Showing papers on "Yeast published in 1980"
TL;DR: The DNA sequence of a 1.45 kb EcoRI fragment from the yeast (Saccharomyces cerevisiae) TRP1 region has been determined and theTRP1 gene has been located on the fragment by analysis of potential initiation and termination codons in the DNA sequence.
547 citations
TL;DR: Recombinant plasmids capable of complementing cdc28 mutations were isolated by transformation of a cDC28ts strain and selection for clones capable of growth at the restrictive temperature, confirming the identity of the cloned sequences.
Abstract: cdc28, one of several genes required for cell division in the yeast Saccharomyces cerevisiae, has been isolated on recombinant plasmids. A recombinant plasmid pool containing the entire yeast genome was constructed by partial digestion of yeast DNA with the four-base recognition restriction endonuclease Sau3A to give the equivalent of random fragments, size selection on sucrose gradients, and introduction of the fragments into the yeast vector YRp7 by use of the homology of Sau3A ends with those generated in the vector by cleavage with BamHI. Recombinant plasmids capable of complementing cdc28 mutations were isolated by transformation of a cdc28ts strain and selection for clones capable of growth at the restrictive temperature. Plasmids responsible for complementing the cdc28ts phenotype were shown to recombine specifically with the chromosomal cdc28 locus, confirming the identity of the cloned sequences. In addition, one of the recombinant plasmids was capable of complementing a mutation in tyr1, a gene genetically linked to cdc28. This method of gene isolation and identification should be applicable to all yeast genes for which there are readily scorable mutants.
430 citations
TL;DR: A versatile stain has been developed for demonstrating pollen, fungal hyphae and spores, bacteria and yeasts, and the concentration of lactic acid in the stain mixture plays an important role in the differential staining of pollen.
Abstract: A versatile stain has been developed for demonstrating pollen, fungal hyphae and spores, bacteria and yeasts. The mixture is made by compounding in the following order: ethanol, 20 ml; 1% malachite...
357 citations
TL;DR: The nucleotide sequence of the yeast DNA plasmid from Saccharomyces cerevisiae strain A364A D5 has been determined and possible functions are suggested, and attributes of an improved vector for cloning foreign DNAs in yeast are discussed.
Abstract: The nucleotide sequence of the yeast DNA plasmid (2µ circle) from Saccharomyces cerevisiae strain A364A D5 has been determined. The plasmid contains 6,318 base pairs, including two identical inverted repeats of 599 base pairs. Possible functions are suggested, and attributes of an improved vector for cloning foreign DNAs in yeast are discussed.
333 citations
TL;DR: A partially purified fraction of the enzymes, easily obtained with a single purification step, had a high lytic specific activity and was superior to commercial preparations in regard to nuclease, protease, and chitinase contamination.
Abstract: Yeast lytic activity was purified from the culture supernatant of Oerskovia xanthineolytica grown on minimal medium with insoluble yeast glucan as the carbon source. The lytic activity was found to consist of two synergistic enzyme activities which copurified on carboxymethyl cellulose and Sephadex G-150, but were resolved on Bio-Gel P-150. The first component was a beta-1,3-glucanase with a molecular weight of 55,000. The K(m) for yeast glucan was 0.4 mg/ml; that for laminarin was 5.9 mg/ml. Hydrolysis of beta-1,3-glucans was endolytic, yielding a mixture of products ranging from glucose to oligomers of 10 or more. The size distribution of products was pH dependent, smaller oligomers predominating at the lower pH. The glucanase was unable to lyse yeast cells without 2-mercaptoethanol or the second lytic component, an alkaline protease. Neither of these agents had any effect on the glucanase activity on polysaccharide substrates. The protease had a molecular weight of 30,000 and hydrolyzed Azocoll and a variety of denatured proteins. The enzyme was unusual in that it had an affinity for Sephadex. Although the activity was insensitive to most protease inhibitors, it was affected by polysaccharides; yeast mannan was a potent inhibitor. The enzyme did not have any mannanase activity, however. Neither pronase nor trypsin could substitute for this protease in promoting yeast cell lysis. A partially purified fraction of the enzymes, easily obtained with a single purification step, had a high lytic specific activity and was superior to commercial preparations in regard to nuclease, protease, and chitinase contamination. Lyticase has been applied in spheroplast, membrane, and nucleic acid isolation, and has proved useful in yeast transformation procedures.
286 citations
TL;DR: An immunological screening technique has been used for the detection of a specific antigen-producing clone in a bank of bacterial colonies containing hybrid plasmids, identified an Escherichia coli colony, containing a yeast DNA insert in plasmid ColE1, that produces antigen which combines with antibody directed against purified yeast 3-phosphoglycerate kinase.
217 citations
TL;DR: The living cells in the gel were maintained at the high level of 109 per ml of gel and continuous production of ethanol using the complete medium containing 10% glucose was carried out with a retention time of 1 h.
Abstract: Immobilized growing yeast cells were prepared in kappa-carra-geenan gel. Gel beads containing a small number of cells were incubated in a complete medium. The cells grew very well in the gel and the number of living cells per ml of gel increased to over 10 times that of free cells per ml of culture medium. After growing in the gel, the cells formed a dense layer of cells near the gel surface and produced large amounts of ethanol. The conditions for continuous production of ethanol using immobilized growing yeast cells were investigated. The supply of appropriate nutrients for growth was essential for the continuous production. The living cells in the gel were maintained at the high level of 109 per ml of gel and continuous production of ethanol using the complete medium containing 10% glucose was carried out with a retention time of 1 h. In this operation, a stable steady state was maintained for longer than 3 months. The ethanol concentration was 50 mg/ml and the conversion of glucose utilized to ethanol produced was almost 100% of the theoretical yield.
191 citations
TL;DR: The synthesis of the major linkage found in yeast cell wall structural polysaccharides, glucosyl-beta-(1 leads to 3)-glucosyl, was studied with a membrane preparation from Saccharomyces cerevisiae and suggests localized and reversible activation of the enzyme during the cell cycle.
191 citations
TL;DR: Labeling kinetics of RNA and measurements of the polyadenylated fractions showed that RNA hybridizing to the pBR322 plasmid was polyadenyated to the same extent as RNA hybridization to the ura3+ gene.
164 citations
TL;DR: The present work presents a meta-analysis of yeast chromosomol genes that highlights the importance of knowing the carrier and removal status of canine coronavirus, as a source of infection for other animals.
Abstract: PROSPECTIVES AND SUMMARY 845 GENETIC ANALYSIS IN yEAST 846 Tetrad Analysis 846 New Advances in Genetic Analysis 850 Construction of recombinant DNA molecules 850 Identification of cloned genes 850 Yeast transformation 854 ORGANIZATION AND REPLICATION OF THE YEAST CHROMOSOMES 856 Chromosome Structure 858 Chromosome Replication 860 Structure of replicating nuclear DNA molecules 860 Control of DNA synthesis 862 Yeast DNA polymerases .. ... ... ........ ... ..... 864 ANALYSIS OF SPECIFIC CLASSES OF CHROMOSOMAL GENES 864 Structural Analysis of Single-Copy Genes 865 Repeating Chromosomal Genes 866 Yeast ribosomal DNA genes 866 Structure of yeast tRNA genes 867 Other repeating yeast chromosomol genes 868 STRUCTURE AND REPLICATION OF THE 2p.m PLASMIDS 869 CONCLUSIONS AND FUTURE PROSPECTS 871
142 citations
TL;DR: A procedure is presented for the rapid isolation of mitochondrial DNA (mtDNA) in high yield from Saccharomyces cerevisiae by a single cycle of bisbenzimide-CsCl buoyant density centrifugation.
TL;DR: The use of the newly developed technique of yeast transformation to isolate recombinant plasmids that contain the structural gene for ADHI (ADC1) is described, selected from a pool of plasmid containing yeast DNA by virtue of their ability to complement the growth defect of yeast strains completely lacking ADH activity.
Abstract: Alcohol dehydrogenase (ADH) catalyses the interconversion of acetaldehyde and ethanol. This reaction is the final step during alcoholic fermentation in yeast and the initial step in the metabolism of ethanol in a wide variety of organisms. The regulation of ADH is now being studied in several organisms, notably yeast1–4, Drosophila5, maize6 and man7,8. These organisms contain multiple ADH isozymes which are differentially expressed. The budding yeast Saccharomyces cerevisiae is particularly suitable for study because of the potential for genetic analysis of enzyme regulation and structure. Yeast possesses three distinct ADH isozymes2. In the presence of glucose ADHI (the ‘classical’ yeast ADH) is expressed whereas ADHII is repressed. A third enzyme, mADH, is associated with mitochondria. Mutants lacking each of these enzymes have been isolated by virtue of their greater tolerance to allyl alcohol1, which is converted by ADH to the toxin, acrolein. There is no evidence as to whether the regulation of any one of these enzymes is under transcriptional control. Availability of recombinant DNA clones containing the respective genes would be of obvious value in resolving this problem. We describe here the use of the newly developed technique of yeast transformation9–11 to isolate recombinant plasmids that contain the structural gene for ADHI (ADC1). These plasmids were selected from a pool of plasmids containing yeast DNA by virtue of their ability to complement the growth defect of yeast strains completely lacking ADH activity. Using DNA sequencing, they have been shown to contain the structural gene for ADHI.
TL;DR: The rate of production of ethanol from glucose by Saccharomyces cerevisia in a packed column is examined and these rates are compared with those for the same reaction in a stirred reactor.
Abstract: Ethanol is produced from cellulosic material such as corn stover by treating the cellulosic material in a first hydrolysis stage with a dilute acid solution to hydrolyze pentosans to xylose, separating solids from the resultant hydrolysate, treating the solids in a second hydrolysis stage with a concentrated acid solution to hydrolyze hexosans to glucose, and fermenting the glucose to ethanol by passing a solution of the glucose over a fixed film of yeast prepared by attaching yeast with a polyfunctional agent to a proteinaceous material coated on a solid support. The use of a first hydrolysis stage avoids the production of furfural which is toxic and inhibits yeast fermentation. Xylose produced from the first stage may also be fermented to ethanol with the fixed film of yeast.
TL;DR: During growth on wood beta-1,4-xylans the yeast Cryptococcus albidus produced at least two enzymes which convert the polysaccharide to xylose catabolized by the cells, and both crude and purified beta-xylanase were unable to hydrolyze cellulose or its soluble derivatives.
Abstract: During growth on wood β-1,4-xylans the yeast Cryptococcus albidus produced at least two enzymes which convert the polysaccharide to xylose catabolized by the cells. The enzyme almost completely secreted into culture fluid was identified as an endo-1,4-β-xylanase.The function of the extracellular β-xylanase is to hydrolyze xylan to oligosaccharides, mainly to xylobiose and xylotriose, which enter the cell where they are split by the second identified enzyme, a cell-bound β-xylosidase (xylobiase). Aryl β-xylosidase activity detected in the culture fluid was shown to be due to low affinity of β-xylanase for p-nitrophenyl β-D-xylopyranoside. This property of β-xylanase was preserved after purification of the enzyme by chromatography on DEAE-cellulose, CM-Sephadex and Biogel A 1.5 m or Biogel P 100. Purified β-xylanase exhibited certain microheterogeneity after polyacrylamide gel electrophoresis.
Both extracellular β-xylanase and intracellular β-xylosidase were produced in much lower amounts by the cells grown on glucose than by the cells grown on xylan. This suggested that they are not produced constitutively.
The investigated strain was not able to grow on cellulose and the crude and purified β-xylanase were unable to hydrolyze cellulose or its soluble derivatives.
TL;DR: Inhibition studies with these subunit-specific antibodies showed that the common subunits are not always similarly exposed to antibody attack within the three enzymes, which indicates the structural similarity, organization and evolution of eukaryotic RNA polymerases.
TL;DR: The integrity of the ATPase activity is not appreciably modified by its solubilization by lysolecithin and the subsequent purification and lipid reconstitution procedures, indicating classic Michaelis-Menten mechanisms and noninteracting binding sites for the substrate MgATP.
TL;DR: An examination of the difference spectra observed upon the addition of carbon tetrachloride, halothane and trichloroethylene to whole-cell or microsomal suspensions of yeast suggested that cytochrome P-450 mediated the metabolism of the hydrocarbons tested to cytotoxic and genetically active compounds.
Abstract: Cells of Saccharomyces cerevisiae , harvested from log-phase cultures, contain cytochrome P-450 and are capable of metabolizing promutagens to genetically active products. The activities of 7 halogenated aliphatic hydrocarbons in the yeast system have been investigated. All of the compounds tested (methylene chloride, halothane, chloroform, carbon tetrachloride, trichloroethylene, tetrachloroethylene and s -tetrachloroethane) induced mitotic gene convertants and recombinants and, to a lesser extent, gene revertants when incubated with logphase cells of the yeast strain D7. An examination of the difference spectra observed upon the addition of carbon tetrachloride, halothane and trichloroethylene to whole-cell or microsomal suspensions of yeast suggested that cytochrome P-450 mediated the metabolism of the hydrocarbons tested to cytotoxic and genetically active compounds.
TL;DR: When the cells of a lager brewing yeast Saccharomyces uvarum (carlsbergensis) were grown in minimal media containing sucrose and a non-metabolized sugar sorbitol, significant levels of intracellular ethanol were obtained.
Abstract: When the cells of a lager brewing yeast Saccharomyces uvarum (carlsbergensis) were grown in minimal media containing sucrose and a non-metabolized sugar sorbitol, significant levels of intracellular ethanol were obtained. Intracellular ethanol concentration decreased as the osmotic pressure of the medium was lowered and the proportion of extracellular ethanol increased. A reduction in cell viability occurred when there were high levels of intracellular ethanol. The total amount of glycerol produced increased with increased osmotic pressure, but glycerol diffused out of the cells faster than ethanol.
TL;DR: The Escherichia coli R factor-derived chloramphenicol resistance (camr) gene is functionally expressed in the yeast Saccharomyces cerevisiae and segregate with the yeast marker LEU2, carried by the transforming plasmid, during both mitotic growth and meiotic division.
Abstract: The Escherichia coli R factor-derived chloramphenicol resistance (camr) gene is functionally expressed in the yeast Saccharomyces cerevisiae. the gene was introduced by transformation into yeast cells as part of a chimeric plasmid, pYT11-LEU2, constructed in vitro. The plasmide vector consists of the E. coli plasmid pBR325 (carrying the camr gene), the yeast 2-micron DNA plasmid, and the yeast LEU2 structural gene. Yeast cells harboring pYT11-LEU2 acquire resistance to chloramphenicol and cell-free extracts prepared from such cells contain chloramphenicol acetyltransferase (acetyl-CoA: chloramphenicol 3-O-acetyltransferase, EC 2.3.1.28), the enzyme specified by the camr gene in E. coli. Resistance to chloramphenicol and the presence of chloramphenicol acetyltransferase activity segregate with the yeast marker LEU2, carried by the transforming plasmid, during both mitotic growth and meiotic division.
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TL;DR: A library of DNA from the yeast was constructed in phage lambda Charon 4 vector and then screened by differential plaque filter hybridization for genes induced by phosphate starvation to examine mRNA levels of these genes in regulatory mutants of acid phosphatase.
Abstract: A library of DNA from the yeast, Saccharomyces cerevisiae, was constructed in phage lambda Charon 4 vector and then screened by differential plaque filter hybridization for genes induced by phosphate starvation. Two EcoRI fragments of 7.9 and 5.0 kilobase pairs that contained such genes were isolated. These cloned fragments may each carry one of the several copies of the genes for the repressible acid phosphatase of yeast. The fragments were use to examine mRNA levels of these genes in regulatory mutants of acid phosphatase.
TL;DR: The rate of N-glycosylation in vitro is approximately 25-fold faster with Glc3Man9GlcNAc2-P- P-dolichol than with Man9Gl cNAc 2-P -P-D-dollichol, and the apparent Km value for the glucosylation species is 75 nM, while that for the unglucosylated glycolipid is 55 nM.
TL;DR: The extent of change in cytoplasmic proteins which accompanies yeast-to-mycelium morphogenesis of Candida albicans was analyzed by two-dimensional gel electrophoresis and suggests that proteins unique to either phase may serve a regulatory function.
Abstract: The extent of change in cytoplasmic proteins which accompanies yeast-to-mycelium morphogenesis of Candida albicans was analyzed by two-dimensional gel electrophoresis. Pure cultures of yeasts and true hyphae (i.e., without concomitant production of pseudohyphae) were grown in a synthetic low-sulfate medium. The two strains selected for this study were strain 4918, which produces pure mycelial cultures in low-sulfate medium at 37 degrees C and yeast cells at 24 degrees C, and strain 2252, which produces yeast cells exclusively at both 24 and 37 degrees C in low-sulfate medium. The proteins of both strains were labeled at both temperatures with [35S]sulfate, cytoplasmic fractions were prepared by mechanical disruption and ultracentrifugation, and the labeled proteins were analyzed by two-dimensional electrophoresis. Highly reproducible protein spot patterns were obtained which defined hundreds of proteins in each extract. Ten protein spots were identified on the two-dimensional gels of the 4918 mycelial-phase extract which were not present in the 4918 yeast-phase extract. These proteins appeared to be modifications of preexisting yeast-phase proteins rather than proteins synthesized de novo in the mycelial cells because 5 were absorbed by rabbit anti-yeast-phase immunoglobulin and each of the 10 was also present in extracts of strain 2252 grown at 24 and 37 degrees C, indicating that they were neither unique to filamentous cells nor sufficient for induction or maintenance of the mycelial morphology. Thirty-three proteins were identified in the 4918 yeast-phase extract which were not present in the 4918 mycelial-phase extract. Pulse-chase experiments revealed the synthesis of new proteins during yeast-to-mycelial conversion, but none of these was unique to mycelial cells. No differences in the major cytoplasmic proteins of any of the yeast- or mycelial-phase extracts were identified. This finding suggests that the major structural proteins of the cytoplasm are not extensively modified and argues instead that proteins unique to either phase may serve a regulatory function.
TL;DR: In grape must of high sugar concentration, yeast growth, the viability rate of "resting" yeast cells, and fermentation activity were stimulated under certain conditions of aeration and temperature, which might be interpreted as being a result of the yeast cell sterol content.
Abstract: In grape must of high sugar concentration, yeast growth, the viability rate of "resting" yeast cells, and fermentation activity were stimulated under certain conditions of aeration and temperature. This stimulation might be interpreted as being a result of the yeast cell sterol content. The addition of certain sterols to the fermenting medium was able to increase this sterol content. According to aeration conditions of the medium, which determined the sterol content of yeasts, the sterols added in the medium acted as (i) growth factors, (ii) fermentation inhibitors, and (iii) survival factors for the yeast.
TL;DR: The present contribution is the first description of a peroxisomal amine oxidase, which catalyses the oxidation of methylamine by molecular oxygen into ammonia, formaldehyde and hydrogen peroxide in organisms studied.
Abstract: The metabolism of methylamine as the nitrogen source for growth of the non-methylotrophic yeast Candida utilis and the methylotrophic yeast Hansenula polymorpha was investigated. Grwoth of both organisms in media with glucose and methylamine was associated with the presence of an amine oxidase in these cells. The enzyme catalyses the oxidation of methylamine by molecular oxygen into ammonia, formaldehyde and hydrogen peroxide and it is considered to be the key enzyme in methylamine metabolism in the organisms studied.
TL;DR: Two strains are suggested, one forming yeasts and one forming true mycelium in low-sulfate medium at 37 degrees C, thus providing controls for both strain variation and for molecular changes induced by environmental change but unrelated to morphogenesis.
Abstract: A low-sulfate synthetic medium was developed in which pure cultures of yeast- and mycelial-phase Candida albicans could be cultivated for investigations of the molecular biology of dimorphism. The medium contained ammonium ions, phosphate buffer, salts, glucose, and biotin. Morphogenesis was found to be dependent upon the strain of C. albicans. Of six strains tested in the low-sulfate medium at 37 degrees C, three formed mixed cultures of yeasts, true mycelium and pseudomycelium, two formed pure cultures of true mycelium, and one maintained yeast growth. All six strains produced pure cultures of yeasts at 24 degrees C. The buffering capacity of the medium maintained the pH at 6.9 even at high-density cell growth. The low concentration of sulfate and the absence of amino acids in the medium provided conditions in which to radiolabel cellular constituents with [35S]sulfate. For molecular investigations, the use of two strains is suggested, one forming yeasts and one forming true mycelium in low-sulfate medium at 37 degrees C, thus providing controls for both strain variation and for molecular changes induced by environmental change but unrelated to morphogenesis.
TL;DR: Methyl β-D-xylopyranoside was found to serve as a nonutilizable inducer of β-xylanase, and the enzyme induced by the glycoside appears to be identical with that produced by the cells during growth on xylan.
Abstract: Extracellular endo-1,4-β-xylanase synthesis in the yeast Cryptococcus albidus is largely inducible. During growth on wood xylans the yeast produces the enzyme in amounts two orders of magnitude greater than on other carbon sources, including xylose. The enzyme can be induced in washed glucose-grown cells by xylan and β-1,4-xylooligosaccharides. Among the oligosaccharides only xylobiose was not degraded extracellularly, therefore it appears to be natural inducer of the enzyme. Xylobiose as a metabolisable inducer is effective at low concentrations and constant availability to cells. At high concentration of xylobiose the inductive effect is less pronounced because of catabolic repression by degration products. Methyl β-D-xylopyranoside was found to serve as a nonutilizable inducer of β-xylanase. The enzyme induced by the glycoside appears to be identical with that produced by the cells during growth on xylan.