used as an electron acceptor for the reduction reaction in the catholyte. Power production with biofuel cell was coupled with a desalination process. The maximum current density produced by the cell was 88 mA.m-2. In those conditions, it was found that concentration of salt was removed 64% from initial 0.6 M after 1-month operation. This result proves that yeast fuel cells can be used to remove salt through electrically driven membrane processes and demonstrated that could be applied for energy production and desalination. Further developments are in progress to improve power output to make yeast fuel cells applicable for water treatment.

Yeasts for Global Happiness: report of the 14th International Congress on Yeasts (ICY14) held in Awaji Island.

PubMed

Watanabe, Daisuke; Takagi, Hiroshi

2017-02-01

The 14th International Congress on Yeasts (ICY14) was held at Awaji Yumebutai International Conference Center (Awaji, Hyogo) in Japan from 11 to 15 September 2016. The main slogan of ICY14 was 'Yeasts for Global Happiness', which enabled us to acknowledge the high-potential usefulness of yeasts contributing to the global happiness in terms of food/beverage, health/medicine and energy/environment industries, as well as to basic biosciences. In addition, two more concepts were introduced: 'from Japan to the world' and 'from senior to junior'. As it was the first ICY meeting held in Japan or other Asian countries, ICY14 provided a good opportunity to widely spread the great achievements by Japanese and Asian yeast researchers, such as those by the 2016 Nobel Laureate Dr. Yoshinori Ohsumi, and also, to convey the fun and importance of yeasts to the next generation of researchers from Asia and all over the world. As a result, a total of 426 yeast lovers from 42 countries (225 overseas and 201 domestic participants) with different generations attended ICY14 to share the latest knowledge of a wide range of yeast research fields and to join active and constructive scientific discussions. © 2017 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

Yeasts are essential for cocoa bean fermentation.

PubMed

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.

Yeast Genetics and Biotechnological Applications

NASA Astrophysics Data System (ADS)

Mishra, Saroj; Baranwal, Richa

Yeast can be recognized as one of the very important groups of microorganisms on account of its extensive use in the fermentation industry and as a basic eukaryotic model cellular system. The yeast Saccharomyces cerevisiae has been extensively used to elucidate the genetics and regulation of several key functions in the cell such as cell mating, electron transport chain, protein trafficking, cell cycle events and others. Even before the genome sequence of the yeast was out, the structural organization and function of several of its genes was known. With the availability of the origin of replication from the 2 μm plasmid and the development of transformation system, it became the host of choice for expression of a number of important proteins. A large number of episomal and integrative shuttle vectors are available for expression of mammalian proteins. The latest developments in genomics and micro-array technology have allowed investigations of individual gene function by site-specific deletion method. The application of metabolic profiling has also assisted in understanding the cellular network operating in this yeast. This chapter is aimed at reviewing the use of this system as an experimental tool for conducting classical genetics. Various vector systems available, foreign genes expressed and the limitations as a host will be discussed. Finally, the use of various yeast enzymes in biotechnology sector will be reviewed.

Functional adaptation between yeast actin and its cognate myosin motors.

PubMed

Stark, Benjamin C; Wen, Kuo-Kuang; Allingham, John S; Rubenstein, Peter A; Lord, Matthew

2011-09-02

We employed budding yeast and skeletal muscle actin to examine the contribution of the actin isoform to myosin motor function. While yeast and muscle actin are highly homologous, they exhibit different charge density at their N termini (a proposed myosin-binding interface). Muscle myosin-II actin-activated ATPase activity is significantly higher with muscle versus yeast actin. Whether this reflects inefficiency in the ability of yeast actin to activate myosin is not known. Here we optimized the isolation of two yeast myosins to assess actin function in a homogenous system. Yeast myosin-II (Myo1p) and myosin-V (Myo2p) accommodate the reduced N-terminal charge density of yeast actin, showing greater activity with yeast over muscle actin. Increasing the number of negative charges at the N terminus of yeast actin from two to four (as in muscle) had little effect on yeast myosin activity, while other substitutions of charged residues at the myosin interface of yeast actin reduced activity. Thus, yeast actin functions most effectively with its native myosins, which in part relies on associations mediated by its outer domain. Compared with yeast myosin-II and myosin-V, muscle myosin-II activity was very sensitive to salt. Collectively, our findings suggest differing degrees of reliance on electrostatic interactions during weak actomyosin binding in yeast versus muscle. Our study also highlights the importance of native actin isoforms when considering the function of myosins.

Genomic Evolution of the Ascomycete Yeasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Riley, Robert; Haridas, Sajeet; Salamov, Asaf

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 amore » 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.« less

Specialist nectar-yeasts decline with urbanization in Berlin

NASA Astrophysics Data System (ADS)

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.

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

PubMed Central

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

Yeast Can Affect Behavior and Learning.

ERIC Educational Resources Information Center

Crook, William G.

1984-01-01

A pediatrician recounts his experiences in diagnosing and treating allergies to common yeast germs that may result in behavior and learning problems. He lists characteristics that may predispose children to yeast-connected health problems. (CL)

Subunit mass fingerprinting of mitochondrial complex I.

PubMed

Morgner, Nina; Zickermann, Volker; Kerscher, Stefan; Wittig, Ilka; Abdrakhmanova, Albina; Barth, Hans-Dieter; Brutschy, Bernhard; Brandt, Ulrich

2008-10-01

We have employed laser induced liquid bead ion desorption (LILBID) mass spectrometry to determine the total mass and to study the subunit composition of respiratory chain complex I from Yarrowia lipolytica. Using 5-10 pmol of purified complex I, we could assign all 40 known subunits of this membrane bound multiprotein complex to peaks in LILBID subunit fingerprint spectra by comparing predicted protein masses to observed ion masses. Notably, even the highly hydrophobic subunits encoded by the mitochondrial genome were easily detectable. Moreover, the LILBID approach allowed us to spot and correct several errors in the genome-derived protein sequences of complex I subunits. Typically, the masses of the individual subunits as determined by LILBID mass spectrometry were within 100 Da of the predicted values. For the first time, we demonstrate that LILBID spectrometry can be successfully applied to a complex I band eluted from a blue-native polyacrylamide gel, making small amounts of large multiprotein complexes accessible for subunit mass fingerprint analysis even if they are membrane bound. Thus, the LILBID subunit mass fingerprint method will be of great value for efficient proteomic analysis of complex I and its assembly intermediates, as well as of other water soluble and membrane bound multiprotein complexes.

Immobilization of Yarrowia lipolytica Lipase on Macroporous Resin Using Different Methods: Characterization of the Biocatalysts in Hydrolysis Reaction.

PubMed

Sun, Jingjing; Chen, Yiling; Sheng, Jun; Sun, Mi

2015-01-01

To improve the reusability and organic solvent tolerance of microbial lipase and expand the application of lipase (hydrolysis, esterification, and transesterification), we immobilized marine microbial lipase using different methods and determined the properties of immobilized lipases. Considering the activity and cost of immobilized lipase, the concentration of lipase was fixed at 2 mg/mL. The optimal temperature of immobilized lipases was 40°C and 5°C higher than free lipase. The activities of immobilized lipases were much higher than free lipase at alkaline pH (more than 50% at pH 12). The free lipase lost most activity (35.3%) and immobilized lipases retained more than 46.4% of their initial activity after 3 h heat treatment at 70°C. At alkaline pH, immobilized lipases were more stable than free lipase (more than 60% residue activity at pH 11 for 3 h). Immobilized lipases retained 80% of their activity after 5 cycles and increased enzyme activity (more than 108.7%) after 3 h treatment in tert-butanol. Immobilization of lipase which improved reusability of lipase and provided a chance to expand the application of marine microbial lipase in organic system expanded the application range of lipase to catalyze hydrolysis and esterification in harsh condition.

Yeast as a model for Ras signalling.

PubMed

Tisi, Renata; Belotti, Fiorella; Martegani, Enzo

2014-01-01

For centuries yeast species have been popular hosts for classical biotechnology processes, such as baking, brewing, and wine making, and more recently for recombinant proteins production, thanks to the advantages of unicellular organisms (i.e., ease of genetic manipulation and rapid growth) together with the ability to perform eukaryotic posttranslational modifications. Moreover, yeast cells have been used for few decades as a tool for identifying the genes and pathways involved in basic cellular processes such as the cell cycle, aging, and stress response. In the budding yeast S. cerevisiae the Ras/cAMP/PKA pathway is directly involved in the regulation of metabolism, cell growth, stress resistance, and proliferation in response to the availability of nutrients and in the adaptation to glucose, controlling cytosolic cAMP levels and consequently the cAMP-dependent protein kinase (PKA) activity. Moreover, Ras signalling has been identified in several pathogenic yeasts as a key controller for virulence, due to its involvement in yeast morphogenesis. Nowadays, yeasts are still useful for Ras-like proteins investigation, both as model organisms and as a test tube to study variants of heterologous Ras-like proteins.

Flor Yeast: New Perspectives Beyond Wine Aging

PubMed Central

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

Physiological and environmental control of yeast prions

PubMed Central

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

2014-01-01

Prions are self-perpetuating protein isoforms that cause fatal and incurable neurodegenerative disease in mammals. Recent evidence indicates that a majority of human proteins involved in amyloid and neural inclusion disorders possess at least some prion properties. In lower eukaryotes, such as yeast, prions act as epigenetic elements, which increase phenotypic diversity by altering a range of cellular processes. While some yeast prions are clearly pathogenic, it is also postulated that prion formation could be beneficial in variable environmental conditions. Yeast and mammalian prions have similar molecular properties. Crucial cellular factors and conditions influencing prion formation and propagation were uncovered in the yeast models. Stress-related chaperones, protein quality control deposits, degradation pathways and cytoskeletal networks control prion formation and propagation in yeast. Environmental stresses trigger prion formation and loss, supposedly acting via influencing intracellular concentrations of the prion-inducing proteins, and/or by localizing prionogenic proteins to the prion induction sites via heterologous ancillary helpers. Physiological and environmental modulation of yeast prions points to new opportunities for pharmacological intervention and/or prophylactic measures targeting general cellular systems rather than the properties of individual amyloids and prions. PMID:24236638

Isolation and characterization of ethanol tolerant yeast strains

PubMed Central

Tikka, Chiranjeevi; Osuru, Hari Prasad; Atluri, Navya; Raghavulu, Praveen Chakravarthi Veera; yellapu, Nanda Kumar; Mannur, Ismail Shaik; Prasad, Uppu Venkateswara; Aluru, Sudheer; K, Narasimha Varma; Bhaskar, Matcha

2013-01-01

Yeast strains are commonly associated with sugar rich environments. Various fruit samples were selected as source for isolating yeast cells. The isolated cultures were identified at Genus level by colony morphology, biochemical characteristics and cell morphological characters. An attempt has been made to check the viability of yeast cells under different concentrations of ethanol. Ethanol tolerance of each strain was studied by allowing the yeast to grow in liquid YEPD (Yeast Extract Peptone Dextrose) medium having different concentrations of ethanol. A total of fifteen yeast strains isolated from different samples were used for the study. Seven strains of Saccharomyces cerevisiae obtained from different fruit sources were screened for ethanol tolerance. The results obtained in this study show a range of tolerance levels between 7%-12% in all the stains. Further, the cluster analysis based on 22 RAPD (Random Amplified polymorphic DNA) bands revealed polymorphisms in these seven Saccharomyces strains. PMID:23750092

Phylogenetics of Saccharomycetales, the ascomycete yeasts.

PubMed

Suh, Sung-Oui; Blackwell, Meredith; Kurtzman, Cletus P; Lachance, Marc-André

2006-01-01

Ascomycete yeasts (phylum Ascomycota: subphylum Saccharomycotina: class Saccharomycetes: order Saccharomycetales) comprise a monophyletic lineage with a single order of about 1000 known species. These yeasts live as saprobes, often in association with plants, animals and their interfaces. A few species account for most human mycotic infections, and fewer than 10 species are plant pathogens. Yeasts are responsible for important industrial and biotechnological processes, including baking, brewing and synthesis of recombinant proteins. Species such as Saccharomyces cerevisiae are model organisms in research, some of which led to a Nobel Prize. Yeasts usually reproduce asexually by budding, and their sexual states are not enclosed in a fruiting body. The group also is well defined by synapomorphies visible at the ultrastructural level. Yeast identification and classification changed dramatically with the availability of DNA sequencing. Species identification now benefits from a constantly updated sequence database and no longer relies on ambiguous growth tests. A phylogeny based on single gene analyses has shown the order to be remarkably divergent despite morphological similarities among members. The limits of many previously described genera are not supported by sequence comparisons, and multigene phylogenetic studies are under way to provide a stable circumscription of genera, families and orders. One recent multigene study has resolved species of the Saccharomycetaceae into genera that differ markedly from those defined by analysis of morphology and growth responses, and similar changes are likely to occur in other branches of the yeast tree as additional sequences become available.

Yeast species associated with wine grapes in China.

PubMed

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

Antimicrobial activity of yeasts against some pathogenic bacteria

PubMed Central

Younis, Gamal; Awad, Amal; Dawod, Rehab E.; Yousef, Nehal E.

2017-01-01

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. PMID:28919693

Genomic evolution of the ascomycetous yeasts

USDA-ARS?s Scientific Manuscript database

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 Taphr...

Repeat-Associated Fission Yeast-Like Regional Centromeres in the Ascomycetous Budding Yeast Candida tropicalis

PubMed Central

Chatterjee, Gautam; Sankaranarayanan, Sundar Ram; Guin, Krishnendu; Thattikota, Yogitha; Padmanabhan, Sreedevi; Siddharthan, Rahul; Sanyal, Kaustuv

2016-01-01

The centromere, on which kinetochore proteins assemble, ensures precise chromosome segregation. Centromeres are largely specified by the histone H3 variant CENP-A (also known as Cse4 in yeasts). Structurally, centromere DNA sequences are highly diverse in nature. However, the evolutionary consequence of these structural diversities on de novo CENP-A chromatin formation remains elusive. Here, we report the identification of centromeres, as the binding sites of four evolutionarily conserved kinetochore proteins, in the human pathogenic budding yeast Candida tropicalis. Each of the seven centromeres comprises a 2 to 5 kb non-repetitive mid core flanked by 2 to 5 kb inverted repeats. The repeat-associated centromeres of C. tropicalis all share a high degree of sequence conservation with each other and are strikingly diverged from the unique and mostly non-repetitive centromeres of related Candida species—Candida albicans, Candida dubliniensis, and Candida lusitaniae. Using a plasmid-based assay, we further demonstrate that pericentric inverted repeats and the underlying DNA sequence provide a structural determinant in CENP-A recruitment in C. tropicalis, as opposed to epigenetically regulated CENP-A loading at centromeres in C. albicans. Thus, the centromere structure and its influence on de novo CENP-A recruitment has been significantly rewired in closely related Candida species. Strikingly, the centromere structural properties along with role of pericentric repeats in de novo CENP-A loading in C. tropicalis are more reminiscent to those of the distantly related fission yeast Schizosaccharomyces pombe. Taken together, we demonstrate, for the first time, fission yeast-like repeat-associated centromeres in an ascomycetous budding yeast. PMID:26845548

Comparative genomics of biotechnologically important yeasts

USDA-ARS?s Scientific Manuscript database

Ascomycete yeasts are metabolically diverse, with great potential for biotechnology. Here, we report the comparative genome analysis of 29 taxonomically and biotechnologically important yeasts, including 16 newly sequenced. We identify a genetic code change, CUG-Ala, in Pachysolen tannophilus in the...

Counting Yeast.

ERIC Educational Resources Information Center

Bealer, Jonathan; Welton, Briana

1998-01-01

Describes changes to a traditional study of population in yeast colonies. Changes to the procedures include: (1) only one culture per student team; (2) cultures are inoculated only once; and (3) the same tube is sampled daily. (DDR)

Yeast diversity and native vigor for flavor phenotypes.

PubMed

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.

Making Sense of the Yeast Sphingolipid Pathway.

PubMed

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.

Novel brewing yeast hybrids: creation and application.

PubMed

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.

Improving industrial yeast strains: exploiting natural and artificial diversity

PubMed Central

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

MALDI-TOF MS as a tool to identify foodborne yeasts and yeast-like fungi.

PubMed

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 value<1.7. Two strains could not be identified at first as they represented a mix of two different species. These mixes were Rhodotorula babjevae with Meyerozyma caribbica and Clavispora lusitaniae with Debaryomyces hansenii. After separation, all four species could be correctly identified with scores>1.7. 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.

Comparative genomics of biotechnologically important yeasts

USDA-ARS?s Scientific Manuscript database

Saccharomyces cerevisiae, is used in the vast majority of the world’s bioprocesses, and its economic significance is unchallenged. It, however, represents only a small slice of yeast physiological diversity. Many other yeasts, are used in lesser known, but commercially important processes that take ...

The yeast Golgi apparatus: insights and mysteries

PubMed Central

Papanikou, Effrosyni; Glick, Benjamin S.

2009-01-01

The Golgi apparatus is known to modify and sort newly synthesized secretory proteins. However, fundamental mysteries remain about the structure, operation, and dynamics of this organelle. Important insights have emerged from studying the Golgi in yeasts. For example, yeasts have provided direct evidence for Golgi cisternal maturation, a mechanism that is likely to be broadly conserved. Here, we highlight features of the yeast Golgi as well as challenges that lie ahead. PMID:19879270

Evolution and variation of the yeast (Saccharomyces) genome.

PubMed

Mortimer, R K

2000-04-01

In this review we describe the role of the yeast Saccharomyces in the development of human societies including the use of this organism in the making of wine, bread, beer, and distilled beverages. We also discuss the tremendous diversity of yeast found in natural (i.e., noninoculated) wine fermentations and the scientific uses of yeast over the past 60 years. In conclusion, we present ideas on the model of "genome renewal" and the use of this model to explain the mode by which yeast has evolved and how diversity can be generated.

Candida lipolytica UCP0988 Biosurfactant: Potential as a Bioremediation Agent and in Formulating a Commercial Related Product

PubMed Central

Santos, Danyelle K. F.; Resende, Ana H. M.; de Almeida, Darne G.; Soares da Silva, Rita de Cássia F.; Rufino, Raquel D.; Luna, Juliana M.; Banat, Ibrahim M.; Sarubbo, Leonie A.

2017-01-01

The aim of the present study was to investigate the potential application of the biosurfactant from Candida lipolytica grown in low-cost substrates, which has previously been produced and characterized under optimized conditions as an adjunct material to enhance the remediation processes of hydrophobic pollutants and heavy metals generated by the oil industry and propose the formulation of a safe and stable remediation agent. In tests carried out with seawater, the crude biosurfactant demonstrated 80% oil spreading efficiency. The dispersion rate was 50% for the biosurfactant at a concentration twice that of the CMC. The biosurfactant removed 70% of motor oil from contaminated cotton cloth in detergency tests. The crude biosurfactant also removed 30–40% of Cu and Pb from standard sand, while the isolated biosurfactant removed ~30% of the heavy metals. The conductivity of solutions containing Cd and Pb was sharply reduced after biosurfactants' addition. A product was prepared through adding 0.2% potassium sorbate as preservative and tested over 120 days. The formulated biosurfactant was analyzed for emulsification and surface tension under different pH values, temperatures, and salt concentrations and tested for toxicity against the fish Poecilia vivipara. The results showed that the formulation had no toxicity and did not cause significant changes in the tensoactive capacity of the biomolecule while maintaining activity demonstrating suitability for potential future commercial product formulation. PMID:28507538

Brewer's/baker's yeast (Saccharomyces cerevisiae) and preventive medicine: Part II.

PubMed

Moyad, Mark A

2008-02-01

Yeast is the term generally applied to a unicellular fungus, and there are hundreds of species now identified. One of the most notable and well-known species of yeast in health and wellness is known as Saccharomyces cerevisiae, which is also known by its more common names, brewer's yeast or baker's yeast. Typically, brewer's yeast is used as a protein supplement, energy booster, immune enhancer, or other vehicle where other compounds can be inserted to create a commercialized health product. For example, one of the most notable positive findings was the encouraging results from a large randomized trial of adults recently vaccinated for seasonal influenza who also received an over-the-counter daily adjuvant modified brewer's yeast-based product (EpiCor) to prevent colds and flu symptoms. The modified yeast-based product significantly reduced the incidence and duration of this common condition. Yeast-based technology is also being used as a molecular mechanistic model of caloric restriction (CR) with the goal of improving the human life span. The current and potential impact of yeast-based technology in medicine is encouraging and should receive more attention, but the recent preliminary positive results of CR in humans may be in part due to what has been already learned from brewer's yeast.

The yeast spectrum of the 'tea fungus Kombucha'.

PubMed

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

[Yeast species in vulvovaginitis candidosa].

PubMed

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.

21 CFR 172.381 - Vitamin D2 bakers yeast.

Code of Federal Regulations, 2014 CFR

2014-04-01

... conventional bakers yeast. (c) The additive may be used in yeast-leavened baked goods and baking mixes and yeast-leavened baked snack foods at levels not to exceed 400 International Units of vitamin D2 per 100...

Anaerobic digestion of food waste using yeast.

PubMed

Suwannarat, Jutarat; Ritchie, Raymond J

2015-08-01

Fermentative breakdown of food waste seems a plausible alternative to feeding food waste to pigs, incineration or garbage disposal in tourist areas. We determined the optimal conditions for the fermentative breakdown of food waste using yeast (Saccharomyces cerevisiae) in incubations up to 30days. Yeast efficiently broke down food waste with food waste loadings as high as 700g FW/l. The optimum inoculation was ≈46×10(6)cells/l of culture with a 40°C optimum (25-40°C). COD and BOD were reduced by ≈30-50%. Yeast used practically all the available sugars and reduced proteins and lipids by ≈50%. Yeast was able to metabolize lipids much better than expected. Starch was mobilized after very long term incubations (>20days). Yeast was effective in breaking down the organic components of food waste but CO2 gas and ethanol production (≈1.5%) were only significant during the first 7days of incubations. Copyright © 2015 Elsevier Ltd. All rights reserved.

Nectar yeasts: a natural microcosm for ecology.

PubMed

Chappell, Callie R; Fukami, Tadashi

2018-06-01

The species of yeasts that colonize floral nectar can modify the mutualistic relationships between plants and pollinators by changing the chemical properties of nectar. Recent evidence supporting this possibility has led to increased interest among ecologists in studying these fungi as well as the bacteria that interact with them in nectar. Although not fully explored, nectar yeasts also constitute a promising natural microcosm that can be used to facilitate development of general ecological theory. We discuss the methodological and conceptual advantages of using nectar yeasts from this perspective, including simplicity of communities, tractability of dispersal, replicability of community assembly, and the ease with which the mechanisms of species interactions can be studied in complementary experiments conducted in the field and the laboratory. To illustrate the power of nectar yeasts as a study system, we discuss several topics in community ecology, including environmental filtering, priority effects, and metacommunity dynamics. An exciting new direction is to integrate metagenomics and comparative genomics into nectar yeast research to address these fundamental ecological topics. Copyright © 2018 John Wiley & Sons, Ltd.

Kinetics of growth and sugar consumption in yeasts.

PubMed

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.

The ecology of the Drosophila-yeast mutualism in wineries

PubMed Central

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

The ecology of the Drosophila-yeast mutualism in wineries.

PubMed

Quan, Allison S; Eisen, Michael B

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.

Improving industrial yeast strains: exploiting natural and artificial diversity.

PubMed

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.

Chromosomal Aneuploidy Improves the Brewing Characteristics of Sake Yeast.

PubMed

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

Chromosomal Aneuploidy Improves the Brewing Characteristics of Sake Yeast

PubMed Central

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

2017-01-01

ABSTRACT 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

21 CFR 172.325 - Bakers yeast protein.

Code of Federal Regulations, 2010 CFR

2010-04-01

... harmful microbial toxin. (d) The ingredient is used in food as a nutrient supplement as defined in § 170.3... Special Dietary and Nutritional Additives § 172.325 Bakers yeast protein. Bakers yeast protein may be...

21 CFR 172.325 - Bakers yeast protein.

Code of Federal Regulations, 2012 CFR

2012-04-01

... harmful microbial toxin. (d) The ingredient is used in food as a nutrient supplement as defined in § 170.3... Special Dietary and Nutritional Additives § 172.325 Bakers yeast protein. Bakers yeast protein may be...

21 CFR 172.325 - Bakers yeast protein.

Code of Federal Regulations, 2011 CFR

2011-04-01

... harmful microbial toxin. (d) The ingredient is used in food as a nutrient supplement as defined in § 170.3... Special Dietary and Nutritional Additives § 172.325 Bakers yeast protein. Bakers yeast protein may be...

21 CFR 172.325 - Bakers yeast protein.

Code of Federal Regulations, 2013 CFR

2013-04-01

... harmful microbial toxin. (d) The ingredient is used in food as a nutrient supplement as defined in § 170.3... Special Dietary and Nutritional Additives § 172.325 Bakers yeast protein. Bakers yeast protein may be...

Mammalian amyloidogenic proteins promote prion nucleation in yeast.

PubMed

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.

Inventions on baker's yeast storage and activation at the bakery plant.

PubMed

Gélinas, Pierre

2010-01-01

Baker's yeast is the gas-forming ingredient in bakery products. Methods have been invented to properly handle baker's yeast and optimize its activity at the bakery plant. Over the years, incentives for inventions on yeast storage and activation have greatly changed depending on trends in the baking industry. For example, retailer's devices for cutting bulk pressed yeast and techniques for activating dry yeast have now lost their importance. Review of patents for invention indicates that activation of baker's yeast activity has been a very important issue for bakers, for example, with baking ingredients called yeast foods. In the recent years and especially for highly automated bakeries, interest has moved to equipments and processes for optimized storage of liquid cream yeast to thoroughly control dough fermentation and bread quality.

Guidelines and recommendations on yeast cell death nomenclature.

PubMed

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 accepted set of concepts and terms is still missing. Here, we propose unified criteria for the defi-nition 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 differ-ential definition of terms including apoptosis, regulated necrosis, and autophagic cell death, as we refer to additional cell death rou-tines 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 au-thors, 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 pro-gress of this vibrant field of research.

Guidelines and recommendations on yeast cell death nomenclature

PubMed Central

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

Applications of yeast surface display for protein engineering

PubMed Central

Cherf, Gerald M.; Cochran, Jennifer R.

2015-01-01

The method of displaying recombinant proteins on the surface of Saccharomyces cerevisiae via genetic fusion to an abundant cell wall protein, a technology known as yeast surface display, or simply, yeast display, has become a valuable protein engineering tool for a broad spectrum of biotechnology and biomedical applications. This review focuses on the use of yeast display for engineering protein affinity, stability, and enzymatic activity. Strategies and examples for each protein engineering goal are discussed. Additional applications of yeast display are also briefly presented, including protein epitope mapping, identification of protein-protein interactions, and uses of displayed proteins in industry and medicine. PMID:26060074

The primary structures of two yeast enolase genes. Homology between the 5' noncoding flanking regions of yeast enolase and glyceraldehyde-3-phosphate dehydrogenase genes.

PubMed

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

[The yeast biofilm in human medicine].

PubMed

Růzicka, Filip; Holá, Veronika; Votava, Miroslav

2007-08-01

In recent years, the role of Candida yeasts as causative agents of nosocomial infections has increased. One of the important virulence factors contributing to the development of such infections is biofilm production. This virulence factor enables yeast to colonize both native surfaces and artificial implants. The most common sources of infection are patients themselves, in particular the gastrointestinal tract and skin. The vectors of exogenous yeast infections are predominantly the hands of the health personnel and contaminated medical instruments. The adhesion of yeasts to the implant surfaces is determined both by implant surface and yeast characteristics. This is followed by proliferation and production of microcolonies and extracellular matrix. The final biofilm structure is also influenced by the production of hyphae and pseudohyphae. The entire process of biofilm production is controlled by numerous regulatory systems, with the key role being played by the quorum sensing system. Like the adhered bacterial cultures, candidas growing in the form of a biofilm are highly resistant to antimicrobial therapy. Resistance of yeast biofilms to antifungals is a complex process with multiple contributing factors. These are especially increased gene expression (e.g. genes encoding the so called multidrug efflux pumps), limited penetration of substances through the extracellular matrix, inhibited cell growth and altered microenvironment in deeper biofilm layers. The concentrations of antifungals able to effectively affect the biofilm cells exceed, by several orders of magnitude, the values of conventionally determined MICs. High biofilm resistance results in ineffective antifungal therapy of biofilm infections. Therefore, if possible, the colonized implant should be removed. Conservative therapy should involve antifungals with a proven effect on the biofilm (e.g. caspofungin). The most effective measure in fighting biofilm infections is prevention, especially adhering to

Heavy metal removal by caustic-treated yeast immobilized in alginate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Y.; Wilkins, E.

1995-12-31

Saccharomyces cerevisiae yeast biomass was treated with hot alkali to increase its biosorption capacity for heavy metals and then was immobilized in alginate gel. Biosorption capacities for Cu{sup 2+}, Cd{sup 2+}, and Zn{sup 2+} on alginate gel, native yeast, native yeast immobilized in alginate gel, and caustic-treated yeast immobilized in alginate gel were all compared. Immobilized yeasts could be reactivated and reused in a manner similar to the ion exchange resins. Immobilized caustic-treated yeast has high heavy metal biosorption capacity and high metal removal efficiency in a rather wide acidic pH region. The biosorption isotherm of immobilized caustic-treated yeast wasmore » studied, and empirical equations were obtained. The initial pH of polluted water affected the metal removal efficiency significantly, and the equilibrium biosorption capacity seemed to be temperature independent at lower initial metal concentrations.« less

A Proteome-wide Fission Yeast Interactome Reveals Network Evolution Principles from Yeasts to Human.

PubMed

Vo, Tommy V; Das, Jishnu; Meyer, Michael J; Cordero, Nicolas A; Akturk, Nurten; Wei, Xiaomu; Fair, Benjamin J; Degatano, Andrew G; Fragoza, Robert; Liu, Lisa G; Matsuyama, Akihisa; Trickey, Michelle; Horibata, Sachi; Grimson, Andrew; Yamano, Hiroyuki; Yoshida, Minoru; Roth, Frederick P; Pleiss, Jeffrey A; Xia, Yu; Yu, Haiyuan

2016-01-14

Here, we present FissionNet, a proteome-wide binary protein interactome for S. pombe, comprising 2,278 high-quality interactions, of which ∼ 50% were previously not reported in any species. FissionNet unravels previously unreported interactions implicated in processes such as gene silencing and pre-mRNA splicing. We developed a rigorous network comparison framework that accounts for assay sensitivity and specificity, revealing extensive species-specific network rewiring between fission yeast, budding yeast, and human. Surprisingly, although genes are better conserved between the yeasts, S. pombe interactions are significantly better conserved in human than in S. cerevisiae. Our framework also reveals that different modes of gene duplication influence the extent to which paralogous proteins are functionally repurposed. Finally, cross-species interactome mapping demonstrates that coevolution of interacting proteins is remarkably prevalent, a result with important implications for studying human disease in model organisms. Overall, FissionNet is a valuable resource for understanding protein functions and their evolution. Copyright © 2016 Elsevier Inc. All rights reserved.

Actin and Endocytosis in Budding Yeast

PubMed Central

Goode, Bruce L.; Eskin, Julian A.; Wendland, Beverly

2015-01-01

Endocytosis, the process whereby the plasma membrane invaginates to form vesicles, is essential for bringing many substances into the cell and for membrane turnover. The mechanism driving clathrin-mediated endocytosis (CME) involves > 50 different protein components assembling at a single location on the plasma membrane in a temporally ordered and hierarchal pathway. These proteins perform precisely choreographed steps that promote receptor recognition and clustering, membrane remodeling, and force-generating actin-filament assembly and turnover to drive membrane invagination and vesicle scission. Many critical aspects of the CME mechanism are conserved from yeast to mammals and were first elucidated in yeast, demonstrating that it is a powerful system for studying endocytosis. In this review, we describe our current mechanistic understanding of each step in the process of yeast CME, and the essential roles played by actin polymerization at these sites, while providing a historical perspective of how the landscape has changed since the preceding version of the YeastBook was published 17 years ago (1997). Finally, we discuss the key unresolved issues and where future studies might be headed. PMID:25657349

Organoleptic Analysis of Doughs Fermented with Yeasts From A Nigerian Palm Wine (Elaeis guineensis) and Certain Commercial Yeasts

PubMed Central

B, Boboye; I, Dayo-Owoyemi; F. A, Akinyosoye

2008-01-01

Yeasts isolated from a freshly tapped palm wine obtained from Akure, Nigeria were identified as Schizosaccharomyces pombe, Saccharomyces cerevisiae, Debaryomyces hansenii, Geotrichum lactis and Zygosaccharomyces rouxii. Each of the isolates was used to ferment wheat flour dough and baked. Sensory analysis of the doughs was carried out on leavening, texture, aroma, taste and appearance. Saccharomyces cerevisiae performed best in leavening the dough while Debaryomyces hansenii produced doughs with the best taste and aroma. Appearances of the doughs made with all the isolated yeasts did not differ significantly (P<0.05) from that of the dough that lacked yeast. PMID:19088921

[Malassezia yeasts and their significance in dermatology].

PubMed

Hort, W; Nilles, M; Mayser, P

2006-07-01

Yeasts of the genus Malassezia belong to the normal microflora of the human skin. In addition they are known to cause a variety of skin diseases; the most frequent of which is pityriasis versicolor. Malassezia yeasts are also thought to be associated with seborrheic dermatitis, dandruff and Malassezia folliculitis. Recently the significance of Malassezia yeasts as a trigger factor for atopic dermatitis of the head and neck region has been pointed out. The role of the Malassezia yeasts in these different diseases has been controversial in the past and remains an issue because of difficulties in isolation, culture and differentiation of the organism. Thanks to molecular techniques, 10 species can actually be differentiated. The article presents the different Malassezia-associated diseases, their clinical picture, diagnosis and appropriate therapy. In addition the speciation of Malassezia is reviewed.

Yeast as a tool to identify anti-aging compounds

PubMed Central

Zimmermann, Andreas; Hofer, Sebastian; Pendl, Tobias; Kainz, Katharina; Madeo, Frank; Carmona-Gutierrez, Didac

2018-01-01

Abstract In the search for interventions against aging and age-related diseases, biological screening platforms are indispensable tools to identify anti-aging compounds among large substance libraries. The budding yeast, Saccharomyces cerevisiae, has emerged as a powerful chemical and genetic screening platform, as it combines a rapid workflow with experimental amenability and the availability of a wide range of genetic mutant libraries. Given the amount of conserved genes and aging mechanisms between yeast and human, testing candidate anti-aging substances in yeast gene-deletion or overexpression collections, or de novo derived mutants, has proven highly successful in finding potential molecular targets. Yeast-based studies, for example, have led to the discovery of the polyphenol resveratrol and the natural polyamine spermidine as potential anti-aging agents. Here, we present strategies for pharmacological anti-aging screens in yeast, discuss common pitfalls and summarize studies that have used yeast for drug discovery and target identification. PMID:29905792

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

NASA Astrophysics Data System (ADS)

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.

[Treatment of oil-manufacturing wastewater by yeast-SBR system].

PubMed

Lü, Wen-zhou; Liu, Ying; Huang, Yi-zhen

2008-04-01

Eight yeast strains were applied to a sequencing batch reactor (SBR) to treat high-strength oil-containing wastewater. The removal performance, yeast cultivation method and key factors affecting the stability of system were discussed. The results show yeast sludge with MLSS of 19 g/L and SVI of 35 mL/g can be obtained in 6 d in an open system without any molds and bacteria inhibitor addition; In 30 d continuous wastewater treatment, COD and oil removal rate achieve 86.8%-96.9% and above 99.5% respectively under the influent conditions of the COD of 9000-23000 mg/L and oil of 4500-16000 mg/L; Short period of pH impact brings reversible effects on the system and the sludge retention time can affect the SVI of the yeast; Absence of nitrogen induces morphology conversion of some yeast cells from single cell to filamentous one and impairs the settling capability of the yeast.

The Yeast Deletion Collection: A Decade of Functional Genomics

PubMed Central

Giaever, Guri; Nislow, Corey

2014-01-01

The yeast deletion collections comprise >21,000 mutant strains that carry precise start-to-stop deletions of ∼6000 open reading frames. This collection includes heterozygous and homozygous diploids, and haploids of both MATa and MATα mating types. The yeast deletion collection, or yeast knockout (YKO) set, represents the first and only complete, systematically constructed deletion collection available for any organism. Conceived during the Saccharomyces cerevisiae sequencing project, work on the project began in 1998 and was completed in 2002. The YKO strains have been used in numerous laboratories in >1000 genome-wide screens. This landmark genome project has inspired development of numerous genome-wide technologies in organisms from yeast to man. Notable spinoff technologies include synthetic genetic array and HIPHOP chemogenomics. In this retrospective, we briefly describe the yeast deletion project and some of its most noteworthy biological contributions and the impact that these collections have had on the yeast research community and on genomics in general. PMID:24939991

Triacetic acid lactone production in industrial Saccharomyces yeast strains

USDA-ARS?s Scientific Manuscript database

Triacetic acid lactone (TAL) is a potential platform chemical that can be produced in yeast. To evaluate the potential for industrial yeast strains to produce TAL, the g2ps1 gene encoding 2-pyrone synthase was transformed into thirteen industrial yeast strains of varied genetic background. TAL produ...

Yeasts as important agents of onychomycosis: in vitro activity of propolis against yeasts isolated from patients with nail infection.

PubMed

Khosravi, Ali Reza; Shokri, Hojjatollah; Nikaein, Donya; Mansouri, Parvin; Erfanmanesh, Ahmad; Chalangari, Reza; Katalin, Martis

2013-01-01

The purposes of this study were to determine the frequency of the yeast species obtained from patients with clinical features of onychomycosis and the in vitro antifungal susceptibility of the yeast species to propolis. A prospective study was carried out at the Mycology Research Center in Iran from 2010 to 2011. Clinical diagnosis was performed by direct microscopic examination and culture. Different yeast species were identified by morphological and biochemical tests. An antifungal susceptibility test to fluconazole (FLU) and propolis by the broth microdilution method was performed on each isolate. One hundred and twenty-eight fungal isolates were obtained. The most prevalent fungi were yeasts (81, 63.2%), dermatophytes (36, 28.1%), and nondermatophyte fungi (11, 8.6%). Fingernails were more affected than toenails (65.4% vs. 19.8%, respectively). The most frequently found species was Candida albicans (38.5%), followed by Candida spp. (23.1%), C. tropicalis (10.8%), C. kefyr (6.2%), C. krusei (3.1%), Malassezia globosa (4.6%), M. slooffiae (4.6%), and M. pachydermatis (1.5%). Of all yeast isolates (65), seven showed resistance to FLU. The average MIC of propolis for FLU-susceptible isolates was 5.8 μg/mL, whereas this value was 12.25 μg/mL for FLU-resistant isolates. Our results proved that the propolis inhibits the growth of pathogenic yeasts and confirmed the efficiency of propolis as an anti-Candida and anti-Malassezia agent.

Astaxanthinogenesis in the yeast Phaffia rhodozyma - optimization of low-cost culture media and yeast cell-wall lysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fontana, J.D.; Baron, M.; Guimaraes, M.F.

Astaxanthin is a diketo-dihydroxy-carotenoid produced by Phaffia rhodozyma, a basidiomicetous yeast. A low-cost fermentation medium consisting of raw sugarcane juice and urea was developed to exploit the active sucrolytic/urelolytic enzyme apparatus inherent to the yeast. As compared to the beneficial effect of 0.1 g% urea, a ready nitrogen source, mild phosphoric pre inversion of juice sucrose to glucose and fructose, promptly fermentable carbon sources, resulted in smaller benefits. Corn steep liquor (CSL) was found to be a valuable supplement for both yeast biomass yield (9.2 g dry cells/L) and astaxanthin production (1.3 mg/g cells). Distillery effluent (vinace), despite only amore » slightly positive effect on yeast growth, allowed for the highest pigment productivity (1.9 mg/g cells). Trace amounts of Ni{sup 2} (1 mg/L, as a cofactor for urease) resulted in controversial effects, namely, biomass decrease and astaxanthin increase, with no effect on the release (and uptake) of ammonium ion from urea. 13 refs., 6 figs.« less

The yeast replicative aging model.

PubMed

He, Chong; Zhou, Chuankai; Kennedy, Brian K

2018-03-08

It has been nearly three decades since the budding yeast Saccharomyces cerevisiae became a significant model organism for aging research and it has emerged as both simple and powerful. The replicative aging assay, which interrogates the number of times a "mother" cell can divide and produce "daughters", has been a stalwart in these studies, and genetic approaches have led to the identification of hundreds of genes impacting lifespan. More recently, cell biological and biochemical approaches have been developed to determine how cellular processes become altered with age. Together, the tools are in place to develop a holistic view of aging in this single-celled organism. Here, we summarize the current state of understanding of yeast replicative aging with a focus on the recent studies that shed new light on how aging pathways interact to modulate lifespan in yeast. Copyright © 2018. Published by Elsevier B.V.

New lager yeast strains generated by interspecific hybridization.

PubMed

Krogerus, Kristoffer; Magalhães, Frederico; Vidgren, Virve; Gibson, Brian

2015-05-01

The interspecific hybrid Saccharomyces pastorianus is the most commonly used yeast in brewery fermentations worldwide. Here, we generated de novo lager yeast hybrids by mating a domesticated and strongly flocculent Saccharomyces cerevisiae ale strain with the Saccharomyces eubayanus type strain. The hybrids were characterized with respect to the parent strains in a wort fermentation performed at temperatures typical for lager brewing (12 °C). The resulting beers were analysed for sugar and aroma compounds, while the yeasts were tested for their flocculation ability and α-glucoside transport capability. These hybrids inherited beneficial properties from both parent strains (cryotolerance, maltotriose utilization and strong flocculation) and showed apparent hybrid vigour, fermenting faster and producing beer with higher alcohol content (5.6 vs 4.5 % ABV) than the parents. Results suggest that interspecific hybridization is suitable for production of novel non-GM lager yeast strains with unique properties and will help in elucidating the evolutionary history of industrial lager yeast.

Interactions between yeast lees and wine polyphenols during simulation of wine aging. II. Analysis of desorbed polyphenol compounds from yeast lees.

PubMed

Mazauric, Jean-Paul; Salmon, Jean-Michel

2006-05-31

In the first part of this work, the analysis of the polyphenolic compounds remaining in the wine after different contact times with yeast lees during simulation of red wine aging was undertaken. To achieve a more precise view of the wine polyphenols adsorbed on lees during red wine aging and to establish a clear balance between adsorbed and remnant polyphenol compounds, the specific analysis of the chemical composition of the adsorbed polyphenolic compounds (condensed tannins and anthocyanins) after their partial desorbtion from yeast lees by denaturation treatments was realized in the second part of the study. The total recovery of polyphenol compounds from yeast lees was not complete, since a rather important part of the initial wine colored polyphenols, especially those with a dominant blue color component, remained strongly adsorbed on yeast lees, as monitored by color tristimulus and reflectance spectra measurements. All anthocyanins were recovered at a rather high percentage (about 62%), and it was demonstrated that they were not adsorbed in relation with their sole polarity. Very few monomeric phenolic compounds were extracted from yeast lees. With the use of drastic denaturing treatments, the total recovery of condensed tannins reached 83%. Such tannins extracted from yeast lees exhibited very high polymeric size and a rather high percentage of galloylated residues by comparison with initial wine tannins, indicating that nonpolar tannins were preferentially desorbed from yeast lees by the extraction treatments.

Vaginal Yeast Infections

MedlinePlus

... for sure if yogurt with Lactobacillus or other probiotics can prevent or treat vaginal yeast infections. If ... Chen, H., et al. (2013). Impact of eating probiotic yogurt on colonization by Candida species of the ...

Oxidative Stress and Programmed Cell Death in Yeast

PubMed Central

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. PMID:22737670

[Yeast microbiota in artisanal cheeses from Corrientes, Argentina].

PubMed

Cardozo, Marina C; Fusco, Ángel J V; Carrasco, Marta S

The artisanal cheese from Corrientes (from the Spanish acronym QAC-Queso Artesanal de Corrientes/Artisanal Cheese from Corrientes) is a soft cheese elaborated with raw cow milk and an artisanal coagulant agent. Lactic bacteria contitute the main flora of this cheese although yeasts are also present in high quantities as secondary microbiota and might play a relevant role in cheese ripening. The aim of this work was to evaluate yeast occurrence during QAC elaboration and ripening, and the effect of seasonal variation. Yeasts were isolated and purified from raw materials and cheese at different ripening stagesl elaborated during the different seasons. Yeast sample counts were in the order of 10 3 - 10 7 UFC/ml o UFC/g. Ninety yeast strains were classified: 9 from milk, 28 from the coagulant agent, 10 from curd and 43 from cheese. Candida predominated in milk samples while other yeast genera had low incidence. Candida also predominated in the coagulant agent samples, followed by genera Myxozyma and Debaryomyces. The isolates obtained from cheese belonged to the same genera predominating in the coagulant agent, and showed the same order of prevalence. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

A Comparison of the Beneficial Effects of Live and Heat-Inactivated Baker's Yeast on Nile Tilapia: Suggestions on the Role and Function of the Secretory Metabolites Released from the Yeast.

PubMed

Ran, Chao; Huang, Lu; Liu, Zhi; Xu, Li; Yang, Yalin; Tacon, Philippe; Auclair, Eric; Zhou, Zhigang

2015-01-01

Yeast is frequently used as a probiotic in aquaculture with the potential to substitute for antibiotics. In this study, the involvement and extent to which the viability of yeast cells and thus the secretory metabolites released from the yeast contribute to effects of baker's yeast was investigated in Nile tilapia. No yeast, live yeast or heat-inactivated baker's yeast were added to basal diets high in fishmeal and low in soybean (diet A) or low in fishmeal and high in soybean (diet B), which were fed to fish for 8 weeks. Growth, feed utilization, gut microvilli morphology, and expressions of hsp70 and inflammation-related cytokines in the intestine and head kidney were assessed. Intestinal microbiota was investigated using 16S rRNA gene pyrosequencing. Gut alkaline phosphatase (AKP) activity was measured after challenging the fish with Aeromonas hydrophila. Results showed that live yeast significantly improved FBW and WG (P < 0.05), and tended to improve FCR (P = 0.06) of fish compared to the control (no yeast). No significant differences were observed between inactivated yeast and control. Live yeast improved gut microvilli length (P < 0.001) and density (P < 0.05) while inactivated yeast did not. The hsp70 expression level in both the intestine and head kidney of fish was significantly reduced by live yeast (P < 0.05) but not inactivated yeast. Live yeast but not inactivated yeast reduced intestinal expression of tnfα (P < 0.05), tgfβ (P < 0.05 under diet A) and il1β (P = 0.08). Intestinal Lactococcus spp. numbers were enriched by both live and inactivated yeast. Lastly, both live and inactivated yeast reduced the gut AKP activity compared to the control (P < 0.001), indicating protection of the host against infection by A. hydrophila. In conclusion, secretory metabolites did not play major roles in the growth promotion and disease protection effects of yeast. Nevertheless, secretory metabolites were the major contributing factor towards improved gut

The impact of yeast fermentation on dough matrix properties.

PubMed

Rezaei, Mohammad N; Jayaram, Vinay B; Verstrepen, Kevin J; Courtin, Christophe M

2016-08-01

Most studies on dough properties are performed on yeastless dough to exclude the complicating, time-dependent effect of yeast. Baker's yeast, however, impacts dough matrix properties during fermentation, probably through the production of primary (CO2 and ethanol) and secondary (glycerol, acetic acid and succinic acid) metabolites. The aim of this study is to obtain a better understanding of the changes in yeasted dough behavior introduced by fermentation, by investigating the impact of yeast fermentation on Farinograph dough consistency, dough spread, Kieffer rig dough extensibility and gluten agglomeration behavior in a fermented dough-batter gluten starch separation system. Results show that fermentation leads to a dough with less flow and lower extensibility that breaks more easily under stress and strain. The dough showed less elastic and more plastic deformation behavior. Gluten agglomerates were smaller for yeasted dough than for the unyeasted control. These changes probably have to be attributed to metabolites generated during fermentation. Indeed, organic acids and also ethanol in concentrations produced by yeast were previously shown to have similar effects in yeastless dough. These findings imply the high importance of yeast fermentation metabolites on dough matrix properties in industrial bread production. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Yeast as a model system for mammalian seven-transmembrane segment receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jeansonne, N.E.

1994-05-01

Investigators have used the budding yeast Saccharomyces cerevisiae as a model system in which to study the {beta}-adrenergic receptor, the T-cell receptor pathway, initiation of mammalian DNA replication, initiation of mammalian transcription, secretion, the CDC2 kinase system, cell cycle control, and aging, as well as the function of oncogenes. This list continues to growth with the discovery of an immunoglobulin heavy-chain binding homologue in yeast, an Rb binding protein homologue, and a possible yeast arrestin. Yeast is relatively easy to maintain, to grow, and to genetically manipulate. A single gene can be overexpressed, selectively mutated or deleted from its chromosomalmore » location. In this way, the in vivo function of a gene can be studied. It has become reasonable to consider yeast as a model system for studying the seven transmembrane segments (7-TMS) receptor family. Currently, subtypes of the {beta}-adrenergic receptor are being studied in yeast. The receptor and its G{sub {alpha}}-G-protein, trigger the mating pheromone receptor pathway. This provides a powerful assay for determining receptor function. Studies expressing the muscarinic cholinergic receptor in yeast are underway. The yeast pheromone receptor belongs to this receptor family, sharing sequences and secondary structure homology. An effective strategy has been to identify a yeast pathway or process which is homologous to a mammalian system. The pathway is delineated in yeast, identifying other genetic components. Then yeast genes are used to screen for human homologues of these components. The putative human homologues are then expressed in yeast and in mammalian cells to determine function. When this type of {open_quotes}mixing and matching{close_quotes} works, yeast genetics can be a powerful tool. 115 refs.« less

Nonlinear Dielectric Properties of Yeast Cells Cultured in Different Environmental Conditions

NASA Astrophysics Data System (ADS)

Kawanishi, Gomon; Fukuda, Naoki; Muraji, Masafumi

The harmonics of the electric current through yeast suspensions, the nonlinear dielectric properties of yeast cells, have particular patterns according to the biological activity of the cells and the measurement of these patterns is a technique for determining the activity of living cells. The concentration of glucose and oxygen in yeast culture medium influences the manifestation of fermentation or respiration of yeast cells. Measurements were made with yeast cells (Saccharomyces cerevisiae) cultured aerobically and anaerobically in sufficient glucose concentration, aerobic fermentation and anaerobic fermentation, and aerobically in limited glucose concentration, respiration. The results showed that the harmonics were barely apparent for yeast cells in aerobic fermentation and respiratory; however, cells in the anaerobic fermentation displayed substantial third and fifth harmonics. We can say that environmental condition affects the yeast cells' nonlinear properties, from another viewpoint, the measurements of the nonlinear properties are available to determine the activity of yeast cells adjusted to the conditions of their cultivation.

Diversity and killer activity of yeasts in Malaysian fermented food samples.

PubMed

Lim, S L; Tay, S T

2011-08-01

The biodiversity and the killer activity of yeasts isolated from various types of fermented food in Malaysia were investigated in this study. Of 252 yeasts isolated from 48 fermented food samples in this study, 19 yeast species were identified based on sequence analysis of the ITS1-5.8S-ITS2 partial fragments of the yeasts. A total of 29 (11.5%) of the yeast isolates demonstrated killer activity to at least one Candida species tested in this study; including 22 isolates of Trichosporon asahii, 4 isolates of Pichia anomala, and one isolate each of Pichia norvegensis, Pichia fermentans and Issatchenkia orientalis, respectively. The presence of killer yeasts reflects antagonism that occurs during microbial interaction in the fermented food, whereby certain yeasts produce killer toxins and possibly other toxic substances in competition for limited nutrients and space. The anti-Candida activity demonstrated by killer yeasts in this study should be further explored for development of alternative therapy against candidiasis.

Yeast Infection Test

MedlinePlus

... infections of the skin and genitals. Serious yeast infections occur more often in hospital patients and in people with weakened immune systems. References Centers for Disease Control and Prevention [Internet]. Atlanta: U.S. Department of Health ...

Biocavity laser spectroscopy of genetically altered yeast cells and isolated yeast mitochondria

NASA Astrophysics Data System (ADS)

Gourley, Paul L.; Hendricks, Judy K.; McDonald, Anthony E.; Copeland, R. Guild; Naviaux, Robert K.; Yaffe, Michael P.

2006-02-01

We report an analysis of 2 yeast cell mutants using biocavity laser spectroscopy. The two yeast strains differed only by the presence or absence of mitochondrial DNA. Strain 104 is a wild-type (ρ +) strain of the baker's yeast, Saccharomyces cerevisiae. Strain 110 was derived from strain 104 by removal of its mitochondrial DNA (mtDNA). Removal of mtDNA causes strain 110 to grow as a "petite" (ρ -), named because it forms small colonies (of fewer cells because it grows more slowly) on agar plates supplemented with a variety of different carbon sources. The absence of mitochondrial DNA results in the complete loss of all the mtDNA-encoded proteins and RNAs, and loss of the pigmented, heme-containing cytochromes a and b. These cells have mitochondria, but the mitochondria lack the normal respiratory chain complexes I, III, IV, and V. Complex II is preserved because its subunits are encoded by genes located in nuclear DNA. The frequency distributions of the peak shifts produced by wild-type and petite cells and mitochondria show striking differences in the symmetry and patterns of the distributions. Wild-type ρ + cells (104) and mitochondria produced nearly symmetric, Gaussian distributions. The ρ - cells (110) and mitochondria showed striking asymmetry and skew that appeared to follow a Poisson distribution.

Efforts to make and apply humanized yeast

PubMed Central

Laurent, Jon M.; Young, Jonathan H.; Kachroo, Aashiq H.

2016-01-01

Despite a billion years of divergent evolution, the baker’s yeast Saccharomyces cerevisiae has long proven to be an invaluable model organism for studying human biology. Given its tractability and ease of genetic manipulation, along with extensive genetic conservation with humans, it is perhaps no surprise that researchers have been able to expand its utility by expressing human proteins in yeast, or by humanizing specific yeast amino acids, proteins or even entire pathways. These methods are increasingly being scaled in throughput, further enabling the detailed investigation of human biology and disease-specific variations of human genes in a simplified model organism. PMID:26462863

The Influence of Heating Mains on Yeast Communities in Urban Soils

NASA Astrophysics Data System (ADS)

Tepeeva, A. N.; Glushakova, A. M.; Kachalkin, A. V.

2018-04-01

The number and species diversity of yeasts in urban soils (urbanozems) affected by heating mains and in epiphytic yeast complexes of grasses growing above them were studied. The number of yeasts in the soil reached 103-104 CFU/g; on the plants, 107 CFU/g. Significant (by an order of magnitude) increase in the total number of soil yeasts in the zone of heating mains in comparison with the surrounding soil was found in winter period. Overall, 25 species of yeasts were isolated in our study. Yeast community of studied urbanozems was dominated by the Candida sake, an eurybiont of the temperate zone and other natural ecotopes with relatively low temperatures, but its share was minimal in the zone of heating mains. In general, the structure of soil and epiphytic yeast complexes in the zones of heating mains differed from that in the surrounding area by higher species diversity and a lower share of pigmented species among the epiphytic yeasts. The study demonstrated that the number and species structure of soil yeast communities in urban soils change significantly under the influence of the temperature factor and acquire a mosaic distribution pattern.

Genetic Polymorphism in Wine Yeasts: Mechanisms and Methods for Its Detection

PubMed Central

Guillamón, José M.; Barrio, Eladio

2017-01-01

The processes of yeast selection for using as wine fermentation starters have revealed a great phenotypic diversity both at interspecific and intraspecific level, which is explained by a corresponding genetic variation among different yeast isolates. Thus, the mechanisms involved in promoting these genetic changes are the main engine generating yeast biodiversity. Currently, an important task to understand biodiversity, population structure and evolutionary history of wine yeasts is the study of the molecular mechanisms involved in yeast adaptation to wine fermentation, and on remodeling the genomic features of wine yeast, unconsciously selected since the advent of winemaking. Moreover, the availability of rapid and simple molecular techniques that show genetic polymorphisms at species and strain levels have enabled the study of yeast diversity during wine fermentation. This review will summarize the mechanisms involved in generating genetic polymorphisms in yeasts, the molecular methods used to unveil genetic variation, and the utility of these polymorphisms to differentiate strains, populations, and species in order to infer the evolutionary history and the adaptive evolution of wine yeasts, and to identify their influence on their biotechnological and sensorial properties. PMID:28522998

Nectar yeasts warm the flowers of a winter-blooming plant

PubMed Central

Herrera, Carlos M.; Pozo, María I.

2010-01-01

Yeasts are ubiquitous in terrestrial and aquatic microbiota, yet their ecological functionality remains relatively unexplored in comparison with other micro-organisms. This paper formulates and tests the novel hypothesis that heat produced by the sugar catabolism of yeast populations inhabiting floral nectar can increase the temperature of floral nectar and, more generally, modify the within-flower thermal microenvironment. Two field experiments were designed to test this hypothesis for the winter-blooming herb Helleborus foetidus (Ranunculaceae). In experiment 1, the effect of yeasts on the within-flower thermal environment was tested by excluding them from flowers, while in experiment 2 the test involved artificial inoculation of virgin flowers with yeasts. Nectary temperature (Tnect), within-flower air temperature (Tflow) and external air temperature (Tair) were measured on experimental and control flowers in both experiments. Experimental exclusion of yeasts from the nectaries significantly reduced, and experimental addition of yeasts significantly increased, the temperature excess of nectaries (ΔTnect = Tnect − Tair) and the air space inside flowers in relation to the air just outside the flowers. In non-experimental flowers exposed to natural pollinator visitation, ΔTnect was linearly related to log yeast cell density in nectar, and reached +6°C in nectaries with the densest yeast populations. The warming effect of nectar-dwelling yeasts documented in this study suggests novel ecological mechanisms potentially linking nectarivorous microbes with winter-blooming plants and their insect pollinators. PMID:20147331

Glycosylceramide modifies the flavor and metabolic characteristics of sake yeast.

PubMed

Ferdouse, Jannatul; Yamamoto, Yuki; Taguchi, Seiga; Yoshizaki, Yumiko; Takamine, Kazunori; Kitagaki, Hiroshi

2018-01-01

In the manufacture of sake, Japanese traditional rice wine, sake yeast is fermented with koji, which is steamed rice fermented with the non-pathogenic fungus Aspergillus oryzae . During fermentation, sake yeast requires lipids, such as unsaturated fatty acids and sterols, in addition to substances provided by koji enzymes for fermentation. However, the role of sphingolipids on the brewing characteristics of sake yeast has not been studied. In this study, we revealed that glycosylceramide, one of the sphingolipids abundant in koji, affects yeast fermentation. The addition of soy, A. oryzae , and Grifola frondosa glycosylceramide conferred a similar effect on the flavor profiles of sake yeast. In particular, the addition of A. oryzae and G. frondosa glycosylceramide were very similar in terms of the decreases in ethyl caprylate and ethyl 9-decenoate. The addition of soy glycosylceramide induced metabolic changes to sake yeast such as a decrease in glucose, increases in ethanol and glycerol and changes in several amino acids and organic acids concentrations. Tricarboxylic acid (TCA) cycle, pyruvate metabolism, starch and sucrose metabolism, and glycerolipid metabolism were overrepresented in the cultures incubated with sake yeast and soy glycosylceramide. This is the first study of the effect of glycosylceramide on the flavor and metabolic profile of sake yeast.

Brewer's/baker's yeast (Saccharomyces cerevisiae) and preventive medicine: part I.

PubMed

Moyad, Mark A

2007-12-01

Yeast is the term generally applied to a unicellular fungus, and there are hundreds of species now identified. One of the most notable and well-known species of yeast in health and wellness is known as Saccharomyces cerevisiae, which is also known by its more common names, brewer's yeast or baker's yeast. It is usually grown on hops or another substrate similar to the plant utilized in the beer-making industry, after which it is harvested and killed. The final product is generally half composed of protein, as well as a large amount of B vitamins and minerals, and depending on the technology, a diverse number of other healthy compounds. Typically, brewer's yeast is used as a protein supplement, energy booster, immune enhancer, or other vehicle where other compounds can be inserted to create a commercialized health product. A more extensive review of the preventive medical aspects of yeast will be covered in Part 2 of this article to be published in a future issue of Urologic Nursing. Yeast-based technology is also being used as a molecular mechanistic model of caloric restriction with the goal of improving the human life span. The current and potential impact of yeast-based technology in medicine is encouraging.

Glycosylceramide modifies the flavor and metabolic characteristics of sake yeast

PubMed Central

Taguchi, Seiga; Yoshizaki, Yumiko; Takamine, Kazunori

2018-01-01

In the manufacture of sake, Japanese traditional rice wine, sake yeast is fermented with koji, which is steamed rice fermented with the non-pathogenic fungus Aspergillus oryzae. During fermentation, sake yeast requires lipids, such as unsaturated fatty acids and sterols, in addition to substances provided by koji enzymes for fermentation. However, the role of sphingolipids on the brewing characteristics of sake yeast has not been studied. In this study, we revealed that glycosylceramide, one of the sphingolipids abundant in koji, affects yeast fermentation. The addition of soy, A. oryzae, and Grifola frondosa glycosylceramide conferred a similar effect on the flavor profiles of sake yeast. In particular, the addition of A. oryzae and G. frondosa glycosylceramide were very similar in terms of the decreases in ethyl caprylate and ethyl 9-decenoate. The addition of soy glycosylceramide induced metabolic changes to sake yeast such as a decrease in glucose, increases in ethanol and glycerol and changes in several amino acids and organic acids concentrations. Tricarboxylic acid (TCA) cycle, pyruvate metabolism, starch and sucrose metabolism, and glycerolipid metabolism were overrepresented in the cultures incubated with sake yeast and soy glycosylceramide. This is the first study of the effect of glycosylceramide on the flavor and metabolic profile of sake yeast. PMID:29761062

Dielectric modelling of cell division for budding and fission yeast

NASA Astrophysics Data System (ADS)

Asami, Koji; Sekine, Katsuhisa

2007-02-01

The frequency dependence of complex permittivity or the dielectric spectrum of a system including a cell in cell division has been simulated by a numerical technique based on the three-dimensional finite difference method. Two different types of cell division characteristic of budding and fission yeast were examined. The yeast cells are both regarded as a body of rotation, and thus have anisotropic polarization, i.e. the effective permittivity of the cell depends on the orientation of the cell to the direction of an applied electric field. In the perpendicular orientation, where the rotational axis of the cell is perpendicular to the electric field direction, the dielectric spectra for both yeast cells included one dielectric relaxation and its intensity depended on the cell volume. In the parallel orientation, on the other hand, two dielectric relaxations appeared with bud growth for budding yeast and with septum formation for fission yeast. The low-frequency relaxation was shifted to a lower frequency region by narrowing the neck between the bud and the mother cell for budding yeast and by increasing the degree of septum formation for fission yeast. After cell separation, the low-frequency relaxation disappeared. The simulations well interpreted the oscillation of the relative permittivity of culture broth found for synchronous cell growth of budding yeast.

A new methodology to obtain wine yeast strains overproducing mannoproteins.

PubMed

Quirós, Manuel; Gonzalez-Ramos, Daniel; Tabera, Laura; Gonzalez, Ramon

2010-04-30

Yeast mannoproteins are highly glycosylated proteins that are covalently bound to the beta-1,3-glucan present in the yeast cell wall. Among their outstanding enological properties, yeast mannoproteins contribute to several aspects of wine quality by protecting against protein haze, reducing astringency, retaining aroma compounds and stimulating growth of lactic-acid bacteria. The development of a non-recombinant method to obtain enological yeast strains overproducing mannoproteins would therefore be very useful. Our previous experience on the genetic determinants of the release of these molecules by Saccharomyces cerevisiae has allowed us to propose a new methodology to isolate and characterize wine yeast that overproduce mannoproteins. The described methodology is based on the resistance of the killer 9 toxin produced by Williopsis saturnus, a feature linked to an altered biogenesis of the yeast cell wall. Copyright 2010 Elsevier B.V. All rights reserved.

Biosorption of nickel by yeasts in an osmotically unsuitable environment.

PubMed

Breierová, Emilia; Certík, Milan; Kovárová, Annamaria; Gregor, Tomas

2008-01-01

The tolerance, sorption of nickel(II) ions, and changes in the production and composition of exopolymers of eight yeast strains grown under nickel presence with/without NaCl were studied. Strains of Pichia anomala and Candida maltosa known as the most resistant yeasts against nickel tolerated up to 3 mM Ni2+. NaCl addition decreased both the resistance of the yeast strains toward nickel ions and the sorption of metal ions into cells. All yeasts absorbed nickel predominantly into exopolymers (glycoproteins) and on the surface of cells. However, while the amount of polysaccharide moieties of exoglycoproteins of most of the resistant yeasts was induced by stress conditions, the ratio polysaccharide/protein in the exopolymers remained unchanged in the sensitive species Cystofilobasidium. The exopolymer composition might play a key role in yeast adaptation to stress conditions caused by heavy metal ions.

Cellodextrin transport in yeast for improved biofuel production.

PubMed

Galazka, Jonathan M; Tian, Chaoguang; Beeson, William T; Martinez, Bruno; Glass, N Louise; Cate, Jamie H D

2010-10-01

Fungal degradation of plant biomass may provide insights for improving cellulosic biofuel production. We show that the model cellulolytic fungus Neurospora crassa relies on a high-affinity cellodextrin transport system for rapid growth on cellulose. Reconstitution of the N. crassa cellodextrin transport system in Saccharomyces cerevisiae promotes efficient growth of this yeast on cellodextrins. In simultaneous saccharification and fermentation experiments, the engineered yeast strains more rapidly convert cellulose to ethanol when compared with yeast lacking this system.

Yeast: An Experimental Organism for Modern Biology.

ERIC Educational Resources Information Center

Botstein, David; Fink, Gerald R.

1988-01-01

Discusses the applicability and advantages of using yeasts as popular and ideal model systems for studying and understanding eukaryotic biology at the cellular and molecular levels. Cites experimental tractability and the cooperative tradition of the research community of yeast biologists as reasons for this success. (RT)

Characterization of Hyaluronan-Degrading Enzymes from Yeasts.

PubMed

Smirnou, Dzianis; Krčmář, Martin; Kulhánek, Jaromír; Hermannová, Martina; Bobková, Lenka; Franke, Lukáš; Pepeliaev, Stanislav; Velebný, Vladimír

2015-10-01

Hyaluronidases (HAases) from yeasts were characterized for the first time. The study elucidated that hyaluronate 4-glycanohydrolase and hyaluronan (HA) lyase can be produced by yeasts. Six yeasts producing HAases were found through express screening of activities. The extracellular HAases from two of the yeast isolates, Pseudozyma aphidis and Cryptococcus laurentii, were characterized among them. P. aphidis HAase hydrolyzed β-1,4 glycosidic bonds of HA, yielding even-numbered oligosaccharides with N-acetyl-D-glucosamine at the reducing end. C. laurentii produced hyaluronan lyase, which cleaved β-1,4 glycosidic bonds of HA in β-elimination reaction, and the products of HA degradation were different-sized even-numbered oligosaccharides. The shortest detected HA oligomer was dimer. The enzymes' pH and temperature optima were pH 3.0 and 37-45 °C (P. aphidis) and pH 6.0 and 37 °C (C. laurentii), respectively. Both HAases showed good thermostability.

Nuclear Magnetic Resonance Spectroscopy-Based Identification of Yeast.

PubMed

Himmelreich, Uwe; Sorrell, Tania C; Daniel, Heide-Marie

2017-01-01

Rapid and robust high-throughput identification of environmental, industrial, or clinical yeast isolates is important whenever relatively large numbers of samples need to be processed in a cost-efficient way. Nuclear magnetic resonance (NMR) spectroscopy generates complex data based on metabolite profiles, chemical composition and possibly on medium consumption, which can not only be used for the assessment of metabolic pathways but also for accurate identification of yeast down to the subspecies level. Initial results on NMR based yeast identification where comparable with conventional and DNA-based identification. Potential advantages of NMR spectroscopy in mycological laboratories include not only accurate identification but also the potential of automated sample delivery, automated analysis using computer-based methods, rapid turnaround time, high throughput, and low running costs.We describe here the sample preparation, data acquisition and analysis for NMR-based yeast identification. In addition, a roadmap for the development of classification strategies is given that will result in the acquisition of a database and analysis algorithms for yeast identification in different environments.

Responses of Yeast Biocontrol Agents to Environmental Stress

PubMed Central

Sui, Yuan; Wisniewski, Michael; Droby, Samir

2015-01-01

Biological control of postharvest diseases, utilizing wild species and strains of antagonistic yeast species, is a research topic that has received considerable attention in the literature over the past 30 years. In principle, it represents a promising alternative to chemical fungicides for the management of postharvest decay of fruits, vegetables, and grains. A yeast-based biocontrol system is composed of a tritrophic interaction between a host (commodity), a pathogen, and a yeast species, all of which are affected by environmental factors such as temperature, pH, and UV light as well as osmotic and oxidative stresses. Additionally, during the production process, biocontrol agents encounter various severe abiotic stresses that also impact their viability. Therefore, understanding the ecological fitness of the potential yeast biocontrol agents and developing strategies to enhance their stress tolerance are essential to their efficacy and commercial application. The current review provides an overview of the responses of antagonistic yeast species to various environmental stresses, the methods that can be used to improve stress tolerance and efficacy, and the related mechanisms associated with improved stress tolerance. PMID:25710368

Oxygen requirements of yeasts. [Saccharomyces cerevisiae; Candida tropicalis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Visser, W.; Scheffers, W.A.; Batenburg-Van Der Vegte, W.H.

1990-12-01

Type species of 75 yeast genera were examined for their ability to grow anaerobically in complex and mineral media. To define anaerobic conditions, we added a redox indicator, resazurin, to the media to determine low redox potentials. All strains tested were capable of fermenting glucose to ethanol in oxygen-limited shake-flask cultures, even those of species generally regarded as nonfermentative. However, only 23% of the yeast species tested grew under anaerobic conditions. A comparative study with a number of selected strains revealed that Saccharomyces cerevisiae stands out as a yeast capable of rapid growth at low redox potentials. Other yeasts, suchmore » as Torulaspora delbrueckii and Candida tropicalis, grew poorly ({mu}{sub max}, 0.03 and 0.05 h{sup {minus}1}, respectively) under anaerobic conditions in mineral medium supplemented with Tween 80 and ergosterol. The latter organisms grew rapidly under oxygen limitation and then displayed a high rate of alcoholic fermentation. It can be concluded that these yeasts have hitherto-unidentified oxygen requirements for growth.« less

Vegemite Beer: yeast extract spreads as nutrient supplements to promote fermentation.

PubMed

Kerr, Edward D; Schulz, Benjamin L

2016-01-01

Vegemite is an iconic Australian food spread made from spent brewers' yeast extract, which has been reported to be used as an ingredient in illegal home brewing. In this study, we tested the utility of Vegemite and the similar spread Marmite in promoting fermentation. We could not culture microorganisms from either Vegemite or Marmite, consistent with these food-grade spreads being essentially sterile. To test if the addition of Vegemite or Marmite could assist in fermentation when additional viable yeast was also present, solutions containing glucose and a range of concentrations of either Vegemite or Marmite were inoculated with brewers' yeast. No fermentation occurred in any condition without addition of extra brewer's yeast. Fermentation did not occur when yeast was inoculated into solutions containing only glucose, but progressed efficiently with when Vegemite or Marmite was also added. Gas Chromatography confirmed that ethanol was present at ∼3% v/v post-fermentation in all samples which contained glucose, Vegemite or Marmite, and brewers' yeast. Trace amounts of methanol were also detected. Mass spectrometry proteomics identified abundant intracellular yeast proteins and barley proteins in Vegemite and Marmite, and abundant secreted yeast proteins from actively growing yeast in those samples to which extra brewers' yeast had been added. We estimate that the real-world cost of home brewed "Vegemite Beer" would be very low. Our results show that Vegemite or other yeast extract spreads could provide cheap and readily available sources of nutrient supplementation to increase the efficiency of fermentation in home brewing or other settings.

Yeast Biomass Production in Brewery's Spent Grains Hemicellulosic Hydrolyzate

NASA Astrophysics Data System (ADS)

Duarte, Luís C.; Carvalheiro, Florbela; Lopes, Sónia; Neves, Ines; Gírio, Francisco M.

Yeast single-cell protein and yeast extract, in particular, are two products which have many feed, food, pharmaceutical, and biotechnological applications. However, many of these applications are limited by their market price. Specifically, the yeast extract requirements for culture media are one of the major technical hurdles to be overcome for the development of low-cost fermentation routes for several top value chemicals in a biorefinery framework. A potential biotechnical solution is the production of yeast biomass from the hemicellulosic fraction stream. The growth of three pentose-assimilating yeast cell factories, Debaryomyces hansenii, Kluyveromyces marxianus, and Pichia stipitis was compared using non-detoxified brewery's spent grains hemicellulosic hydrolyzate supplemented with mineral nutrients. The yeasts exhibited different specific growth rates, biomass productivities, and yields being D. hansenii as the yeast species that presented the best performance, assimilating all sugars and noteworthy consuming most of the hydrolyzate inhibitors. Under optimized conditions, D. hansenii displayed a maximum specific growth rate, biomass yield, and productivity of 0.34 h-1, 0.61 g g-1, and 0.56 g 1-1 h-1, respectively. The nutritional profile of D. hansenii was thoroughly evaluated, and it compares favorably to others reported in literature. It contains considerable amounts of some essential amino acids and a high ratio of unsaturated over saturated fatty acids.

Formulation and evaluation of dried yeast tablets using different techniques.

PubMed

Al-Mohizea, Abdullah M; Ahmed, Mahrous O; Al-jenoobi, Fahad I; Mahrous, Gamal M; Abdel-Rahman, Aly A

2007-08-01

The aim of this study was to prepare and evaluate dried yeast tablets using both direct compression and dry granulation techniques in comparison with the conventional wet granulation as well as commercial product. Wet granulation technique is not favorable for producing the yeast tablets due to the problems of color darkening and the reduction of the fermentation power of the yeast as a result of the early start of the fermentation process due to the presence of moisture. Twenty six formulae of dried yeast tablets were prepared and evaluated. Certain directly compressible vehicles were employed for preparing these tablets. The quality control tests (weight uniformity, friability, disintegration time and hardness) of the prepared dried yeast tablets were performed according to B.P. 1998 limits. All batches of the prepared tablets complied with the B.P. limits of weight uniformity. Moreover, small values of friability % (1% or less) were obtained for all batches of dried yeast tablets with acceptable hardness values, indicating good mechanical properties which can withstand handling. On the other hand, not all batches complied with the limit of disintegration test which may be attributed to various formulation component variables. Therefore, four disintegrating agents were investigated for their disintegrating effect. It was found that the method of preparation, whether it is direct compression, dry granulation or wet granulation, has an effect on disintegration time of these dried yeast tablets and short disintegration times were obtained for some of the formulae. The shortest disintegration time was obtained with those tablets prepared by direct compression among the other techniques. Therefore, the direct compression is considered the best technique for preparation of dried yeast tablets and the best formula (which showed shorter disintegration time and better organoleptic properties than the available commercial yeast tablets) was chosen. Drug content for dried

Cross-referencing yeast genetics and mammalian genomes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hieter, P.; Basset, D.; Boguski, M.

1994-09-01

We have initiated a project that will systematically transfer information about yeast genes onto the genetic maps of mice and human beings. Rapidly expanding human EST data will serve as a source of candidate human homologs that will be repeatedly searched using yeast protein sequence queries. Search results will be automatically reported to participating labs. Human cDNA sequences from which the ESTs are derived will be mapped at high resolution in the human and mouse genomes. The comparative mapping information cross-references the genomic position of novel human cDNAs with functional information known about the cognate yeast genes. This should facilitatemore » the initial identification of genes responsible for mammalian mutant phenotypes, including human disease. In addition, the identification of mammalian homologs of yeast genes provides reagents for determining evolutionary conservation and for performing direct experiments in multicellular eukaryotes to enhance study of the yeast protein`s function. For example, ESTs homologous to CDC27 and CDC16 were identified, and the corresponding cDNA clones were obtained from ATTC, completely sequenced, and mapped on human and mouse chromosomes. In addition, the CDC17hs cDNA has been used to raise antisera to the CDC27Hs protein and used in subcellular localization experiments and junctional studies in mammalian cells. We have received funding from the National Center for Human Genome Research to provide a community resource which will establish comprehensive cross-referencing among yeast, human, and mouse loci. The project is set up as a service and information on how to communicate with this effort will be provided.« less

21 CFR 172.381 - Vitamin D2 bakers yeast.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Vitamin D2 bakers yeast. 172.381 Section 172.381... CONSUMPTION Special Dietary and Nutritional Additives § 172.381 Vitamin D2 bakers yeast. Vitamin D2 bakers yeast may be used safely in foods as a source of vitamin D2 and as a leavening agent in accordance with...

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

PubMed Central

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

Independent Evolution of Winner Traits without Whole Genome Duplication in Dekkera Yeasts.

PubMed

Guo, Yi-Cheng; Zhang, Lin; Dai, Shao-Xing; Li, Wen-Xing; Zheng, Jun-Juan; Li, Gong-Hua; Huang, Jing-Fei

2016-01-01

Dekkera yeasts have often been considered as alternative sources of ethanol production that could compete with S. cerevisiae. The two lineages of yeasts independently evolved traits that include high glucose and ethanol tolerance, aerobic fermentation, and a rapid ethanol fermentation rate. The Saccharomyces yeasts attained these traits mainly through whole genome duplication approximately 100 million years ago (Mya). However, the Dekkera yeasts, which were separated from S. cerevisiae approximately 200 Mya, did not undergo whole genome duplication (WGD) but still occupy a niche similar to S. cerevisiae. Upon analysis of two Dekkera yeasts and five closely related non-WGD yeasts, we found that a massive loss of cis-regulatory elements occurred in an ancestor of the Dekkera yeasts, which led to improved mitochondrial functions similar to the S. cerevisiae yeasts. The evolutionary analysis indicated that genes involved in the transcription and translation process exhibited faster evolution in the Dekkera yeasts. We detected 90 positively selected genes, suggesting that the Dekkera yeasts evolved an efficient translation system to facilitate adaptive evolution. Moreover, we identified that 12 vacuolar H+-ATPase (V-ATPase) function genes that were under positive selection, which assists in developing tolerance to high alcohol and high sugar stress. We also revealed that the enzyme PGK1 is responsible for the increased rate of glycolysis in the Dekkera yeasts. These results provide important insights to understand the independent adaptive evolution of the Dekkera yeasts and provide tools for genetic modification promoting industrial usage.