Showing papers on "Yeast published in 1986"
TL;DR: Fusion of the yeast CYC1 gene to lacZ in one of the vectors allowed detection of regulated expression of this gene when cells were grown under conditions of catabolite repression or derepression.
561 citations
Journal Article•
TL;DR: The central role of the redox couples NAD+/ NADH and NADP+/NADPH in the metabolism of sugars by yeasts is discussed in relation to energy metabolism and product formation.
Abstract: 1. SUMMARY The central role of the redox couples NAD+/ NADH and NADP+/NADPH in the metabolism of sugars by yeasts is discussed in relation to energy metabolism and product formation. Besides their physical compartmentation in cytosol and mitochondria, the two coenzyme systems are separated by chemical compartmentation as a consequence of the absence of transhydrogenase activity. This has considerable consequences for the redox balances of both coenzyme systems and hence for sugar metabolism in yeasts. As examples, the competition between respiration and fermentation of glucose, the Crabtree effect, the Custers effect, adaptation to anaerobiosis, the activities of the hexose monophosphate pathway, and the fermentation of xylose in yeast are discussed.
559 citations
TL;DR: In this article, the central role of the redox couples NAD+/NADH and NADP+ /NADPH in the metabolism of sugars by yeast is discussed in relation to energy metabolism and product formation.
Abstract: The central role of the redox couples NAD+/NADH and NADP+/NADPH in the metabolism of sugars by yeasts is discussed in relation to energy metabolism and product formation. Besides their physical compartmentation in cytosol and mitochondria, the two coenzyme systems are separated by chemical compartmentation as a consequence of the absence of transhydrogenase activity. This has considerable consequences for the redox balances of both coenzyme systems and hence for sugar metabolism in yeasts.
As examples, the competition between respiration and fermentation of glucose, the Crabtree effect, the Custers effect, adaptation to anaerobiosis, the activities of the hexose monophosphate pathway, and the fermentation of xylose in yeast are discussed.
544 citations
TL;DR: Yeast calmodulin is a small, heat-stable, acidic, retained by hydrophobic matrices in a Ca2-dependent manner, exhibits a pronounced Ca2+-induced shift in electrophoretic mobility, and binds 45Ca2+.
384 citations
TL;DR: A PRC1-SUC2 gene fusion that directs the synthesis in Saccharomyces cerevisiae of a hybrid polypeptide consisting of a 433-residue amino-terminal domain derived from the yeast vacuolar protease carboxypeptidase Y is constructed.
Abstract: We have constructed a PRC1-SUC2 gene fusion that directs the synthesis in Saccharomyces cerevisiae of a hybrid polypeptide consisting of a 433-residue amino-terminal domain derived from the yeast vacuolar protease carboxypeptidase Y (CPY; EC 3.4.16.1) and a 511-residue carboxyl-terminal domain derived from the secreted yeast enzyme invertase (EC 3.2.1.26). Fractionation data indicated that this amount of CPY primary sequence is sufficient to quantitatively divert invertase to the yeast vacuole. The phenotypic consequence of localizing active invertase to the vacuole has enabled us to select for mutants that "mislocalize" the hybrid protein to the cell surface. The corresponding mutations that lead to this effect are all trans-acting and recessive, and they define at least eight complementation groups. These vacuolar protein targeting (vpt) mutants also exhibit hybrid protein independent defects in wild-type CPY delivery to the yeast vacuole. Precursor forms of CPY accumulate in the mutants and are secreted into the yeast periplasm and extracellular medium. The vpt mutants should provide useful information pertaining to the mechanisms by which yeast cells regulate vacuolar protein traffic.
362 citations
Journal Article•
Abstract: The most probable mechanisms involved in the formation of carbonyl compounds, fusel alcohols, fatty acid esters, and free fatty acids by yeast in fermentation are discussed. Isopentyl alcohol, phenethyl alcohol, and their acetates in addition to the ethyl esters from hexanoate to laurate formed by different yeasts in sugar fermentation were determined. The bivariate distributions of the concentrations indicate that the mutual relationships of the compounds depend significantly on the yeast used and on the capacity of the yeast to produce alcohols and esters. The wine yeast used in this work produced smaller amounts of ethyl esters of fatty acids from acetic acid to octanoic acid under aerobic fermentation conditions than under strictly anaerobic conditions; on the other hand, ethyl laurate and ethyl 9-hexadecenoate were formed more abundantly in anaerobic conditions. Furthermore, qualitatively-similar flavor compositions were obtained in a Spanish sherry and a Finnish berry wine of sherry type. Hence, the formation of the most dominant compounds occurring in beverages depend more on the yeast selected than the raw materials used in fermentation.
351 citations
TL;DR: A method is described for the purification of chromosomal and plasmid DNA from the yeast Saccharomyces cerevisiae that is rapid, gives 75% of theoretical yield, and produces DNA that can be cut with restriction endonucleases.
312 citations
TL;DR: The gag-pol gene of HTLV-III (human T-lymphotropic virus), the virus linked to AIDS, was expressed in yeast, and processing of the gag precursor into proteins of the same size as those in the virion was observed.
Abstract: The gag-pol gene of HTLV-III (human T-lymphotropic virus), the virus linked to AIDS (acquired immune deficiency syndrome), was expressed in yeast, and processing of the gag precursor into proteins of the same size as those in the virion was observed. Processing of the gag gene in yeast cells mimics the process that naturally occurs in mammalian cells during maturation of virions. Therefore it was possible to perform mutational analysis of the virus genome to localize the gene that codes for the protease function to the amino terminal coding region of the pol gene. Since this region overlaps the gag gene, it is likely that ribosomal frameshifting occurs from gag to pol. Antibodies in all of the AIDS patients' sera tested recognized the yeast synthesized gag proteins, although the sera showed differences in relative reactivity to the individual gag proteins and the precursor. This yeast system should be valuable not only for production of viral proteins for diagnostic or vaccine purposes but also for analysis of the genetics and biochemistry of viral gene functions--parameters that are difficult to study otherwise with this virus.
308 citations
TL;DR: The in vitro synthesized precursor of the alpha-factor pheromone, prepro-alpha-factor, of Saccharomyces cerevisiae was translocated across yeast microsomal membranes in either a homologous or a wheat germ cell free system.
300 citations
TL;DR: It is reported here that PDE2 encodes a high-affinity cAMPosphodiesterase that shares sequence homology with animal cell phosphodiesterases and implies that the effects of RAS2Val19 are mediated through its changes in cAMP concentration.
Abstract: A gene, PDE2, has been cloned from the yeast Saccharomyces cerevisiae that, when present in high copy, reverses the phenotypic effects of RAS2Val19, a mutant form of the RAS2 gene that renders yeast cells sensitive to heat shock and starvation. It has previously been shown that the RAS proteins are potent activators of yeast adenylate cyclase. We report here that PDE2 encodes a high-affinity cAMP phosphodiesterase that shares sequence homology with animal cell phosphodiesterases. These results therefore imply that the effects of RAS2Val19 are mediated through its changes in cAMP concentration.
282 citations
TL;DR: It is suggested that ATP hydrolysis may supply the energy required for translocation of proteins across the endoplasmic reticulum.
Abstract: We describe an in vitro system with all components derived from the yeast Saccharomyces cerevisiae that can translocate a yeast secretory protein across microsomal membranes. In vitro transcribed prepro-alpha-factor mRNA served to program a membrane-depleted yeast translation system. Translocation and core glycosylation of prepro-alpha-factor were observed when yeast microsomal membranes were added during or after translation. A membrane potential is not required for translocation. However, ATP is required for translocation and nonhydrolyzable analogues of ATP cannot serve as a substitute. These findings suggest that ATP hydrolysis may supply the energy required for translocation of proteins across the endoplasmic reticulum.
TL;DR: The observed changes in immobilized cell metabolism and composition may have arisen from disturbance to the yeast cell cycle by the cell attachment, causing alterations in the normal pattern of yeast bud development, DNA replication, and synthesis of cell wall components.
Abstract: The kinetic properties of Saccharomyces cerevisiae immobilized on crosslinked gelatin were found to be substantially different from those of the suspended yeast. Batch fermentation experiments conducted in a gradientless reaction system allowed comparison of immobilized cell and suspended cell performance. The specific rate of ethanol production by the immobilized cells was 40-50% greater than for the suspended yeast. The immobilized cells consumed glucose twice as fast as the suspended cells, but their specific growth rate was reduced by 45%. Yields of biomass from the immobilized cell population were lower at one-third the value for the suspended cells. Cellular composition was also affected by mobilization. Measurements of intracellular poly-saccharide levels showed that the immobilized yeast stored larger quantities of reserve carbohydrates and contained more structural polysaccharide than did suspended cells. Flow cytometry was used to obtain DNA, RNA, and protein frequency functions for immobilized and suspended cell populations. These data showed that the immobilized cells have higher ploidy than cells in suspension. The observed changes in immobilized cell metabolism and composition may have arisen from disturbance to the yeast cell cycle by cell attachment, causing alterations in the normal pattern of yeast bud development, DNA replication, and synthesis of cell wall components.
TL;DR: A DNA fragment called suc1 has been found to rescue cells mutated in the cell cycle control gene cdc2 of the fission yeast Schizosaccharomyces pombe, suggesting that it interacts directly with the cDC2 gene function.
Abstract: A DNA fragment called suc1 has been found to rescue cells mutated in the cell cycle control gene cdc2 of the fission yeast Schizosaccharomyces pombe. The suppressing activity of suc1 is observed when it is present on a multicopy number plasmid. The gene does not hybridise to cdc2 and maps elsewhere in the genome. Its effect is cdc2 allele specific suggesting that it interacts directly with the cdc2 gene function.
TL;DR: Aromatic monomers in steam-exploded poplar were quantitatively analyzed and their influences on ethanol fermentation by the yeast Saccharomyces cerecisiae were studied as mentioned in this paper.
TL;DR: A homologous cell-free system has been derived from the yeast Saccharomyces cerevisiae that allows the translation, translocation, and glycosylation of the precursors of yeast mating factor alpha and invertase.
TL;DR: In this paper, the formation of 4-ethyl and 4-vinyl derivatives of guaiacol, phenol and syringol from ferulic acid,p-coumaric acid and sinapic acid, respectively, by Brettanomyces sp. in a synthetic medium was studied by gas chromatography-mass spectrometry.
Abstract: The formation of 4-ethyl and 4-vinyl derivatives of guaiacol, phenol and syringol from ferulic acid,p-coumaric acid and sinapic acid, respectively, byBrettanomyces sp. in a synthetic medium was studied by gas chromatography-mass spectrometry. Some of these metabolites possess strong spicy, smoke-like, medicinal, clove-like, woody or phenolic odours and their role as spoilage compounds in wine is discussed. Their formation appears to be characteristic of this yeast genus and its sporulating formDekkera, suggesting these yeasts are Pof+. This paper attempts to clarify the distinctive and ‘characteristic’ odours which have long been attributed toBrettanomyces yeast metabolism.
TL;DR: The fermentation by Candida shehatae and Pichia stipitis of xylitol and the various sugars which are liberated upon hydrolysis of lignocellulosic biomass was investigated and P. stipitis exhibited a higher volumetric rate and yield of ethanol production.
Abstract: The fermentation by Candida shehatae and Pichia stipitis of xylitol and the various sugars which are liberated upon hydrolysis of lignocellulosic biomass was investigated. Both yeasts produced ethanol from d-glucose, d-mannose, d-galactose and d-xylose. Only P. stipitis fermented d-cellobiose, producing 6.5 g·l-1 ethanol from 20 g·l-1 cellobiose within 48 h. No ethanol was produced from l-arabinose, l-rhamnose or xylitol. Diauxie was evident during the fermentation of a sugar mixture. Following the depletion of glucose, P. stipitis fermented galactose, mannose, xylose and cellobiose simultaneously with no noticeable preceding lag period. A similar fermentation pattern was observed with C. shehatae, except that it failed to utilize cellobiose even though it grew on cellobiose when supplied as the sole sugar. P. stipitis produced considerably more ethanol from the sugar mixture than C. shehatae, primarily due to its ability to ferment cellobiose. In general P. stipitis exhibited a higher volumetric rate and yield of ethanol production. This yeast fermented glucose 30–50% more rapidly than xylose, whereas the rates of ethanol production from these two sugars by C. shehatae were similar. P. stipitis had no absolute vitamin requirement for xylose fermentation, but biotin and thiamine enhanced the rate and yield of ethanol production significantly.
TL;DR: This chapter describes the regulation of glucose metabolism in Saccharomyces cerevisiae and related yeasts in view of recent data.
Abstract: Publisher Summary This chapter describes the regulation of glucose metabolism in Saccharomyces cerevisiae and related yeasts in view of recent data. Metabolism of growing yeasts depends on the yeast strain, the carbon source, and the physicochemical factors in the environment. Glucose breakdown in Saccharomyces cerevisiae and related yeast proceeds via different pathways. Principally respirative, fermentative (anaerobic), and respiro-fermentative glucose metabolism have to be considered, but growth on ethanol is also considered due to the diauxic growth pattern observed in batch cultures. Ethanol accumulated during the first growth phase with respiro-fermentative glucose catabolism is used for subsequent growth. The type of glucose metabolism depends on cultivation conditions. Respirative glucose catabolism is possible in feed-controlled systems only. The shift to respiro-fermentative glucose breakdown is governed by the respirative capacity of cells, and therefore occurs above a certain glucose-feed rate. Respiro-fermentative glucose metabolism is primarily the consequence of an overflow reaction at the level of pyruvate when respiration is saturated.
TL;DR: Two alpha-tubulin genes from the budding yeast Saccharomyces cerevisiae were identified and cloned by cross-species DNA homology and it was revealed that they encoded gene products of 447 and 445 amino acids that are highly homologous toalpha-tubulins from other species.
Abstract: Two alpha-tubulin genes from the budding yeast Saccharomyces cerevisiae were identified and cloned by cross-species DNA homology. Nucleotide sequencing studies revealed that the two genes, named TUB1 and TUB3, encoded gene products of 447 and 445 amino acids, respectively, that are highly homologous to alpha-tubulins from other species. Comparison of the sequences of the two genes revealed a 19% divergence between the nucleotide sequences and a 10% divergence between the amino acid sequences. Each gene had a single intervening sequence, located at an identical position in codon 9. Cell fractionation studies showed that both gene products were present in yeast microtubules. These two genes, along with the TUB2 beta-tubulin gene, probably encode the entire complement of tubulin in budding yeast cells.
TL;DR: The mechanism of translocation indicates that a protein of 18 000 daltons is capable of crossing an endoplasmic reticulum‐derived membrane post‐translationally, and seems to be restricted to prepro‐α‐factor in the yeast in vitro system.
Abstract: In an in vitro system comprising a yeast cell-free translation system, yeast microsomes and mRNA encoding prepro-alpha-factor, the translocation of this protein across the membrane of the microsomal vesicle and its glycosylation could b uncoupled from its translation. Such post-translational processing is dependent upon the presence of ATP in the system. It is not, however, affected by a variety of uncouplers, ionophores or inhibitors, including carbonyl cyanide m-chlorophenyl hydrazone (CCCP), valinomycin, nigericin, dinitrophenol (DNP), potassium cyanide (KCN) or N-ethyl maleimide (NEM). This mechanism of translocation is significant as it indicates that a protein of 18 000 daltons is capable of crossing an endoplasmic reticulum-derived membrane post-translationally. For the moment, this phenomenon seems to be restricted to prepro-alpha-factor in the yeast in vitro system. Neither invertase nor IgG chi light chain could be translocated post-translationally in yeast, nor was such processing observed for prepro-alpha-factor in a wheat germ system supplemented with canine pancreatic microsomes.
TL;DR: The rate of ethanol production per milligram of cell protein begins to decline in the early stage of batch fermentation before high concentrations of ethanol have accumulated in yeast extract-peptone medium, and the addition of magnesium restored the ability of glucose-reconstituted medium to support vigorous growth.
Abstract: The rate of ethanol production per milligram of cell protein begins to decline in the early stage of batch fermentation before high concentrations of ethanol have accumulated. In yeast extract-peptone medium (20% glucose), this initial decline appears to be related to growth and to result in part from a nutrient deficiency. The addition of yeast extract, peptone, and ashed preparations of these restored the ability of glucose-reconstituted medium (in which cells had been previously grown) to support vigorous growth. Magnesium was identified as the active component. Supplementing fermentations with 0.5 mM magnesium prolonged exponential growth, resulting in increased yeast cell mass. The addition of magnesium also reduced the decline in fermentative activity (micromoles of CO2 evolved per hour per milligram of protein) during the completion of batch fermentations. These two effects reduced the time required for the conversion of 20% glucose into ethanol by 1/3 with no measurable loss in ethanol yield (98% of theoretical maximum yield). It is possible that some of the reported beneficial effects of complex nutrients (soy flour and yeast extract) for ethanol production also result from the correction of a simple inorganic ion deficiency, such as magnesium.
TL;DR: The isolation, by an immunological method, of a second nuclear gene encoding citrate synthase (CIT2) is reported, which showed that disruption of both genes in the yeast genome was necessary to produce classical citate synthase-deficient phenotypes: glutamate auxotrophy and poor growth on rich medium containing lactate, a nonfermentable carbon source.
Abstract: The tricarboxylic acid cycle occurs within the mitochondria of the yeast Saccharomyces cerevisiae. A nuclear gene encoding the tricarboxylic acid cycle enzyme citrate synthase has previously been isolated (M. Suissa, K. Suda, and G. Schatz, EMBO J. 3:1773-1781, 1984) and is referred to here as CIT1. We report here the isolation, by an immunological method, of a second nuclear gene encoding citrate synthase (CIT2). Disruption of both genes in the yeast genome was necessary to produce classical citrate synthase-deficient phenotypes: glutamate auxotrophy and poor growth on rich medium containing lactate, a nonfermentable carbon source. Therefore, the citrate synthase produced from either gene was sufficient for these metabolic roles. Transcription of both genes was maximally repressed in medium containing both glucose and glutamate. However, transcription of CIT1 but not of CIT2 was derepressed in medium containing a nonfermentable carbon source. The significance of the presence of two genes encoding citrate synthase in S. cerevisiae is discussed.
TL;DR: To determine the fraction of the yeast Saccharomyces cerevisiae genome that is required for normal cell growth and division, diploid strains that were heterozygous for random single disruptions were constructed and the phenotype of the resulting haploid strains was examined.
TL;DR: A glucoamylase gene has been cloned from a Rhizopus genomic DNA library using synthetic oligonucleotides corresponding to the amino acid sequence of the glu coamylasing gene by recombination of both genes.
Abstract: A glucoamylase gene has been cloned from a Rhizopus genomic DNA library using synthetic oligonucleotides corresponding to the amino acid sequence of the glucoamylase. Since this glucoamylase gene was not expressed in yeast cells, we have cloned a glucoamylase gene from a cDNA library prepared from Rhizopus mRNA. Sequence analysis of both glucoamylase genes revealed that the genomic gene contained 4 intervening sequences and the cDNA gene lacked 145 nucleotides corresponding to the N-terminal region. The glucoamylase consists of 604 amino acids including a putative signal peptide and its molecular weight was calculated to be 65, 000. The glucoamylase gene to be expressed in yeast cells was constructed by recombination of both genes. The yeast cells containing this constructed glucoamylase gene secreted the glucoamylase into the culture fluid and grew at almost the normal rate on a medium containing starch as the sole carbon source.
TL;DR: In this paper, the dominant sugar in red oak acid prehydrolysate, xylose, was fermented to ethanol, and a maximal ethanol concentration of 9.9 g l −1 was obtained from an acid pre-hydrolyate containing 21.7 g l−1 of xyloses.
TL;DR: The ten-nucleotide-long sequence have been omitted while sequencing the 18S rRNA gene from yeast Saccharomyces cerevisiae and this GAAGAUGAUC sequence and some other minor corrections are reintroduced into the yeast 18SrRNA primary structure.
TL;DR: The killer phenomenon, which was previously considered to be restricted to yeasts, was found to occur among unrelated microorganisms.
Abstract: The killer effect of 36 Hansenula, Pichia, Saccharomyces, and Candida species on 26 hyphomycetes isolates, 1 isolate of the achlorophyllous microorganism Prototheca, 4 isolates of the lipophilic yeast Malassezia, 1 isolate of the aerobic actinomycete Nocardia, and 19 isolates of bacteria was studied. The killer phenomenon, which was previously considered to be restricted to yeasts, was found to occur among unrelated microorganisms.
TL;DR: The results strongly suggest that glucose stimulates inositolphospholipid turnover, Ca2+ mobilization, and subsequent cell proliferation in a manner similar to that of growth factors with mammalian cells, and that RAS-encoded proteins are involved in regulation of this glucose-induced inositylinositolipids turnover in yeast.
Abstract: Incubation of yeast Saccharomyces cerevisiae at very low (0.02%) glucose levels led to arrest of the cell cycle at the G0/G1 phase. Readdition of glucose to these "starved" yeast resulted in cell proliferation. In glucose-starved yeast, glucose stimulated 32P incorporation into phosphatidic acid, phosphatidylinositol, phosphatidylinositol monophosphate, and phosphatidylinositol bisphosphate but not into phosphatidylethanolamine and phosphatidylcholine. Preincubation of yeast with [3H]inositol and subsequent exposure to glucose resulted in rapid formation of [3H]inositol monophosphate and [3H]inositol trisphosphate, presumably derived from phosphatidylinositol and phosphatidylinositol bisphosphate. Under similar conditions, glucose elicited both efflux and influx of Ca2+ in yeast. Glucose-induced 32P incorporation into inositolphospholipids and formation of [3H]inositol phosphates were more pronounced in RAS-related mutants such as ras1, ras1 ras2 bcy1, and RAS2Val19 than in the wild-type strain. These results strongly suggest that glucose stimulates inositolphospholipid turnover, Ca2+ mobilization, and subsequent cell proliferation in a manner similar to that of growth factors with mammalian cells, and that RAS-encoded proteins are involved in regulation of this glucose-induced inositolphospholipid turnover in yeast.
TL;DR: It is found that the cloned fission yeast TOP2 gene can complement the budding yeast top2 mutation, although the fission fungi TOP2 protein sequence is only 50% homologous to the recently determined sequence of budding yeast.
Abstract: We have determined the complete nucleotide sequence of a 5.3-kb long genomic DNA fragment of the fission yeast Schizosaccharomyces pombe that encodes DNA topoisomerase II. It contains a 4293 bp long single open reading frame. The predicted polypeptide has 1431 residues (mol. wt 162,000) and shows three characteristic domains; the large C-terminal region, which consists of alternating acidic-basic stretches and might be a chromatin-binding domain, the NH2 half domain homologous to the ATP-binding gyrB subunit of bacterial gyrase and the central-to-latter part which is homologous to the NH2 domain of the catalytic gyrA subunit, suggesting a possible evolutionary consequence of the gene fusion of the bacterial gyrase subunits into the eucaryotic DNA topoisomerase II gene. We have found that the cloned fission yeast TOP2 gene can complement the budding yeast top2 mutation, although the fission yeast TOP2 protein sequence is only 50% homologous to the recently determined sequence of budding yeast (J.C. Wang, personal communication). Conversely, the budding yeast TOP2 gene can complement the fission yeast top2 mutations, indicating that their DNA topoisomerase II genes are functionally exchangeable.
TL;DR: In yeast, the ends of the chromosomes (telomeres) terminate in repeated poly(C1-3A) sequences, despite their different primary sequences, yet does not bind specifically to telomeric repeats, such aspoly(C4A2), poly( C4A4), and poly (C 1-8T) from other lower eukaryotes.
Abstract: In yeast, the ends of the chromosomes (telomeres) terminate in repeated poly(C1-3A) sequences. We have identified a yeast activity that binds specifically to these poly(C1-3A) repeats. An agarose gel binding assay was used to detect and characterize this activity in cell extracts using both cloned telomere DNA and yeast genomic DNA as substrates. The activity appears to bind specifically to poly(C1-3A) sequences, despite their different primary sequences, yet does not bind specifically to telomeric repeats, such as poly(C4A2), poly(C4A4), and poly (C1-8T) from other lower eukaryotes.