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Research was conducted to examine the ability of certain yeast strains to prevent spoilage of maize silage fermentation due to aerobic degradation of lactic acid. Results show yeast strain, PCK27, which is unable to grow on lactic acid, improves the aerobic stability of the process.

Chromosome VI of Saccharomyces cerevisiae has been shown to contain nine origins of DNA replication differing in initiation frequency, and replicating sequentially during the cell-cycle S-phase. There are also links between activation of the multiple origins and regulation of S-phase progress. The effects of methyl methane sulphonate (MMS), a DNA-damaging agent were studied, and it was found that MMS slows S-phase progression, as well as selectively blocking initiation from late origins.

The mother cell and bud in Saccharomyces cerevisiae are linked by a narrow neck, which is closed during cytokinesis. The role of the actomyosin ring in the process is reported, and it was found that the contractile actomyosin ring is not essential for cytokinesis which can be completed by a process that requires septin function, facilitated by F-actin.

A Saccharomyces cerevisiae glycerol-3-phosphate dehydrogenase mutant, gpd2-delta, shows a large decrease in the carbon dioxide evolution rate (CER) after transition from aerobic to anaerobic conditions. Another mutant, gpd1-delta, and wild-type cells show no change in the CER under anaerobic conditions during the exponential growth phase. The gpd1-delta gpd2-delta double mutant exhibits a decrease in the fermentation rate on transition to anaerobic conditions. The addition of acetoin under anaerobic conditions enhances the CER value due to a its reduction to butanediol.

The rate of glucose and xylose metabolism is higher in aerobic than in anoxic cell suspensions of Pichia stipitis, while ethanol production is slightly lower. This may account for the higher intracellular pH in P. stipitis in the presence of oxygen. However, Saccharomyces cerevisiae shows identical rates of glucose consumption and ethanol production under both conditions. Agarose-immobilized S. cerevisiae shows an increase in the rate of carbohydrate metabolism, but P. stipitis does not. P-31 and C-13 nuclear magnetic resonance spectroscopy is used to analyze the two species.

The septins present in different genes of Saccharomyces cerevisiae participate in the spore formation process. A new septin-gene, SPR3, has been isolated that encodes a fifth septin of S. cerevisiae. Results of gene deletion vary for different genetic backgrounds, suggesting overlapping functions for the septins. The septin Spr3p has two predicted coiled-coil domains separated by about 50 amino acids near its COOH terminus where most other septins contain a continuous coiled-coildomain.

The highly active xylanase (XynA) signal peptide of the fungus Aureobasidium pullulans is an efficient posttranslational processor of the xynA product and helps in the secretion of xylanase in Saccharomyces cerevisiae. The peptide is about three times more effective than the S. cerevisiae invertase or mating alpha-factor pheromone. XynA increases the efficiency of secretion while transcription and translation are unaffected. The S. cerevisiae clones with cDNA and genomic DNA have no activity in culture media and a little xylanase activity in cell-associated fractions.

An evaluation of the resolution of the polymerase chain reaction (PCR)-mediated typing techniques indicates that no single technique is sufficient for the discrimination of Saccharomyces cerevisiae strains. The random amplified polymorphic assay using the OPA-11 and 24 primers and PCR fingerprinting allow for the study of genetic diversity between the strains. Restriction enzyme analysis of PCR-amplified units is useful in determining the intraspecies-specific patterns.

The BED1 gene of Saccharomyces cerevisiae is essential for polarized growth and efficient budding. BED1 encodes an alpha-1,2-galactosyltransferase homologue, an integral membrane protein present in the endoplasmic reticulum. The BED1 mutant cell neither exhibits bud emergence nor possesses the ellipsoidal shape specific for wild-types, and are bigger than the wild-type cells. An intact morphogenesis checkpoint control helps in the survival of the mutants. Polarized growth and bud emergence depend on the glycosylation of specific proteins or lipids.

The interactions established at the 5'-splice site during spliceosome assembly are likely to be important for both precise recognition of the upstream intron boundary and for positioning this site in the active center of the spliceosome. Definition of the RNA-RNA and the RNA-protein interactions at the 5' splice site would be facilitated by the use of a small substrate amenable to modification during chemical synthesis. We describe a trans-splicing reaction performed in Saccharomyces cerevisiae extracts in which the 5' splice site and the 3' splice site are on separate molecules. The RNA contributing the 5' splice site is only 20 nucleotides long and was synthesized chemically. The trans-splicing reaction is accurate and has the same sequence, ATP, and [Mg.sup.2+] requirements as cis-splicing. We also report how deoxy substitutions around the 5'-splice site affect trans-splicing efficiency.

Deletion of the clathrin heavy-chain gene, CHC1, in the budding yeast Saccharomyces cerevisiae results in growth, morphological, and membrane trafficking defects, and in some strains chc1-[delta] is lethal. A previous study identified five genes which, in multicopy, rescue inviable strains of Chc- yeast. Now we report that one of the suppressor loci, BMH2/SCD3, encodes a protein of the 14-3-3 family. The 14-3-3 proteins are abundant acidic proteins of [is nearly equal to] 30 kDa with numerous isoforms and a diverse array of reported functions. The Bmh2 protein is >70% identical to the mammalian [epsilon]-isoform and >90% identical to a previously reported yeast 14-3-3 protein encoded by BMH1. Single deletions of BMH1 or BMH2 have no discernible phenotypes, but deletion of both BMH1 and BMH2 is lethal. High-copy BMHI also rescues inviable strains of [Chc.sup.-] yeast, although not as well as BMH2. In addition, the slow growth of viable strains of [Chc.sup.-] yeast is further impaired when combined with single bmh mutations, often resulting in lethality. Overexpression of BMH genes also partially suppresses the temperature sensitivity of the cdc25-1 mutant, and high-copy TPK1, encoding a cAMP-dependent protein kinase, restores [Bmh.sup.-] yeast to viability. High-copy TPK1 did not rescue [Chc.sup.-] yeast. These genetic interactions suggest that budding-yeast 14-3-3 proteins are multifunctional and may play a role in both vesicular transport and Ras signaling pathways.

Alkalinization and anaerobiosis influence the pH buffering and phosphagenic activities of polyphosphate, and the catabolite secretion in chemostat-cultivated Saccharomyces cerevisiae. Cultures with high concentrations of polyphosphate have increased cytosolic pH and accelerated degradation of polyphosphate to small chains. The extracellular pH is normalized with reduced degradation of polyphosphate in dilute media. Complete polyphosphate hydrolysis occurs during anaerobiosis.

Saccharomyces cerevisiae strains containing the Pichia stipitis XYL1 and XYL2 genes and overexpressing the TKL1 and TAL1 genes show increased growth on xylose as compared to those expressing only the XYL1 and XYL2 genes. The overexpression of only TKL1 has no effect on the bacterial growth. The rate of xylose metabolism increases with the increase in fermentation of a xylose-glucose mixture during oxygen-limited conditions. TKL1 and TAL1 encode transketolase and transaldolase, respectively, which are involved in xylose metabolism by the pentose phosphate pathway.

The effects of the rotational information of DNA in determining the in vitro localization of nucleosomal core particles (ncps) have been studied in the Saccharomyces cerevisiae 5S rRNA repeat gene. We have altered the distribution of the phased series of flexibility signals present on this DNA by inserting a 25-bp tract, and we have analyzed the effects of this mutation on the distribution and on the frequencies of ncps, as compared with the wild type and a reference 21-bp insertion mutant. The variation of the standard free energy of nucleosome reconstitution was determined. The results show that the DNA rotational information is a major determinant of ncps positioning, define how many rotationally phased signals are required for the formation of a stable particle, and teach how to modify their distribution through the alteration of the rotational signals.

The di-lysine motif present in the type-I transmembrane Emp47 protein of Saccharomyces cerevisiae EMP47 gene is required for its localization in the Golgi body. Emp47 is a Golgi protein at steady-state despite the fact that the di-lysine motif is generally present in endoplasmic reticulum (ER) proteins. Alpha-COP is necessary for the ER-localization of Emp47p but not for Golgi-localization. The lumenal site of Emp47p is similar in structure to that of the mammalian ER-Golgi intermediate compartment-53 and VIP36 proteins located in the secretory pathway.

Eukaryotic genomes contain tracts of DNA in which a single base or a small number of bases are repeated (microsatellites). Mutations in the yeast DNA mismatch repair genes MSH2, PMS1, and MLH1 increase the frequency of mutations for normal DNA sequences and destabilize microsatellites. Mutations of human homologs of MSH2, PMSI, and MLH1 also cause microsatellite instability and result in certain types of cancer. We find that a mutation in the yeast gene MSH3 that does not substantially affect the rate of spontaneous mutations at several loci increases microsatellite instability about 40-fold, preferentially causing deletions. We suggest that MSH3 has different substrate specificities than the other mismatch repair proteins and that the human MSH3 homolog (MRP1) may be mutated in some tumors with microsatellite instability.

The characterization of the catalytic region of Ira2p, a GTPase-activating protein, in Saccharomyces cerevisiae reveals that the active site is highly conserved. Monovalent and divalent salts suppress the activity of Ira2p-383 fragment. The enzyme is similar to mammalian p120-GAP neurofibromin, and incorporates a binding site for tubulin, which inhibits both Ira2p and neurofibromin. This binding site extends to the N-terminal flanking region of the enzyme.

Despite evidence that DNA topoisomerase I is required to relieve torsional stress during DNA replication and transcription, yeast strains with a top1 null mutation are viable and display no gross defects in DNA or RNA synthesis, possibly because other proteins provide overlapping functions. We isolated mutants whose inviability or growth defect is relieved when TOP1 is expressed [trfmutants (topoisomerase one-requiring function)] . The TRF genes define at least four complementation groups. TRF3 is allelic to TOP2. TRF1 is allelic to HPR1, previously shown to be homologous to TOP1 over two short regions. TRF4 encodes a novel 584-amino acid protein with homology to the N-terminus of Saccharomyces cerevisiae topo I. Like top1 mutants, trf4 mutants have elevated rDNA recombination and fail to shut off RNA polymerase II transcription in stationary phase. trf4 null mutants are cs for viability, display reduced expression of GALI and Cell Cycle Box UAS::LacZ fusions, and are inviable in combination with trfI null mutants, indicating that both proteins may share a common function with DNA topoisomerase I. The existence of multiple TRF complementation groups suggests that not all biological functions of topo I can be carried out by topo II.

The break down of the maltose transporter in the yeast Saccharomyces cerevisiae appears to depend on endocytosis, occur in the vacuole, and be independent of preoteosome function. Researchers studied the break down of the transporter in mutants deficient in endocytosis, proteosome activity, and vacuolar proteolysis. The reaction, which occurs during fermentation in the absence of nitrogen, also required Sec18p.

Transport of maltose by the maltose transporter in Saccharomyces cerevisiae appears to be monophasic and reversible and the transporter facilitates movement of maltose in either direction across the membrane. Researchers analyzed maltose transport in S. cerevisiae with different mutations in the gene for the maltose transporter protein Mal161p. They developed a method for creating membranes that could be used for such transporter analysis.

The primary and secondary structure of the cyanide adduct of a triple mutant iso-1-cytochrome c of Saccharomyces cerevisiae obtained from an analyses of the (super 1)H NMR spectrum is given. The quality of the mutant protein as determined by the RMSD values is similar to the ferrocytochrome c of the horse heart. The secondary structure and folding of the mutant is similar to that of the wild-type protein. A distal ligand-binding cavity is formed by if alanine is present instead of the axial ligand. Due to four prolines which do not contain amide NH there is no connectivity in the amino acid sequence.

The Plasmodium falciparum multiple drug resistance gene (pfmdr1) acts as transport molecule in resisting drugs and is complementary with the ste6 gene of Saccharomyces cerevisiae, that codes a mating pheromone a-factor export molecule. Mutations in the pfmdr1 gene obtained with chloroquine resistance removes this function indicating that genetic alterations affect the transport function. The pfmdr1 gene belongs to the ATP-binding gene family, the members of which act as transport molecules.

The cauliflower mosaic virus (CaMV) gene VI that codes for a plant virus translational transactivator, acts as an intermediary in the release of a reporter gene in bakers' yeast, Saccharomyces cerevisiae. The CaMP gene VI also enhances the activity of chloramphenicol acetyltransferase to a great extent. CaMV is a plant virus that can express six of its genes.

The detection of a human protein kinase, K35, a cyclin-dependent protein kinase 8 that forms a complex with cyclin C in vitro and in vivo has been possible, which is probably involved in transcription in mammals. Cyclin is found to occur in K35 immunoprecipitates obtained from HeLa cells, suggesting the two proteins form a complex in vivo. The K35-cyclin C complex resembles the SRB10-SRB11 complex in Saccharomyces cerevisiae that is a part of the RNA polymerase II holoenzyme.

The synthesis of glycosylphosphatidylinositol (GPI) anchored membrane proteins involves the posttranslational linking of glycolipids to glycoproteins. During the biosynthesis, the mutants are decreased because each of them gathers an abnormal, incomplete GPI glycolipid with either zero, two or four mannoses. The GPI anchored proteins stabilize the yeast cell wall.

The yeast, Saccharomyces cerevisiae, recapitulates retroviral -1 frameshifting, as detected by the assays in the yeast that screen host mutants. A two-times increase in -1 frameshifting levels in yeast mutants is noticed, by using a model substrate of the mouse mammary tumor virus gag-pro frameshift site. The haploid cells devoid of 82% of the IFSI open reading frame are viable and phenotypically similar to isf1-1 mutants.

In Saccharomyces cerevisiae, the processes of meiosis I and meiosis II are regulated by a protein kinase called Cdc28. The active kinase has a catalytic subunit and a regulatory subunit called a cyclin. Nuclear division in eukaryotes may be of three types, meiosis I, meiosis II and mitosis, the differences in the processes lie in the ways in which the sister chromatids are separated. The cell has many cyclins which seem to play specialized roles in meiosis I and meiosis II.

Est1p, a protein coded by the protein, EST1, may be a subunit of yeast telomerase or else it is a protein associated with telomerase that is necessary for its activity in vitro. Telomerase is a ribonucleoprotein that uses an RNA template in the absence of a DNA template to increase the telomeric DNA strand. Telomerases are necessary for the maintenance of Saccharomyces chromosomes and their function is to prevent loss of DNA that would occur if a linear DNA molecule were replicated by a DNA polymerase.

Beta1,3-coupled disaccharides comprising the type I and type IV core disaccharide of carbohydrate-containing antigens are obtained by the glycosidase-mediated coupling of glycal acceptors with galactose. The beta1, 3-coupled disaccharide serves as an intermediary in chemical and enzymatic manipulation. An enzyme, beta1,6-N-acetylglucosaminyltransferase, facilitates the modification of the type IV disaccharide to a mucin-type trisaccharide, by a transfer of N-acetylglucosamine, to the 6-hydroxy position of N-acetylgalactosamine or galactose-reducing terminus.

Enrichment cultures of yeast isolates obtained from spent sulfite liquor yielded two types of yeast: Saccharomyces cerevisiae and Pichia membranaefaciens. Two strains of S. cerevisiae produced more ethanol than baker's yeast at pH < 6.0 and in the presence of acetic acid. One of the S. cerevisiae strains showed catabolite repression on glucose and galactose in the absence of acetic acid. However, in the presence of acetic acid, the isolate fermented glucose and galactose simultaneously. Although none of the isolates fermented xylose, low levels of xylose reductase and xylitol dehydrogenase activities were detected.

Slower step-pressure pulses are most effective in the ultra high pressure (UPH) pasteurization of pineapple juice. The decrease in the count of Saccharomyces cerevisiae is more with slower UPH step-pressure cycles than with static-pressure treatments. The inactivation of yeast is more with longer on-pressure and shorter off-pressure time.

The first description of entire, uniform isotopic labeling of a eukaryotic cytochrome c, iso-1-cytochrome c from the Saccharomyces cerevisiae yeast, is presented. Heteronuclear NMR methods could be used to assign resonances, measure amide hydrogen exchange rates and calculate relaxation parameters, due to sufficient quantities of super15N-labeled protein.

A five-step synthetic strategy helps prepare the antifungal agent (+)-preussin from t-Boc-(S)-phenylalanine. The stereoselective synthesis of (+)-preussin involves a Hg(II)-mediated 5-endo-dig ring closure of a ynone. Examination of (+)-preussin with a strain of Saccharomyces cerevisiae, which is deficient in recombinational DNA repair, reveals pressuin to be a DNA damaging agent in yeast.