記述言語：日本語   出版者・発行元：Bioscience, Biotechnology and Biochemistry  

In this study, we developed an efficient gene targeting system for the osmophilic fungus Aspergillus chevalieri, which is commonly used in the production of a dried bonito, katsuobushi. Specifically, we utilized the clustered regularly interspaced short palindromic repeats/Cas9 system to disrupt the ATP sulfurylase encoding sC gene. This results in methionine auxotroph and selenate-resistance. Additionally, we disrupted the DNA ligase IV encoding ligD gene, which is required for nonhomologous end joining. Using the sC marker and selenate-resistance as a selection pressure, we were able to rescue the sC marker and generate a ligD sC strain. We determined that the gene targeting efficiency of the ligD sC strain was significantly higher than that of the parental sC strain, which indicates that this strain provides efficient genetic recombination for the genetic analysis of A. chevalieri.

DOI： 10.1093/bbb/zbad033

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記述言語：日本語   出版者・発行元：Journal of Bioscience and Bioengineering  

Kokuto-shochu is a traditional Japanese spirit prepared from kokuto, obtained by evaporating water from sugarcane (Saccharum officinarum L.) juice. To clarify the effects of sugarcane cultivars on the sensory quality of kokuto-shochu, we investigated the flavor characteristics and composition of volatiles in kokuto-shochu prepared from kokuto using three different sugarcane cultivars, NiF8, Ni15, and RK97-14. Furthermore, experiments were conducted by using the cultivars collected between 2018 and 2020 to observe annual variations in their properties. The amino acid content of the three kokuto varieties did not differ significantly, but the amino acid content of NiF8 was two to five times higher than that of RK97-14, which was the same for all samples collected in the selected years. The browning degrees of kokuto were also higher in NiF8, and they were positively correlated to the amino acid contents of kokuto. The kokuto-like aroma of shochu made from Ni15 was stronger than that of shochu made from RK97-14. The concentration of ethyl lactate in shochu made from Ni15 was higher, however, the concentration of guaiacol was the lowest in the three cultivars’ products. Shochu made from NiF8 had the highest levels of Maillard reaction products (MRPs; pyrazines and furans), β-damascenone, and guaiacol amounts. In contrast, shochu made from RK97-14 tended to have a fruity flavor, and lower MRP levels than those made from NiF8. Thus, it was shown that sugarcane cultivars affect the sensory characteristics and volatiles in kokuto-shochu.

DOI： 10.1016/j.jbiosc.2023.03.010

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記述言語：日本語   出版者・発行元：Journal of Bioscience and Bioengineering  

In shochu-making, a small amount of fermenting moromi is added to a koji/water mixture instead of yeast culture to initiate fermentation. This is a characteristic process called Sashi-moto. It is known that shochu yeast is replaced by wild yeast upon repetition of Sashi-moto. The shochu yeast strains Kagoshima No. 2 (K2), Kagoshima No. 4 (C4), and Kagoshima No. 5 (H5), but not Kagoshima No. 6 (A6), were replaced by wild yeast (strain No. S5-g). K2 and C4 were easily replaced compared to H5, and the specific growth rates of K2 and C4 were lower than that of S5-g under higher osmotic pressure. Although the specific growth rate of H5 was higher than that of S5-g, its yeast population at the stationary phase was smaller than S5-g. On the other hand, both the specific growth rate and yeast population of A6 were higher than those of S5-g. The specific growth rate of yeast would be affected by osmotic tolerance and specific characters of the yeast strain.

DOI： 10.1016/j.jbiosc.2022.07.011

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記述言語：日本語   出版者・発行元：Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids  

Autophagy is regulated by phosphoinositides. We have previously shown that phosphatidylinositol 4-phosphate (PtdIns(4)P) is localized in the autophagosomal membrane. Additionally, in yeast cells, phosphatidylinositol 4-kinases Pik1p and Stt4p play important roles in the formation of the autophagosome and its fusion with the vacuole, respectively. In this study, we analyzed the primary role of PtdIns(4)P phosphatases in yeast autophagy. The PtdIns(4)P labeling densities in the membranes of the vacuoles, mitochondria, nucleus, endoplasmic reticulum, and plasma membrane dramatically increased in the phosphatase deletion mutants sac1∆ and sjl3∆, and the temperature-sensitive mutant sac1ts/sjl3∆ at the restrictive temperature. GFP-Atg8 processing assay indicated defective autophagy in the sac1∆ and sac1ts/sjl3∆ mutants. In contrast to the localization of PtdIns(4)P in the luminal leaflet of autophagosomal membranes in the wild-type yeast, PtdIns(4)P was localized in both the luminal and cytoplasmic leaflets of the autophagosomal membranes in the sac1∆ strain. In addition, the number of autophagic bodies in the vacuole significantly decreased in the sac1∆ strain, although autophagosomes were present in the cytoplasm. In the sac1ts/sjl3∆ strain, the number of autophagosomes in the cytoplasm dramatically decreased at the restrictive temperature. Considering that the numbers of autophagosomes and autophagic bodies in the sjl3∆ strain were comparable to those in the wild-type yeast, we found that the autophagosome could not be formed when PtdIns(4)P phosphatase activities of both Sac1p and Sjl3p were diminished. Together, these results indicate that the turnover of PtdIns(4)P by phosphatases is essential for autophagosome biogenesis.

DOI： 10.1016/j.bbalip.2022.159184

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記述言語：英語   出版者・発行元：John Wiley & Sons Australia, Ltd  

Massa Medicata Fermentata(MMF)はAspergillus属やRhizopus属による微生物発酵でコムギと数種のハーブから調製されるので、微生物発酵の影響について検討した。National Standard for Chinese Patent Drugsに従ってMMFの原材料を混合し、Aspergillus oryzae培養で調製(MMF-A)またはRhizopus oryzae培養で調製(MMF-R)した。MMFの調製により酵素活性は上昇した。MMF-AはMMF-Rよりもα-アミラーゼとリパーゼの活性が高かった。プロテアーゼ活性はMMF-Rがわずかに高かった。テルペノイドは発酵で低下したが、2,3-ブタンジオール、アセトイン、グアヤコールはMMFだけで検出された。1-オクテン-3-オールのC8化合物はMMF-RよりもMMF-Aで高く、4-ビニルグアヤコールとピラジンの芳香族化合物はMMF-Rで高かった。

記述言語：日本語   出版者・発行元：Traditional and Kampo Medicine  

Aim: Massa Medicata Fementata (MMF) is a crude drug used in East Asia to treat anorexia and dyspepsia. It is prepared from wheat and several herbs through microbial fermentation using Aspergillus sp. and Rhizopus sp. There is great difference in the quality of commercial MMF, and the microbes of MMF are suggested to affect its quality. We investigated the effects of microbial fermentation on the quality of MMF. Methods: Raw materials of MMF were mixed according to the ratio listed in the National Standard for Chinese Patent Drugs, and MMF was prepared using pure cultures of Aspergillus oryzae or Rhizopus oryzae. Digestive enzyme activities (α-amylase, protease, and lipase) and volatile compounds were measured using an analytical kit and GC–MS, respectively. Results: Enzyme activity increased in MMF. MMF prepared with A. oryzae (MMF-A) showed higher α-amylase and lipase activities than that prepared with R. oryzae (MMF-R). Protease activity was marginally higher in MMF-R than in MMF-A. GC–MS analysis revealed that terpenoids decreased with fermentation; however, 2,3-butanediol, acetoin, and guaiacol were detected in MMF only. C8 compounds such as 1-octen-3-ol were higher in MMF-A than MMF-R; however, aromatic compounds such as 4-vinylguaiacol and pyrazines were higher in MMF-R than MMF-A. Conclusion: Microbial fermentation contributes to increased enzyme activity and changes in MMF volatiles. These properties of MMF were considerably affected by the microbes used, and it is proposed in this study that it is important to have microbial control in the production of commercial MMF.

DOI： 10.1002/tkm2.1303

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記述言語：英語  

DOI： 10.1016/j.dib.2022.107888

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記述言語：日本語   出版者・発行元：Food Bioscience  

In this study, we compared the flavor profiles Chinese rice-flavor baijiu and Japanese awamori—traditional liquors with similar manufacturing processes. Citric acid and lactic acid were the major acid constituents in the mash of awamori and rice-flavor baijiu, respectively. The amino acid content of awamori mash was greater than that of rice-flavor baijiu owing to the higher proteolytic activities in the former. The high amino acid content also induced yeast growth at the early stage of fermentation in awamori mash, thus increasing the fermentation rate. Sensory evaluation revealed that rice-flavor baijiu had a significantly stronger alcoholic, floral, and fruity aroma, whereas the awamori had a strong sour, koji-like, oily, and cereal-like aromas. We determined the aroma threshold of (+)-ethyl D-lactate (15,400 μg/L) and (−)-ethyl L-lactate (12,500 μg/L). Both compounds have a common sweet, fruity, and green leaves-like aroma and a different nuance; (+)-ethyl D-lactate has a peach- and citrus-like aromas, and (−)-ethyl L-lactate has an apple-like and milky aromas. In rice-flavor baijiu, (−)-ethyl L-lactate constitutes the major isoform, as evident from the higher concentration of L (+)-lactic acid in the mash. Higher alcohols, (−)-ethyl L-lactate, and isoamyl acetate contents were greater in rice-flavor baijiu, while 1-octen-3-ol and acetic acid were greater in awamori. These results showed that solid-state saccharification contributed to the production of ethyl lactate by supplying in the mash a large amount of lactic acid produced by Rhizopus oryzae and koji contributed to the production of 1-octen-3-ol via the high fatty acid oxygenase activity produced by Aspergillus luchuensis.

DOI： 10.1016/j.fbio.2021.101375

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記述言語：日本語   出版者・発行元：Journal of Food Science  

Rice-flavor baijiu is a traditional Chinese liquor. The flavor profile and volatiles presented with or without the solid-state saccharification (SSS) were investigated to reveal the effects of SSS process on the quality of rice-flavor baijiu. The liquor prepared with SSS had a sweet flavor. It contained significantly higher contents of β-phenylethyl alcohol, β-phenylethyl acetate, and ethyl lactate with odor active value of >1. The liquor prepared without SSS had a cheese-like flavor. It was confirmed that the cheese-like flavor derived from butanoic acid was only detected in the liquor prepared without SSS. SSS facilitated the biosynthesis of β-phenylethyl alcohol and ethyl lactate by supplying a large amount of phenylalanine and lactic acid at the initial stage of fermentation, and it prevented contamination. These results indicated that the SSS process contributed to produce the characteristic flavor compounds of rice-flavor baijiu. Practical Application: Solid-state saccharification (SSS) process of rice-flavor baijiu contributes not only in brewing, but also in the production of the characteristic flavor compounds and the repression of the off-flavor derived from the contamination. Therefore, SSS is a critical process to control the flavor of rice-flavor baijiu.

DOI： 10.1111/1750-3841.15935

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記述言語：日本語   出版者・発行元：Microbiology Resource Announcements  

In this study, we report the chromosome-level genome sequence of the osmophilic filamentous fungus Aspergillus chevalieri M1, which was isolated from a dried bonito, katsuobushi. This fungus plays a significant role in the fermentation and ripening process. Thus, elucidating the sequence data for this fungus will aid in subsequent genomic research on the fungi involved in katsuobushi production.

DOI： 10.1128/MRA.00385-21

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記述言語：日本語   出版者・発行元：Microbiology Resource Announcements  

Aspergillus puulaauensis strain MK2 was isolated from a dead hard tick (Haemaphysalis longicornis). Here, we determined the chromosome-level genome sequence of A. puulaauensis MK2.

DOI： 10.1128/MRA.00372-21

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記述言語：日本語   出版者・発行元：Microbiology Resource Announcements  

Aspergillus luchuensis is used for the production of awamori and shochu, which are traditional Japanese distilled alcoholic beverages. Here, we determined the chromosome-level genome sequence of A. luchuensis RIB2601.

DOI： 10.1128/MRA.00384-21

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記述言語：日本語   出版者・発行元：Bioscience, biotechnology, and biochemistry  

Escherichia coli strain, whose gene is one of the subunits of succinate dehydrogenase (sdhA), and gene of the transcriptional repressor of isocitrate lyase (iclR) were disrupted, accumulated 6.6 times as much intracellular succinate as the wild-type MG1655 strain in aerobic growth, but succinate was not found in the culture medium. E. coli citT gene that encodes a citrate transporter was cloned under the control of the lacI promoter in pBR322-based plasmid and the above strain was transformed. This transformant, grown under aerobic condition in M9-tryptone medium with citrate, accumulated succinate in the medium while no succinate was found in the medium without citrate. CitT was active as a succinate transporter for 168 h by changing the culture medium or for 24 h in fed-batch culture. This study suggests that the CitT transporter functions as a succinate exporter in E. coli for succinate production in the presence of citrate.

DOI： 10.1093/bbb/zbaa109

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記述言語：日本語   出版者・発行元：LWT  

We investigated the contribution of the complex expressions of two α-amylases, namely acid-stable α-amylase (AS-amylase) and acid-labile α-amylase (AL-amylase), from Aspergillus kawachii to the microstructure of koji and the brewing. AL-amylase was found to be the primary contributor to the decomposition of starch in the early stage of koji making. From the middle stage, both α-amylases decomposed the starch in a coordinated manner, and at the final stage, acid-stable α-amylase and glucoamylase decomposed the starch granules. Characterization of koji prepared by the single disruptions of AS-amylase or AL-amylase genes, double disruption of both α-amylase genes, and triple disruption of two α-amylase and glucoamylase genes in A. kawachii cells indicated that both α-amylases can work in a synergistic manner to decompose starches during koji making. AL-amylase was found, for the first time, to play an important role in starch decomposition in koji. The speed of alcohol fermentation and ester contents of the fermented mash were higher the mash prepared the control strain, followed by single, double, and triple disruptants. These results indicate that the microstructure of koji plays a role in promoting alcohol fermentation and flavor development.

DOI： 10.1016/j.lwt.2020.110580

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記述言語：日本語   出版者・発行元：Journal of Food Science  

Abstract: Additional moisture in preparing red koji, Monascus-fermented rice, is a characteristic production process. To determine how additional moisture affects red koji preparation as per quality, we compared the growth of Monascus purpureus, enzyme and pigment production, and related gene expressions using our findings. We considered two kinds of red koji: one prepared with additional moisture at the middle part of the preparation and the other prepared without additional moisture. Our results showed that additional moisture did not promote the growth of M. purpureus, but it was significantly increased the pigment (red and yellow) and tended to increase the α-amylase level and saccharification power. Although adding a high amount of moisture (approximately 60% moisture content) promoted pigment production, it slightly repressed enzyme production. In contrast, adding approximately 50% moisture content promoted enzyme production. These findings showed that the additional moisture can affect the quality of red koji on the purpose. The expression of 10 pigment biosynthetic gene clusters and two glycohydrolase genes in red koji after adding moisture was analyzed through real-time qPCR. Eight genes were upregulated within 1 hr after adding water, with mppR2 being the first upregulated gene within 30 min. The expression of genes as per pigment production quickly responded to additional moisture during solid-state fermentation. Moreover, acetyl-CoA, which is a starting substrate for pigment content in red koji was increased within 3 hr after adding water. This study first described the relationship between additional moisture and expression of pigment biosynthetic genes by Monascus spp. during red koji preparation.

DOI： 10.1111/1750-3841.15610

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記述言語：日本語   出版者・発行元：Journal of Bioscience and Bioengineering  

Citrate exporter CexA plays a key role in the production of citric acid in fungi; however, its role in intracellular metabolism has remained unclear. In this study, we comparably characterized homologous cexA genes in the white koji fungus Aspergillus luchuensis mut. kawachii and the yellow koji fungus Aspergillus oryzae, which exhibit high and low abilities, respectively, to produce citric acid. Disruption of cexA caused a significant decline of both extracellular and intracellular citric acid accumulation in Aspergillus kawachii, while overexpression of the A. kawachii cexA gene (AkcexA) into A. oryzae significantly enhanced both extracellular and intracellular citric acid accumulation in A. oryzae to a level comparable to that of A. kawachii. In addition, overexpression of two intrinsic cexA homologs (AocexA and AocexB) in A. oryzae also enhanced its extracellular and intracellular citric acid accumulation. Comprehensive analysis of intracellular metabolites from an AkcexA-overexpressing strain of A. oryzae compared with its control strain identified metabolic changes associated with intracellular citric acid accumulation via the glycolytic pathway, pentose phosphate pathway, and tricarboxylic acid cycle. Our results indicate that citric acid export enhances not only extracellular citric acid accumulation but also intracellular metabolic fluxes to generate citric acid.

DOI： 10.1016/j.jbiosc.2020.09.002

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記述言語：日本語   出版者・発行元：Journal of the Institute of Brewing  

DOI： 10.1002/jib.674

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出版者・発行元：Journal of Bioscience and Bioengineering  

DOI： 10.1016/j.jbiosc.2020.05.006

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.2323/jgam.2019.09.003.

記述言語：日本語   掲載種別：研究論文（学術雑誌）  

出版者・発行元：(公社)日本生物工学会  

白コウジカビAspergillus luchuensis mut. kawachiiの固相培養における、デンプン分解酵素とクエン酸産生の調節に対するサーチュインの役割を解析した。5種のサーチュイン遺伝子破壊株の性質解析により、これらの遺伝子がデンプン分解活性と酸性化に関与し、特にsirD破壊株では酸安定なα-アミラーゼ活性とクエン酸産生が低下した。sirD破壊株は菌糸の色が変化し、細胞壁合成阻害剤に高い感受性を示し、分生子形成が低下した。キャップ分析遺伝子発現と定量的RT-PCRを行ったところ、sirD破壊株において酸安定性α-アミラーゼ遺伝子とクエン酸エクスポーターの転写体レベルが低下していた。SirDはA.kawachiiの固相発酵において、酸安定性α-アミラーゼとクエン酸の産生を含む全体の転写調節に重要な役割を果たしていると考えられた。

出版者・発行元：Applied and Environmental Microbiology  

DOI： 10.1128/AEM.01950-19

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DOI： 10.3389/fnut.2020.00115

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出版者・発行元：公益社団法人 日本食品科学工学会  

<p>This study aimed to reveal the chemical and flavor profiles of rice-flavor <i>baijiu</i> by comparison with <i>awamori</i> and <i>kome-shochu</i>, traditional Japanese liquors. Rice-flavor <i>baijiu</i> is similar to <i>awamori</i> and <i>kome-shochu</i> with regard to ingredients and the fermentation starter. Of the 15 rice-flavor <i>baijiu</i> samples in this study, 3 had a light yellow to dark-brown color. Dark-brown samples had high glucose and low amino acid contents. Lactic acid was detected in all rice-flavor <i>baijiu</i> samples. Compared to <i>awamori</i> and <i>kome-shochu</i>, rice-flavor <i>baijiu</i> contained more acetic acid. Thirty-four volatile compounds in rice-flavor <i>baijiu</i> were identified and quantified. In all, 18 compounds in rice-flavor <i>baijiu</i>, 15 in <i>awamori</i> and 13 in <i>kome-shochu</i> had an odor activity value (OAV) of >1. Of these, 11 compounds showed a three-fold higher OAV in rice-flavor <i>baijiu</i> than in <i>awamori</i> and <i>kome-shochu</i>. Principal component analysis revealed that ethyl lactate is a key volatile compound that is distinct to rice-flavor <i>baijiu</i>.</p>

DOI： 10.3136/fstr.26.411

出版者・発行元：Bioscience, Biotechnology and Biochemistry  

DOI： 10.1080/09168451.2020.1792761

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出版者・発行元：Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids  

DOI： 10.1016/j.bbalip.2019.05.004

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出版者・発行元：Letters in Applied Microbiology  

DOI： 10.1111/lam.13148

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出版者・発行元：Applied and Environmental Microbiology  

DOI： 10.1128/AEM.03136-18

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出版者・発行元：European Journal of Cell Biology  

DOI： 10.1016/j.ejcb.2018.03.007

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：American Society for Microbiology  

出版者・発行元：Yeast  

DOI： 10.1002/yea.3243

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出版者・発行元：Food Chemistry  

DOI： 10.1016/j.foodchem.2016.12.005

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出版者・発行元：一般社団法人 日本生薬学会  

<p>"<i>Shinkiku</i>" is a traditional digestive drug prepared by the fermentation of wheat and some herbs with fermentative microbes. <i>Shinkiku</i> is manufactured in China and Korea, and also used in Japanese <i>Kampo</i> medicine as a component of <i>Hangebyakujutsutemmato</i>. However, there are currently no quality standards for <i>shinkiku</i>, and thus, the quality of <i>shinkiku</i> has considerable variation depending on its manufacturer. Although these variations would be partially derived from the differences in fermentative microbes, there are no studies about microbial diversities or chemical constituents in commercial <i>shinkiku</i>. Thus, we investigated the microbial diversity and chemical constituents of 15 commercial <i>shinkiku</i> samples to standardize its quality. PCR-denaturing gradient gel electrophoresis of 16S rDNA and ITS1 sequences revealed that different microbes such as <i>Lactobacillus</i> sp. and <i>Candida</i> sp. were present in each <i>shinkiku</i> sample. On the other hand, most <i>shinkiku</i> samples showed amylase (12/15 samples) and lipase activities (9/15 samples) that behave as digestants. In addition, all samples commonly contained ferulic acid (>10 nmol/g), which has anti-inflammatory and anti-oxidant activities. Thus, enzyme activities and ferulic acid were suggested to be one of the candidates for use as reference standards for the quality control of <i>shinkiku</i>. Exceptional <i>shinkiku</i> samples without enzyme activities showed a baked brown color, and ferulic acid content was inversely related with the brightness color of <i>shinkiku</i> (R²=0.47). Therefore, it seems that color indices would be effective to predict the quality of <i>shinkiku</i> such as enzyme activities and ferulic acid.</p>

DOI： 10.24684/jspharm.71.1_41

出版者・発行元：公益財団法人　応用微生物学・分子細胞生物学研究奨励会  

<p><i>Aspergillus luchuensis</i> is a kuro (black) koji fungus that has been used as a starch degrader for the awamori- and shochu-making industries in Japan. In this study, we investigated the effect of ion beam irradiation on <i>A. luchuensis</i> RIB2601 and obtained a high starch-degrading mutant strain U1. Strain U1 showed reduced growth rate, whereas it showed higher α-amylase, glucoamylase, and α-glucosidase activities on a mycelial mass basis than the wild type (wt) strain both on agar plates and in rice koji. In addition, strain U1 showed higher <i>N</i>-acetylglucosamine content in the cell wall and higher sensitivity to calcofluor white, suggesting a deficiency in cell wall composition. Interestingly, produced protein showed higher expression of acid-labile α-amylase (AmyA) and glucoamylase (GlaA) in strain U1, although real-time RT-PCR indicated no significant change in the transcription of the <i>amyA</i> or <i>glaA</i> gene. These results suggested that the high amylolytic activity of strain U1 is attributable to a high AmyA and GlaA production level, but the elevated production is not due to transcriptional regulation of the corresponding genes. Furthermore, RNA-seq analysis indicated that strain U1 shows transcriptional changes in at least 604 genes related to oxidation-reduction, transport, and glucosamine-containing compound metabolic processes, which may be involved in the deficient cell wall composition of strain U1.</p>

DOI： 10.2323/jgam.2017.02.004

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：日本語   掲載種別：研究論文（学術雑誌）  

記述言語：日本語   掲載種別：研究論文（学術雑誌）  

記述言語：日本語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）  

記述言語：英語   掲載種別：研究論文（学術雑誌）