Showing papers on "Yeast published in 2002"
TL;DR: The yeast Saccharomyces cerevisiae is now recognized as a model system representing a simple eukaryote whose genome can be easily manipulated and made particularly accessible to gene cloning and genetic engineering techniques.
Abstract: Publisher Summary The yeast Saccharomyces cerevisiae is now recognized as a model system representing a simple eukaryote whose genome can be easily manipulated. Yeast has only a slightly greater genetic complexity than bacteria and shares many of the technical advantages that permitted rapid progress in the molecular genetics of prokaryotes and their viruses. Some of the properties that make yeast particularly suitable for biological studies include rapid growth, dispersed cells, the ease of replica plating and mutant isolation, a well-defined genetic system, and most important, a highly versatile DNA transformation system. Being nonpathogenic, yeast can be handled with little precautions. Large quantities of normal baker's yeast are commercially available and can provide a cheap source for biochemical studies. The development of DNA transformation has made yeast particularly accessible to gene cloning and genetic engineering techniques. Structural genes corresponding to virtually any genetic trait can be identified by complementation from plasmid libraries. Plasmids can be introduced into yeast cells either as replicating molecules or by integration into the genome. In contrast to most other organisms, integrative recombination of transforming DNA in yeast proceeds exclusively via homologous recombination. Cloned yeast sequences, accompanied by foreign sequences on plasmids, can therefore be directed at will to specific locations in the genome.
3,547 citations
TL;DR: It is concluded that Yor197w indeed functions as a bona fide caspase in yeast and the name Yeast Caspase-1 is proposed, pointing to a physiological role in elimination of overaged cells.
866 citations
TL;DR: It is concluded that neither storage lipids nor lipid bodies are essential for growth in yeast.
502 citations
TL;DR: A novel gene with sequence similarity to plant expansins, isolated from the cellulolytic fungus Trichoderma reesei, is described, which has an N-terminal fungal type cellulose binding domain connected by a linker region to the expansin-like domain.
Abstract: Plant cell wall proteins called expansins are thought to disrupt hydrogen bonding between cell wall polysaccharides without hydrolyzing them. We describe here a novel gene with sequence similarity to plant expansins, isolated from the cellulolytic fungus Trichoderma reesei. The protein named swollenin has an N-terminal fungal type cellulose binding domain connected by a linker region to the expansin-like domain. The protein also contains regions similar to mammalian fibronectin type III repeats, found for the first time in a fungal protein. The swollenin gene is regulated in a largely similar manner as the T. reesei cellulase genes. The biological role of SWOI was studied by disrupting the swo1 gene from T. reesei. The disruption had no apparent effect on the growth rate on glucose or on different cellulosic carbon sources. Non-stringent Southern hybridization of Trichoderma genomic DNA with swo1 showed the presence of other swollenin-like genes, which could substitute for the loss of SWOI in the disruptant. The swollenin gene was expressed in yeast and Aspergillus niger var. awamori. Activity assays on cotton fibers and filter paper were performed with concentrated SWOI-containing yeast supernatant that disrupted the structure of the cotton fibers without detectable formation of reducing sugars. It also weakened filter paper as assayed by an extensometer. The SWOI protein was purified from A. niger var. awamori culture supernatant and used in an activity assay with Valonia cell walls. It disrupted the structure of the cell walls without producing detectable amounts of reducing sugars.
438 citations
TL;DR: Glass DNA microarrays used to investigate the transcription profiles of 6333 predicted ORFs in cells undergoing this transition and their responses to changes in temperature and culture medium identified transcripts that are induced before germ tube initiation and shut off later in the developmental process.
Abstract: The ability of the pathogenic fungus Candida albicans to switch from a yeast to a hyphal morphology in response to external signals is implicated in its pathogenicity. We used glass DNA microarrays to investigate the transcription profiles of 6333 predicted ORFs in cells undergoing this transition and their responses to changes in temperature and culture medium. We have identified several genes whose transcriptional profiles are similar to those of known virulence factors that are modulated by the switch to hyphal growth caused by addition of serum and a 37°C growth temperature. Time course analysis of this transition identified transcripts that are induced before germ tube initiation and shut off later in the developmental process. A strain deleted for the Efg1p and Cph1p transcription factors is defective in hyphae formation, and its response to serum and increased temperature is almost identical to the response of a wild-type strain grown at 37°C in the absence of serum. Thus Efg1p and Cph1p are needed for the activation of the transcriptional program that is induced by the presence of serum.
369 citations
TL;DR: These and other new findings suggest a common mechanism for membrane fission events that has been conserved and adapted during eukaryotic evolution.
361 citations
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TL;DR: Even though yeast lack much of the molecular machinery that is responsible for apoptosis in metazoans, they can be a powerful tool in apoptosis research and help to discover new genes and chemical compounds that modulate the cell-death pathways of higher eukaryotes.
Abstract: Even though yeast lack much of the molecular machinery that is responsible for apoptosis in metazoans, they can be a powerful tool in apoptosis research. The ectopic expression of several animal apoptosis proteins in yeast can help us to discover new genes -- and chemical compounds -- that modulate the cell-death pathways of higher eukaryotes.
333 citations
TL;DR: It is shown here that DGA1, the sole member of this gene family in yeast, has a physiological role in triglyceride synthesis, and three enzymes can account for diacylglycerol esterification in yeast.
321 citations
TL;DR: Xylose uptake does not determine the xylose flux under the conditions and in the yeast strains investigated, and several heterologous monosaccharide transporters from bacteria and plant cells did not confer sufficient uptake activity to restore growth on xylOSE.
Abstract: For an economically feasible production of ethanol from plant biomass by microbial cells, the fermentation of xylose is important. As xylose uptake might be a limiting step for xylose fermentation by recombinant xylose-utilizing Saccharomyces cerevisiae cells a study of xylose uptake was performed. After deletion of all of the 18 hexose-transporter genes, the ability of the cells to take up and to grow on xylose was lost. Reintroduction of individual hexose-transporter genes in this strain revealed that at intermediate xylose concentrations the yeast high- and intermediate-affinity transporters Hxt4, Hxt5, Hxt7 and Gal2 are important xylose-transporting proteins. Several heterologous monosaccharide transporters from bacteria and plant cells did not confer sufficient uptake activity to restore growth on xylose. Overexpression of the xylose-transporting proteins in a xylose-utilizing PUA yeast strain did not result in faster growth on xylose under aerobic conditions nor did it enhance the xylose fermentation rate under anaerobic conditions. The results of this study suggest that xylose uptake does not determine the xylose flux under the conditions and in the yeast strains investigated.
317 citations
TL;DR: It is shown that, like Mcm2–7p, Cdt1p accumulates in the nucleus during G1 phase and is excluded from the nucleus later in the cell cycle by cyclin dependent kinases (cdks).
Abstract: Cdt1 is essential for loading Mcm2–7 proteins into prereplicative complexes (pre-RCs) during replication licensing and has been found in organisms as diverse as fission yeast and humans. We have identified a homologue of Cdt1 in Saccharomyces cerevisiae, which is required for pre-RC assembly. We show that, like Mcm2–7p, Cdt1p accumulates in the nucleus during G1 phase and is excluded from the nucleus later in the cell cycle by cyclin dependent kinases (cdks). Cdt1p interacts with the Mcm2–7p complex, and the nuclear accumulation of these proteins during G1 is interdependent. This coregulation of Cdt1p and Mcm2–7p represents a novel level of pre-RC control.
313 citations
TL;DR: The effects of including lyophilised whole yeast, Saccharomyces cerevisiae, in the diet on the seabream innate immune response showed that yeast supplements enhanced all the latter responses, but not the humoral response.
TL;DR: Evidence is provided that induced resistance against postharvest decay of citrus fruit should be considered an important component of the multiple modes of action of the yeast Candida oleophila.
Abstract: The yeast Candida oleophila, the base of the commercial product Aspire, is recommended for the control of postharvest decay in citrus and pome fruit Its modes of action include nutrient competition, site exclusion, and direct mycoparasitism In the present study, we showed that application of Candida oleophila to surface wounds or to intact ‘Marsh Seedless’ grapefruit elicited systemic resistance against Penicillium digitatum, the main postharvest pathogen of citrus fruit The induction of pathogen resistance in fruit was already pronounced 24 h after elicitation; it was distance, concentration, and time dependent and restricted to the peel tissue closely surrounding the yeast application site The induction of pathogen resistance required viable yeast cells at concentrations of 108 to 109 cells ml-1 Nonviable autoclaved or boiled yeast cells or lower yeast concentrations were ineffective in enhancing fruit disease resistance Application of Candida oleophila cell suspensions to grapefruit peel
TL;DR: Evidence is presented that salt induces PCD in yeast and plants because of an ionic, rather than osmotic, etiology and both salt-sensitive mutants of yeast and Arabidopsis exhibit substantially more profound PCD symptoms, indicating that salt-induced PCD is mediated by ion disequilibrium.
Abstract: Programmed cell death (PCD) is a fundamental cellular process conserved in metazoans, plants and yeast. Evidence is presented that salt induces PCD in yeast and plants because of an ionic, rather than osmotic, etiology. In yeast, NaCl inhibited growth and caused a time-dependent reduction in viability that was preceded by DNA fragmentation. NaCl also induced the cytological hallmarks of lysigenous-type PCD, including nuclear fragmentation, vacuolation and lysis. The human anti-apoptotic protein Bcl-2 increased salt tolerance of wild-type yeast strain and calcineurin-deficient yeast mutant (cnb1Delta) that is defective for ion homeostasis, but had no effect on the NaCl or sorbitol sensitivity of the osmotic hypersensitive hog1Delta mutant -- results that further link PCD in the response to the ion disequilibrium under salt stress. Bcl-2 suppression of cnb1Delta salt sensitivity was ENA1 (P-type ATPase gene)-dependent, due in part to transcriptional activation. Salt-induced PCD (TUNEL staining and DNA laddering) in primary roots of both Arabidopsis thaliana wild type (Col-1 gl1) and sos1 (salt overly sensitive) mutant seedlings correlated positively with treatment lethality. Wild-type plants survived salt stress levels that were lethal to sos1 plants because secondary roots were produced from the shoot/root transition zone. PCD-mediated elimination of the primary root in response to salt shock appears to be an adaptive mechanism that facilitates the production of roots more able to cope with a saline environment. Both salt-sensitive mutants of yeast (cnb1Delta) and Arabidopsis (sos1) exhibit substantially more profound PCD symptoms, indicating that salt-induced PCD is mediated by ion disequilibrium.
TL;DR: The construction of a novel cellulose-degrading yeast strain by genetically codisplaying two cellulolytic enzymes on the cell surface of Saccharomyces cerevisiae indicates that efficient simultaneous saccharification and fermentation of cellulose to ethanol are carried out by a recombinant yeast cells displaying cellulolytics enzymes.
Abstract: For direct and efficient ethanol production from cellulosic materials, we constructed a novel cellulose-degrading yeast strain by genetically codisplaying two cellulolytic enzymes on the cell surface of Saccharomyces cerevisiae. By using a cell surface engineering system based on α-agglutinin, endoglucanase II (EGII) from the filamentous fungus Trichoderma reesei QM9414 was displayed on the cell surface as a fusion protein containing an RGSHis6 (Arg-Gly-Ser-His6) peptide tag in the N-terminal region. EGII activity was detected in the cell pellet fraction but not in the culture supernatant. Localization of the RGSHis6-EGII-α-agglutinin fusion protein on the cell surface was confirmed by immunofluorescence microscopy. The yeast strain displaying EGII showed significantly elevated hydrolytic activity toward barley β-glucan, a linear polysaccharide composed of an average of 1,200 glucose residues. In a further step, EGII and β-glucosidase 1 from Aspergillus aculeatus No. F-50 were codisplayed on the cell surface. The resulting yeast cells could grow in synthetic medium containing β-glucan as the sole carbon source and could directly ferment 45 g of β-glucan per liter to produce 16.5 g of ethanol per liter within about 50 h. The yield in terms of grams of ethanol produced per gram of carbohydrate utilized was 0.48 g/g, which corresponds to 93.3% of the theoretical yield. This result indicates that efficient simultaneous saccharification and fermentation of cellulose to ethanol are carried out by a recombinant yeast cells displaying cellulolytic enzymes.
TL;DR: In these experiments, brewers dried yeast and carbadox had minimal effects on growth, microbial populations, and intestinal health traits of early-weaned pigs, but certain serum immunological traits were enhanced by feeding yeast.
Abstract: Brewers dried yeast, a source of mannan oligosaccharides (MOS), was assessed as an alternative to an antimicrobial agent (carbadox) for young pigs in two experiments. The yeast contained 5.2% MOS. Agglutination tests confirmed adsorption of several serovars of E. coli and Salmonella spp. onto the yeast product. In Exp. 1, seven replicates (five pigs per pen) of 22-d-old pigs were fed a nonmedicated basal diet or the basal diet with carbadox (55 mg/kg), yeast (3%), or a combination of 3% yeast and 2% citric acid for 28 d. Carbadox did not improve growth performance. Growth rate and feed intake were depressed (P 0.05) among treatments, but fecal shedding of carbadox-resistant coliforms was higher (P < 0.01) during the 9-d period in pigs fed carbadox. Total fecal coliforms were consistently lower throughout the postinoculation period in pigs fed yeast (P < 0.05). Yeast reduced colonization oftotal coliforms in the duodenum,jejunum, cecum, and colon, but it did not have a consistent effect on colonization of E. coli K88. Pigs fed yeast tended (P < 0.10) to have higher serum IgG levels than controls. In these experiments, brewers dried yeast and carbadox had minimal effects on growth, microbial populations, and intestinal health traits of early-weaned pigs, but certain serum immunological traits were enhanced by feeding yeast.
TL;DR: It is found that the SSU1-R allele, which confers sulfite resistance to yeast cells, is the product of a reciprocal translocation between chromosomes VIII and XVI due to unequal crossing-over mediated by microhomology between very short sequences on the 5' upstream regions of theSSU1 and ECM34 genes.
Abstract: The unaware use of yeast for winemaking by the first agricultural civilizations has been reported as far back as 7400 years ago. Until the middle of the last millennium, wines were mainly produced around the Mediterranean Sea and the Caucasus. Since then, winemaking has spread with the European colonizers throughout the temperate regions of the world (Pretorius 2000).
Although different genera and species of yeasts are found in musts, the species Saccharomyces cerevisiae is mainly responsible for the transformation of musts into wines. The origin of S. cerevisiae is controversial. Some authors propose that this species is a “natural” organism present in plant fruits (Mortimer and Polsinelli 1999). Others argue that S. cerevisiae is a domesticated species originated from its closest relative S. paradoxus, a wild species found all around the world (Vaughan-Martini and Martini 1995). This debate is important in postulating the original genome of S. cerevisiae and how the strong selective pressure applied since its first unconscious use in controlled fermentation processes has reshaped it. Useful phenotypic traits such as fast growth in sugar-rich media, high alcohol production and tolerance, and good flavor production selected for billions of generations have had strong influences on the S. cerevisiae genome.
In contrast to most S. cerevisiae strains used in the laboratory, which are either haploid or diploid and have a constant chromosome electrophoretic profile, wine yeast strains are mainly diploid, aneuploid, or polyploid, homothallic, and highly heterozygous (Bakalinsky and Snow 1990; Barre et al. 1993; Codon et al. 1995), and show a high level of chromosome length polymorphisms (Bidenne et al. 1992; Rachidi et al. 1999). Moreover, wine yeast strains seem not to remain genetically uniform (Pretorius 2000). Their exacerbated capacity to reorganize its genome by chromosome rearrangements such as Ty-promoted chromosomal translocations (Longo and Vezinhet 1993; Rachidi et al. 1999), mitotic crossing-over (Aguilera et al. 2000), and gene conversion (Puig et al. 2000) promotes a faster adaptation to environmental changes than spontaneous mutations, which occur at comparatively very low rates. The ploidy of the wine yeasts may confer advantages in adapting to variable external environments or increasing the dosage of some genes important for fermentation (Bakalinsky and Snow 1990; Salmon 1997).
In addition, the possibility of adaptive gross genomic changes occurring during laboratory growth conditions has been demonstrated with DNA chip technology by Hughes et al. (2000). Those authors showed in multiple cases that the deletion of a gene strongly favors the acquisition of a second copy of a whole chromosome or a chromosomal segment containing a compensatory copy of a close homolog of the deleted gene.
In a comparative study of transcriptomes, we found that SSU1, a gene that mediates sulfite efflux in S. cerevisiae and, hence, confers sulfite resistance (Park and Bakalinsky 2000), showed a significantly higher expression in the T73 wine yeast strain than in a laboratory strain (Hauser et al. 2001). In contrast to the allele present in the laboratory strains, a highly sulfite-resistant wine strain exhibited a translocation involving the promoter region of the gene (SSU1-R allele), which produces an increase in the sulfite resistance (Goto-Yamamoto et al. 1998). In the present study, we explored the organization of this gene at the molecular level in different wine yeast strains.
TL;DR: It is shown that when yeast cells are exposed to Cd(2+), most of the sulfur assimilated by the cells is converted into glutathione, a thiol-metabolite essential for detoxification.
TL;DR: Glutathione (GSH: L-gamma-glutamyl-L-cysteinylglycine) is present in high concentrations up to 10 mM in yeast cells, giving this tripeptide the properties of a cellular redox buffer.
Abstract: Glutathione (GSH: L-gamma-glutamyl-L-cysteinylglycine) is present in high concentrations up to 10 mM in yeast cells. Its very low redox potential (E'(o)=-240 mV for thiol disulfide exchange) gives this tripeptide the properties of a cellular redox buffer. In Saccharomyces cerevisiae and non-conventional yeasts (NCY), GSH may be involved in basic cellular functions such as the maintenance of mitochondrial and membrane integrity. GSH also assumes pivotal roles in (i) response to sulfur and nitrogen starvation; (ii) detoxification of endogenous toxic metabolites, such as excess formaldehyde produced during the growth of the methylotrophic yeasts Hansenula polymorpha, Candida boidinii and Kloeckera sp.; (iii) protection against oxidative stress provoked by exposure of the cells to reactive oxygen species including peroxides and hydroperoxides; (iv) detoxification of xenobiotics such as halogenated aromatics, alkylating agents and arsenite; (v) resistance to heavy-metal stress exemplified by the responses of S. cerevisiae and Schizosaccharomyces pombe to cadmium salts; (vi) yeast mycelium transition in Candida and Aureobasidium sp.
TL;DR: Analysis of wine fermentation microbial ecology by using both culture-dependent and -independent methods reveals the complexity of yeast interactions enriched during spontaneous fermentations.
Abstract: Culture-dependent and -independent methods were used to examine the yeast diversity present in botrytis-affected ("botrytized") wine fermentations carried out at high ( approximately 30 degrees C) and ambient ( approximately 20 degrees C) temperatures. Fermentations at both temperatures possessed similar populations of Saccharomyces, Hanseniaspora, Pichia, Metschnikowia, Kluyveromyces, and Candida species. However, higher populations of non-Saccharomyces yeasts persisted in ambient-temperature fermentations, with Candida and, to a lesser extent, Kluyveromyces species remaining long after the fermentation was dominated by Saccharomyces. In general, denaturing gradient gel electrophoresis profiles of yeast ribosomal DNA or rRNA amplified from the fermentation samples correlated well with the plating data. The direct molecular methods also revealed a Hanseniaspora osmophila population not identified in the plating analysis. rRNA analysis also indicated a large population (>10(6) cells per ml) of a nonculturable Candida strain in the high-temperature fermentation. Monoculture analysis of the Candida isolate indicated an extreme fructophilic phenotype and correlated with an increased glucose/fructose ratio in fermentations containing higher populations of Candida. Analysis of wine fermentation microbial ecology by using both culture-dependent and -independent methods reveals the complexity of yeast interactions enriched during spontaneous fermentations.
TL;DR: The utility of the DNA binding dye, SYTOX Green, in the cell cycle analysis of yeast is shown, with better coefficients of variation, improved linearity between DNA content and fluorescence, and decreased peak drift associated with changes in dye concentration, growth conditions or cell size.
Abstract: The budding yeast, Saccharomyces cerevisiae has been a remarkably useful model system for the study of eukaryotic cell cycle regulation. Flow cytometric analysis of DNA content in budding yeast has become a standard tool for the analysis of cell cycle progression. However, popular protocols utilizing the DNA binding dye, propidium iodide, suffer from a number of drawbacks that confound accurate analysis by flow cytometry. Here we show the utility of the DNA binding dye, SYTOX Green, in the cell cycle analysis of yeast. Samples analyzed using SYTOX Green exhibited better coefficients of variation, improved linearity between DNA content and fluorescence, and decreased peak drift associated with changes in dye concentration, growth conditions or cell size.
TL;DR: The analysis of the S. cerevisiae strains showed that indigenous strains competed with commercial strains inoculated in other fermentation tanks of the cellar, resulting in the continuous use of commercial yeasts reduced the diversity and importance of the indigenous S. continentaliae strains.
TL;DR: Results from studies on S. cerevisiae have been used to understand metal transport in other species of yeast as well as in higher eukaryotes.
Abstract: All eukaryotes and most prokaryotes require transition metals. In recent years there has been an enormous advance in our understanding of how these metals are transported across the plasma membrane. Much of this understanding has resulted from studies on the budding yeast Saccharomyces cerevisiae. A variety of genetic and biochemical approaches have led to a detailed understanding of how transition metals such as iron, copper, manganese, and zinc are acquired by cells. The regulation of metal transport has been defined at both the transcriptional and posttranslational levels. Results from studies on S. cerevisiae have been used to understand metal transport in other species of yeast as well as in higher eukaryotes.
TL;DR: Yeast colonies isolated from vineyard and cellar substrates were analysed and Saccharomyces cerevisiae was not the major species isolated as Candida sorbosa represented 76% of the species isolated.
TL;DR: The YMR318C gene product has been purified to homogeneity from overexpressing yeast cells, and found to be a homodimeric ADH, composed of 40 kDa subunits and with a pI of 5.0-5.4, to be the first NADPH-dependent medium-chain ADH to be characterized in S. cerevisiae.
Abstract: YMR318C represents an open reading frame from Saccharomyces cerevisiae with unknown function. It possesses a conserved sequence motif, the zinc-containing alcohol dehydrogenase (ADH) signature, specific to the medium-chain zinc-containing ADHs. In the present study, the YMR318C gene product has been purified to homogeneity from overexpressing yeast cells, and found to be a homodimeric ADH, composed of 40 kDa subunits and with a pI of 5.0-5.4. The enzyme was strictly specific for NADPH and was active with a wide variety of substrates, including aliphatic (linear and branched-chain) and aromatic primary alcohols and aldehydes. Aldehydes were processed with a 50-fold higher catalytic efficiency than that for the corresponding alcohols. The highest k(cat)/K(m) values were found with pentanal>veratraldehyde > hexanal > 3-methylbutanal >cinnamaldehyde. Taking into consideration the substrate specificity and sequence characteristics of the YMR318C gene product, we have proposed this gene to be called ADH6. The disruption of ADH6 was not lethal for the yeast under laboratory conditions. Although S. cerevisiae is considered a non lignin-degrading organism, the catalytic activity of ADHVI can direct veratraldehyde and anisaldehyde, arising from the oxidation of lignocellulose by fungal lignin peroxidases, to the lignin biodegradation pathway. ADHVI is the only S. cerevisiae enzyme able to significantly reduce veratraldehyde in vivo, and its overexpression allowed yeast to grow under toxic concentrations of this aldehyde. The enzyme may also be involved in the synthesis of fusel alcohols. To our knowledge this is the first NADPH-dependent medium-chain ADH to be characterized in S. cerevisiae.
TL;DR: This screening survey could constitute the first approach in selecting yeast strains of environmental origin potentially exploitable as enzyme producers in tropical environments of the Brazilian rain forest.
Abstract: Aims: The objective of this study was to investigate the extracellular enzymatic activity (EEA) profile of yeasts isolated from tropical environments of the Brazilian rain forest. This screening survey could constitute the first approach in selecting yeast strains of environmental origin potentially exploitable as enzyme producers.
Methods and Results: In this study, 348 yeast (193 ascomycetes and 155 basidiomycetes) and 46 yeast-like strains (Aureobasidium pullulans) were screened for their EEA profile. The spread occurrence of extracellular amylases, esterases, lipases, proteases, pectinases and chitinases appeared to be a strain-related character.
Conclusions: Yeasts isolated from tropical environments could represent a promising source of EEA. Selected strains showed maximum levels of EEA under acidic or neutral conditions.
Significance and Impact of the Study: This study demonstrated the potential for yeasts isolated from extreme environments as sources of industrially relevant enzymes for biotechnological purposes.
TL;DR: Interestingly, the yeast cells displaying the FLProROL protein showed strong flocculation, even though the glycosylphosphatidylinositol anchor attachment signal and cell-membrane-anchoring region of Flo1p had been deleted from this gene.
Abstract: We constructed a novel cell-surface display system, using as a new type of cell-wall anchor 3,297 or 4,341 bp of the 3' region of the FLO1 gene (FS or FL gene, respectively), which encodes the flocculation functional domain of Flo1p. In this system, the N terminus of the target protein was fused to the FS or FL protein and the fusion proteins were expressed under the control of the inducible promoter UPR-ICL (5' upstream region of the isocitrate lyase of Candida tropicalis). Using this new system, recombinant lipase with a pro sequence from Rhizopus oryzae (rProROL), which has its active site near the C terminus, was displayed on the cell surface. Cell-surface display of the FSProROL and FLProROL fusion proteins was confirmed by immunofluorescence microscopy and immunoblotting. Lipase activity reached 145 IU/liter (61.3 IU/g [dry cell weight]) on the surface of the yeast cells, which successfully catalyzed the methanolysis reaction. Using these whole-cell biocatalysts, methylesters synthesized from triglyceride and methanol reached 78.3% after 72 h of reaction. To our knowledge, this is the first example of cell-surface display of lipase with high activity. Interestingly, the yeast cells displaying the FLProROL protein showed strong flocculation, even though the glycosylphosphatidylinositol anchor attachment signal and cell-membrane-anchoring region of Flo1p had been deleted from this gene. The cell-surface display system based on FL thus endows the yeast strain with both novel enzyme display and strong flocculation ability.
TL;DR: This review focuses on recent advances on the roles of cortical tags, GTPases and the cytoskeleton in the generation and maintenance of cell polarity in yeast.
TL;DR: The results of this study provide a global description of transcriptional response for adaptation to low temperature in yeast cells and reveal three phases: early, middle, and late phases.
TL;DR: The range of intracellular transcript abundance in yeast is examined using kinetically monitored, reverse transcriptase-initiated PCR (kRT-PCR) and the results show that transcript availability in yeast varies over six orders of magnitude.
TL;DR: The functional characterization of yeast tsa2Δ mutants and the comparison of TSA1 with TSA2 implicate the cooperation of Tsa1p and Tsa2p in the cellular defense against reactive oxygen and nitrogen species.