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PubMed2001

A Saccharomyces servazzii clone homologous to Saccharomyces cerevisiae chromosome III spanning KAR4, ARS 304 and SPB1 lacks the recombination enhancer but contains an unknown ORF.

In order to learn about the evolutionary conservation of the recombination enhancer (RE) that controls donor preference during mating type switching in Saccharomyces cerevisiae, we have cloned a 13 kb region from S. servazzii. We find that the order of four genes surrounding the RE in S. cerevisiae (PRD1, KAR4, SPB1 and PBN1) is preserved in S.servazzii. However, there is an additional ORF in S. servazzii between PRD1 and KAR4 that is not homologous to any gene in S. cerevisiae or to genes in other organisms. Despite a 75-79% amino acid identity for KAR4 and SPB1, respectively, the S. servazzii sequence did not carry a well-conserved RE sequence and these sequences lacked RE function when introduced into S. cerevisiae. The S. servazzii region contains a sequence that supports autonomous DNA replication in S. cerevisiae and may represent a homologue of ARS304. The S. servazziii sequence has Genbank Accession No. BankIt359091 AF307954.

Amino Acid Sequence↗

PubMed2002

Breeding of brewer's yeast by hybridization between a top-fermenting yeast Saccharomyces cerevisiae and a cryophilic yeast Saccharomyces bayanus.

To improve the fermentability of a top-fermenting yeast at low-temperature, we performed hybridization trials between four top-fermenting Saccharomyces cerevisiae strains and a cryophilic yeast Saccharomyces bayanus YM84 with good fermentability at low-temperature. The hybrids selected using 5-bromo-4-chloro-3-indolyl-alpha-D-galactopyranoside were checked with pulsed-field gel electrophoresis and their brewing performance at the low-temperature of 10.5 degrees C was observed using small-scale (2 l) fermentation trials.

Journal Article↗

PubMed1997

Secondary products formation as a tool for discriminating non-Saccharomyces wine strains. Strain diversity in non-Saccharomyces wine yeasts.

A total of 78 strains of non-Saccharomyces yeasts were isolated: 30 strains of Kloeckera apiculata, 20 of Candida stellata, 8 of Candida valida and 20 of Zygosaccharomyces fermentati. The diversity of yeast species and strains was monitored by determining the formation of secondary products of fermentation, such as acetaldehyde, ethyl acetate and higher alcohols. Within each species, the strains were distinguishable in phenotypes through the production of different amounts of by-products. In particular, a great variability was found in C. stellata, where six different phenotypes were identified by means of the production of acetaldehyde, ethyl acetate, isobutanol and isoamyl alcohol. At different stages of the spontaneous fermentation different phenotypes of the non-Saccharomyces yeasts were represented, characterized by consistent differences in some by-products involved in the wine bouquet, such as acetaldehyde.

Acetaldehyde↗

PubMed1983

Effects of Fusariotoxin T-2 on Saccharomyces cerevisiae and Saccharomyces carlsbergensis.

A Fusarium metabolite, T-2 toxin, inhibits the growth of Saccharomyces carlsbergensis and Saccharomyces cerevisiae. The growth inhibitory concentrations of T-2 toxin were 40 and 100 mug/ml, respectively, for exponentially growing cultures of the two yeasts. S. carlsbergensis was more sensitive to the toxin and exhibited a biphasic dose-response curve. Addition of the toxin at 10 mug/ml of S. carlsbergensis culture resulted in a retardation of growth as measured turbidimetrically, after only 30 to 40 min. This action was reversible upon washing the cells free of the toxin. The sensitivity of the yeasts to the toxin was dependent upon the types and concentrations of carbohydrates used in the growth media. The sensitivity of the cells to the toxin decreased in glucose-repressed cultures. These results suggest that T-2 toxin interferes with mitochondrial functions of these yeasts.

Journal Article↗

PubMed2003

Quantitative comparison of transient growth of Saccharomyces cerevisiae, Saccharomyces kluyveri, and Kluyveromyces lactis.

A multitude of metabolic regulations occur in yeast, particularly under dynamic process conditions, such as under sudden glucose excess. However, quantification of regulations and classification of yeast strains under these conditions have yet to be elucidated, which requires high-frequency and consistent quantification of the metabolic response. The present study aimed at quantifying the dynamic regulation of the central metabolism of strains Saccharomyces cerevisiae, S. kluyveri, and Kluyveromyces lactis upon sudden glucose excess, accomplished by a shift-up in dilution rate inside of the oxidative region using a small metabolic flux model. It was found that, under transient growth conditions, S. kluyveri behaved like K. lactis, while classification using steady-state conditions would position S. kluyveri close to S. cerevisiae. For transient conditions and based on the observation whether excess glucose is initially used for catabolism (energy) or anabolism (carbon), we propose to classify strains into energy-driven, such as S. cerevisiae, and carbon-driven, such as S. kluyveri and K. lactis, strains. Furthermore, it was found that the delayed onset of fermentative catabolism in carbon-driven strains is a consequence of low catabolic flux and the initial shunt of glucose in non-nitrogen-containing biomass constituents. The MFA model suggests that energy limitation forced the cell to ultimately increase catabolic flux, while the capacity of oxidative catabolism is not sufficient to process this flux oxidatively. The combination of transient experiments and its exploitation with reconciled intrinsic rates using a small metabolic model could corroborate earlier findings of metabolic regulations, such as tight glucose control in carbon-driven strains and transient changes in biomass composition, as well as explore new regulations, such as assimilation of ethanol before glucose. The benefit from using small metabolic flux models is the richness of information and the enhanced insight into intrinsic metabolic pathways without a priori knowledge of adaptation kinetics. Used in an online context, this approach serves as an efficient tool for strain characterization and physiological studies.

Adaptation, Physiological↗

PubMed1993

Molecular genetics in Saccharomyces kluyveri: the HIS3 homolog and its use as a selectable marker gene in S. kluyveri and Saccharomyces cerevisiae.

We cloned the Saccharomyces kluyveri HIS3 homolog, k-HIS3, and made a partial deletion of the gene. The k-HIS3 gene complemented a HIS3 deletion in S. cerevisiae. The DNA sequences of the open reading frames (ORFs) of the HIS3 homologs are 70% identical at the DNA level and 83% identical at the deduced amino acid level. The ORF upstream of the k-HIS3 gene is related to the PET56 gene of S. cerevisiae found upstream of the HIS3 gene of S. cerevisiae. The ORF downstream from the k-HIS3 gene is not related to the DED1 gene found downstream of the HIS3 gene in S. cerevisiae.

Amino Acid Sequence↗

PubMed1994

Comparative analysis of the region of the mitochondrial genome containing the ATPase subunit 9 gene in the two related yeast species Saccharomyces douglasii and Saccharomyces cerevisiae.

We have determined the nucleotide sequence of a region of the mitochondrial genome of the yeast Saccharomyces douglasii which contains the ATPase subunit 9 gene and part of the intergenic sequences that surround it. The gene is 228 nucleotides long and encodes a polypeptide of 76 aa. A comparison of the coding sequence with that of S. cerevisiae reveals the presence of three silent transitions. A high level of similarity is also found between regions involved in the initiation of transcription and mRNA processing. More interestingly, a region of similarity situated outside the known regulatory regions has been identified. As the intergenic regions are generally highly divergent, the remarkable conservation of these non-coding sequences suggests that their structure may be relevant to the expression of this region of the mitochondrial DNA.

Adenosine Triphosphatases↗

PubMed1998

Reciprocal translocation at duplicated RPL2 loci might cause speciation of Saccharomyces bayanus and Saccharomyces cerevisiae.

By a genomic comparison of two sibling yeasts, Saccharomyces bayanus and S. cerevisiae, we previously demonstrated that chromosomes II and IV of S. cerevisiae were rearranged into chromosomes 12 and 14 of S. bayanus or vice versa. In the present study we have delimited the translocation break sites in chromosomes II and IV by Southern hybridization using DNA fragments of S. cerevisiae cosmid clones as probes. The results suggest that the reciprocal translocation of chromosomes II and IV had occurred at duplicated RPL2 loci. Furthermore, the translocation sites in S. bayanus were confirmed by the cloning and sequence analysis of the regions flanking RPL2 loci. Several genes in the regions flanking the RPL2 loci were present in the order expected for a translocation at these loci between the two species. These results indicated that the reciprocal translocation between chromosomes II and IV was generated by homologous recombination at duplicated RPL2 loci on the two chromosomes. Therefore, we propose that duplicated genes or duplicated regions play an important role in altering genomic organization during the speciation of S. bayanus and S. cerevisiae.

Base Sequence↗

PubMed2005

Saccharomyces cerevisiae GTPase complex: Gtr1p-Gtr2p regulates cell-proliferation through Saccharomyces cerevisiae Ran-binding protein, Yrb2p.

A Gtr1p GTPase, the GDP mutant of which suppresses both temperature-sensitive mutants of Saccharomyces cerevisiae RanGEF/Prp20p and RanGAP/Rna1p, was presently found to interact with Yrb2p, the S. cerevisiae homologue of mammalian Ran-binding protein 3. Gtr1p bound the Ran-binding domain of Yrb2p. In contrast, Gtr2p, a partner of Gtr1p, did not bind Yrb2p, although it bound Gtr1p. A triple mutant: yrb2delta gtr1delta gtr2delta was lethal, while a double mutant: gtr1delta gtr2delta survived well, indicating that Yrb2p protected cells from the killing effect of gtr1delta gtr2delta. Recombinant Gtr1p and Gtr2p were purified as a complex from Escherichia coli. The resulting Gtr1p-Gtr2p complex was comprised of an equal amount of Gtr1p and Gtr2p, which inhibited the Rna1p/Yrb2 dependent RanGAP activity. Thus, the Gtr1p-Gtr2p cycle was suggested to regulate the Ran cycle through Yrb2p.

Cell Proliferation↗

PubMed2003

Saccharomyces cerevisiae fungemia after Saccharomyces boulardii treatment in immunocompromised patients.

Saccharomyces cerevisiae is widely used as a probiotic compound. Clinical data suggest that this agent is safe and effective. We report two cases of fungemia caused by S. cerevisiae occurring in immunosuppressed patients treated orally with S. boulardii Molecular typing confirmed clonality in isolate strains from patients and the capsule. Physicians caring for immunosuppressed patients must be aware of this potential serious complication of probiotic use.

Adult↗

PubMed1984

Plasmids resembling 2-micrometers DNA in the osmotolerant yeasts Saccharomyces bailii and Saccharomyces bisporus.

Five of eight strains of Saccharomyces bailii and one of 13 strains of S. bisporus were found to harbour DNA plasmids. pSB1 and pSB2 plasmids were isolated from S. bailii strains IFO 0488 and IFO 1047, respectively, and pSB3 and pSB4 from S. bisporus strain IFO 1730. All four plasmids resemble 2-micrometers DNA of S. cerevisiae in that their molecular sizes are about 6 kb, each molecule possesses a pair of inverted repeats, they exist as a mixture of two isomers and their copy numbers in the native host are similar. None of them showed homology with 2-micrometers DNA or with each other by Southern hybridization under moderately stringent conditions, but pSB4 hybridized with the pSR1 DNA, which was found previously in a strain of S. rouxii. Each of the pSB plasmids has DNA sequence(s) effective for autonomous replication in S. cerevisiae. In S. cerevisiae, pSB3 and pSB4 showed intramolecular recombination but neither supported isomerization of 2-micrometers DNA.

DNA Restriction Enzymes↗

PubMed1996

Development of a polymerase chain reaction/restriction fragment length polymorphism method for Saccharomyces cerevisiae and Saccharomyces bayanus identification in enology.

Several yeast strains of the species Saccharomyces cerevisiae, S. bayanus and S. paradoxus, first identified by hybridization experiments and measurements of DNA/DNA homology, were characterized using polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) analysis of the MET2 gene. There was no exception to the agreement between this method and classical genetic analyses for any of the strains examined, so PCR/RFLP of the MET2 gene is a reliable and fast technique for delimiting S. cerevisiae and S. bayanus. Enological strains classified as S. bayanus, S. chevalieri, and S. capensis gave S. cerevisiae restriction patterns, whereas most S. uvarum strains belong to S. bayanus. Enologists should no longer use the name of S. bayanus for S. cerevisiae Gal strains, and should consider S. bayanus as a distinct species.

Base Sequence↗

PubMedDate not supplied

[Component activity of the isoenzyme spectra of Saccharomyces cerevisiae, Saccharomyces carlsbergensis and their hybrids].

The object of this work was to study the activity and the isozyme spectra of hexokinase (the triggering enzyme of glycolysis), glucose-6-phosphate dehydrogenase (the key enzyme of the pentose-phosphate shunt), malate dehydrogenase and isocitrate dehydrogenase (the enzymes of the citric acid cycle) and alcohol dehydrogenase (the enzyme involved in the first steps of ethanol oxidation) in Saccharomyces cerevisiae, race Ya, S. carlsbergensis, race 4228, and their hybrid 67. The parent organisms and their hybrid were shown to differ from one another in the qualitative composition and the activity of the isozyme spectra of the above enzymes.

Alcohol Oxidoreductases↗

PubMed2003

Viable Saccharomyces cerevisiae cells at high concentrations cause early growth arrest of non-Saccharomyces yeasts in mixed cultures by a cell-cell contact-mediated mechanism.

The growth of Kluyveromyces thermotolerans and Torulaspora delbrueckii was examined in mixed cultures with Saccharomyces cerevisiae in YPD modified for wine fermentations. Although the three yeasts had similar maximum specific growth rates in these fermentations, K. thermotolerans and T. delbrueckii arrested growth earlier than S. cerevisiae, thereby obtaining lower stationary phase cell concentrations than S. cerevisiae. Various single and mixed culture fermentations with the three yeasts were carried out in order to find an explanation for this phenomenon. The early growth arrests of K. thermotolerans and T. delbrueckii were absent in single cultures of the two yeasts, and they seemed to be due neither to nutrient limitations nor to the presence of growth-inhibitory compounds. Rather, they seemed to be due to a cell-cell contact mechanism dependent on the presence of viable S. cerevisiae cells at high concentrations. These results contribute to an increased understanding of why K. thermotolerans and T. delbrueckii arrest growth before S. cerevisiae during wine fermentations.

Cell Communication↗

PubMed2003

Characterization of early deaths of non- Saccharomyces yeasts in mixed cultures with Saccharomyces cerevisiae.

The survival of Kluyveromyces thermotolerans and Torulaspora delbrueckii in mixed cultures with Saccharomyces cerevisiae was examined at low oxygen availability in a defined grape juice medium. In these fermentations, K. thermotolerans and T. delbrueckii died off earlier than S. cerevisiae, and K. thermotolerans and T. delbrueckii exhibited parabolic death kinetics. Furthermore, the early deaths seemed to be non-apoptotic in nature. In order to understand the mechanism causing the early deaths, various single- and mixed-culture fermentations were carried out. The early deaths could not be explained by nutrient depletion or the presence of toxic compounds. Rather, they seemed to be mediated by a cell-to-cell contact mechanism at high cell densities of S. cerevisiae, and to a lesser ability of K. thermotolerans and T. delbrueckii to compete for space, as compared to S. cerevisiae. These results contribute to an increased understanding of why K. thermotolerans and T. delbrueckii die off before S. cerevisiae in wine fermentations.

Cell Communication↗

PubMed1989

Influence of different factors on the induction of chromosome malsegregation in Saccharomyces cerevisiae D61.M by bavistan and assessment of its genotoxic property in the Ames test and in Saccharomyces cerevisiae D7.

Bavistan is known to be a potent inducer of chromosome malsegregation in Saccharomyces cerevisiae. The influence of different factors on the induction of chromosome malsegregation in S. cerevisiae D61.M was investigated. With both standard protocols used (16 h overnight incubation and cold treatment protocol) bavistan, in a concentration range of 2.5-20 micrograms/ml, induced malsegregants to the same extent. The frequencies of malsegregants obtained were not influenced by the plating volume used on selective medium. Induction of malsegregants and toxicity became stronger with increasing supplementation of the incubation medium with yeast extract and peptone. The effects of bavistan on chromosome malsegregation were more pronounced at 28 degrees C--the normal temperature for yeast growth--as compared to 33 and 37 degrees C. A study of the time dependence of the induction of chromosome loss showed that malsegregants can already be detected after 8 h and 1.5 h (second incubation period) using the incubation protocols without and with cold treatment, respectively. To clarify whether a selection towards malsegregants occurs, the growth of mixed cultures of red, cycloheximide-sensitive cells and white, cycloheximide-resistant, leucine-auxotrophic cells prepared at different ratios was compared. A strong selection towards red cells and against the malsegregants was observed. In addition, bavistan was tested for genotoxic activity in Salmonella (Ames test) and in yeast S. cerevisiae D7. No mutagenic activity was detected using S. cerevisiae D7 (gene conversion, reverse mutation, mitotic crossing-over) with and without rat-liver S9. In contrast bavistan induced histidine revertants in the frameshift strains TA1537, TA1538, TA97 and TA98 of Salmonella typhimurium after addition of an exogenous metabolic activation system.

Benzimidazoles↗

PubMed1983

Molecular cloning of the rDNA of Saccharomyces rosei and comparison of its transcription initiation region with that of Saccharomyces carlsbergensis.

We have cloned one complete repeating unit of rDNA from Saccharomyces rosei and determined its physical and genetic organization. Heteroduplex analysis of the rDNA units from S. rosei and S. carlsbergensis shows that the nontranscribed spacers are largely nonhomologous in sequence, whereas the transcribed regions are essentially homologous. We also determined the transcription initiation site for the 37S precursor RNA on S. rosei rDNA. Sequence comparison of the region surrounding the site of transcription initiation for the 37S RNA with the corresponding region of S. carlsbergensis revealed extensive homology from position -9 downstream into the external transcribed spacer. Very little homology was observed between position -9 and -55, but some homologous tracts are present upstream from position -55.

Base Sequence↗

PubMed2001

The effect of oxygen on the survival of non-Saccharomyces yeasts during mixed culture fermentations of grape juice with Saccharomyces cerevisiae.

AIMS: The effect of oxygen on the survival of Torulaspora delbrueckii and Kluyveromyces thermotolerans during mixed culture fermentations in grape juice with Saccharomyces cerevisiae was investigated. METHODS AND RESULTS: Fermentations were carried out in two simple fermentation systems differing in the availability of oxygen. At low available oxygen conditions, T. delbrueckii and K. thermotolerans began to die off after two days of mixed culture fermentation. In filtrates from 2-day-old mixed cultures, single cultures of T. delbrueckii and K. thermotolerans survived and actively produced ethanol to concentrations of approx. 65 and 70 g l-1, respectively, at low available oxygen conditions. Oxygen clearly increased the survival time and decreased the death rate of T. delbrueckii and K. thermotolerans in mixed cultures, whereas it did not affect the growth and survival of S. cerevisiae. CONCLUSION: Our results show that the deaths of T. delbrueckii and K. thermotolerans in mixed cultures at low available oxygen conditions are not due to toxic metabolites produced by the yeasts but rather to the lack of oxygen. Furthermore, they indicate that T. delbrueckii and K. thermotolerans are less tolerant to low available oxygen conditions than S. cerevisiae. SIGNIFICANCE AND IMPACT OF THE STUDY: Our study reveals new knowledge on the mechanisms underlying the succession of yeasts during wine fermentations. This knowledge may be of importance when creating defined, mixed starter cultures for the controlled production of wines with a wide range of flavour compositions.

Carbohydrate Metabolism↗