ORCID Profile
0000-0001-9127-3175
Current Organisations
University of the Free State
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Macquarie University
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Biochemistry and Cell Biology | Synthetic Biology
Publisher: Springer Science and Business Media LLC
Date: 05-1993
DOI: 10.1007/BF00228603
Publisher: American Society for Microbiology
Date: 09-2012
DOI: 10.1128/AEM.01279-12
Abstract: Saccharomyces cerevisiae has evolved a highly efficient strategy for energy generation which maximizes ATP energy production from sugar. This adaptation enables efficient energy generation under anaerobic conditions and limits competition from other microorganisms by producing toxic metabolites, such as ethanol and CO 2 . Yeast fermentative and flavor capacity forms the biotechnological basis of a wide range of alcohol-containing beverages. Largely as a result of consumer demand for improved flavor, the alcohol content of some beverages like wine has increased. However, a global trend has recently emerged toward lowering the ethanol content of alcoholic beverages. One option for decreasing ethanol concentration is to use yeast strains able to ert some carbon away from ethanol production. In the case of wine, we have generated and evaluated a large number of gene modifications that were predicted, or known, to impact ethanol formation. Using the same yeast genetic background, 41 modifications were assessed. Enhancing glycerol production by increasing expression of the glyceraldehyde-3-phosphate dehydrogenase gene, GPD1 , was the most efficient strategy to lower ethanol concentration. However, additional modifications were needed to avoid negatively affecting wine quality. Two strains carrying several stable, chromosomally integrated modifications showed significantly lower ethanol production in fermenting grape juice. Strain AWRI2531 was able to decrease ethanol concentrations from 15.6% (vol/vol) to 13.2% (vol/vol), whereas AWRI2532 lowered ethanol content from 15.6% (vol/vol) to 12% (vol/vol) in both Chardonnay and Cabernet Sauvignon juices. Both strains, however, produced high concentrations of acetaldehyde and acetoin, which negatively affect wine flavor. Further modifications of these strains allowed reduction of these metabolites.
Publisher: Springer Science and Business Media LLC
Date: 15-08-2000
Abstract: The xynB gene encoding the Bacillus pumilus beta-xylosidase was expressed separately and jointly with the Trichoderma reesei beta-xylanase (xyn2) gene in the yeast Saccharomyces cerevisiae. Both genes were placed under the transcriptional control of the glucose-derepressible alcohol dehydrogenase 2 promoter (ADH2p) and terminator (ADH2T) sequences. The xynB gene was fused in frame to the yeast mating factor alpha1 secretion sequence (MFalpha1s) to effect secretion in S. cerevisiae. The fusion protein was designated Xlo1. Xlo1 produced in S. cerevisiae exhibited low affinity for xylobiose, but eventually hydrolyzed xylobiose and xylotriose to the monomeric constituent, D-xylose. Coproduction of Xyn2 and Xlo1 by S. cerevisiae led to a 25% increase in the amount of reducing sugars released from birchwood xylan compared to S. cerevisiae producing only the Xyn2 beta-xylanase. However, no D-xylose was produced from birchwood xylan, presumably due to very low Xlo1 beta-xylosidase activity and its low affinity for xylobiose.
Publisher: Oxford University Press (OUP)
Date: 2005
DOI: 10.1534/GENETICS.104.033704
Abstract: The invasive and filamentous growth forms of Saccharomyces cerevisiae are adaptations to specific environmental conditions, under particular conditions of limited nutrient availability. Both growth forms are dependent on the expression of the FLO11 gene, which encodes a cell-wall-associated glycoprotein involved in cellular adhesion. A complex regulatory network consisting of signaling pathways and transcription factors has been associated with the regulation of FLO11. Mss11p has been identified as a transcriptional activator of this gene, and here we present an extensive genetic analysis to identify functional relationships between Mss11p and other FLO11 regulators. The data show that Mss11p is absolutely required for the activation of FLO11 by most proteins that have previously been shown to affect FLO11 expression, including the signaling proteins Ras2p, Kss1p, and Tpk2p, the activators Tec1p, Flo8p, and Phd1p, and the repressors Nrg1p, Nrg2p, Sok2p, and Sfl1p. The genetic evidence furthermore suggests that Mss11p activity is not dependent on the presence of any of the above-mentioned factors and that the protein also regulates other genes involved in cellular adhesion phenotypes. Taken together, the data strongly suggest a central role for Mss11p in the regulatory network controlling FLO11 expression, invasive growth, and pseudohyphal differentiation.
Publisher: Springer Science and Business Media LLC
Date: 03-2005
DOI: 10.1007/S10529-005-1014-8
Abstract: To illustrate the effect of a cellulose-binding domain (CBD) on the enzymatic characteristics of non-cellulolytic exoglucanases, 10 different recombinant enzymes were constructed combining the Saccharomyces cerevisiae exoglucanases, EXG1 and SSG1, with the CBD2 from the Trichoderma reesei cellobiohydrolase, CBH2, and a linker peptide. The enzymatic activity of the recombinant enzymes increased with the CBD copy number. The recombinant enzymes, CBD2-CBD2-L-EXG1 and CBD2-CBD2-SSG1, exhibited the highest cellobiohydrolase activity (17.5 and 16.3 U mg(-1) respectively) on Avicel cellulose, which is approximately 1.5- to 2-fold higher than the native enzymes. The molecular organisation of CBD in these recombinant enzymes enhanced substrate affinity, molecular flexibility and synergistic activity, contributing to their elevated action on the recalcitrant substrates as characterised by adsorption, kinetics, thermostability and scanning electron microscopic analysis.
Publisher: Springer Science and Business Media LLC
Date: 1998
Abstract: Saccharomyces cerevisiae T206 K+R+, a K2 killer yeast, was differentiated from other NCYC killer strains of S. cerevisiae on the basis of CHEF-karyotyping and mycoviral RNA separations. Genomic DNA of strain T206 was resolved into 13 chromosome bands, ranging from approximately 0.2 to 2.2 Mb. The resident virus in strain T206 yielded L and M RNA species of approximately 5.1 kb and 2.0 kb, respectively. In micro-scale vinifications, strain T206 showed a lethal effect on a K-R- mesophilic wine yeast. Metabolite accumulation and toxin activity were measured over a narrow pH range of 3.2 to 3.5. Contrary to known fermentation trends, the challenged fermentations were neither stuck nor protracted although over 70% of the cell population was killed. Toxin-sensitive cells showed cytosolic efflux.
Publisher: Oxford University Press (OUP)
Date: 02-2015
DOI: 10.1534/GENETICS.114.173633
Abstract: The Saccharomyces sensu stricto group encompasses species ranging from the industrially ubiquitous yeast Saccharomyces cerevisiae to those that are confined to geographically limited environmental niches. The wealth of genomic data that are now available for the Saccharomyces genus is providing unprecedented insights into the genomic processes that can drive speciation and evolution, both in the natural environment and in response to human-driven selective forces during the historical “domestication” of these yeasts for baking, brewing, and winemaking.
Publisher: Elsevier
Date: 2005
Publisher: American Chemical Society (ACS)
Date: 14-11-2019
Abstract: Despite being used chiefly for fermenting the sugars of grape must to alcohol, wine yeasts (most prominently
Publisher: American Society for Microbiology
Date: 09-2003
DOI: 10.1128/AEM.69.9.5228-5237.2003
Abstract: Volatile aroma-active esters are responsible for the fruity character of fermented alcoholic beverages such as beer and wine. Esters are produced by fermenting yeast cells in an enzyme-catalyzed intracellular reaction. In order to investigate and compare the roles of the known Saccharomyces cerevisiae alcohol acetyltransferases, Atf1p, Atf2p and Lg-Atf1p, in volatile ester production, the respective genes were either deleted or overexpressed in a laboratory strain and a commercial brewing strain. Subsequently, the ester formation of the transformants was monitored by headspace gas chromatography and gas chromatography combined with mass spectroscopy (GC-MS). Analysis of the fermentation products confirmed that the expression levels of ATF1 and ATF2 greatly affect the production of ethyl acetate and isoamyl acetate. GC-MS analysis revealed that Atf1p and Atf2p are also responsible for the formation of a broad range of less volatile esters, such as propyl acetate, isobutyl acetate, pentyl acetate, hexyl acetate, heptyl acetate, octyl acetate, and phenyl ethyl acetate. With respect to the esters analyzed in this study, Atf2p seemed to play only a minor role compared to Atf1p. The atf1 Δ atf2 Δ double deletion strain did not form any isoamyl acetate, showing that together, Atf1p and Atf2p are responsible for the total cellular isoamyl alcohol acetyltransferase activity. However, the double deletion strain still produced considerable amounts of certain other esters, such as ethyl acetate (50% of the wild-type strain), propyl acetate (50%), and isobutyl acetate (40%), which provides evidence for the existence of additional, as-yet-unknown ester synthases in the yeast proteome. Interestingly, overexpression of different alleles of ATF1 and ATF2 led to different ester production rates, indicating that differences in the aroma profiles of yeast strains may be partially due to mutations in their ATF genes.
Publisher: MDPI AG
Date: 10-05-2023
Abstract: Traditional kombucha is a functional tea-based drink that has gained attention as a low or non-alcoholic beverage. The fermentation is conducted by a community of different microorganisms, collectively called SCOBY (Symbiotic Culture of Bacteria and Yeast) and typically consists of different acetic acid bacteria and fermenting yeast, and in some cases lactic acid bacteria that would convert the sugars into organic acids—mostly acetic acid. In this study, the effect of including a Pichia kluyveri starter culture in a kombucha fermentation was investigated. P. kluyveri additions led to a quicker accumulation of acetic acid along with the production of several acetate esters including isoamyl acetate and 2-phenethyl acetate. A subsequent tasting also noted a significant increase in the fruitiness of the kombucha. The significant contribution to the aroma content shows the promise of this yeast in future microbial formulations for kombucha fermentations.
Publisher: Springer Science and Business Media LLC
Date: 05-1995
DOI: 10.1007/BF00132014
Publisher: Elsevier BV
Date: 04-1991
DOI: 10.1016/0378-1119(91)90353-D
Abstract: A glucoamylase-encoding gene (STA2) from Saccharomyces diastaticus and an alpha-amylase-encoding gene (AMY) from Bacillus amyloliquefaciens were cloned separately into a yeast-integrating shuttle vector (YIp5), generating recombinant plasmids pSP1 and pSP2, respectively. The STA2 and AMY genes were jointly cloned into YIp5, generating plasmid pSP3. Subsequently, the dominant selectable marker APH1, encoding resistance to Geneticin G418 (GtR), was cloned into pSP3, resulting in pSP4. For enhanced expression of GtR, the APH1 gene was fused to the GAL10 promoter and terminated by the URA3 terminator, resulting in pSP5. Plasmid pSP5 was converted to a circular minichromosome (pSP6) by the addition of the ARS1 and CEN4 sequences. Laboratory strains of Saccharomyces cerevisiae transformed with plasmids pSP1 through pSP6, stably produced and secreted glucoamylase and/or alpha-amylase. Brewers' and distillers' yeast transformed with pSP6 were also capable of secreting amylolytic enzymes. Yeast transformants containing pSP1, pSP2 and pSP3 assimilated soluble starch with an efficiency of 69%, 84% and 93%, respectively. The major starch hydrolysis products produced by crude amylolytic enzymes found in the culture broths of the pSP1-, pSP2- and pSP3-containing transformants, were glucose, glucose and maltose (1:1), and glucose and maltose (3:1), respectively. These results confirmed that co-expression of the STA2 and AMY genes synergistically enhanced starch degradation.
Publisher: Elsevier BV
Date: 2003
Publisher: Wiley
Date: 28-10-2003
DOI: 10.1002/BIT.10797
Abstract: Lipomyces kononenkoae and Saccharomycopsis fibuligera possess highly efficient alpha-amylase and/or glucoamylase activities that enable both of these yeasts to utilize raw starch as a carbon source. Eight constructs containing the L. kononenkoae alpha-amylase genes (LKA1 and LKA2), and the S. fibuligera alpha-amylase (SFA1) and glucoamylase (SFG1) genes were prepared. The first set of constructs comprised four single gene cassettes each containing one of the in idual amylase coding sequences (LKA1, LKA2, SFA1 or SFG1) under the control of the phosphoglycerate kinase gene (PGK1) promoter and terminator, while the second set comprised two single cassettes containing SFA1 and SFG1 linked to their respective native promoters and terminators. The third set of constructs consisted of two double-gene cassettes, one containing LKA1 plus LKA2 under the control of the PGK1 promoter and terminator, and the other SFA1 plus SFG1 controlled by their respective native promoters and terminators. These constructs were transformed into a laboratory strain Saccharomyces cerevisiae (Sigma1278b). Southern-blot analysis confirmed the stable integration of the different gene constructs into the S. cerevisiae genome and plate assays revealed amylolytic activity. The strain expressing LKA1 and LKA2 resulted in the highest levels of alpha-amylase activity in liquid media. This strain was also the most efficient at starch utilization in batch fermentations, utilizing 80% of the available starch and producing 0.61g/100 mL of ethanol after 6 days of fermentation. The strain expressing SFG1 under the control of the PGK1 expression cassette gave the highest levels of glucoamylase activity. It was shown that the co-expression of these heterologous alpha-amylase and glucoamylase genes enhance starch degradation additively in S. cerevisiae. This study has resulted in progress towards laying the foundation for the possible development of efficient starch-degrading S. cerevisiae strains that could eventually be used in consolidated bioprocessing, and in the brewing, whisky, and biofuel industries.
Publisher: MDPI AG
Date: 15-08-2022
DOI: 10.3390/FERMENTATION8080395
Abstract: Enabling technologies in synthetic biology now present the opportunity to engineer wine yeast for enhanced novel aromas. In doing so, improved wine products will increase the desirability of wine for the consumer and add value to the winemaker. The action of the enzyme carotenoid cleavage dioxygenase 1 (CCD1) on β-carotene to produce β-ionone is of interest to improve the aroma and flavour of the wine. Engineering the yeast, Saccharomyces cerevisiae, to produce higher concentrations of CCD1 in grape-must presents an opportunity to increase the levels of this volatile organic compound, thus enhancing the organoleptic properties of wine. To this end, four phylogenetically erse plant CCD1 genes were synthesised with a secretion signal peptide and transformed into S. cerevisiae. The relative ability of each enzyme secreted into the yeast supernatant to cleave the deep orange C40 β-carotene was determined by spectrophotometry furthermore, the by-product of such cleavage, the highly aromatic C13 β-ionone, was assessed by head-space solid-phase micro-extraction, with analysis and detection by GCMS. Reduction in β-carotene levels and release of β-ionone from the supernatant were validated by LCMS detection of CCD1. These experiments demonstrated that expression in yeast of the CCD1s derived from Petunia hybrida and Vitis vinifera and their subsequent secretion into the medium provided superior efficacy in both β-carotene reduction and β-ionone liberation. We anticipate this knowledge being of benefit to future winemakers in producing a vinous product with enhanced organoleptic properties.
Publisher: Oxford University Press (OUP)
Date: 11-2004
DOI: 10.1016/J.FEMSLE.2004.09.022
Abstract: The volatile thiol 4-mercapto-4-methylpentan-2-one (4MMP) is a potent contributor to wine aroma. In grape juice, 4MMP is bound to cysteine as a non-volatile compound and requires the action of yeast during fermentation to release the aroma active thiol. A method was developed to measure 4MMP release from the precursor by headspace solid-phase microextraction and separation by gas chromatography with atomic emission detection to screen the ability of wine yeast to release 4MMP. Yeast commonly used in white wine making were grown with the precursor at two different temperatures, and the amount of 4MMP released was measured. The results demonstrate that yeast strain selection and fermentation temperature can provide an important tool to enhance or modulate the grape-derived aromas formed during wine fermentation.
Publisher: Elsevier BV
Date: 02-2004
Publisher: American Chemical Society (ACS)
Date: 12-07-2003
DOI: 10.1021/JF026224D
Abstract: Phenolic acids, which are generally esterified with tartaric acid, are natural constituents of grape must and wine and can be released as free acids (principally p-coumaric, caffeic, and ferulic acids) by certain cinnamoyl esterase activities during the wine-making process. Some of the microorganisms present in grape can metabolize the free phenolic acids into 4-vinyl and 4-ethyl derivatives. These volatile phenols contribute to the aroma of wine. The Saccharomyces cerevisiae phenyl acrylic acid decarboxylase gene (PAD1) is steadily transcribed, but its encoded product, Pad1p, shows low activity. In contrast, the phenolic acid decarboxylase (PADC) from Bacillus subtilis and the p-coumaric acid decarboxylase (PDC) from Lactobacillus plantarum display substrate-inducible decarboxylating activity in the presence of phenolic acids. In an attempt to develop wine yeasts with optimized decarboxylation activity on phenolic acids, the padc, pdc, and PAD1 genes were cloned under the control of S. cerevisiae's constitutive phosphoglyceratekinase I gene promoter (PGK1(P)()) and terminator (PGK1(T)()) sequences. These gene constructs were integrated into the URA3 locus of a laboratory strain of S. cerevisiae, Sigma1278b. The overexpression of the two bacterial genes, padc and pdc, in S. cerevisiae showed high enzyme activity. However, this was not the case for PAD1. The padc and pdc genes were also integrated into an industrial wine yeast strain, S. cerevisiae VIN13. As an additional control, both alleles of PAD1 were disrupted in the VIN13 strain. In microvinification trials, all of the laboratory and industrial yeast transformants carrying the padc and pdc gene constructs showed an increase in volatile phenol formation as compared to the untransformed host strains (Sigma1278b and VIN13). This study offers prospects for the development of wine yeast starter strains with optimized decarboxylation activity on phenolic acids and the improvement of wine aroma in the future.
Publisher: Springer Berlin Heidelberg
Date: 2009
Publisher: Wiley
Date: 05-01-2006
Publisher: Oxford University Press (OUP)
Date: 12-2003
Publisher: Oxford University Press (OUP)
Date: 27-02-2007
DOI: 10.1007/S10295-007-0213-9
Abstract: Enzyme engineering was performed to link the beta-glucosidase enzyme (BGL1) from Saccharomycopsis fibuligera to the cellulose-binding domain (CBD2) of Trichoderma reesei cellobiohydrolase (CBHII) to investigate the effect of a fungal CBD on the enzymatic characteristics of this non-cellulolytic yeast enzyme. Recombinant enzymes were constructed with single and double copies of CBD2 fused at the N-terminus of BGL1 to mimic the two-domain organization displayed by cellulolytic enzymes in nature. The engineered S. fibuligera beta-glucosidases were expressed in Saccharomyces cerevisiae under the control of phosphoglycerate-kinase-1 promoter (PGK1 ( P )) and terminator (PGK1 ( T )) and yeast mating pheromone alpha-factor secretion signal (MFalpha1 ( S )). The secreted enzymes were purified and characterized using a range of cellulosic and non-cellulosic substrates to illustrate the effect of the CBD on their enzymatic activity. The results indicated that the recombinant enzymes of BGL1 displayed a 2-4-fold increase in their hydrolytic activity toward cellulosic substrates like avicel, amorphous cellulose, bacterial microcrystalline cellulose, and carboxy methyl cellulose in comparison with the native enzyme. The organization of the CBD in these recombinant enzymes also resulted in enhanced substrate affinity, molecular flexibility and synergistic activity, thereby improving the ability of the enzymes to act on and hydrolyze cellulosic substrates, as characterized by adsorption, kinetics, thermal stability, and scanning electron microscopic analyses.
Publisher: Elsevier
Date: 2015
Publisher: American Society for Microbiology
Date: 09-2002
DOI: 10.1128/MMBR.66.3.506-577.2002
Abstract: Fundamental features of microbial cellulose utilization are examined at successively higher levels of aggregation encompassing the structure and composition of cellulosic biomass, taxonomic ersity, cellulase enzyme systems, molecular biology of cellulase enzymes, physiology of cellulolytic microorganisms, ecological aspects of cellulase-degrading communities, and rate-limiting factors in nature. The methodological basis for studying microbial cellulose utilization is considered relative to quantification of cells and enzymes in the presence of solid substrates as well as apparatus and analysis for cellulose-grown continuous cultures. Quantitative description of cellulose hydrolysis is addressed with respect to adsorption of cellulase enzymes, rates of enzymatic hydrolysis, bioenergetics of microbial cellulose utilization, kinetics of microbial cellulose utilization, and contrasting features compared to soluble substrate kinetics. A biological perspective on processing cellulosic biomass is presented, including features of pretreated substrates and alternative process configurations. Organism development is considered for “consolidated bioprocessing” (CBP), in which the production of cellulolytic enzymes, hydrolysis of biomass, and fermentation of resulting sugars to desired products occur in one step. Two organism development strategies for CBP are examined: (i) improve product yield and tolerance in microorganisms able to utilize cellulose, or (ii) express a heterologous system for cellulose hydrolysis and utilization in microorganisms that exhibit high product yield and tolerance. A concluding discussion identifies unresolved issues pertaining to microbial cellulose utilization, suggests approaches by which such issues might be resolved, and contrasts a microbially oriented cellulose hydrolysis paradigm to the more conventional enzymatically oriented paradigm in both fundamental and applied contexts.
Publisher: Proceedings of the National Academy of Sciences
Date: 06-08-1996
Abstract: Pseudohyphal differentiation in Saccharomyces cerevisiae was first described as a response of diploid cells to nitrogen limitation. Here we report that haploid and diploid starch-degrading S. cerevisiae strains were able to switch from a yeast form to a filamentous pseudohyphal form in response to carbon limitation in the presence of an le supply of nitrogen. Two genes, MSS10 and MUC1, were cloned and shown to be involved in pseudohyphal differentiation and invasive growth. The deletion of MSS10 resulted in extremely reduced amounts of pseudohyphal differentiation and invasive growth, whereas the deletion of MUC1 abolished pseudohyphal differentiation and invasive growth completely. Mss10 appears to be a transcriptional activator that responds to nutrient limitation and coregulates the expression of MUC1 and the STA1-3 glucoamylase genes, which are involved in starch degradation. MUC1 encodes a 1367-amino acid protein, containing several serine/threonine-rich repeats. Muc1 is a putative integral membrane-bound protein, similar to mammalian mucin-like membrane proteins that have been implicated to play a role in the ability of cancer cells to invade other tissues.
Publisher: Elsevier BV
Date: 12-2019
Publisher: Elsevier
Date: 2022
Publisher: American Society for Microbiology
Date: 12-2002
Publisher: Springer Berlin Heidelberg
Date: 2009
Publisher: Springer Science and Business Media LLC
Date: 07-1988
DOI: 10.1007/BF00405846
Publisher: EMBO
Date: 12-11-2010
Publisher: MDPI AG
Date: 08-07-2022
DOI: 10.3390/APP12146919
Abstract: In this study, the impact of the apiculate yeast Hanseniaspora occidentalis as a co-partner with Saccharomyces cerevisiae was investigated in a sequential-type mixed-culture fermentation of Muscaris grape must. As with other fermentation trials using Hanseniaspora strains, a significant increase in ethyl acetate was observed, but most intriguing was the almost complete abolition of malic acid (from 2.0 g/L to 0.1 g/L) in the wine. Compared to the pure S. cerevisiae inoculum, there was also a marked increase in the concentrations of the other acetate esters. Modulation of some of the varietal elements, such as rose oxide, was also observed. This work shows the promising use of H. occidentalis in a mixed-culture must fermentation, especially in the acid modulation of fruit juice matrices.
Publisher: Zenodo
Date: 2019
Publisher: Elsevier BV
Date: 04-1991
DOI: 10.1016/0378-1119(91)90354-E
Abstract: We have determined the complete nucleotide (nt) sequence of a 5070-bp DNA fragment containing a glucoamylase-encoding gene (STA2) from Saccharomyces diastaticus. The 5' transcription start points for STA1, STA2 and STA3 were determined by primer extension of their respective mRNAs using reverse transcriptase. The sequence data show one major open reading frame (ORF) of 767 amino acids encoding GAII with a calculated Mr of 82,514. The 5' region in the ORF contains two ATG sequences within 30 nt of each other. The upstream region of STA2 was lified by the polymerase chain reaction (PCR) and fused to the Escherichia coli lacZ gene. Some of the PCR products contained mutations in ATG1 and/or ATG2. Results indicated that both ATG1 and ATG2 encode functional translation start codons, but ATG2 was shown to encode the stronger initiator. The upstream region of STA2 contains a canonical sequence that is homologous to known sites of repression by the MATa/MAT alpha-encoded repressor. Also, consensus RAP1 (Repressor-Activator Protein 1)-binding sites are located in the 5' upstream region and within the coding region of STA2.
Publisher: Wiley
Date: 15-07-2006
DOI: 10.1002/YEA.1382
Abstract: The fruity odours of wine are largely derived from the synthesis of esters and higher alcohols during yeast fermentation. The ATF1- and ATF2-encoded alcohol acetyltransferases of S. cerevisiae are responsible for the synthesis of ethyl acetate and isoamyl acetate esters, while the EHT1-encoded ethanol hexanoyl transferase is responsible for synthesizing ethyl caproate. However, esters such as these might be degraded by the IAH1-encoded esterase. The objectives of this study were: (a) to overexpress the genes encoding ester-synthesizing and ester-degrading enzymes in wine yeast (b) to prepare Colombard table wines and base wines for distillation using these modified strains and (c) to analyse and compare the ester concentrations and aroma profiles of these wines and distillates. The overexpression of ATF1 significantly increased the concentrations of ethyl acetate, isoamyl acetate, 2-phenylethyl acetate and ethyl caproate, while the overexpression of ATF2 affected the concentrations of ethyl acetate and isoamyl acetate to a lesser degree. The overexpression of IAH1 resulted in a significant decrease in ethyl acetate, isoamyl acetate, hexyl acetate and 2-phenylethyl acetate. The overexpression of EHT1 resulted in a marked increase in ethyl caproate, ethyl caprylate and ethyl caprate. The flavour profile of the wines and distillates prepared using the modified strains were also significantly altered as indicated by formal sensory analysis. This study offers prospects for the development of wine yeast starter strains with optimized ester-producing capability that could assist winemakers in their effort to consistently produce wine and distillates such as brandy to definable flavour specifications and styles.
Publisher: Springer Science and Business Media LLC
Date: 10-1995
DOI: 10.1007/BF00310817
Abstract: Circadian rhythmicity in transcriptomic profiles has been shown in many physiological processes, and the disruption of circadian patterns has been found to associate with several diseases. In this paper, we developed a series of likelihood-based methods to detect (i) circadian rhythmicity (denoted as LR_rhythmicity) and (ii) differential circadian patterns comparing two experimental conditions (denoted as LR_diff). In terms of circadian rhythmicity detection, we demonstrated that our proposed LR_rhythmicity could better control the type I error rate compared to existing methods under a wide variety of simulation settings. In terms of differential circadian patterns, we developed methods in detecting differential litude, differential phase, differential basal level and differential fit, which also successfully controlled the type I error rate. In addition, we demonstrated that the proposed LR_diff could achieve higher statistical power in detecting differential fit, compared to existing methods. The superior performance of LR_rhythmicity and LR_diff was demonstrated in four real data applications, including a brain aging data (gene expression microarray data of human postmortem brain), a time-restricted feeding data (RNA sequencing data of human skeletal muscles) and a scRNAseq data (single cell RNA sequencing data of mouse suprachiasmatic nucleus). An R package for our methods is publicly available on GitHub iffCircadian/diffCircadian.
Publisher: Springer Science and Business Media LLC
Date: 07-1993
DOI: 10.1007/BF00324662
Publisher: Springer Science and Business Media LLC
Date: 07-1988
DOI: 10.1007/BF00405847
Abstract: With increased interest in integrating environmental justice into the process for developing environmental regulations in the United States, analysts and decision makers are confronted with the question of what methods and data can be used to assess disproportionate environmental health impacts. However, as a first step to identifying data and methods, it is important that analysts understand what information on equity impacts is needed for decision making. Such knowledge originates from clearly stated equity objectives and the reflection of those objectives throughout the analytical activities that characterize Regulatory Impact Analysis (RIA), a process that is traditionally used to inform decision making. The framework proposed in this paper advocates structuring analyses to explicitly provide pre-defined output on equity impacts. Specifically, the proposed framework emphasizes: (a) defining equity objectives for the proposed regulatory action at the onset of the regulatory process, (b) identifying specific and related sub-objectives for key analytical steps in the RIA process, and (c) developing explicit analytical/research questions to assure that stated sub-objectives and objectives are met. In proposing this framework, it is envisioned that information on equity impacts informs decision-making in regulatory development, and that this is achieved through a systematic and consistent approach that assures linkages between stated equity objectives, regulatory analyses, selection of policy options, and the design of compliance and enforcement activities.
Publisher: Elsevier BV
Date: 11-2002
DOI: 10.1016/S0167-7799(02)02058-9
Abstract: Grapevine biotechnology is one of the most promising developments in the global wine industry, which is increasingly faced with conflicting demands from markets, consumers and environmentalists. In the grapevine industries, this technology and its supporting disciplines entail the establishment of stress tolerant and disease resistant varieties of Vitis vinifera, with increased productivity, efficiency, sustainability and environmental friendliness, especially regarding improved pest and disease control, water use efficiency and grape quality. The implementation and successful commercialisation of genetically improved grapevine varieties will only be realized if an array of hurdles, both scientific and otherwise, can be overcome.
Publisher: Springer Science and Business Media LLC
Date: 23-04-2019
Publisher: Springer Science and Business Media LLC
Date: 1983
DOI: 10.1007/BF00499499
Publisher: Oxford University Press (OUP)
Date: 12-2005
DOI: 10.1016/J.FEMSYR.2005.05.004
Abstract: The FLO11-encoded flocculin is required for a variety of important phenotypes in Saccharomyces cerevisiae, including flocculation, adhesion to agar and plastic, invasive growth, pseudohyphae formation and biofilm development. We present evidence that Flo11p belongs to the Flo1-type class of flocculins rather than to the NewFlo class. Both Flo1-type and NewFlo yeast flocculation are inhibited by mannose. NewFlo flocculation, however, is also inhibited by several other carbohydrates including glucose, maltose and sucrose. These differences have in at least one case been shown to reflect differences in the structure of the carbohydrate-binding site of the flocculins. We report that Flo11p-dependent flocculation is inhibited by mannose, but not by glucose, maltose or sucrose. Furthermore, Flo11p contains a peptide sequence highly similar to one that has been shown to characterise Flo1-type flocculins. Further characterisation of the properties of Flo11p-dependent flocculation revealed that it is dependent on calcium, occurs only at cell densities greater than 1 x 10(8) ml(-1), and only occurs at acidic pH.
Publisher: Elsevier BV
Date: 07-2012
Publisher: Informa UK Limited
Date: 1991
DOI: 10.3109/10409239109081720
Abstract: Saccharomyces cerevisiae has been used widely both as a model system for unraveling the biochemical, genetic, and molecular details of gene expression and the secretion process, and as a host for the production of heterologous proteins of biotechnological interest. The potential of starch as a renewable biological resource has stimulated research into amylolytic enzymes and the broadening of the substrate range of S. cerevisiae. The enzymatic hydrolysis of starch, consisting of linear (amylose) and branched glucose polymers (amylopectin), is catalyzed by alpha- and beta-amylases, glucoamylases, and debranching enzymes, e.g., pullulanases. Starch utilization in the yeast S. cerevisiae var. diastaticus depends on the expression of the three unlinked genes, STA1 (chr. IV), STA2 (chr. II), and STA3 (chr. XIV), each encoding one of the extracellular glycosylated glucoamylases isozymes GAI, GAII, or GAIII, respectively. The restriction endonuclease maps of STA1, STA2, and STA3 are identical. These genes are absent in S. cerevisiae, but a related gene, SGA1, encoding an intracellular, sporulation-specific glucoamylase (SGA), is present. SGA1 is homologous to the middle and 3' regions of the STA genes, but lacks a 5' sequence that encodes the domain for secretion of the extracellular glucoamylases. The STA genes are positively regulated by the presence of three GAM genes. In addition to positive regulation, the STA genes are regulated negatively at three levels. Whereas strains of S. diastaticus are capable of expressing the STA genes, most strains of S. cerevisiae contain STA10, whose presence represses the expression of the STA genes in an undefined manner. The STA genes are also repressed in diploid cells, presumably by the MATa/MAT alpha-encoded repressor. STA gene expression is reduced in liquid synthetic media, it is carbon catabolite repressed by glucose, and is inhibited in petite mutants.
Publisher: American Chemical Society (ACS)
Date: 12-02-2002
DOI: 10.1021/JF0111514
Abstract: Juice of the Sclerocarya birrea subsp. caffra (marula) fruit was fermented by indigenous microflora and different commercial Saccharomyces cerevisiae yeast strains at different temperatures, namely, 15 and 30 degrees C. Volatile acids, esters, and higher alcohols were quantified in the wine and distillates, and the results were interpreted using a multivariate analysis of variance and an average linkage cluster analysis. Significant differences between 15 and 30 degrees C and also among yeasts with respect to volatile compounds were observed. Yeast strains VIN7 and FC consistently produced wines and final distillates significantly different from the other strains. A panel of tasters and marula and brandy producers was asked to select wines and distillates that had an acceptable and typical marula "nose". They were also asked to detect the differences among wines and distillates fermented with the same yeast strain at different temperatures.
Publisher: Elsevier BV
Date: 08-2004
Publisher: Springer Science and Business Media LLC
Date: 08-04-2023
DOI: 10.1038/S41467-023-37748-7
Abstract: Naturally evolved organisms typically have large genomes that enable their survival and growth under various conditions. However, the complexity of genomes often precludes our complete understanding of them, and limits the success of biotechnological designs. In contrast, minimal genomes have reduced complexity and therefore improved engineerability, increased biosynthetic capacity through the removal of unnecessary genetic elements, and less recalcitrance to complete characterisation. Here, we review the past and current genome minimisation and re-functionalisation efforts, with an emphasis on the latest advances facilitated by synthetic genomics, and provide a critical appraisal of their potential for industrial applications.
Publisher: American Chemical Society (ACS)
Date: 13-05-2004
DOI: 10.1021/JF035431Q
Abstract: Principal component analysis (PCA) was used to identify the main sources of variation in the Fourier transform infrared (FT-IR) spectra of 329 wines of various styles. The FT-IR spectra were gathered using a specialized WineScan instrument. The main sources of variation included the reducing sugar and alcohol content of the s les, as well as the stage of fermentation and the maturation period of the wines. The implications of the variation between the different wine styles for the design of calibration models with accurate predictive abilities were investigated using glycerol calibration in wine as a model system. PCA enabled the identification and interpretation of s les that were poorly predicted by the calibration models, as well as the detection of in idual s les in the s le set that had atypical spectra (i.e., outlier s les). The Soft Independent Modeling of Class Analogy (SIMCA) approach was used to establish a model for the classification of the outlier s les. A glycerol calibration for wine was developed (reducing sugar content 8% v/v) with satisfactory predictive ability (SEP = 0.40 g/L). The RPD value (ratio of the standard deviation of the data to the standard error of prediction) was 5.6, indicating that the calibration is suitable for quantification purposes. A calibration for glycerol in special late harvest and noble late harvest wines (RS 31-147 g/L, alcohol > 11.6% v/v) with a prediction error SECV = 0.65 g/L, was also established. This study yielded an analytical strategy that combined the careful design of calibration sets with measures that facilitated the early detection and interpretation of poorly predicted s les and outlier s les in a s le set. The strategy provided a powerful means of quality control, which is necessary for the generation of accurate prediction data and therefore for the successful implementation of FT-IR in the routine analytical laboratory.
Publisher: Informa UK Limited
Date: 14-02-2023
Publisher: American Society for Microbiology
Date: 10-2008
DOI: 10.1128/AEM.00394-08
Abstract: In many industrial fermentation processes, the Saccharomyces cerevisiae yeast should ideally meet two partially conflicting demands. During fermentation, a high suspended yeast count is required to maintain a satisfactory rate of fermentation, while at completion, efficient settling is desired to enhance product clarification and recovery. In most fermentation industries, currently used starter cultures do not satisfy this ideal, probably because nonflocculent yeast strains were selected to avoid fermentation problems. In this paper, we assess molecular strategies to optimize the flocculation behavior of S. cerevisiae . For this purpose, the chromosomal copies of three dominant flocculation genes, FLO1 , FLO5 , and FLO11 , of the haploid nonflocculent, noninvasive, and non-flor-forming S. cerevisiae FY23 strain were placed under the transcriptional control of the promoters of the ADH2 and HSP30 genes. All six promoter-gene combinations resulted in specific flocculation behaviors in terms of timing and intensity. The strategy resulted in stable expression patterns providing a platform for the direct comparison and assessment of the specific impact of the expression of in idual dominant FLO genes with regard to cell wall characteristics, such as hydrophobicity, biofilm formation, and substrate adhesion properties. The data also clearly demonstrate that the flocculation behavior of yeast strains can be tightly controlled and fine-tuned to satisfy specific industrial requirements.
Publisher: Wiley
Date: 11-05-2007
DOI: 10.1002/YEA.1493
Abstract: Volatile thiols, such as 4-mercapto-4-methylpentan-2-one (4MMP), 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA), are among the most potent aroma compounds found in wine and can have a significant effect on wine quality and consumer preferences. At optimal concentrations in wine, these compounds impart flavours of passionfruit, grapefruit, gooseberry, blackcurrant, lychee, guava and box hedge. The enzymatic release of aromatic thiols from grape-derived, non-volatile cysteinylated precursors (Cys-4MMP and Cys-3MH) and the further modification thereof (conversion of 3MH into 3MHA) during fermentation, enhance the varietal characters of wines such as Sauvignon Blanc. Wine yeast strains have limited and varying capacities to produce aroma-enhancing thiols from their non-volatile counterparts in grape juice. Even under optimal fermentation conditions, the most efficient thiol-releasing Saccharomyces cerevisiae wine strain known realizes less than 5% of the thiol-related flavour potential of grape juice. The objective of this study was to develop a wine yeast able to unleash the untapped thiol aromas in grape juice during winemaking. To achieve this goal, the Escherichia coli tnaA gene, encoding a tryptophanase with strong cysteine-beta-lyase activity, was cloned and overexpressed in a commercial wine yeast strain under the control of the regulatory sequences of the yeast phosphoglycerate kinase I gene (PGK1). This modified strain expressing carbon-sulphur lyase activity released up to 25 times more 4MMP and 3MH in model ferments than the control host strain. Wines produced with the engineered strain displayed an intense passionfruit aroma. This yeast offers the potential to enhance the varietal aromas of wines to predetermined market specifications.
Publisher: Wiley
Date: 08-1993
DOI: 10.1111/J.1365-2672.1993.TB02760.X
Abstract: A 1209-base pair (bp) DNA fragment containing the endopolygalacturonase-encoding gene (peh1) from Erwinia carotovora subsp. carotovora was lified by the polymerase chain reaction (PCR) technique and expressed in Escherichia coli. The nucleotide sequence of the PCR product was determined and found to be highly homologous to the primary structures of other polygalacturonase-encoding genes. The peh1 DNA fragment encoding the mature polygalacturonase was inserted between two different yeast expression-secretion cassettes and a yeast gene terminator, generating recombinant yeast-integrating shuttle plasmids pAMS10 and pAMS11. These YIp5-derived plasmids were transformed and stably integrated into the genome of a laboratory strain of Saccharomyces cerevisiae. Transcription initiation signals present in these expression-secretion cassettes were derived from the yeast alcohol dehydrogenase (ADC1P) or mating pheromone alpha-factor (MF alpha 1P) gene promoters. The transcription termination signals were derived from the yeast tryptophan synthase gene terminator (TRP5T). Secretion of polygalacturonase was directed by the signal sequence of the yeast mating pheromone alpha-factor (MF alpha 1S). Northern blot analysis revealed the presence of peh1 mRNA in the yeast transformants and a polypectate agarose test was used to monitor polygalacturonase production.
Publisher: Apple Academic Press
Date: 24-02-2014
DOI: 10.1201/B16586
Publisher: Springer Science and Business Media LLC
Date: 27-10-2006
DOI: 10.1007/S11248-006-9019-1
Abstract: Polygalacturonase-inhibiting proteins (PGIPs) selectively inhibit polygalacturonases (PGs) secreted by invading plant pathogenic fungi. PGIPs display differential inhibition towards PGs from different fungi, also towards different isoforms of PGs originating from a specific pathogen. Recently, a PGIP-encoding gene from Vitis vinifera (Vvpgip1) was isolated and characterised. PGIP purified from grapevine was shown to inhibit crude polygalacturonase extracts from Botrytis cinerea, but this inhibitory activity has not yet been linked conclusively to the activity of the Vvpgip1 gene product. Here we use a transgenic over-expression approach to show that the PGIP encoded by the Vvpgip1 gene is active against PGs of B. cinerea and that over-expression of this gene in transgenic tobacco confers a reduced susceptibility to infection by this pathogen. A calculated reduction in disease susceptibility of 47-69% was observed for a homogeneous group of transgenic lines that was statistically clearly separated from untransformed control plants following infection with Botrytis over a 15-day-period. VvPGIP1 was subsequently purified from transgenic tobacco and used to study the specific inhibition profile of in idual PGs from Botrytis and Aspergillus. The heterologously expressed and purified VvPGIP1 selectively inhibited PGs from both A. niger and B. cinerea, including BcPG1, a PG from B. cinerea that has previously been shown to be essential for virulence and symptom development. Altogether our data confirm the antifungal nature of the VvPGIP1, and the in vitro inhibition data suggest at least in part, that the VvPGIP1 contributed to the observed reduction in disease symptoms by inhibiting the macerating action of certain Botrytis PGs in planta. The ability to correlate inhibition profiles to in idual PGs provides a more comprehensive analysis of PGIPs as antifungal genes with biotechnological potential, and adds to our understanding of the importance of PGIP:PG interactions during disease and symptom development in plants.
Publisher: Wiley
Date: 2000
DOI: 10.1002/1097-0061(20000615)16:8<675::AID-YEA585>3.0.CO;2-B
Publisher: Wiley
Date: 30-06-1995
Abstract: The polymorphic extracellular glucoamylase-encoding genes STA1 (chr. IV), STA2 (chr. II) and STA3 (chr. XIV), from Saccharomyces cerevisiae var. diastaticus probably evolved by genomic rearrangement of DNA regions (S1, S2 and SGA1) present in S. cerevisiae, and subsequent translocation to unlinked regions of chromosomal regions. S1, encoding a homologue to the threonine/serine-rich domain of STA glucoamylases (GAI-III), mapped to the right arm of chromosome IX. S2, encoding the hydrophobic leader peptide of GAI-III), was also mapped on the right arm of chromosome IX, next to S1, close to DAL81. The SGA1 sporulation-specific, intracellular glucoamylase-encoding gene is located on the left arm of chromosome IX, 32 kb proximal of HIS5.
Publisher: Springer Science and Business Media LLC
Date: 09-2012
Publisher: Oxford University Press (OUP)
Date: 23-04-2002
Publisher: Informa UK Limited
Date: 1995
DOI: 10.3109/07435809509030445
Abstract: Cytochrome P450c17 (P450c17), together with cytochrome P450c21 (P450c21), plays an important role in progesterone metabolism in the mammalian adrenal cortex. Low levels of expression and the presence of other steroidogenic enzymes in adrenal cortex endoplasmic reticulum (ER) impedes purification and characterisation of wild type as well as mutant forms of the hemoprotein. Heterologous gene expression systems have previously been used successfully to express active P450c17. Heterologous expression can also be used for the preparation of anti-P450c17-IgG. For antibody production larger amounts of pure P450c17 peptide, rather than the active protein, is, however, desirable. If the expressed protein can be affinity tagged and secreted into the medium, isolation and purification will be facilitated. Saccharomyces cerevisiae, YPH259, was transformed with a modified YCplac111 yeast expression-secretion vector (pPRL2). The gene coding for a truncated human P450c17 (signal anchor sequence 1-18 was removed) was inserted, in reading frame, downstream from the leader sequence MF alpha. A histidine tag was incorporated at the C-terminus. The modified yeast expression vector was expressed in yeast, the secreted P450c17-peptide purified by affinity chromatography and identified by immunoblot analysis.
Publisher: MDPI AG
Date: 16-02-2021
DOI: 10.3390/IJMS22041943
Abstract: Apiculate yeasts belonging to the genus Hanseniaspora are commonly isolated from viticultural settings and often dominate the initial stages of grape must fermentations. Although considered spoilage yeasts, they are now increasingly becoming the focus of research, with several whole-genome sequencing studies published in recent years. However, tools for their molecular genetic manipulation are still lacking. Here, we report the development of a tool for the genetic modification of Hanseniaspora uvarum. This was employed for the disruption of the HuATF1 gene, which encodes a putative alcohol acetyltransferase involved in acetate ester formation. We generated a synthetic marker gene consisting of the HuTEF1 promoter controlling a hygromycin resistance open reading frame (ORF). This new marker gene was used in disruption cassettes containing long-flanking (1000 bp) homology regions to the target locus. By increasing the antibiotic concentration, transformants were obtained in which both alleles of the putative HuATF1 gene were deleted in a diploid H. uvarum strain. Phenotypic characterisation including fermentation in Müller-Thurgau must showed that the null mutant produced significantly less acetate ester, particularly ethyl acetate. This study marks the first steps in the development of gene modification tools and paves the road for functional gene analyses of this yeast.
Publisher: Oxford University Press (OUP)
Date: 10-2003
Publisher: MDPI AG
Date: 16-07-2018
DOI: 10.3390/FERMENTATION4030054
Abstract: Yeast—especially Saccharomyces cerevisiae—have long been a preferred workhorse for the production of numerous recombinant proteins and other metabolites. S. cerevisiae is a noteworthy aroma compound producer and has also been exploited to produce foreign bioflavour compounds. In the past few years, important strides have been made in unlocking the key elements in the biochemical pathways involved in the production of many aroma compounds. The expression of these biochemical pathways in yeast often involves the manipulation of the host strain to direct the flux towards certain precursors needed for the production of the given aroma compound. This review highlights recent advances in the bioengineering of yeast—including S. cerevisiae—to produce aroma compounds and bioflavours. To capitalise on recent advances in synthetic yeast genomics, this review presents yeast as a significant producer of bioflavours in a fresh context and proposes new directions for combining engineering and biology principles to improve the yield of targeted aroma compounds.
Publisher: American Chemical Society (ACS)
Date: 23-10-2008
DOI: 10.1021/JF801695H
Abstract: Alcoholic fermentation using Saccharomyces wine yeast is an effective means of modulating wine aroma. This study investigated the impact of coinoculating commercial yeast strains (Vin7, QA23, Vin13) on the volatile composition and sensory profile of Sauvignon Blanc wines. Small-scale replicated fermentations were conducted using single-strain and coinoculations of Vin7 with QA23 and with Vin13. The results showed that the chemical and sensory profiles of the coinoculated wines were different from both the single-strain wines and equal blends of the single-strain wines. Volatile thiol analysis indicated that the Vin7/QA23 coinoculated wines were highest in 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA), although this pattern was not observed for the Vin7/Vin13 yeast combination. The negative white vinegar aroma and high volatile acidity measured in the Vin7 single-strain wines were not present in the coinoculated wines. This study demonstrates that coinoculations can modify the aroma profile of wines, when complementary yeasts are used.
Publisher: MDPI AG
Date: 27-06-2018
DOI: 10.20944/PREPRINTS201806.0451.V1
Abstract: Abstract: Yeast & ndash especially Saccharomyces cerevisiae & ndash have long been a preferred workhorse for the production of numerous recombinant proteins and other metabolites. S. cerevisiae is a noteworthy aroma compound producer, and has also been exploited to produce foreign bioflavour compounds. In the past few years, important strides have been made in unlocking the key elements in the biochemical pathways involved in the production of many aroma compounds. The expression of these biochemical pathways in yeast often involves the manipulation of the host strain to direct the flux towards certain precursors needed for the production of the given aroma compound. This review highlights recent advances in the bioengineering of yeast & ndash including S. cerevisiae & ndash to produce aroma compounds and bioflavours. To capitalise on recent advances in synthetic yeast genomics, this review presents yeast as a significant producer of bioflavours in a fresh context and proposes new directions for combining engineering and biology principles to improve the yield of targeted aroma compounds.
Publisher: Cold Spring Harbor Laboratory
Date: 04-11-2020
DOI: 10.1101/2020.11.03.367615
Abstract: The unfolded protein response (UPR) is a highly conserved cellular response in eukaryotic cells to counteract endoplasmic reticulum (ER) stress, typically triggered by unfolded protein accumulation. In addition to its relevance to human diseases like cancer cell development, the induction of the UPR has a significant impact on recombinant protein production yields in microbial cell factories, including the industrial workhorse Saccharomyces cerevisiae. Being able to accurately detect and measure this ER stress response in single cells, enables the rapid optimisation of protein production conditions and high-throughput strain selection strategies. Current methodologies to monitor the UPR in S. cerevisiae are often temporally and spatially removed from the cultivation stage, or lack updated systematic evaluation. To this end we constructed and systematically evaluated a series of high-throughput UPR sensors by different designs, incorporating either yeast native UPR promoters or novel synthetic minimal UPR promoters. The native promoters of DER1 and ERO1 were identified to have suitable UPR biosensor properties and served as an expression level guide for orthogonal sensor benchmarking. Our best synthetic minimal sensor, SM1, was only 98 bp in length, had minimal homology to other native yeast sequences and displayed superior sensor characteristics. Using this synthetic minimal UPR sensor, we demonstrate its ability to accurately discriminate between cells expressing different heterologous proteins and at varying production levels. Our sensor is thus a novel high-throughput tool for determining expression/engineering strategies for optimal heterologous protein production.
Publisher: MDPI AG
Date: 07-05-2020
DOI: 10.3390/FERMENTATION6020049
Abstract: Recently, there has been a growing interest in the role of non-Saccharomyces yeast (NSY) as a coculturing partner with Saccharomyces cerevisiae during grape must fermentation. We investigated three new strains, namely Nakazawaea ishiwadae, Candida railenensis and Debaryomyces hansenii, for their oenological potential in mixed-culture micro-vinifications with S. cerevisiae Vin13 using Muscaris grape must. None of the NSY strains impeded the fermentation performance as all the mixed-culture experiments finished at the same time. Coculturing with N. ishiwadae yielded significantly higher concentrations of ethyl and acetate esters in the final wine product. Apart from higher acetic acid levels, wines produced with C. railenensis and D. hansenii yielded much lower esters concentrations. The concentrations of certain terpenes and norisoprenoids were also significantly modulated in the mixed-culture fermentations. This study reveals the rarely reported species of N. ishiwadae as a promising coculturing partner for increasing aroma-active compounds in a wine.
Publisher: Springer Science and Business Media LLC
Date: 04-2007
DOI: 10.1007/S00253-006-0828-1
Abstract: Sulfur compounds in wine can be a 'double-edged sword'. On the one hand, certain sulfur-containing volatile compounds such as hydrogen sulfide, imparting a rotten egg-like aroma, can have a negative impact on the perceived quality of the wine, and on the other hand, some sulfur compounds such as 3-mercaptohexanol, imparting fruitiness, can have a positive impact on wine flavor and aroma. Furthermore, these compounds can become less or more attractive or repulsive depending on their absolute and relative concentrations. This presents an interesting challenge to the winemaker to modulate the concentrations of these quality-determining compounds in wine in accordance with consumer preferences. The wine yeast Saccharomyces cerevisiae plays a central role in the production of volatile sulfur compounds. Through the sulfate reduction sequence pathway, the HS(-) is formed, which can lead to the formation of hydrogen sulfide and various mercaptan compounds. Therefore, limiting the formation of the HS(-) ion is an important target in metabolic engineering of wine yeast. The wine yeast is also responsible for the transformation of non-volatile sulfur precursors, present in the grape, to volatile, flavor-active thiol compounds. In particular, 4-mercapto-4-methylpentan-2-one, 3-mercaptohexanol, and 3-mercaptohexyl acetate are the most important volatile thiols adding fruitiness to wine. This paper briefly reviews the metabolic processes involved in the production of important volatile sulfur compounds and the latest strategies in the pursuit of developing wine yeast strains as tools to adjust wine aroma to market specifications.
Publisher: Oxford University Press (OUP)
Date: 04-2005
DOI: 10.1111/J.1365-2672.2004.02549.X
Abstract: The objective of this study was to investigate the effects of free molecular and bound forms of sulphur dioxide and oxygen on the viability and culturability of a selected strain of Acetobacter pasteurianus and a selected strain of Brettanomyces bruxellensis in wine. Acetic acid bacteria and Brettanomyces/Dekkera yeasts associated with wine spoilage were isolated from bottled commercial red wines. One bacterium, A. pasteurianus strain A8, and one yeast, B. bruxellensis strain B3a, were selected for further study. The resistance to sulphur dioxide and the effect of oxygen addition on these two selected strains were determined by using plating and epifluorescence techniques for monitoring cell viability in wine. Acetobacter pasteurianus A8 was more resistant to sulphur dioxide than B. bruxellensis B3a, with the latter being rapidly affected by a short exposure time to free molecular form of sulphur dioxide. As expected, neither of these microbial strains was affected by the bound form of sulphur dioxide. The addition of oxygen negated the difference observed between plate and epifluorescence counts for A. pasteurianus A8 during storage, while it stimulated growth of B. bruxellensis B3a. Acetobacter pasteurianus A8 can survive under anaerobic conditions in wine in the presence of sulphur dioxide. Brettanomyces bruxellensis B3a is more sensitive to sulphur dioxide than A. pasteurianus A8, but can grow in the presence of oxygen. Care should be taken to exclude oxygen from contact with wine when it is being transferred or moved. Wine spoilage can be avoided by preventing growth of undesirable acetic acid bacteria and Brettanomyces/Dekkera yeasts through the effective use of sulphur dioxide and the management of oxygen throughout the winemaking process.
Publisher: Elsevier BV
Date: 11-2006
DOI: 10.1016/J.MIMET.2006.03.019
Abstract: A rapid screening method for the evaluation of the major fermentation products of Saccharomyces wine yeasts was developed using Fourier transform infrared spectroscopy and principal component factor analysis. Calibration equations for the quantification of volatile acidity, glycerol, ethanol, reducing sugar and glucose concentrations in fermented Chenin blanc and synthetic musts were derived from the Fourier transform infrared spectra of small-scale fermentations. The accuracy of quantification of volatile acidity in both Chenin blanc and synthetic must was excellent, and the standard error of prediction was 0.07 g l(-1) and 0.08 g l(-1), respectively. The respective standard error of prediction in Chenin blanc and synthetic musts for ethanol was 0.32% v/v and 0.31% v/v, for glycerol was 0.38 g l(-1) and 0.32 g l(-1), for reducing sugar in Chenin blanc must was 0.56 g l(-1) and for glucose in synthetic must was 0.39 g l(-1). These values were in agreement with the accuracy obtained by the respective reference methods used for the quantification of the components. The screening method was applied to quantify the fermentation products of glycerol-overproducing hybrid yeasts and commercial wine yeasts. Principal component factor analysis of the fermentation data facilitated an overall comparison of the fermentation profiles (in terms of the components tested) of the strains. The potential of Fourier transform infrared spectroscopy as a tool to rapidly screen the fermentative properties of wine yeasts and to speed up the evaluation processes in the initial stages of yeast strain development programs is shown.
Publisher: Elsevier BV
Date: 10-1990
Publisher: Elsevier BV
Date: 12-2019
Publisher: Canadian Science Publishing
Date: 08-2009
DOI: 10.1139/W09-039
Abstract: The use and release of genetically modified organisms (GMOs) is an issue of intense public concern and, in the case of food and beverages, products containing GMOs or products thereof carry the risk of consumer rejection. The recent commercialization of 2 GM wine yeasts in the United States and Canada has made research and development of risk assessments for GM microorganisms a priority. The purpose of this study was to take a first step in establishing a risk-assessment process for future use and potential release of GM wine yeasts into the environment. The behaviour and spread of a GM wine yeast was monitored in saturated sand columns, saturated sand flow cells, and conventional flow cells. A widely used commercial Saccharomyces cerevisiae wine yeast, VIN13, a VIN13 transgenic strain (LKA1, which carries the LKA1 α-amylase gene of Lipomyces kononenkoae ), a soil bacterium ( Dyadobacter fermentens ), and a nonwine soil-borne yeast ( Cryptococcus laurentii ) were compared in laboratory-scale microcosm systems designed to monitor microbial mobility behaviour, survival, and attachment to surfaces. It was found that LKA1 cells survived in saturated sand columns, but showed little mobility in the porous matrix, suggesting that the cells attached with high efficiency to sand. There was no significant difference between the mobility patterns of LKA1 and VIN13. All 3 yeasts (VIN13, LKA1, and C. laurentii) were shown to form stable biofilms the 2 S. cerevisiae strains either had no difference in biofilm density or the LKA1 biofilm was less dense than that of VIN13. When co-inoculated with C. laurentii, LKA1 had no negative influence on the breakthrough of the Cryptococcus yeast in a sand column or on its ability to form biofilms. In addition, LKA1 did not successfully integrate into a stable mixed-biofilm community, nor did it disrupt the biofilm community. Overall, it was concluded that the LKA1 transgenic yeast had the same reproductive success as VIN13 in these 3 microcosms and had no selective advantage over the untransformed parental strain.
Publisher: Elsevier BV
Date: 04-1989
Publisher: Elsevier BV
Date: 04-2013
DOI: 10.1016/J.TIG.2012.10.014
Abstract: Wine is arguably the oldest biotechnological endeavor, with humans having been involved in wine production for at least 7000 years. Despite the artisan nature of its production, work by pioneering scientists such as Antoine-Laurent de Lavoisier and Louis Pasteur placed wine research in a prominent position for the application of cutting-edge biological and chemical sciences, a position it still holds to this day. Technologies such as whole-genome sequencing and systems biology are now revolutionizing winemaking by combining the ability to engineer phenotypes rationally, with a precise understanding of the genetic makeup and key phenotypic drivers of the key organisms that contribute to this age-old industry.
Publisher: Oxford University Press (OUP)
Date: 11-2014
Publisher: Wiley
Date: 31-08-2023
DOI: 10.1002/YEA.3897
Publisher: Oxford University Press (OUP)
Date: 12-12-2019
Abstract: Wine is an archetypal traditional fermented beverage with strong territorial and socio-cultural connotations. Its 7000 year history is patterned by a tradition of innovation. Every value-adding innovation − whether in the vineyard, winery, supply chain or marketplace − that led to the invention of a new tradition spurred progress and created a brighter future from past developments. In a way, wine traditions can be defined as remembered innovations from the distant past − inherited knowledge and wisdom that withstood the test of time. Therefore, it should not be assumed a priori that tradition and innovation are polar opposites. The relations between the forces driven by the anchors of tradition and the wings of innovation do not necessarily involve displacement, conflict or exclusiveness. Innovation can strengthen wine tradition, and the reinvention of a tradition-bound practice, approach or concept can foster innovation. In cases where a paradigm-shifting innovation disrupts a tradition, the process of such an innovation transitioning into a radically new tradition can become protracted while proponents of ergent opinions duke it out. Sometimes these conflicting opinions are based on fact, and sometimes not. The imperfections of such a debate between the ‘ancients’ and the ‘moderns’ can, from time to time, obscure the line between myth and reality. Therefore, finding the right balance between traditions worth keeping and innovations worth implementing can be complex. The intent here is to harness the creative tension between science fiction and science fact when innovation's first-principles challenge the status quo by re-examining the foundational principles about a core traditional concept, such as terroir. Poignant questions are raised about the importance of the terroir (biogeography) of yeasts and the value of the microbiome of grapes to wine quality. This article imagines a metaphorical terroir free from cognitive biases where erse perspectives can converge to uncork the effervescent power of territorial yeast populations as well as ‘nomadic’ yeast starter cultures. At the same time, this paper also engages in mental time-travel. A future scenario is imagined, explored, tested and debated where terroir-less yeast avatars are equipped with designer genomes to safely and consistently produce, in idually or in combination with region-specific wild yeasts and or other starter cultures, high-quality wine according to the preferences of consumers in a range of markets. The purpose of this review is to look beyond the horizon and to synthesize a link between what we know now and what could be. This article informs readers where to look without suggesting what they must see as a way forward. In the context of one of the world's oldest fermentation industries − steeped in a rich history of tradition and innovation − the mantra here is: respect the past, lead the present and secure the future of wine.
Publisher: Springer Science and Business Media LLC
Date: 18-06-2022
Publisher: Oxford University Press (OUP)
Date: 08-2006
DOI: 10.1111/J.1567-1364.2006.00057.X
Abstract: In Saccharomyces cerevisiae, branched-chain amino acid transaminases (BCAATases) are encoded by the BAT1 and BAT2 genes. BCAATases catalyse the transfer of amino groups between those amino acids and alpha-keto-acids. alpha-Keto-acids are precursors for the biosynthesis of higher alcohols, which significantly influence the aroma and flavour of yeast-derived fermentation products. The objective of this study was to investigate the influence of BAT-gene expression on general yeast physiology, on aroma and flavour compound formation and on the sensory characteristics of wines and distillates. For this purpose, the genes were overexpressed and deleted in a laboratory strain, BY4742, and overexpressed in an industrial wine yeast strain, VIN13. The data show that, with the exception of a slow growth phenotype observed for the BAT1 deletion strain, the fermentation behaviour of the strains was unaffected by the modifications. The chemical and sensory analysis of fermentation products revealed a strong correction between BAT gene expression and the formation of many aroma compounds. The data suggest that the adjustment of BAT gene expression could play an important role in assisting winemakers in their endeavour to produce wines with specific flavour profiles.
Publisher: MDPI AG
Date: 12-2020
DOI: 10.3390/MICROORGANISMS8121914
Abstract: Genome-scale engineering and custom synthetic genomes are reshaping the next generation of industrial yeast strains. The Cre-recombinase-mediated chromosomal rearrangement mechanism of designer synthetic Saccharomyces cerevisiae chromosomes, known as SCRaMbLE, is a powerful tool which allows rapid genome evolution upon command. This system is able to generate millions of novel genomes with potential valuable phenotypes, but the excessive loss of essential genes often results in poor growth or even the death of cells with useful phenotypes. In this study we expanded the versatility of SCRaMbLE to industrial strains, and evaluated different control measures to optimize genomic rearrangement, whilst limiting cell death. To achieve this, we have developed RED (rapid evolution detection), a simple colorimetric plate-assay procedure to rapidly quantify the degree of genomic rearrangements within a post-SCRaMbLE yeast population. RED-enabled semi-synthetic strains were mated with the haploid progeny of industrial yeast strains to produce stress-tolerant heterozygous diploid strains. Analysis of these heterozygous strains with the RED-assay, genome sequencing and custom bioinformatics scripts demonstrated a correlation between RED-assay frequencies and physical genomic rearrangements. Here we show that RED is a fast and effective method to evaluate the optimal SCRaMbLE induction times of different Cre-recombinase expression systems for the development of industrial strains.
Publisher: Oxford University Press (OUP)
Date: 11-11-2013
Abstract: 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.
Publisher: Wiley
Date: 15-01-1998
DOI: 10.1002/(SICI)1097-0061(19980115)14:1<67::AID-YEA200>3.0.CO;2-T
Publisher: Elsevier BV
Date: 07-2020
Publisher: American Society for Microbiology
Date: 02-2000
DOI: 10.1128/AEM.66.2.744-753.2000
Abstract: The distinctive flavor of wine, brandy, and other grape-derived alcoholic beverages is affected by many compounds, including esters produced during alcoholic fermentation. The characteristic fruity odors of the fermentation bouquet are primarily due to a mixture of hexyl acetate, ethyl caproate (apple-like aroma), iso-amyl acetate (banana-like aroma), ethyl caprylate (apple-like aroma), and 2-phenylethyl acetate (fruity, flowery flavor with a honey note). The objective of this study was to investigate the feasibility of improving the aroma of wine and distillates by overexpressing one of the endogenous yeast genes that controls acetate ester production during fermentation. The synthesis of acetate esters by the wine yeast Saccharomyces cerevisiae during fermentation is ascribed to at least three acetyltransferase activities, namely, alcohol acetyltransferase (AAT), ethanol acetyltransferase, and iso-amyl AAT. To investigate the effect of increased AAT activity on the sensory quality of Chenin blanc wines and distillates from Colombar base wines, we have overexpressed the alcohol acetyltransferase gene ( ATF1 ) of S. cerevisiae . The ATF1 gene, located on chromosome XV, was cloned from a widely used commercial wine yeast strain of S. cerevisiae , VIN13, and placed under the control of the constitutive yeast phosphoglycerate kinase gene ( PGK1 ) promoter and terminator. Chromoblot analysis confirmed the integration of the modified copy of ATF1 into the genome of three commercial wine yeast strains (VIN7, VIN13, and WE228). Northern blot analysis indicated constitutive expression of ATF1 at high levels in these yeast transformants. The levels of ethyl acetate, iso-amyl acetate, and 2-phenylethyl acetate increased 3- to 10-fold, 3.8- to 12-fold, and 2- to 10-fold, respectively, depending on the fermentation temperature, cultivar, and yeast strain used. The concentrations of ethyl caprate, ethyl caprylate, and hexyl acetate only showed minor changes, whereas the acetic acid concentration decreased by more than half. These changes in the wine and distillate composition had a pronounced effect on the solvent or chemical aroma (associated with ethyl acetate and iso-amyl acetate) and the herbaceous and heads-associated aromas of the final distillate and the solvent or chemical and fruity or flowery characters of the Chenin blanc wines. This study establishes the concept that the overexpression of acetyltransferase genes such as ATF1 could profoundly affect the flavor profiles of wines and distillates deficient in aroma, thereby paving the way for the production of products maintaining a fruitier character for longer periods after bottling.
Publisher: Wiley
Date: 15-06-1999
DOI: 10.1002/(SICI)1097-0061(19990615)15:8<647::AID-YEA409>3.0.CO;2-5
Publisher: Springer Science and Business Media LLC
Date: 07-1994
DOI: 10.1007/BF00136479
Publisher: Oxford University Press (OUP)
Date: 23-12-2011
DOI: 10.1111/J.1567-1364.2011.00773.X
Abstract: The vast majority of wine fermentations are performed principally by Saccharomyces cerevisiae. However, there are a growing number of instances in which other species of Saccharomyces play a predominant role. Interestingly, the presence of these other yeast species generally occurs via the formation of interspecific hybrids that contain genomic contributions from both S. cerevisiae and non-S. cerevisiae species. However, despite the large number of wine strains that are characterized at the genomic level, there remains limited information regarding the detailed genomic structure of hybrids used in winemaking. To address this, we describe the genome sequence of the thiol-releasing commercial wine yeast hybrid VIN7. VIN7 is shown to be an almost complete allotriploid interspecific hybrid that is comprised of a heterozygous diploid complement of S. cerevisiae chromosomes and a haploid Saccharomyces kudriavzevii genomic contribution. Both parental strains appear to be of European origin, with the S. cerevisiae parent being closely related to, but distinct from, the commercial wine yeasts QA23 and EC1118. In addition, several instances of chromosomal rearrangement between S. cerevisiae and S. kudriavzevii sequences were observed that may mark the early stages of hybrid genome consolidation.
Publisher: Elsevier BV
Date: 03-1987
DOI: 10.1016/0147-619X(87)90025-4
Abstract: A new procedure was used to isolate 11 plasmids from eight Leuconostoc oenos strains. Plasmid DNA was not detected in 34 other strains of this species. Plasmid sizes ranged from 2.47 to 4.61 kilobase pairs. This is the first report of extrachromosomal elements in L. oenos.
Publisher: Springer Science and Business Media LLC
Date: 11-03-2021
Publisher: Springer Science and Business Media LLC
Date: 19-10-2022
DOI: 10.1038/S41467-022-33974-7
Abstract: Human enterprises through the solar system will entail long-duration voyages and habitation creating challenges in maintaining healthy diets. We discuss consolidating multiple sensory and nutritional attributes into microorganisms to develop customizable food production systems with minimal inputs, physical footprint, and waste. We envisage that a yeast collection bioengineered for one-carbon metabolism, optimal nutrition, and erse textures, tastes, aromas, and colors could serve as a flexible food-production platform. Beyond its potential for supporting humans in space, bioengineered microbial-based food could lead to a new paradigm for Earth’s food manufacturing that provides greater self-sufficiency and removes pressure from natural ecosystems.
Publisher: Springer Science and Business Media LLC
Date: 11-07-2006
DOI: 10.1007/S00294-006-0084-Z
Abstract: In Saccharomyces cerevisiae the Ras/cAMP/PKA signalling pathway controls multiple metabolic pathways, and alterations in the intracellular concentrations of cAMP through modification of signalling pathway factors can be lethal or result in severe growth defects. In this work, the important role of Ras2p in metabolic regulation during growth on the non-fermentable carbon source glycerol is further investigated. The data show that the overexpression of RAS2 suppresses the growth defect of the glyoxylate cycle citrate synthase mutant, cit2Delta. The overexpression results in enhanced proliferation and biomass yield when cells are grown on glycerol as sole carbon source, and increases citrate synthase activity and intracellular citrate concentration. Interestingly, the suppression of cit2Delta and the enhanced proliferation and biomass yield are only observed when RAS2 is overexpressed and not in strains containing the constitutively active allele RAS2(val19). However, both RAS2 and RAS2(val19)upregulated citrate synthase activity. We propose that the RAS2 overexpression results in a combination of general upregulation of respiratory growth capacity and an increase in mitochondrial citrate/citrate synthases, which together, complement the metabolic requirements of the cit2Delta mutant. The data therefore provide new evidence for the role of Ras2p as a powerful modulator of metabolism during growth on a non-fermentable carbon source.
Publisher: Elsevier
Date: 2021
Publisher: MDPI AG
Date: 02-06-2020
DOI: 10.3390/FOODS9060717
Abstract: Speeding up grape must fermentation would be of great economic benefit. We subjected Saccharomyces cerevisiae VIN13 and two recombinant VIN13-strains expressing ATF1 alleles under two different promoters (either PGK1 or HXT7) to four styles of grape must fermentations we then assessed the effect of constantly stirring a must fermentation (isomixing). The four different fermentation setups were as follows: isomixed, closed in an ANKOM Rf Gas productions system isomixed, open in a stirred tall tube cylinder static, closed constituting a conventional fermentation in a wine bottle equipped with an airlock and static and static, open in a tall tube cylinder (without stirring). We report on major fermentation parameters and the volatile aroma compositions generated in the finished wines. The primary fermentations of the strains subjected to constant stirring finished after 7 days, whereas the static fermentations reached dryness after 19 days. The wines derived from isomixed fermentations produced approximately 0.7% less ethanol compared to the unstirred fermentations. The speed that the isomixed fermentation took to reach completion may provide an alternative to static fermentations in the preparation of base wines for sparkling wine production. The observed increase of volatiles of isomixed fermentations merits further investigation.
Publisher: Springer Science and Business Media LLC
Date: 11-02-1998
Abstract: Accurate transcription by RNA polymerase II is usually dependent on the presence of a TATA element, and/or an initiator element, in the promoters of protein-encoding genes. The STA1-3 genes, encoding three glucoamylase isozymes (Sta1p, Sta2p and Sta3p, respectively) responsible for starch hydrolysis in the yeast Saccharomyces cerevisiae, have been shown to contain long and complex promoters with several regulatory regions. These promoters are also virtually identical to the yeast MUC1 gene promoter this gene encodes a mucin-like protein and is evolutionary linked to, and transcriptionally co-regulated with, STA1-3. The STA1-3 genes contain two putative TATA sequences one conforming to the typical TATA box sequence, TATAAA, and another with the sequence of TATAAT. Here we present a study into the functional relevance of these putative TATA sequences and their effects on the transcription of the STA2 gene (as a representative model of the STA1-3 multigene family) and, by analogy, the MUC1 gene. We show that the TATAAA motif is the functional TATA box for STA2 and influences transcript levels, transcript initiation sites, and glucoamylase activities.
Publisher: Springer Science and Business Media LLC
Date: 04-03-2016
Publisher: Wiley
Date: 03-2004
DOI: 10.1002/YEA.1100
Abstract: The yeast alcohol acetyl transferase I, Atf1p, is responsible for the major part of volatile acetate ester production in fermenting Saccharomyces cerevisiae cells. Some of these esters, such as ethyl acetate and isoamyl acetate, are important for the fruity flavours of wine, beer and other fermented beverages. In order to reveal the subcellular localization of Atf1p and further unravel the possible physiological role of this protein, ATF1::GFP fusion constructs were overexpressed in brewer's yeast. The transformant strain showed a significant increase in acetate ester formation, similar to that of an ATF1 overexpression strain, indicating that the Atf1p-GFP fusion protein was active. UV fluorescence microscopy revealed that the fusion protein was localized in small, sphere-like organelles. These organelles could be selectively stained by the fluorescent dye Nile red, indicating that they contained high amounts of neutral lipids and/or sterols, a specific characteristic of yeast lipid particles. Purification of lipid particles from wild type and ATF1 deletion cells confirmed that the Atf1p-GFP fusion protein was located in these organelles. Furthermore, a clear alcohol acetyl transferase activity could be measured in the purified lipid particles of both wild type and transformed cells. The localization of Atf1p in lipid particles may indicate that Atf1p has a specific role in the lipid and/or sterol metabolism that takes place in these particles.
Publisher: Wiley
Date: 13-01-2017
Publisher: MDPI AG
Date: 24-01-2023
DOI: 10.3390/FERMENTATION9020109
Abstract: In this study, the aroma-production profiles of seven different Hanseniaspora strains, namely H. guilliermondii, H. meyeri, H. nectarophila, H. occidentalis, H. opuntiae, H. osmophila and H. uvarum were determined in a simultaneous co-inoculation with the wine yeast Saccharomyces cerevisiae Ch agne Epernay Geisenheim (Uvaferm CEG). All co-inoculated fermentations with Hanseniaspora showed a dramatic increase in ethyl acetate levels except the two (H. occidentalis and H. osmophila) that belong to the so-called slow-evolving clade, which had no meaningful difference, compared to the S. cerevisiae control. Other striking observations were the almost complete depletion of lactic acid in mixed-culture fermentations with H. osmophila, the more than 3.7 mg/L production of isoamyl acetate with H. guilliermondii, the significantly lower levels of glycerol with H. occidentalis and the increase in certain terpenols, such as citronellol with H. opuntiae. This work allows for the direct comparison of wines made with different Hanseniapora spp. showcasing their oenological potential, including two (H. meyeri and H. nectarophila) previously unexplored in winemaking experiments.
Publisher: Springer Berlin Heidelberg
Date: 2006
Publisher: Oxford University Press (OUP)
Date: 2017
Abstract: Biosensors are valuable and versatile tools in synthetic biology that are used to modulate gene expression in response to a wide range of stimuli. Ligand responsive transcription factors are a class of biosensor that can be used to couple intracellular metabolite concentration with gene expression to enable dynamic regulation and high-throughput metabolite producer screening. We have established the Saccharomyces cerevisiae WAR1 transcriptional regulator and PDR12 promoter as an organic acid biosensor that can be used to detect varying levels of para-hydroxybenzoic acid (PHBA) production from the shikimate pathway and output green fluorescent protein (GFP) expression in response. The dynamic range of GFP expression in response to PHBA was dramatically increased by engineering positive-feedback expression of the WAR1 transcriptional regulator from its target PDR12 promoter. In addition, the noise in GFP expression at the population-level was controlled by normalising GFP fluorescence to constitutively expressed mCherry fluorescence within each cell. These biosensor modifications increased the high-throughput screening efficiency of yeast cells engineered to produce PHBA by 5,000-fold, enabling accurate fluorescence activated cell sorting isolation of producer cells that were mixed at a ratio of 1 in 10,000 with non-producers. Positive-feedback, ratiometric transcriptional regulator expression is likely applicable to many other transcription-factor romoter pairs used in synthetic biology and metabolic engineering for both dynamic regulation and high-throughput screening applications.
Publisher: Springer Science and Business Media LLC
Date: 2003
Abstract: The Saccharomyces cerevisiae chitinase, encoded by the CTS1-2 gene has recently been confirmed by in vitro tests to possess antifungal abilities. In this study, the CTS1-2 gene has been evaluated for its in planta antifungal activity by constitutive overexpression in tobacco plants to assess its potential to increase the plant's defence against fungal pathogens. Transgenic tobacco plants, generated by Agrobacterium-mediated transformation, showed stable integration and inheritance of the transgene. Northern blot analyses conducted on the transgenic tobacco plants confirmed transgene expression. Leaf extracts from the transgenic lines inhibited Botrytis cinerea spore germination and hyphal growth by up to 70% in a quantitative in vitro assay, leading to severe physical damage on the hyphae. Several of the F1 progeny lines were challenged with the fungal pathogen, B. cinerea, in a detached leaf infection assay, showing a decrease in susceptibility ranging from 50 to 70%. The plant lines that showed increased disease tolerance were also shown to have higher chitinase activities.
Publisher: Oxford University Press (OUP)
Date: 10-2002
DOI: 10.1046/J.1472-765X.2002.01181.X
Abstract: The identification, differentiation and characterization of indigenous Saccharomyces sensu stricto strains isolated from Croatian vineyards and the evaluation of their oenological potential. A total of 47 Saccharomyces sensu stricto strains were isolated from Chardonnay grapes and identified by physiological and molecular genetic methods. By using the standard physiological and biochemical tests, six isolates were identified as Saccharomyces cerevisiae and 41 as Saccharomyces paradoxus. However, PCR-RFLP analyses of the internal transcribed spacer (ITS1) region of the 18S ribosomal DNA identified 12 of the isolates as S.cerevisiae and 35 as S. paradoxus. Fermentation trials in a grape juice medium showed that these isolates ferment vigorously at 18 degrees C and display tolerance to high levels of ethanol. None of these isolates appeared to produce either hydrogen sulphide or killer toxins. Saccharomyces paradoxus, possessing potentially important oenological characteristics, occurs in much higher numbers than S. cerevisiae in the indigenous population of Saccharomyces sensu stricto strains in Croatian vineyards. This study forms an essential step towards the preservation and exploitation of the hidden oenological potential of the untapped wealth of yeast bio ersity in the Croatian grape-growing regions. The results obtained demonstrate the value of using molecular genetic methods, such as PCR-RFLP analyses, in conjunction with the traditional taxonomic methods based on phenotypic characteristics in such ecotaxonomic surveys. The results also shed some light on the ecology and oenological potential of S.paradoxus, which is considered to be the natural parent species of the domesticated species of the Saccharomyces sensu stricto group.
Publisher: Oxford University Press (OUP)
Date: 02-2013
Abstract: Acetic acid, a byproduct formed during yeast alcoholic fermentation, is the main component of volatile acidity (VA). When present in high concentrations in wine, acetic acid imparts an undesirable 'vinegary' character that results in a significant reduction in quality and sales. Previously, it has been shown that saké yeast strains resistant to the antifungal cerulenin produce significantly lower levels of VA. In this study, we used a classical mutagenesis method to isolate a series of cerulenin-resistant strains, derived from a commercial diploid wine yeast. Four of the selected strains showed a consistent low-VA production phenotype after small-scale fermentation of different white and red grape musts. Specific mutations in YAP1, a gene encoding a transcription factor required for oxidative stress tolerance, were found in three of the four low-VA strains. When integrated into the genome of a haploid wine strain, the mutated YAP1 alleles partially reproduced the low-VA production phenotype of the diploid cerulenin-resistant strains, suggesting that YAP1 might play a role in (regulating) acetic acid production during fermentation. This study offers prospects for the development of low-VA wine yeast starter strains that could assist winemakers in their effort to consistently produce wine to definable quality specifications.
Publisher: Elsevier BV
Date: 10-2006
DOI: 10.1016/J.JBIOTEC.2006.03.029
Abstract: Commercial polysaccharase preparations are applied to winemaking to improve wine processing and quality. Expression of polysaccharase-encoding genes in Saccharomyces cerevisiae allows for the recombinant strains to degrade polysaccharides that traditional commercial yeast strains cannot. In this study, we constructed recombinant wine yeast strains that were able to degrade the problem-causing grape polysaccharides, glucan and xylan, by separately integrating the Trichoderma reesei XYN2 xylanase gene construct and the Butyrivibrio fibrisolvens END1 glucanase gene cassette into the genome of the commercial wine yeast strain S. cerevisiae VIN13. These genes were also combined in S. cerevisiae VIN13 under the control of different promoters. The strains that were constructed were compared under winemaking conditions with each other and with a recombinant wine yeast strain expressing the endo-beta-1,4-glucanase gene cassette (END1) from B. fibrisolvens and the endo-beta-1,4-xylanase gene cassette (XYN4) from Aspergillus niger, a recombinant strain expressing the pectate lyase gene cassette (PEL5) from Erwinia chrysanthemi and the polygalacturonase-encoding gene cassette (PEH1) from Erwinia carotovora. Wine was made with the recombinant strains using different grape cultivars. Fermentations with the recombinant VIN13 strains resulted in significant increases in free-flow wine when Ruby Cabernet must was fermented. After 6 months of bottle ageing significant differences in colour intensity and colour stability could be detected in Pinot Noir and Ruby Cabernet wines fermented with different recombinant strains. After this period the volatile composition of Muscat d'Alexandria, Ruby Cabernet and Pinot Noir wines fermented with different recombinant strains also showed significant differences. The Pinot Noir wines were also sensorial evaluated and the tasting panel preferred the wines fermented with the recombinant strains.
Publisher: Springer New York
Date: 2019
Publisher: American Society for Microbiology
Date: 12-2001
DOI: 10.1128/AEM.67.12.5512-5519.2001
Abstract: The β-xylosidase-encoding xlnD gene of Aspergillus niger 90196 was lified by the PCR technique from first-strand cDNA synthesized on mRNA isolated from the fungus. The nucleotide sequence of the cDNA fragment was verified to contain a 2,412-bp open reading frame that encodes a 804-amino-acid propeptide. The 778-amino-acid mature protein, with a putative molecular mass of 85.1 kDa, was fused in frame with the Saccharomyces cerevisiae mating factor α1 signal peptide (MFα1 s ) to ensure correct posttranslational processing in yeast. The fusion protein was designated Xlo2. The recombinant β-xylosidase showed optimum activity at 60°C and pH 3.2 and optimum stability at 50°C. The K i (app) value for d -xylose and xylobiose for the recombinant β-xylosidase was determined to be 8.33 and 6.41 mM, respectively. The XLO2 fusion gene and the XYN2 β-xylanase gene from Trichoderma reesei , located on URA3 -based multicopy shuttle vectors, were successfully expressed and coexpressed in the yeast Saccharomyces cerevisiae under the control of the alcohol dehydrogenase II gene ( ADH2 ) promoter and terminator. These recombinant S. cerevisiae strains produced 1,577 nkat/ml of β-xylanase activity when expressing only the β-xylanase and 860 nkat/ml when coexpressing the β-xylanase with the β-xylosidase. The maximum β-xylosidase activity was 5.3 nkat/ml when expressed on its own and 3.5 nkat/ml when coexpressed with the β-xylanase. Coproduction of the β-xylanase and β-xylosidase enabled S. cerevisiae to degrade birchwood xylan to d -xylose.
Publisher: Springer Science and Business Media LLC
Date: 03-11-2020
DOI: 10.1186/S13068-020-01823-8
Abstract: For the economic production of biofuels and other valuable products from lignocellulosic waste material, a consolidated bioprocessing (CBP) organism is required. With efficient fermentation capability and attractive industrial qualities, Saccharomyces cerevisiae is a preferred candidate and has been engineered to produce enzymes that hydrolyze cellulosic biomass. Efficient cellulose hydrolysis requires the synergistic action of several enzymes, with the optimum combined activity ratio dependent on the composition of the substrate. In vitro SCRaMbLE generated a library of plasmids containing different ratios of a β-glucosidase gene ( CEL3A ) from Saccharomycopsis fibuligera and an endoglucanase gene ( CEL5A ) from Trichoderma reesei . S. cerevisiae , transformed with the plasmid library, displayed a range of in idual enzyme activities and synergistic capabilities. Furthermore, we show for the first time that 4,6-O-(3-ketobutylidene)-4-nitrophenyl-β- d -cellopentaoside (BPNPG5) is a suitable substrate to determine synergistic Cel3A and Cel5A action and an accurate predictive model for this synergistic action was devised. Strains with highest BPNPG5 activity had an average CEL3A and CEL5A gene cassette copy number of 1.3 ± 0.6 and 0.8 ± 0.2, respectively (ratio of 1.6:1). Here, we describe a synthetic biology approach to rapidly optimise gene copy numbers to achieve efficient synergistic substrate hydrolysis. This study demonstrates how in vitro SCRaMbLE can be applied to rapidly combine gene constructs in various ratios to allow screening of synergistic enzyme activities for efficient substrate hydrolysis.
Publisher: Elsevier BV
Date: 11-2009
Publisher: Springer Science and Business Media LLC
Date: 25-03-2006
DOI: 10.1007/S00294-006-0068-Z
Abstract: The ability of many microorganisms to modify adhesion-related properties of their cell surface is of importance for many processes, including substrate adhesion, cell-cell adhesion, invasive growth, pathogenic behaviour and biofilm formation. In the yeast Saccharomyces cerevisiae, a group of structurally related, cell-wall associated proteins encoded by the FLO gene family are directly responsible for many of the cellular adhesion phenotypes displayed by this organism. Previous research has suggested that the differential transcription of FLO genes determines specific adhesion phenotypes. However, the transcriptional regulation of most FLO genes remains poorly understood. Here we show that the transcriptional activator Mss11p, which has previously been shown to be involved in the regulation of starch degradation, the formation of pseudohyphae and haploid invasive growth, also acts as a strong inducer of flocculation. The data indicate that Mss11p induces flocculation together with Flo8p, and that FLO1 is the dominant target gene of the two factors in this process. The deletion of MSS11 leads to a non-flocculent phenotype, and specific domains of Mss11p that are critical for the induction of flocculation are identified. The data clearly show that several essential transcription factors are shared by at least two flocculation genes that control different adhesion phenotypes.
Publisher: Informa UK Limited
Date: 18-08-2016
DOI: 10.1080/07388551.2016.1214945
Abstract: Over the past 15 years, the seismic shifts caused by the convergence of biomolecular, chemical, physical, mathematical, and computational sciences alongside cutting-edge developments in information technology and engineering have erupted into a new field of scientific endeavor dubbed Synthetic Biology. Recent rapid advances in high-throughput DNA sequencing and DNA synthesis techniques are enabling the design and construction of new biological parts (genes), devices (gene networks) and modules (biosynthetic pathways), and the redesign of biological systems (cells and organisms) for useful purposes. In 2014, the budding yeast Saccharomyces cerevisiae became the first eukaryotic cell to be equipped with a fully functional synthetic chromosome. This was achieved following the synthesis of the first viral (poliovirus in 2002 and bacteriophage Phi-X174 in 2003) and bacterial (Mycoplasma genitalium in 2008 and Mycoplasma mycoides in 2010) genomes, and less than two decades after revealing the full genome sequence of a laboratory (S288c in 1996) and wine (AWRI1631 in 2008) yeast strain. A large international project - the Synthetic Yeast Genome (Sc2.0) Project - is now underway to synthesize all 16 chromosomes (∼12 Mb carrying ∼6000 genes) of the sequenced S288c laboratory strain by 2018. If successful, S. cerevisiae will become the first eukaryote to cross the horizon of in silico design of complex cells through de novo synthesis, reshuffling, and editing of genomes. In the meantime, yeasts are being used as cell factories for the semi-synthetic production of high-value compounds, such as the potent antimalarial artemisinin, and food ingredients, such as resveratrol, vanillin, stevia, nootkatone, and saffron. As a continuum of previously genetically engineered industrially important yeast strains, precision genome engineering is bound to also impact the study and development of wine yeast strains supercharged with synthetic DNA. The first taste of what the future holds is the de novo production of the raspberry ketone aroma compound, 4-[4-hydroxyphenyl]butan-2-one, in a wine yeast strain (AWRI1631), which was recently achieved via metabolic pathway engineering and synthetic enzyme fusion. A peek over the horizon is revealing that the future of "Wine Yeast 2.0" is already here. Therefore, this article seeks to help prepare the wine industry - an industry rich in history and tradition on the one hand, and innovation on the other - for the inevitable intersection of the ancient art practiced by winemakers and the inventive science of pioneering "synthetic genomicists". It would be prudent to proactively engage all stakeholders - researchers, industry practitioners, policymakers, regulators, commentators, and consumers - in a meaningful dialog about the potential challenges and opportunities emanating from Synthetic Biology. To capitalize on the new vistas of synthetic yeast genomics, this paper presents wine yeast research in a fresh context, raises important questions and proposes new directions.
Publisher: Wiley
Date: 04-1986
DOI: 10.1111/J.1365-2672.1986.TB01743.X
Abstract: A total of 134 alpha-amylase producing Bacillus isolates and 21 reference strains were ided into 12 groups according to their similarities (% SSM). Phenotypic characteristics determined by the API 20E and API 50CHB galleries, other biochemical tests and morphological characteristics were used for the numerical analysis. The API Computer Service identified 45% of the isolates. The amylase yields of 16 alpha-amylase hyperproducing (AHP) isolates were compared with those of seven amylolytic reference and type strains. The AHP isolates were related to Bacillus subtilis, B. licheniformis and 'B. amyloliquefaciens'.
Publisher: Elsevier BV
Date: 12-2002
Publisher: MDPI AG
Date: 06-11-2020
DOI: 10.20944/PREPRINTS202011.0237.V1
Abstract: Genome-scale engineering and custom synthetic genomes are reshaping the next generation of industrial yeast strains. The Cre-recombinase mediated chromosomal rearrangement mechanism of designer synthetic Saccharomyces cerevisiae chromosomes, known as SCRaMbLE, is a powerful tool which allows rapid genome evolution upon command. This system is able to generate millions of novel genomes with potential valuable phenotypes, but the excessive loss of essential genes often results in poor growth or even the death of cells with useful phenotypes. In this study we expanded the versatility of SCRaMbLE to industrial strains, and evaluated different control measures to optimise genomic rearrangement, whilst limiting cell death. To achieve this, we have developed RED (Rapid Evolution Detection), a simple colorimetric plate-assay procedure to rapidly quantify the degree of genomic rearrangements within a post-SCRaMbLE yeast population. RED-enabled semi-synthetic strains were mated with haploid progeny of industrial yeast strains to produce stress tolerant heterozygous diploid strains. Analysis of these heterozygous strains with the RED-assay, genome sequencing and custom bioinformatics scripts demonstrated a correlation between RED-assay frequencies and physical genomic rearrangements. Here we show that RED is a fast and effective method to evaluate optimal SCRaMbLE induction times of different Cre-recombinse expression systems for the development of industrial strains.
Publisher: American Chemical Society (ACS)
Date: 14-06-2021
DOI: 10.1021/ACSSYNBIO.0C00633
Abstract: The unfolded protein response (UPR) is a highly conserved cellular response in eukaryotic cells to counteract endoplasmic reticulum (ER) stress, typically triggered by unfolded protein accumulation. In addition to its relevance to human diseases like cancer, the induction of the UPR has a significant impact on the recombinant protein production in eukaryotic cell factories, including the industrial workhorse
Publisher: Springer Science and Business Media LLC
Date: 23-08-1996
Abstract: A cDNA fragment encoding the Phanerochaete chrysosporium cellobiohydrolase (cbh1-4) was lified and cloned with the aid of the polymerase chain reaction (PCR) technique. The cbh1-4 gene and the Butyrivibrio fibrisolvens endo-beta-1,4-glucanase (end1) gene were successfully expressed in Saccharomyces cerevisiae under the control of the phosphoglycerate kinase-I (PGK1) and alcohol dehydrogenase-II (ADH2) gene promoters and terminators, respectively. The native P. chrysosporium signal sequence mediated secretion of cellobiohydrolase in S. cerevisiae, whereas secretion of the endo-beta-1,4-glucanase was directed by the signal sequence of the yeast mating pheromone alpha-factor (MFalpha1S). These constructs, designated CBH1 and END1, respectively, were expressed separately and jointly in S. cerevisiae. The construction of fur1 ura3 S. cerevisiae strains allowed for the autoselection of these multicopy URA3-based plasmids in rich medium. Enzyme assays confirmed that co-expression of CBH1 and END1 synergistically enhanced cellulose degradation by S. cerevisiae.
Publisher: Elsevier BV
Date: 10-2010
Publisher: MDPI AG
Date: 06-07-2018
DOI: 10.3390/GENES9070340
Publisher: Elsevier
Date: 2006
Publisher: Springer Science and Business Media LLC
Date: 03-1995
DOI: 10.1007/BF00191185
Publisher: Springer Science and Business Media LLC
Date: 05-1992
DOI: 10.1007/BF00713932
Publisher: Elsevier
Date: 2015
Publisher: Springer Science and Business Media LLC
Date: 24-06-2022
DOI: 10.1038/S41467-022-31305-4
Abstract: The Synthetic Yeast Genome Project ( Sc 2.0) represents the first foray into eukaryotic genome engineering and a framework for designing and building the next generation of industrial microbes. However, the laboratory strain S288c used lacks many of the genes that provide phenotypic ersity to industrial and environmental isolates. To address this shortcoming, we have designed and constructed a neo-chromosome that contains many of these erse pan-genomic elements and which is compatible with the Sc 2.0 design and test framework. The presence of this neo-chromosome provides phenotypic plasticity to the Sc 2.0 parent strain, including expanding the range of utilizable carbon sources. We also demonstrate that the induction of programmable structural variation (SCRaMbLE) provides genetic ersity on which further adaptive gains could be selected. The presence of this neo-chromosome within the Sc 2.0 backbone may therefore provide the means to adapt synthetic strains to a wider variety of environments, a process which will be vital to transitioning Sc 2.0 from the laboratory into industrial applications.
Publisher: Elsevier BV
Date: 08-2007
DOI: 10.1016/J.TIBTECH.2007.05.006
Abstract: Yeast research represents an important nexus between fundamental and applied research. Just as fundamental yeast research transitioned from classical, reductionist strategies to whole-genome techniques, whole-genome studies are advancing to the next level of biological research, referred to as systems biology. Industries that rely on high-performing yeast, such as the wine industry, are therefore poised to reap the many benefits that systems biology can provide. This includes the promise of strain development at speeds and costs which are unobtainable using current techniques. This article reviews the current state of whole-genome techniques available to yeast researchers and outlines how these processes can be used to obtain 'systems-level' information to provide insights into winemaking.
Publisher: Springer Science and Business Media LLC
Date: 1998
Publisher: Oxford University Press (OUP)
Date: 2023
Abstract: Wine is composed of multitudinous flavour components and volatile organic compounds that provide this beverage with its attractive properties of taste and aroma. The perceived quality of a wine can be attributed to the absolute and relative concentrations of favourable aroma compounds hence, increasing the detectable levels of an attractive aroma, such as β-ionone with its violet and berry notes, can improve the organoleptic qualities of given wine styles. We here describe the generation of a new grape-must fermenting strain of Saccharomyces cerevisiae that is capable of releasing β-ionone through the heterologous expression of both the enzyme carotenoid cleavage dioxygenase 1 (CCD1) and its substrate, β-carotene. Haploid laboratory strains of S. cerevisiae were constructed with and without integrated carotenogenic genes and transformed with a plasmid containing the genes of CCD1. These strains were then mated with a sporulated diploid wine industry yeast, VIN13, and four resultant crosses—designated MQ01–MQ04–which were capable of fermenting the must to dryness were compared for their ability to release β-ionone. Analyses of their fermentation products showed that the MQ01 strain produced a high level of β-ionone and offers a fermenting hybrid yeast with the potential to enhance the organoleptic qualities of wine.
Publisher: Springer Science and Business Media LLC
Date: 2002
Publisher: Springer Science and Business Media LLC
Date: 2000
Abstract: A mesophilic wine yeast, Saccharomyces cerevisiae CSIR Y217 K-R- was subjected to the K2 killer effect of Saccharomyces cerevisiae T206 K+R+ in a liquid grape medium. The lethal effect of the K2 mycoviral toxin was confirmed by methylene blue staining. Scanning electron microscopy of cells from challenge experiments revealed rippled cell surfaces, accompanied by cracks and pores, while those unaffected by the toxin, as in the control experiments, showed a smooth surface. Transmission electron microscopy revealed that the toxin damaged the cell wall structure and perturbed cytoplasmic membranes to a limited extent.
Publisher: Public Library of Science (PLoS)
Date: 28-03-2012
Publisher: Springer Science and Business Media LLC
Date: 16-10-2007
DOI: 10.1007/S00253-007-1194-3
Abstract: The yeast Saccharomyces cerevisiae synthesises a variety of volatile aroma compounds during wine fermentation. In this study, the influence of fermentation temperature on (1) the production of yeast-derived aroma compounds and (2) the expression of genes involved in aroma compounds' metabolism (ADH1, PDC1, BAT1, BAT2, LEU2, ILV2, ATF1, ATF2, EHT1 and IAH1) was assessed, during the fermentation of a defined must at 15 and 28 degrees C. Higher concentrations of compounds related to fresh and fruity aromas were found at 15 degrees C, while higher concentrations of flowery related aroma compounds were found at 28 degrees C. The formation rates of volatile aroma compounds varied according to growth stage. In addition, linear correlations between the increases in concentration of higher alcohol and their corresponding acetates were obtained. Genes presented different expression profiles at both temperatures, except ILV2, and those involved in common pathways were co-expressed (ADH1, PDC1 and BAT2 and ATF1, EHT1 and IAH1). These results demonstrate that the fermentation temperature plays an important role in the wine final aroma profile, and is therefore an important control parameter to fine-tune wine quality during winemaking.
Publisher: Springer Science and Business Media LLC
Date: 1995
DOI: 10.1007/BF00518165
Abstract: Bisphenol A is a commercially important chemical used to make polycarbonate plastic, epoxy resins, and other specialty products. Despite an extensive body of in vitro, animal and human observational studies on the effects of exposure to bisphenol A, no authoritative bodies in the U.S. have adopted or recommended occupational exposure limits for bisphenol A. In 2017, the National Institute for Occupational Safety and Health published a Draft process for assigning health-protective occupational exposure bands, i.e., an airborne concentration range, to chemicals lacking an occupational exposure limit. Occupational exposure banding is a systematic process that uses both quantitative and qualitative toxicity information on selected health effect endpoints to assign an occupational exposure band for a chemical. The Draft process proposes three methodological tiers of increasing complexity for assigning an occupational exposure band. We applied Tier 1 (based on the Globally Harmonized System of Classification and Labelling) and Tier 2 (based on authoritative sources/reviews) to assign an occupational exposure band to bisphenol A. Under both Tier 1 and 2, the occupational exposure band for bisphenol A was "E" (<0.01 mg/m
Publisher: Springer Science and Business Media LLC
Date: 14-04-2022
Publisher: Oxford University Press (OUP)
Date: 25-03-2015
DOI: 10.1093/BRAIN/AWV071
Publisher: Springer Science and Business Media LLC
Date: 2008
Publisher: Springer Science and Business Media LLC
Date: 12-1994
DOI: 10.1007/BF00326573
Publisher: MDPI AG
Date: 28-09-2020
DOI: 10.3390/IJMS21197156
Abstract: Some years inspire more hindsight reflection and future-gazing than others. This is even more so in 2020 with its evocation of perfect vision and the landmark ring to it. However, no futurist can reliably predict what the world will look like the next time that a year’s first two digits will match the second two digits—a numerical pattern that only occurs once in a century. As we leap into a new decade, amid uncertainties triggered by unforeseen global events—such as the outbreak of a worldwide pandemic, the accompanying economic hardship, and intensifying geopolitical tensions—it is important to note the blistering pace of 21st century technological developments indicate that while hindsight might be 20/20, foresight is 50/50. The history of science shows us that imaginative ideas, research excellence, and collaborative innovation can, for ex le, significantly contribute to the economic, cultural, social, and environmental recovery of a post-COVID-19 world. This article reflects on a history of yeast research to indicate the potential that arises from advances in science, and how this can contribute to the ongoing recovery and development of human society. Future breakthroughs in synthetic genomics are likely to unlock new avenues of impactful discoveries and solutions to some of the world’s greatest challenges.
Publisher: Wiley
Date: 2001
DOI: 10.1002/YEA.712
Abstract: L-carnitine is required for the transfer of activated acyl-groups across intracellular membranes in eukaryotic organisms. In Saccharomyces cerevisiae, peroxisomal membranes are impermeable to acetyl-CoA, which is produced in the peroxisome when cells are grown on fatty acids as carbon source. In a reversible reaction catalysed by carnitine acetyltransferases (CATs), activated acetyl groups are transferred to carnitine to form acetylcarnitine which can be shuttled across membranes. Here we describe a mutant selection strategy that specifically selects for mutants affected in carnitine-dependent metabolic activities. Complementation of three of these mutants resulted in the cloning of three CAT encoding genes: CAT2, coding for the carnitine acetyltransferase associated with the peroxisomes and the mitochondria YAT1, coding for the carnitine acetyltransferase, which is presumably associated with the outer mitochondrial membrane, and YER024w (YAT2), which encodes a third, previously unidentified carnitine acetyltransferase. The data also show that (a) L-carnitine and all three CATs are essential for growth on non-fermentable carbon sources in a strain with a disrupted CIT2 gene (b) Yat2p contributes significantly to total CAT activity when cells are grown on ethanol and that (c) the carnitine-dependent transfer of activated acetyl groups plays a more important role in cellular processes than previously realised.
Publisher: Springer Science and Business Media LLC
Date: 11-1993
DOI: 10.1007/BF00351848
Abstract: Viral infections are one of the leading causes of human illness. Viruses take over host cell signaling cascades for their replication and infection. Calcium (Ca
Publisher: Springer Science and Business Media LLC
Date: 11-09-2007
DOI: 10.1007/S00253-007-1145-Z
Abstract: Surveys conducted worldwide have shown that a significant proportion of grape musts are suboptimal for yeast nutrients, especially assimilable nitrogen. Nitrogen deficiencies are linked to slow and stuck fermentations and sulphidic off-flavour formation. Nitrogen supplementation of grape musts has become common practice however, almost no information is available on the effects of nitrogen supplementation on wine flavour. In this study, the effect of ammonium supplementation of a synthetic medium over a wide range of nitrogen values on the production of volatile and non-volatile compounds by two high-nitrogen-demand wine fermentation strains of Saccharomyces cerevisiae was determined. To facilitate this investigation, a simplified chemically defined medium that resembles the nutrient composition of grape juice was used. Analysis of variance revealed that ammonium supplementation had significant effects on the concentration of residual sugar, L-malic acid, acetic acid and glycerol but not the ethanol concentration. While choice of yeast strain significantly affected half of the aroma compounds measured, nitrogen concentrations affected 23 compounds, including medium-chain alcohols and fatty acids and their esters. Principal component analysis showed that branched-chain fatty acids and their esters were associated with low nitrogen concentrations, whereas medium-chain fatty esters and acetic acid were associated with high nitrogen concentrations.
Publisher: Elsevier BV
Date: 12-1995
DOI: 10.1016/0378-1119(95)00633-0
Abstract: The yeast Lipomyces kononenkoae (Lk) secretes a highly active raw starch-degrading alpha-amylase (alpha Amy) that liberates reducing groups from glucose polymers containing both alpha-1,4 and alpha-1,6 bonds. The LKA1 gene encoding this industrially important alpha Amy was cloned as a 2261-bp cDNA fragment from a glucose-derepressed mutant (IGC4052B) of Lk and characterized. The nucleotide (nt) sequence of the cDNA fragment was determined, revealing an open reading frame of 1872 bp, encoding a 596 amino-acid (aa) mature protein (LKA1) with a calculated M(r) of 65,706. The similarity between the aa sequence of LKA1 and those of other alpha Amy showed four common conserved regions characteristic of the alpha Amy protein family: (A) 264DIVVNH269, (B) 349GLRIDTVKH357, (B') 376GEVFD380 and (C) 439FLENQD444. The deduced aa sequence revealed significant homology to the aa sequences of the Aspergillus oryzae, Schwanniomyces occidentalis and Saccharomycopsis fibuligera alpha Amy, various bacterial cyclomaltodextrin glucanotransferases, a beta-amylase and the 5'-region of a glucoamylase. LKA1 was expressed in Saccharomyces cerevisiae (Sc) under the control of the phosphoglycerate kinase (PGK1) promoter and Northern blot analysis showed the presence of a single 2.3-kb transcript. The 28-aa signal peptide of the LKA1 protein efficiently directed its secretion into the medium when expressed in Sc.
Publisher: Informa UK Limited
Date: 23-02-2023
Publisher: Elsevier BV
Date: 03-2022
DOI: 10.1016/J.IJFOODMICRO.2022.109549
Abstract: Certain yeast species belonging to the Pichia genus are known to form a distinctive film on grape must and wine. In a mixed-culture type fermentation, Pichia spp. (P. kluyveri in particular) are known to impart beneficial oenological attributes. In this study, we report on an easy isolation method of Pichia spp. from grape must by exploiting their film-forming capacity on media containing 10% ethanol. We isolated and identified two Pichia species, namely Pichia kudriavzevii and Pichia kluyveri, and subsequently co-inoculated them with Saccharomyces cerevisiae to ferment Gewürztraminer musts. Noteworthy differences included a significant increase in the 2-phenethyl acetate levels with the P. kluyveri co-fermentation and a general increase in ethyl esters with the P. kudriavzevii co-fermentation. Both Pichia co-inoculations yielded higher levels of glycerol in the final wines. Based on all the wine parameters we tested, the P. kluyveri strain that was isolated performed similarly to a commercial P. kluyveri strain.
Publisher: Elsevier BV
Date: 15-10-2010
DOI: 10.1016/J.IJFOODMICRO.2010.08.004
Abstract: The objective of this study was to evaluate different infrared spectroscopy methods in combination with chemometrics for the differentiation between Brettanomyces bruxellensis strains. These methods of discrimination were applied to intact yeast cells of B. bruxellensis strains and on wines spoiled by the same strains. Eleven wine isolates of B. bruxellensis were evaluated for volatile phenol production in red wine and their genetic ersity was determined by Restriction Endonuclease Analysis-Pulsed Field Gel Electrophoresis (REA-PFGE). Fourier transform mid-infrared (FTMIR) spectroscopy was used to obtain spectral fingerprints of the spoiled wines. Attenuated total reflectance (ATR) was used to obtain spectral fingerprints from the intact cells of the 11 B. bruxellensis strains. The groupings from the genetic fingerprints obtained with REA-PFGE were used as reference firstly for comparison with the groupings observed with the FTMIR spectral fingerprint of the wines and secondly for the FTIR-ATR spectral fingerprints from the whole cells. Results indicated that ATR-IR spectra obtained by scanning whole cells of B. bruxellensis could be useful for rapid strain typing in comparison or complementary to molecular techniques and FTMIR spectra from wines provide a useful resource for the discrimination between B. bruxellensis contaminated wines.
Publisher: Springer Science and Business Media LLC
Date: 04-11-2020
DOI: 10.1038/S41467-020-19390-9
Abstract: Utilising one-carbon substrates such as carbon dioxide, methane, and methanol is vital to address the current climate crisis. Methylotrophic metabolism enables growth and energy generation from methanol, providing an alternative to sugar fermentation. Saccharomyces cerevisiae is an important industrial microorganism for which growth on one-carbon substrates would be relevant. However, its ability to metabolize methanol has been poorly characterised. Here, using adaptive laboratory evolution and 13 C-tracer analysis, we discover that S. cerevisiae has a native capacity for methylotrophy. A systems biology approach reveals that global rearrangements in central carbon metabolism fluxes, gene expression changes, and a truncation of the uncharacterized transcriptional regulator Ygr067cp supports improved methylotrophy in laboratory evolved S. cerevisiae . This research paves the way for further biotechnological development and fundamental understanding of methylotrophy in the preeminent eukaryotic model organism and industrial workhorse, S. cerevisiae .
Publisher: Elsevier BV
Date: 04-2013
DOI: 10.1016/J.COPBIO.2012.08.006
Abstract: The application of Next Generation sequencing to comparative genomics is enabling in-depth characterization of genetic variation between wine yeast strains used in fermentation starter cultures. Knowledge from this work will be harnessed in strain development programs. As a result, winemakers will soon have at their disposal novel, improved yeast starter cultures displaying increased reliability and providing a means of tailoring wine sensory characteristics for new and ever-changing markets.
Publisher: Springer Science and Business Media LLC
Date: 04-1986
DOI: 10.1007/BF00330381
Abstract: Patients with hepatitis C viral (HCV) may perceive barriers to accessing speciality care for HCV, and these barriers may be related to depressive symptoms. To evaluate the relationship between barriers to care, demographics, and depressive symptoms. A cross-sectional analysis of 126 patients referred for HCV at two speciality HCV clinics. Barriers to care, depressive symptoms and sociodemographics were measured using standardized instruments. A retrospective chart review was conducted to collect clinical outcome data. Depressive symptoms were reported in 26%. Common barriers included lack of personal financial resources lack of HCV knowledge in the community lack of professionals competent in HCV care stigmatization of HCV and long distances to clinics offering care. After we controlled for sociodemographics, depression accounted for an additional 7-18% of variability in all barriers (all p values <0.01). Lower depression, marital and employment status were associated with subsequent receipt of HCV treatment in 38% (45/120) of patients perceived barriers were not. Depression is independently associated with perceived barriers to care. Higher depressive scores, but not perceived barriers, were associated with nontreatment. Healthcare providers who diagnose HCV need to be cognizant of numerous perceived barriers to accessing HCV care, and the impact that depression may have on these perceptions and receipt of treatment.
Publisher: Springer Science and Business Media LLC
Date: 04-1986
DOI: 10.1007/BF00330380
Abstract: We initiated a study at Baylor Jack and Jane Hamilton Heart and Vascular Hospital to compare the sliding scale insulin (SSI) protocol used in 2006 with the SSI protocol currently used to treat diabetic patients admitted for procedures or surgery. An audit of patients' records revealed greater variation in staff compliance with the current protocol than with the previous one. In addition, it seemed that more patients were refusing insulin coverage under the current protocol than under the prior version. Although the study was aborted, the initial findings motivated us to identify obstacles to glucose control and to launch a health care improvement initiative to increase compliance with the SSI protocol. As a result of this process, the hospital has made several changes, including re-educating staff nurses, initiating competency checks of protocol interpretation, promoting patient education, and implementing early identification of inconsistent glucose control.
Publisher: Elsevier BV
Date: 09-1994
DOI: 10.1016/0378-1119(94)90286-0
Abstract: Expression of the glucoamylase-encoding gene (STA2) in Saccharomyces cerevisiae was previously shown to be regulated transcriptionally by both positive and negative factors. The objective of this work was to identify the cis-acting elements responsible for STA2 transcriptional activation as well as the transcriptional repressor effects of STA10 and MATa/MAT alpha. We identified two upstream activation regions (UAS). Three repressor regions responsive to STA10-mediated repression were identified, as well as two regions for down-regulation of STA2 expression. MATa/MAT alpha repression appears to effect STA2 expression either downstream from the translational start site or, indirectly, since no functional a1/alpha 2-responsive sequence was identified in the promoter region.
Publisher: Wiley
Date: 08-1996
DOI: 10.1002/(SICI)1097-0061(199608)12:10<925::AID-YEA987>3.0.CO;2-0
Publisher: Springer Netherlands
Date: 2009
Publisher: Springer Science and Business Media LLC
Date: 02-1999
Abstract: A scanning ribosome will usually initiate translation as soon as it encounters the first favourable AUG codon and only a few eukaryotic transcripts have more complex arrangements. These relatively few complex transcripts are normally characterized by structural features such as multiple AUGs and significant secondary structure. However, the functional relevance of these features has rarely been established. We present here a study of the functional significance of the multiple AUGs in the leader of STA2 transcripts of the budding yeast Saccharomyces cerevisiae, and extrapolate, where applicable, these results to a co-regulated gene, MUC1. The STA2 gene (a representative member of the polymorphic STA1-3 gene family), encodes an extracellular glucoamylase, and is evolutionarily linked to, and transcriptionally co-regulated with, the MUC1 gene, which encodes a mucin-like protein essential for pseudohyphal/invasive growth and cell-adhesion in S. cerevisiae. Each of these genes contains a putative upstream ORF, while STA2 has two additional in-frame AUG codons 5' to the major cistron. We show that utilization of the alternative translational start-sites of STA2 results in glucoamylases that differ at their N-termini, which are associated with differences in their localization patterns. Analysis of mutants revealed the presence of a putative secretion-enhancing signal that might prove to be relevant to the alternative targeting mechanism recently uncovered in S. cerevisiae. We show that a short up-stream ORF present in the leaders of STA1-3 and MUC1 is probably bypassed by a process of leaky scanning.
Publisher: Springer Science and Business Media LLC
Date: 30-10-1997
Abstract: Expression of the STA1-3 glucoamylase genes, responsible for starch degradation in Saccharomyces cerevisiae, is down regulated by the presence of STA10. In order to elucidate the role of STA10 in the regulation of the glucoamylase system, a multicopy genomic library was constructed and screened for genes that enhanced growth of a STA2-STA10 S. cerevisiae strain on starch media. This screen allowed us to clone and characterize a novel activator gene of STA2 (and by extrapolation, STA1 and STA3), designated MSS11. A strain transformed with multiple copies of MSS11 exhibits increased levels of STA2 mRNA and, consequently, increased glucoamylase activity. Deletion of MSS11, located on chromosome XIII, results in media-dependent absence of glucoamylase synthesis. MSS11 has not been cloned previously and the encoded protein, Mss11p, is not homologous to any other known protein. An outstanding feature of Mss11p is that the protein contains regions of 33 asparagine residues interrupted by only three serine residues, and 35 glutamine residues interrupted by a single histidine residue. Epistasis studies showed that deletion of MSS11 abolishes the activation of STA2 caused by the over-expression of MSS10, a previously identified gene. In turn, it was found that deletion of MSS10 still allows activation of STA2 by over-expression of MSS11. Mss11p therefore appears to be positioned below Mss10p in a signal transduction pathway.
Publisher: Elsevier BV
Date: 10-2002
DOI: 10.1016/S0167-7799(02)02049-8
Abstract: Wine producers are facing intensifying competition brought about by a widening gap between wine production and wine consumption, a shift of consumer preferences away from basic commodity wine to top quality wine, and economic globalization. Consequently, they are calling for a total revolution in the 'magical' world of wine. The process of transforming the wine industry from a production- to a market-orientated industry results in an increasing dependence on, amongst others, biotechnological innovation. Market-orientated wine-yeast strains are currently being developed for the cost-competitive production of wine with minimized resource inputs, improved quality and low environmental impact. The emphasis is on the development of Saccharomyces cerevisiae strains with improved fermentation, processing and biopreservation abilities, and capacities for an increase in the wholesomeness and sensory quality of wine.
Publisher: Wiley
Date: 29-11-2002
DOI: 10.1002/YEA.934
Abstract: Lipomyces kononenkoae secretes a battery of highly effective amylases (i.e. alpha-amylase, glucoamylase, isoamylase and cyclomaltodextrin glucanotransferase activities) and is therefore considered as one of the most efficient raw starch-degrading yeasts known. Previously, we have cloned and characterized genomic and cDNA copies of the LKA1 alpha-amylase gene from L. kononenkoae IGC4052B (CBS5608T) and expressed them in Saccharomyces cerevisiae and Schizosaccharomyces pombe. Here we report on the cloning and characterization of the genomic and cDNA copies of a second alpha-amylase gene (LKA2) from the same strain of L. kononenkoae. LKA2 was cloned initially as a 1663 bp cDNA harbouring an open reading frame (ORF) of 1496 nucleotides. Sequence analysis of LKA2 revealed that this ORF encodes a protein (Lka2p) of 499 amino acids, with a predicted molecular weight of 55,307 Da. The LKA2-encoded alpha-amylase showed significant homology to several bacterial cyclomaltodextrin glucanotransferases and also to the alpha-amylases of Aspergillus nidulans, Debaryomyces occidentalis, Saccharomycopsis fibuligera and Sz. pombe. When LKA2 was expressed under the control of the phosphoglycerate kinase gene promoter (PGK1(p)) in S. cerevisiae, it was found that the genomic copy contained a 55 bp intron that impaired the production of biologically active Lka2p in the heterologous host. In contrast to the genomic copy, the expression of the cDNA construct of PGK1p-LKA2 in S. cerevisiae resulted in the production of biologically active alpha-amylase. The LKA2-encoded alpha-amylase produced by S. cerevisiae exhibited a high specificity towards substrates containing alpha-1,4 glucosidic linkages. The optimum pH of Lka2p was found to be 3.5 and the optimum temperature was 60 degrees C. Besides LKA1, LKA2 is only the second L. kononenkoae gene ever cloned and expressed in S. cerevisiae. The cloning, characterization and co-expression of these two genes encoding these highly efficient alpha-amylases form an important part of an extensive research programme aimed at the development of amylolytic strains of S. cerevisiae for the efficient bioconversion of starch into commercially important commodities.
Publisher: MDPI AG
Date: 21-12-2022
DOI: 10.3390/MICROORGANISMS11010014
Abstract: Non-Saccharomyces yeasts are prevalent at the onset of grape must fermentations and can have a significant influence on the final wine product. In contrast to Saccharomyces cerevisiae, the biosynthetic pathways leading to aroma compound formation in these non-conventional yeasts, in particular those that are derived from amino acid metabolism, remains largely unexplored. Within a synthetic must environment, we investigated the amino acid utilization of four species (Hanseniaspora uvarum, Hanseniaspora osmophila, Zygosaccharomyces rouxii, Starmerella bacillaris) and S. cerevisiae. We report on the differential uptake preferences for amino acids with H. uvarum displaying the most rapid uptake of most amino acids. To investigate the fate of amino acids and their direct contribution to aroma synthesis in H. uvarum, H. osmophila and Z. rouxii, musts were supplemented with single amino acids. Aroma profiling undertaken after three days showed the synthesis of specific aroma compounds by the respective yeast was dependent on the specific amino acid supplementation. H. osmophila showed similarities to S. cerevisiae in both amino acid uptake and the synthesis of aroma compounds depending on the nitrogen sources. This study shows how the uptake of specific amino acids contributes to the synthesis of aroma compounds in wine fermentations using different non-Saccharomyces yeasts.
Publisher: Public Library of Science (PLoS)
Date: 20-11-2018
Publisher: Oxford University Press (OUP)
Date: 05-2004
Publisher: Springer Science and Business Media LLC
Date: 15-01-2021
DOI: 10.1038/S41467-020-20764-2
Abstract: The practices of synthetic biology are being integrated into ‘multiscale’ designs enabling two-way communication across organic and inorganic information substrates in biological, digital and cyber-physical system integrations. Novel applications of ‘bio-informational’ engineering will arise in environmental monitoring, precision agriculture, precision medicine and next-generation biomanufacturing. Potential developments include sentinel plants for environmental monitoring and autonomous bioreactors that respond to biosensor signaling. As bio-informational understanding progresses, both natural and engineered biological systems will need to be reimagined as cyber-physical architectures. We propose that a multiple length scale taxonomy will assist in rationalizing and enabling this transformative development in engineering biology.
Publisher: Elsevier BV
Date: 07-2017
DOI: 10.1016/J.IJFOODMICRO.2017.04.006
Abstract: It took several millennia to fully understand the scientific intricacies of the process through which grape juice is turned into wine. This yeast-driven fermentation process is still being perfected and advanced today. Motivated by ever-changing consumer preferences and the belief that the 'best' wine is yet to be made, numerous approaches are being pursued to improve the process of yeast fermentation and the quality of wine. Central to recent enhancements in winemaking processes and wine quality is the development of Saccharomyces cerevisiae yeast strains with improved robustness, fermentation efficiencies and sensory properties. The emerging science of Synthetic Biology - including genome engineering and DNA editing technologies - is taking yeast strain development into a totally new realm of possibility. The first ex le of how future wine strain development might be impacted by these new 'history-making' Synthetic Biology technologies, is the de novo production of the raspberry ketone aroma compound, 4-[4-hydroxyphenyl]butan-2-one, in a wine yeast containing a synthetic DNA cassette. This article explores how this breakthrough and the imminent outcome of the international Yeast 2.0 (or Sc2.0) project, aimed at the synthesis of the entire genome of a laboratory strain of S. cerevisiae, might accelerate the design of improved wine yeasts.
Publisher: Elsevier BV
Date: 05-2016
DOI: 10.1016/J.TIBTECH.2016.02.002
Abstract: Synthetic biology has progressed to the point where genes that encode whole metabolic pathways and even genomes can be manufactured and brought to life. This impressive ability to synthesise and assemble DNA is not yet matched by an ability to predictively engineer biology. These difficulties exist because biological systems are often overwhelmingly complex, having evolved to facilitate growth and survival rather than specific engineering objectives such as the optimisation of biochemical production. A promising and revolutionary solution to this problem is to harness the process of evolution to create microbial strains with desired properties. The tools of systems biology can then be applied to understand the principles of biological design, bringing synthetic biology closer to becoming a predictive engineering discipline.
Publisher: Oxford University Press (OUP)
Date: 2023
Abstract: Lack of gene-function analyses tools limits studying the biology of Hanseniaspora uvarum, one of the most abundant yeasts on grapes and in must. We investigated a rapid PCR-based gene targeting approach for one-step gene replacement in this diploid yeast. To this end, we generated and validated two synthetic antibiotic resistance genes, pFA-hygXL and pFA-clnXL, providing resistance against hygromycin and nourseothricin, respectively, for use with H. uvarum. Addition of short flanking-homology regions of 56–80 bp to these selection markers via PCR was sufficient to promote gene targeting. We report here the deletion of the H. uvarum LEU2 and LYS2 genes with these marker genes via two rounds of consecutive transformations, each resulting in the generation of auxotrophic strains (leu2/leu2 lys2/lys2). The hereby constructed leucine auxotrophic leu2/leu2 strain was subsequently complemented in a targeted manner, thereby further validating this approach. PCR-based gene targeting in H. uvarum was less efficient than in Saccharomyces cerevisiae. However, this approach, combined with the availability of two marker genes, provides essential tools for directed gene manipulations in H. uvarum.
Publisher: No publisher found
Date: 1992
DOI: 10.1016/0378-1119(92)90159-M
Abstract: Nine different expression-secretion cassettes, comprising novel combinations of yeast and bacterial gene promoters and secretion signal sequences, were constructed and evaluated. A pectate lyase-encoding gene (pelE) from Erwinia chrysanthemi was inserted between each one of these expression-secretion cassettes and a yeast gene terminator, generating recombinant yeast-integrating shuttle plasmids pAMS1 through pAMS9. These YIp5-derived plasmids were transformed and stably integrated into the genome of a laboratory strain of Saccharomyces cerevisiae, and the pectate lyase production was monitored. Transcription initiation signals for pelE expression were derived from the yeast alcohol dehydrogenase (ADC1P), the yeast mating pheromone alpha-factor (MF alpha 1P) and the Bacillus amyloliquefaciens alpha-amylase (AMYP) gene promoters. The transcription termination signals were derived from the yeast tryptophan synthase gene terminator (TRP5T). Secretion of pectate lyase (PLe) was directed by the signal sequences of the yeast mating pheromone alpha-factor (MF alpha 1S), B. amyloliquefaciens alpha-amylase (AMYS) and Er. chrysanthemi pectate lyase (pelES). The ADC1P-MF alpha 1S expression-secretion system proved to be the most efficient control cassette for the expression of pelE and the secretion of PLe in S. cerevisiae.
Publisher: Oxford University Press (OUP)
Date: 05-2009
DOI: 10.1111/J.1567-1364.2009.00489.X
Abstract: The production of hydrogen sulfide (H(2)S) during fermentation is a common and significant problem in the global wine industry as it imparts undesirable off-flavors at low concentrations. The yeast Saccharomyces cerevisiae plays a crucial role in the production of volatile sulfur compounds in wine. In this respect, H(2)S is a necessary intermediate in the assimilation of sulfur by yeast through the sulfate reduction sequence with the key enzyme being sulfite reductase. In this study, we used a classical mutagenesis method to develop and isolate a series of strains, derived from a commercial diploid wine yeast (PDM), which showed a drastic reduction in H(2)S production in both synthetic and grape juice fermentations. Specific mutations in the MET10 and MET5 genes, which encode the catalytic alpha- and beta-subunits of the sulfite reductase enzyme, respectively, were identified in six of the isolated strains. Fermentations with these strains indicated that, in comparison with the parent strain, H(2)S production was reduced by 50-99%, depending on the strain. Further analysis of the wines made with the selected strains indicated that basic chemical parameters were similar to the parent strain except for total sulfite production, which was much higher in some of the mutant strains.
Publisher: American Society for Microbiology
Date: 09-2005
DOI: 10.1128/AEM.71.9.5420-5426.2005
Abstract: Volatile thiols, particularly 4-mercapto-4-methylpentan-2-one (4MMP), make an important contribution to the aroma of wine. During wine fermentation, Saccharomyces cerevisiae mediates the cleavage of a nonvolatile cysteinylated precursor in grape juice (Cys-4MMP) to release the volatile thiol 4MMP. Carbon-sulfur lyases are anticipated to be involved in this reaction. To establish the mechanism of 4MMP release and to develop strains that modulate its release, the effect of deleting genes encoding putative yeast carbon-sulfur lyases on the cleavage of Cys-4MMP was tested. The results led to the identification of four genes that influence the release of the volatile thiol 4MMP in a laboratory strain, indicating that the mechanism of release involves multiple genes. Deletion of the same genes from a homozygous derivative of the commercial wine yeast VL3 confirmed the importance of these genes in affecting 4MMP release. A strain deleted in a putative carbon-sulfur lyase gene, YAL012W, produced a second sulfur compound at significantly higher concentrations than those produced by the wild-type strain. Using mass spectrometry, this compound was identified as 2-methyltetrathiophen-3-one (MTHT), which was previously shown to contribute to wine aroma but was of unknown biosynthetic origin. The formation of MTHT in YAL012W deletion strains indicates a yeast biosynthetic origin of MTHT. The results demonstrate that the mechanism of synthesis of yeast-derived wine aroma components, even those present in small concentrations, can be investigated using genetic screens.
Publisher: Elsevier BV
Date: 10-2004
Publisher: Elsevier BV
Date: 2022
DOI: 10.1016/J.TIBTECH.2021.05.001
Abstract: The creative destruction caused by the coronavirus pandemic is yielding immense opportunity for collaborative innovation networks. The confluence of biosciences, information sciences, and the engineering of biology, is unveiling promising bioinformational futures for a vibrant and sustainable bioeconomy. Bioinformational engineering, underpinned by DNA reading, writing, and editing technologies, has become a beacon of opportunity in a world paralysed by uncertainty. This article draws on lessons from the current pandemic and previous agricultural blights, and explores bioinformational research directions aimed at future-proofing the grape and wine industry against biological shocks from global blights and climate change.
Publisher: MDPI AG
Date: 16-06-2020
DOI: 10.3390/MOLECULES25122779
Abstract: Olfactory cues are key drivers of our multisensory experiences of food and drink. For ex le, our perception and enjoyment of the flavour and taste of a wine is primarily steered by its aroma. Making sense of the underlying smells that drive consumer preferences is integral to product innovation as a vital source of competitive advantage in the marketplace, which explains the intense interest in the olfactory component of flavour and the sensory significance of in idual compounds, such as one of the most important apocarotenoids for the bouquet of wine, β-ionone (violet and woody notes). β-Ionone is formed directly from β-carotene as a by-product of the actions of carotenoid cleavage dioxygenases (CCDs). The biological production of CCDs in microbial cell factories is one way that important aroma compounds can be generated on a large scale and with reduced costs, while retaining the ‘natural’ moniker. The CCD family includes the CCD1, CCD2, CCD4, CCD7 and CCD8 however, the functions, co-dependency and interactions of these CCDs remain to be fully elucidated. Here, we review the classification, actions and biotechnology of CCDs, particularly CCD1 and its action on β-carotene to produce the aromatic apocarotenoid β-ionone.
Publisher: Wiley
Date: 1999
DOI: 10.1046/J.1365-2958.1999.01151.X
Abstract: In Saccharomyces cerevisiae, a network of signal transduction pathways governs the switch from yeast-type growth to pseudohyphal and invasive growth that occurs in response to nutrient limitation. Important elements of this network have been identified, including nutrient signal receptors, GTP-binding proteins, components of the pheromone-dependent MAP kinase cascade and several transcription factors. However, the structural and functional mapping of these pathways is far from complete. Here, we present data regarding three genes, MSN1/MSS10, MSS11 and MUC1/FLO11, which form an essential part of the signal transduction network establishing invasive growth. Both MSN1 and MSS11 are involved in the co-regulation of starch degradation and invasive growth. Msn1p and Mss11p act downstream of Mep2p and Ras2p and regulate the transcription of both STA2 and MUC1. We show that MUC1 mediates the effect of Msn1p and Mss11p on invasive growth. In addition, our results suggest that the activity of Msn1p is independent of the invasive growth MAP kinase cascade, but the Mss11p is required for the activation of pseudohyphal and invasive growth by Ste12p. We also show that starch metabolism in S. cerevisiae is subject to regulation by components of the MAP kinase cascade.
Publisher: Oxford University Press (OUP)
Date: 23-07-2007
DOI: 10.1111/J.1365-2672.2007.03474.X
Abstract: The main objective of this study was to develop polysaccharide-degrading wine strains of Saccharomyces cerevisiae, which are able to improve aspects of wine processing and clarification, as well as colour extraction and stabilization during winemaking. Two yeast expression/secretion gene cassettes were constructed, namely (i) a pectinase gene cassette (pPPK) consisting of the endo-polygalacturonase gene (pelE) from Erwinia chrysanthemi and the pectate lyase gene (peh1) from Erwinia carotovora and (ii) a glucanase/xylanase gene cassette (pEXS) containing the endo-beta-1,4-glucanase gene (end1) from Butyrivibrio fibrisolvens and the endo-beta-1,4-xylanase gene (xynC) from Aspergillus niger. The commercial wine yeast strain, VIN13, was transformed separately with these two gene cassettes and checked for the production of pectinase, glucanase and xylanase activities. Pinot Noir, Cinsaut and Muscat d'Alexandria grape juices were fermented using the VIN13[pPPK] pectinase- and the VIN13[pEXS] glucanase/xylanase-producing transformants. Chemical analyses of the resultant wines indicated that (i) the pectinase-producing strain caused a decrease in the concentration of phenolic compounds in Pinot Noir whereas the glucanase/xylanase-producing strain caused an increase in phenolic compounds presumably because of the degradation of the grape skins (ii) the glucanase/xylanase-producing strain caused a decrease in wine turbidity, especially in Pinot Noir wine, as well as a clear increase in colour intensity and (iii) in the Muscat d'Alexandria and Cinsaut wines, the differences between the control wines (fermented with the untransformed VIN3 strain) and the wines produced by the two transformed strains were less prominent showing that the effect of these polysaccharide-degrading enzymes is cultivar-dependent. The recombinant wine yeasts producing pectinase, glucanase and xylanase activities during the fermentation of Pinot Noir, Cinsaut and Muscat d'Alexandria grape juice altered the chemical composition of the resultant wines in a way that such yeasts could potentially be used to improve the clarity, colour intensity and stability and aroma of wine. Aspects of commercial-scale wine processing and clarification, colour extraction and stabilization, and aroma enhancement could potentially be improved by the use of polysaccharide-degrading wine yeasts without the addition of expensive commercial enzyme preparations. This offers the potential to further improve the price:quality ratio of wine according to consumer expectations.
Publisher: Wiley
Date: 03-1990
Abstract: K2 neutral strain Saccharomyces cerevisiae USM12 was identified and characterized. This strain carried an M double-stranded RNA (dsRNA) genome encoding for resistance to K2 toxin. The M dsRNA was larger than the K2 killer yeast M dsRNA and homoduplex analysis of denatured and reannealed K2 neurtal M dsRNA revealed an inverted duplication. Heteroduplex analysis showed that two thirds of the K2 M genome had homology with the K2 neutral M genome. Hybridization showed that the USM12 M dsRNA had significant homology with the K2 M dsRNA. Protein profiles of extracellular proteins from USM12 and a cured strain indicated that USM12 did not secrete any toxin. This is the first time that a K2 neutral yeast strain has been characterized.
Publisher: Wiley
Date: 18-12-2002
DOI: 10.1046/J.1365-2958.2003.03247.X
Abstract: In Saccharomyces cerevisiae , the cell surface protein, Muc1p, was shown to be critical for invasive growth and pseudohyphal differentiation. The transcription of MUC1 and of the co‐regulated STA2 glucoamylase gene is controlled by the interplay of a multitude of regulators, including Ste12p, Tec1p, Flo8p, Msn1p and Mss11p. Genetic analysis suggests that Mss11p plays an essential role in this regulatory process and that it functions at the convergence of at least two signalling cascades, the filamentous growth MAPK cascade and the cAMP‐PKA pathway. Despite this central role in the control of filamentous growth and starch metabolism, the exact molecular function of Mss11p is unknown. We subjected Mss11p to a detailed molecular analysis and report here on its role in transcriptional regulation, as well as on the identification of specific domains required to confer transcriptional activation in response to nutritional signals. We show that Mss11p contains two independent transactivation domains, one of which is a highly conserved sequence that is found in several proteins with unidentified function in mammalian and invertebrate organisms. We also identify conserved amino acids that are required for the activation function.
Publisher: Authorea, Inc.
Date: 22-06-2023
DOI: 10.22541/AU.168747668.85315525/V1
Abstract: Yeast research is entering into a new period of scholarship, with new scientific tools, new questions to ask, and new issues to consider. The politics of emerging and critical technology can no longer be separated from the pursuit of basic science in fields, such as synthetic biology and engineering biology. Given the intensifying race for technological leadership, yeast research is likely to attract significant investment from government, and that it offers huge opportunities to the curious minded from a basic research standpoint. This article provides an overview of new directions in yeast research, and places these trends in their geopolitical context. At the highest level, yeast research is situated within the ongoing convergence of the life sciences with the information sciences. This convergent effect is most strongly pronounced in areas of AI-enabled tools for the life sciences, and the creation of synthetic genomes, minimal genomes, pan-genomes, neochromosomes and metagenomes using computer assisted design tools and methodologies. Synthetic yeast futures encompass basic and applied science questions that will be of intense interest to government and non-government funding sources. It is essential for the yeast research community to map and understand the context of their research in order to ensure their collaborations turn global challenges into research opportunities.
Publisher: Oxford University Press (OUP)
Date: 10-04-2012
DOI: 10.1111/J.1567-1364.2012.00797.X
Abstract: Industrial food-grade yeast strains are selected for traits that enhance their application in quality production processes. Wine yeasts are required to survive in the harsh environment of fermenting grape must, while at the same time contributing to wine quality by producing desirable aromas and flavors. For this reason, there are hundreds of wine yeasts available, exhibiting characteristics that make them suitable for different fermentation conditions and winemaking practices. As wine styles evolve and technical winemaking requirements change, however, it becomes necessary to improve existing strains. This becomes a laborious and costly process when the targets for improvement involve flavor compound production. Here, we demonstrate a new approach harnessing preexisting industrial yeast strains that carry desirable flavor phenotypes - low hydrogen sulfide (H(2) S) production and high ester production. A low-H(2) S Saccharomyces cerevisiae strain previously generated by chemical mutagenesis was hybridized independently with two ester-producing natural interspecies hybrids of S. cerevisiae and Saccharomyces kudriavzevii. Deficiencies in sporulation frequency and spore viability were overcome through use of complementary selectable traits, allowing successful isolation of several novel hybrids exhibiting both desired traits in a single round of selection.
Publisher: Springer Science and Business Media LLC
Date: 03-2015
DOI: 10.1007/BF03175601
Publisher: EMBO
Date: 23-10-2017
Publisher: Oxford University Press (OUP)
Date: 09-2014
Abstract: Brettanomyces bruxellensis, like its wine yeast counterpart Saccharomyces cerevisiae, is intrinsically linked with industrial fermentations. In wine, B. bruxellensis is generally considered to contribute negative influences on wine quality, whereas for some styles of beer, it is an essential contributor. More recently, it has shown some potential for bioethanol production. Our relatively poor understanding of B. bruxellensis biology, at least when compared with S. cerevisiae, is partly due to a lack of laboratory tools. As it is a nonmodel organism, efforts to develop methods for sporulation and transformation have been sporadic and largely unsuccessful. Recent genome sequencing efforts are now providing B. bruxellensis researchers unprecedented access to gene catalogues, the possibility of performing transcriptomic studies and new insights into evolutionary drivers. This review summarises these findings, emphasises the rich data sets already available yet largely unexplored and looks over the horizon at what might be learnt soon through comprehensive population genomics of B. bruxellensis and related species.
Publisher: Springer Science and Business Media LLC
Date: 1997
Publisher: Springer Science and Business Media LLC
Date: 21-03-1997
Abstract: A genomic DNA library of the bacterium Bacillus pumilus PLS was constructed and the beta-xylosidase gene (xynB) was lified from a 3-kb genomic DNA fragment with the aid of the polymerase chain reaction technique. The lified xynB gene was inserted between the yeast alcohol dehydrogenase II gene promoter (ADH2P) and terminator (ADH2T) sequences on a multicopy episomal plasmid (pDLG11). The xynB gene was also fused in-frame to the secretion signal sequence of the yeast mating pheromone alpha-factor (MF alpha 1S) before insertion between the ADH2P and ADH2T sequences on a similar multicopy episomal plasmid (pDLG12). The resulting construct ADH2P-MF alpha 1S-xynB-ADH2T was designated XLO1. Both plasmids pDLG11 and PDLG12 were introduced into Saccharomyces cerevisiae but only the expression of the XLO1 gene yielded biologically functional beta-xylosidase. The total beta-xylosidase activity remained cell-associated with a maximum activity of 0.09 nkat/ml obtained when the recombinant S. cerevisiae strain was grown for 143 h in synthetic medium. The temperature and pH optima of the recombinant Xlo1 enzyme were 45-50 degrees C and pH 6.6 respectively. The enzyme was thermostable at 45 degrees C however, at 60 degrees C most of the Xlo1 was inactive after 5 min.
Publisher: Elsevier BV
Date: 09-2018
DOI: 10.1016/J.YMBEN.2018.08.006
Abstract: Metabolic engineering has been vital to the development of industrial microbes such as the yeast Saccharomyces cerevisiae. However, sequential rounds of modification are often needed to achieve particular industrial design targets. Systems biology approaches can aid in identifying genetic targets for modification through providing an integrated view of cellular physiology. Recently, research into the generation of commercial yeasts that can produce reduced-ethanol wines has resulted in metabolically-engineered strains of S. cerevisiae that are less efficient at producing ethanol from sugar. However, these modifications led to the concomitant production of off-flavour by-products. A combination of transcriptomics, proteomics and metabolomics was therefore used to investigate the physiological changes occurring in an engineered low-ethanol yeast strain during alcoholic fermentation. Integration of 'omics data identified several metabolic reactions, including those related to the pyruvate node and redox homeostasis, as being significantly affected by the low-ethanol engineering methodology, and highlighted acetaldehyde and 2,4,5-trimethyl-1,3-dioxolane as the main off-flavour compounds. Gene remediation strategies were then successfully applied to decrease the formation of these by-products, while maintaining the 'low-alcohol' phenotype. The data generated from this comprehensive systems-based study will inform wine yeast strain development programmes, which, in turn, could potentially play an important role in assisting winemakers in their endeavour to produce low-alcohol wines with desirable flavour profiles.
Publisher: Elsevier BV
Date: 04-2009
Abstract: The main objective of this study was to investigate the effect of different Saccharomyces cerevisiae wine yeast strains on the concentration of aroma-enhancing volatile thiols and fermentation metabolites in Sauvignon Blanc wine. Seven commercial wine yeast strains were selected based on their putative ability to modulate the concentrations of the fruity volatile thiols, 4-mercapto-4-methylpentan-2-one (4MMP), 3-mercapto-hexanol (3MH) and 3-mercapto-hexylacetate (3MHA). Each of these yeasts was used to produce Sauvignon Blanc wines under controlled conditions, in triplicate, in 20-L quantities. The levels of 4MMP, 3MH and 3MHA in these wines were quantified using the p-hydroxymercuribenzoate method. In addition, a total of 24 volatile yeast-derived fermentation aroma compounds were also quantified using headspace solid-phase microextraction coupled with gas chromatography mass spectrometry (HS-SPME-GC-MS). Formal sensory analysis was conducted by 12 trained assessors and, additionally, a panel of 24 experienced wine industry professionals assessed the wines and ranked them in order of preference. The results indicated that the yeast strains varied significantly in terms of their capabilities to (i) produce volatile thiols and fermentation metabolites and (ii) to modulate the varietal characters of Sauvignon Blanc wine. Yeast strains that produced the highest levels of volatile thiols were responsible for wines with the highest perceived intensity of fruitiness, and these wines were preferred by the tasting panels. While the 'green' characters in Sauvignon Blanc wines can be manipulated through vineyard management, the 'tropical fruity' characters appear to be largely dependent on the wine yeast strain used during fermentation. Therefore, the choice of yeast strain offers great potential to modulate wine aroma profiles to definable styles and predetermined consumer market specifications.
Publisher: Oxford University Press (OUP)
Date: 04-2008
DOI: 10.1111/J.1365-2672.2007.03671.X
Abstract: The main objective of this study was to identify amino acid residues in the AGT1-encoded alpha-glucoside transporter (Agt1p) that are critical for efficient transport of maltotriose in the yeast Saccharomyces cerevisiae. The sequences of two AGT1-encoded alpha-glucoside transporters with different efficiencies of maltotriose transport in two Saccharomyces strains (WH310 and WH314) were compared. The sequence variations and discrepancies between these two proteins (Agt1p(WH310) and Agt1p(WH314)) were investigated for potential effects on the functionality and maltotriose transport efficiency of these two AGT1-encoded alpha-glucoside transporters. A 23-amino-acid C-terminal truncation proved not to be critical for maltotriose affinity. The identification of three amino acid differences, which potentially could have been instrumental in the transportation of maltotriose, were further investigated. Single mutations were created to restore the point mutations I505T, V549A and T557S one by one. The single site mutant V549A showed a decrease in maltotriose transport ability, and the I505T and T557S mutants showed complete reduction in maltotriose transport. The amino acids Thr(505) and Ser(557), which are respectively located in the transmembrane (TM) segment TM(11) and on the intracellular segment after TM(12) of the AGT1-encoded alpha-glucoside transporters, are critical for efficient transport of maltotriose in S. cerevisiae. Improved fermentation of starch and its dextrin products, such as maltotriose and maltose, would benefit the brewing and whisky industries. This study could facilitate the development of engineered maltotriose transporters adapted to starch-efficient fermentation systems, and offers prospects for the development of yeast strains with improved maltose and maltotriose uptake capabilities that, in turn, could increase the overall fermentation efficiencies in the beer and whisky industries.
Publisher: Springer Science and Business Media LLC
Date: 11-07-2019
DOI: 10.1038/S41467-019-10862-1
Abstract: The original version of this Comment contained errors in the legend of Figure 2, in which the locations of the fifteenth and sixteenth GBA members were incorrectly given as ‘(15) Australian Genome Foundry, Macquarie University (16) Australian Foundry for Advanced Biomanufacturing, University of Queensland.’. The correct version replaces this with ‘(15) Australian Foundry for Advanced Biomanufacturing (AusFAB), University of Queensland and (16) Australian Genome Foundry, Macquarie University’. This has been corrected in both the PDF and HTML versions of the Comment.
Publisher: Springer Science and Business Media LLC
Date: 09-05-2019
Publisher: American Chemical Society (ACS)
Date: 05-10-2021
Publisher: Oxford University Press (OUP)
Date: 02-2007
DOI: 10.1111/J.1574-6968.2006.00548.X
Abstract: The wine yeast Saccharomyces cerevisiae is central in the production of aroma compounds during fermentation. Some of the most important yeast-derived aroma compounds produced are esters. The esters ethyl acetate and isoamyl acetate are formed from alcohols and acetyl-CoA in a reaction catalysed by alcohol acetyltransferases. The pool of acetyl-CoA available in yeast cells could play a key role in the development of ester aromas. Carnitine acetyltransferases catalyse the reversible reaction between carnitine and acetyl-CoA to form acetylcarnitine and free CoA. This reaction is important in transferring activated acetyl groups to the mitochondria and in regulating the acetyl-CoA/CoA pools within the cell. We investigated the effect of overexpressing CAT2, which encodes the major mitochondrial and peroxisomal carnitine acetyltransferase, on the formation of esters and other flavour compounds during fermentation. We also overexpressed a modified CAT2 that results in a protein that localizes to the cytosol. In general, the overexpression of both forms of CAT2 resulted in a reduction in ester concentrations, especially in ethyl acetate and isoamyl acetate. We hypothesize that overproduction of Cat2p favours the formation of acetylcarnitine and CoA and therefore limits the precursor for ester production. Carnitine acetyltransferase expression could potentially to be used successfully in order to modulate wine flavour.
Publisher: Elsevier BV
Date: 05-1997
DOI: 10.1016/S0168-1656(97)00059-X
Abstract: The EXG1 gene encoding the main Saccharomyces cerevisiae exo-beta-1,3-glucanase was cloned and over-expressed in yeast. The Bacillus subtilis endo-1,3-1,4-beta-glucanase gene (beg1) and the Butyrivibrio fibrisolvens endo-beta-1,4-glucanase gene (end1) were fused to the secretion signal sequence of the yeast mating pheromone alpha-factor (MF alpha 1S) and inserted between the yeast alcohol dehydrogenase II gene promoter (ADH2P) and terminator (ADH2T). Constructs ADH2P-MF alpha 1S-beg1-ADH2T and ADH2P-MF alpha 1S-end 1-ADH2T designated BEG1 and END1, respectively, were expressed separately and jointly with EXG1 in S. cerevisiae. The construction of fur 1 ura3 S. cerevisiae strains allowed for the autoselection of these multicopy URA3-based plasmids in rich medium. Enzyme assays confirmed that co-expression of EXG1, BEG1 and END1 enhanced glucan degradation by S. cerevisiae.
Publisher: MDPI AG
Date: 19-12-2016
Publisher: Springer Science and Business Media LLC
Date: 22-10-1996
Abstract: First-strand cDNA was prepared from mRNA of Aspergillus niger MRC11624 induced on oat spelts xylan. Using the cDNA as a template, the alpha-L-arabinofuranosidase gene (abfB) was lified with the polymerase chain reaction technique. The abfB DNA fragment was inserted between the yeast phosphoglycerate kinase I gene promoter (PGK1P) and terminator (PGK1T) sequences on a multicopy episomal plasmid. The resulting construct PGK1P-abfB-PGK1T was designated ABF2. The ABF2 gene was expressed successfully in Saccharomyces cerevisiae and functional alpha-L-arabinofuranosidase was secreted from the yeast cells. The ABF2 nucleotide sequence was determined and verified to encode a 449-amino-acid protein (Abf2) that is 94% identical to the alpha-L-arabinofuranosidase B of A. niger N400. Maximum alpha-L-arabinofuranosidase activities of 0.020 U/ml and 1.40 U/ml were obtained with autoselective recombinant S. cerevisiae strains when grown for 48 h in synthetic and complex medium respectively.
Publisher: Oxford University Press (OUP)
Date: 11-2008
DOI: 10.1111/J.1567-1364.2008.00434.X
Abstract: Many industrial strains of Saccharomyces cerevisiae have been selected primarily for their ability to convert sugars into ethanol efficiently despite exposure to a variety of stresses. To begin investigation of the genetic basis of phenotypic variation in industrial strains of S. cerevisiae, we have sequenced the genome of a wine yeast, AWRI1631, and have compared this sequence with both the laboratory strain S288c and the human pathogenic isolate YJM789. AWRI1631 was found to be substantially different from S288c and YJM789, especially at the level of single-nucleotide polymorphisms, which were present, on average, every 150 bp between all three strains. In addition, there were major differences in the arrangement and number of Ty elements between the strains, as well as several regions of DNA that were specific to AWRI1631 and that were predicted to encode proteins that are unique to this industrial strain.
Publisher: Public Library of Science (PLoS)
Date: 03-02-2011
Publisher: Oxford University Press (OUP)
Date: 16-03-2018
Publisher: Wiley
Date: 28-02-2008
Publisher: MDPI AG
Date: 10-12-2016
DOI: 10.20944/PREPRINTS201612.0061.V1
Abstract: A perfectly balanced wine can be said to create a symphony in the mouth. To achieve the sublime, both in wine and music, requires imagination and skilled orchestration of artistic craftmanship. For wine, inventiveness starts in the vineyard. Similar to a composer of music, the grapegrower produces grapes through a multitude of specifications to achieve a quality result. Different Vitis vinifera grape varieties allow the creation of wine of different genres. Akin to a conductor of music, the winemaker decides what genre to create and considers resources required to realise the grape& rsquo s potential. A primary consideration is the yeast: inoculate the grape juice or leave it & lsquo wild& rsquo which specific or combined Saccharomyces strain(s) should be used or proceed with a non-Saccharomyces species? Whilst the various Saccharomyces and non-Saccharomyces yeasts perform their role during fermentation, the performance is not over until the & lsquo fat lady& rsquo (S. cerevisiae) has sung (i.e. the grape sugar has been fermented to specified dryness and alcoholic fermentation is complete). Is the wine harmonious or discordant? Will the consumer demand an encore and make a repeat purchase? Understanding consumer needs lets winemakers orchestrate different symphonies (i.e. wine styles) using single- or multi-species ferments. Some consumers will choose the sounds of a philharmonic orchestra comprising a great range of erse instrumentalists (as is the case with wine created from spontaneous fermentation) some will prefer to listen to a smaller ensemble (analogous to wine produced by a selected group of non-Saccharomyces and Saccharomyces yeast) and others will favour the well-known and reliable superstar soprano (i.e. S. cerevisiae). But what if a digital music synthesiser ‒ such as a synthetic yeast& nbsp ‒ becomes available that can produce any music genre with the purest of sounds by the touch of a few buttons? Will synthesisers spoil the character of the music and lead to the loss of the much-lauded romantic mystique? Or will music synthesisers support composers and conductors to create novel compositions and even higher quality performances that will thrill audiences? This article explores these and other relevant questions in the context of winemaking and the role that yeast and its genomics play in the betterment of wine quality.
Start Date: 05-2020
End Date: 12-2023
Amount: $553,035.00
Funder: Australian Research Council
View Funded Activity