Sake yeast strains have difficulty in entering a quiescent state after cell growth cessation.

Sake yeast strains produce a high concentration of ethanol during sake brewing compared to laboratory yeast strains. As ethanol fermentation by yeast cells continues even after cell growth stops, analysis of the physiological state of the stationary phase cells is very important for understanding the mechanism of producing higher concentrations of ethanol. We compared the physiological characteristics of stationary phase cells of both sake and laboratory yeast strains in an aerobic batch culture and under sake brewing conditions.

Aflatoxin non-productivity of Aspergillus oryzae caused by loss of function in the aflJ gene product.

Aspergillus oryzae, although closely related to Aspergillus flavus, does not produce aflatoxin (AF). A. oryzae RIB strains can be classified into three groups (group 1-3) based on the structure of the AF biosynthesis gene homolog cluster (AFHC).

Enhancement of the initial rate of ethanol fermentation due to dysfunction of yeast stress response components Msn2p and/or Msn4p.

Sake yeasts (strains of Saccharomyces cerevisiae) produce high concentrations of ethanol in sake fermentation. To investigate the molecular mechanisms underlying this brewing property, we compared gene expression of sake and laboratory yeasts in sake mash. DNA microarray and reporter gene analyses revealed defects of sake yeasts in environmental stress responses mediated by transcription factors Msn2p and/or Msn4p (Msn2/4p) and stress response elements (STRE).

The concentration of ethyl carbamate in commercial ume (Prunus mume) liqueur products and a method of reducing it.

The ethyl carbamate concentration of commercial ume liqueur products was studied, and a method of reducing it was examined from the viewpoint of antioxidation. The average ethyl carbamate concentration across 38 ume liqueur products was 0.12 mg/l (0.

Contribution of ethanol-tolerant xylanase G2 from Aspergillus oryzae on Japanese sake brewing.

We purified three xylanase isozymes (XynF1, XynF3 and XynG2) from a solid-state Aspergillus oryzae RIB128 culture using chromatography. The results of our sake-brewing experiment, in which we used exogenously supplemented enzymes, revealed that only XynG2 improved the alcohol yield and the material utilization. The alcohol yield of the XynG2 batch displayed an increase of 4.

Simple metabolite extraction method for metabolic profiling of the solid-state fermentation of Aspergillus oryzae.

Solid-state culture of microorganisms is an important style of culture both in the traditional food industry and in the modern fermentation industry. We propose here a simple method for metabolite extraction from the solid-state fermentation of a filamentous fungus, Aspergillus oryzae, which is known as rice-koji. To evaluate the efficiency of metabolite extraction, liquid chromatography-mass spectrometry (LC/MS) was used for simultaneous detection of a wide range of metabolites including amino acids, organic acids and vitamins, which are of interest in rice-koji making.

Contribution of 1,2-dihydroxy-5-(methylsulfinyl)pentan-3-one (DMTS-P1) to the formation of dimethyl trisulfide (DMTS) during the storage of Japanese sake.

Dimethyl trisulfide (DMTS) is involved in the unpalatable aroma of stale Japanese sake, called "hineka". Recently, we isolated one of the precursor compounds of DMTS in sake and identified it as 1,2-dihydroxy-5-(methylsulfinyl)pentan-3-one (DMTS-P1), a previously unknown compound. In this work, the contribution of DMTS-P1 to the formation of DMTS was investigated.

PAD1 and FDC1 are essential for the decarboxylation of phenylacrylic acids in Saccharomyces cerevisiae.

The volatile phenols, to which Saccharomyces cerevisiae converts from phenylacrylic acids including ferulic acid, p-coumaric acid, and cinnamic acid, generate off-flavors in alcoholic beverages such as beer and wine. Using gene disruptants, transformants and cell-free extracts of these strains, we have verified that the adjacent PAD1 (phenylacrylic acid decarboxylase, YDR538W) and FDC1 (ferulic acid decarboxylase, YDR539W) genes are essential for the decarboxylation of phenylacrylic acids in S. cerevisiae.

Effects of sulfur dioxide on formation of fishy off-odor and undesirable taste in wine consumed with seafood.

In order to evaluate sensory compatibility of alcoholic beverages with food, beverages and dried squid, namely, "surume", a common Japanese accompaniment, were consumed together. White wine and dried squid pairings had a more undesirable taste and more fishy off-odor than sake and dried squid pairings. The undesirable taste and fishy off-odor appeared to be caused by degradation of unsaturated fatty acids (e.

Abscisic acid stimulated ripening and gene expression in berry skins of the Cabernet Sauvignon grape.

We investigated the effect of exogenous abscisic acid (ABA) application on the transcriptome as well as the phenolic profiles in the skins of Vitis vinifera cv. Cabernet Sauvignon grape berries grown on the vine and cultured in vitro. ABA application rapidly induced the accumulation of anthocyanin and flavonol.

Aspergillus oryzae atfA controls conidial germination and stress tolerance.

We compared atfA and atfB, the genes encoding the respective ATF/CREB-type transcription factors in Aspergillus oryzae. The germination ratio of DeltaatfA conidia was low without any stress, unlike that of DeltaatfB conidia. The DeltaatfA conidia were more sensitive to oxidative stress than the DeltaatfB conidia, which are also sensitive to oxidative stress.

Disruption of ubiquitin-related genes in laboratory yeast strains enhances ethanol production during sake brewing.

Sake yeast can produce high levels of ethanol in concentrated rice mash. While both sake and laboratory yeast strains belong to the species Saccharomyces cerevisiae, the laboratory strains produce much less ethanol. This disparity in fermentation activity may be due to the strains' different responses to environmental stresses, including ethanol accumulation.

Overexpression of MSN2 in a sake yeast strain promotes ethanol tolerance and increases ethanol production in sake brewing.

To improve the ethanol tolerance of sake yeast we constructed a sake yeast strain that overexpresses MSN2, a transcription factor that is activated by several environmental stresses, including ethanol. We showed that this strain is more ethanol tolerant and produced more ethanol in a sake mash than a control strain.

Color recovery in berries of grape (Vitis vinifera L.) 'Benitaka', a bud sport of 'Italia', is caused by a novel allele at the VvmybA1 locus.

Color mutations in grape berry skin are relatively frequent events, and can be easily seen in the vineyard. Both light-red-skinned 'Ruby Okuyama' and more intense and uniform rosy-skinned 'Benitaka' (Vitis vinifera L.) are bud sports of white-skinned 'Italia'.

Development of VNTR markers for three Aspergillus species used in brewing.

Using a bioinformatics approach, we developed 18 variable number of tandem repeat markers for Aspergillus oryzae for use in population genetic studies. Repeat sequences in the genome sequences of A. oryzae were identified by a tandem repeat finding program.

Ethanol stress stimulates the Ca2+-mediated calcineurin/Crz1 pathway in Saccharomyces cerevisiae.

Environmental stimuli elicit a stress response, which helps to maintain cell survival. In budding yeast Saccharomyces cerevisiae, environmental cues can activate calcineurin, a highly conserved Ca2+-- and calmodulin-dependent protein phosphatase. Calcineurin dephosphorylates the transcription factor Crz1, leading to accumulation of Crz1 in the nuclei and expression of stress responsive genes under the control of a calcineurin-dependent response element (CDRE).

Screening and identification of precursor compounds of dimethyl trisulfide (DMTS) in Japanese sake.

Dimethyl trisulfide (DMTS) is involved in the unpalatable aroma of stale sake, called "hineka"; however, the mechanism underlying the formation of DMTS during the storage of sake has not been elucidated. This paper investigates the precursors of DMTS in sake. An experiment using [methyl-d(3)]-methionine showed that Strecker degradation of methionine plays a minor role in the formation of DMTS.

Inhibition of mitochondrial fragmentation during sake brewing causes high malate production in sake yeast.

We previously demonstrated the presence and fragmentation of mitochondria during alcohol fermentation. Here, we show that Fis1p induces mitochondrial fragmentation, and inhibition of mitochondrial fragmentation causes higher malate production during sake brewing. These findings indicate that mitochondrial morphology affects the metabolism of constituents, providing a breeding strategy for high-malate-producing yeasts.

Aspergillus oryzae atfB encodes a transcription factor required for stress tolerance in conidia.

Using an Aspergillus oryzae EST database, we identified a gene encoding a transcription factor (atfB), which is a member of the ATF/CREB family. Expression of atfB was barely detectable during vegetative growth, but was readily detected during conidiation in solid-state culture. Microarray analyses showed that expression of many other genes, including catalase (catA), were downregulated in an atfB-disruptant.

Multiple functions of ergosterol in the fission yeast Schizosaccharomyces pombe.

Sterols are a major class of membrane lipids in eukaryotes. In Schizosaccharomyces pombe, sterol 24-C-methyltransferase (Erg6p), C-8 sterol isomerase (Erg2p), C-5 sterol desaturase (Erg31p, Erg32p), C-22 sterol desaturase (Erg5p) and C-24 (28) sterol reductase (Sts1p/Erg4p) have been predicted, but not yet determined, to catalyse a sequence of reactions from zymosterol to ergosterol. Disruption mutants of these genes were unable to synthesize ergosterol, and most were tolerant to the polyene drugs amphotericin B and nystatin.

Characterization of peptides generated in proteolytic digest of steamed rice grains by sake koji enzymes.

High-molecular-weight peptides (approximately 10-30 kDa) generated in a digest of steamed rice grains by sake koji enzymes were characterized. Among 13 major spots resolved by 2-D gel electrophoresis, 12 contained peptides having N-termini of rice glutelin as determined by mass fingerprinting analysis and/or MS/MS. The source of these peptides was presumed to be the acidic subunit of rice glutelin.

Mitochondrial dynamics of yeast during sake brewing.

The presence of mitochondria during alcohol fermentation has not been studied. Here, we examined the yeast mitochondrial structure during sake brewing using the green fluorescent protein. Mitochondrial structures were observed throughout brewing and they fragmented as brewing proceeded.

Intraspecies diversity of the industrial yeast strains Saccharomyces cerevisiae and Saccharomyces pastorianus based on analysis of the sequences of the internal transcribed spacer (ITS) regions and the D1/D2 region of 26S rDNA.

We divided industrial yeast strains of Saccharomyces cerevisiae into three groups based on the sequences of their internal transcribed spacer (ITS) regions. One group contained sake yeasts, shochu yeasts, and one bakery yeast, another group contained wine yeasts, and the third group contained beer and whisky yeasts, including seven bakery yeasts. The three groups were distinguished by polymorphisms at two positions, designated positions B and C, corresponding to nucleotide numbers 279 and 301 respectively in the S288C strain.

Effects of accumulated S-adenosylmethionine on growth of yeast cells.

S-adenosylmethionine (SAM) accumulated in cultured yeast cells and affected growth in two ways. High levels of intracellular SAM in yeast inhibited early growth, but increased growth in medium without sources of nitrogen and sulfur. Accumulated SAM in the yeast cells was recycled as a nutritional source depending on the sulfur and nitrogen contents of the medium.

Ethanol-induced death in yeast exhibits features of apoptosis mediated by mitochondrial fission pathway.

Cell death in yeast (Saccharomyces cerevisiae) involves several apoptotic processes. Here, we report the first evidence of the following processes, which are also characteristic of apoptosis, in ethanol-induced cell death in yeast: chromatin condensation and fragmentation, DNA cleavage, and a requirement for de novo protein synthesis. Mitochondrial fission protein, Fis1, appears to mediate ethanol-induced apoptosis and ethanol-induced mitochondrial fragmentation.