2018 Research results

1. Selective adsorption of 1,3-dimethyltrisulfane (DMTS) responsible for aged odour in Japanese sake using supported gold nanoparticles

Author: Haruno Murayama, Yusuke Yamamoto, Misaki Tone, Takayuki Hasegawa, Moemi Kimura, Tamao Ishida, Atsuko Isogai, Tsutomu Fujii, Mitsutaka Okumura, Makoto Tokunaga
Abstract: Gold (Au) nanoparticles (NPs) supported on SiO2 (Au/SiO₂) were prepared by a practical impregnation method and applied as an adsorbent for 1,3-dimethyltrisulfane (DMTS), which is responsible for an unpleasant odour in drinks, especially Japanese sake. Compared with a conventional adsorbent, activated carbon, Au/SiO₂ selectively reduced the DMTS concentration in Japanese sake without decreasing the concentrations of other aromatic components. DFT calculations revealed that the selective adsorption of DMTS occurred through the formation of a stable intermediate. The size of the supported Au NPs was controlled by the preparation conditions and determined from TEM observations and XRD measurements, and the size was ranged from 2.4 nm to 30 nm. Au/SiO₂ having Au NPs with a diameter of 2.4 nm adsorbed DMTS the most efficiently. Smaller Au NPs showed better DMTS adsorption capabilities because larger amounts of Au atoms were exposed on their surfaces in the size range of this study. Langmuir-type monolayer adsorption and one-to-one binding of Au-S are proposed to occur based on an adsorption isotherm experiment. Even though significant differences of the fruity aroma score were not observed in the sensory evaluation between Au/SiO₂ and activated carbon for this less aromatic Japanese sake, Au/SiO₂ selectively decreased the DMTS concentration in the instrumental analysis.
Source: Scientific Reports, 8, 16064 (2018)

2. Development of DNA markers to differentiate good Kyokai sake yeast and the other yeast strains

Author: Takeshi Akao, Zou Yan, Daisuke Watanabe, Naoto Okazaki, and Hitoshi Shimoi
Abstract: Previous studies revealed that a representative sake yeast strain, Kyokai No. 7 (K7), possesses three loss-of-function mutations, rim155067ins, msn4C1540T, Δppt1::Ty2, which enable high fermentation ability of the strain. Since all three were common and specific to K7 and closely related good strains, such as K6, K9, K10 and their derivative strains among approximately 30 strains tested by each mutation, they were expected to work as efficient DNA markers to differentiate those good strains from others. In this study, a rapid and accurate genotyping method, high resolution melting (HRM) analysis, was employed and optimized for typing RIM15⁵⁰⁶⁷ins and MSN4¹⁵⁴⁰, respectively. Typing of these DNA markers was then applied to a broad range of yeast strains (98 strains, 134 stocks) to estimate whether they belonged to the group of K7-realated strains.
Source: J. Brew. Soc. Japan,113 (10), 631-641 (2018) in Japanese

3. Characteristic features of the unique house sake yeast strain Saccharomyces cerevisiae Km67 used for industrial sake brewing

Author: Yoshifumi Takao, Toshinari Takahashi, Tasuku Yamada, Tetsuya Goshima, Atsuko Isogai, Kazuo Sueno, Tsutomu Fujii, and Takeshi Akao
Abstract: For several decades, almost all sake has been brewed with sake yeast Saccharomyces cerevisiae Kyokai no. 7 (K7) group strains. Although the widespread use of these strains has contributed to sake quality improvement, it may have lessened the diversity of sake gustatory properties brought about by house sake yeast (indigenous yeast of sake brewery). Sake yeast S. cerevisiae strain Km67 derives from the house yeast strain of Kiku-masamune Sake Brewing Co., Ltd., and it has been playing a central role in industrial sake brewing for decades. By using DNA sequencing, we revealed that strain Km67 does not possess specific loss-of-function mutations of stress response-related genes, which are characteristic of K7 group strains. Km67 had higher stress tolerance than K7 group strains likely because of the more efficient function of the stress response and heat shock elements in this strain. Sensory evaluation and taste sensor analysis demonstrated that sake brewed with Km67 had characteristically thicker body than sake brewed with K7 group strains. Chemical analysis suggested that unique sensory properties of the sake brewed with Km67 were due to high citramalic acid concentration. Taken together, these results revealed that strain Km67 differs from K7 group strains by genetic background and confers unique chemical composition and taste qualities upon sake it generates. It is expected that sake quality and gustatory properties will be diversified by utilizing house yeast such as strain Km67.
Source: Journal of Bioscience and Bioengineering, 126 (5), 617-623 (2018)

4. Analysis of Sake Components Presented to Sake Contest in 2017

Author: Tsutomu FUJII, Atsuko ISOGAI,Sachiko IIZUKA, Hanae IZU, Ryoko KANDA, Tomokazu HASHIGUCHI and Nami GOTO-YAMAMOTO
Source: Report of the Research Institute of Brewing, 190, 1-18 (2018) in Japanese

5. Review of the 40th Honkaku Shochu and Awamori Contest in 2017

Author: Nobuhiko MUKAI, Muneyoshi KANAI, Masayuki TAKAHASHIO, Nahoko NISHIBORI, Akinori MUTO, Toshihiro YOSHIMURA, and Nami GOTO-YAMAMOTO
Source: Report of the Research Institute of Brewing, 190, 19-34 (2018) in Japanese

6. Anasysis of Beer Components Presented to Craft Beer Contest of Japan Brewers Association in 2018

Author: Kazutaka KUSAKA, Maki KAMIMOTO and Akinori MUTO
Source: Report of the Research Institute of Brewing, 190, 35-40 (2018) in Japanese

7. Classification of Kyokai sake yeast stock strains by brewing charecteristics

Author: Naoto OKAZAKI, Futami NORO, Masaaki INAHASHI, Katsumi NAKAHARA, Hirokazu HASUDA, Takafumi MUTOU, Tomoko MURAKAMI, Tetsuya GOSHIMA, Takeshi AKAO, Yasuzo KIZAKI, and Takeaki ISHIKAWA
Abstract: Twenty brewing characteristics which contribute to classify a Kyokai sake yeast strains were selected by the partition analysis for 24 Kyokai yeast strains which contained 385 stock strains. It was possible to classify these as independent clusters for Kyokai yeast strains using 20 explanatory variables. This narrowed to 344 strains when excluding 41 strains of samples missed from the 24 main clusters. While the total numbers of distinction error strains of a discriminant analyses using 18 explanatory variables for 344 strains were 7 strains, when considered with 16 explanatory variables which excluded the amino acid component left 10 strains. Therefore, we decided to exclude amino acid composition from the analysis items because the contribution was small and there was little influence in cluster analysis and discriminant analysis. Regardless of the old or new period distributions, the brewing characteristics of the 344 strains of K-7 were kept for only one cluster and the K-7 cluster could be roughly divided into 3 sub clusters by the cluster analysis. The reason for being divided into 3 clusters was because of the fermentation ability. In the cluster analyses by 11 brewing characteristics for 24 kinds of all yeast select ion candidates in 1250 strains since 2003, 76 strains of clusters of high aroma generation strains were sorted out and the composition strains were only K-1601, K-1701 and K-1801. Also, all K-1801 distribution strains from 2006 to 2014 were contained in the cluster of K-1801, and the brewing characteristics of the K-1801 were stable.
Source: J. Brew. Soc. Japan,113 (8), 515-524 (2018) in Japanese

8. Effect of acetic acid bacteria on alcoholic liver injury in C57BL/6J mice.

Author: Satoshi SEINO, Hitoshi KURIHARA, Ryosuke MATSUOKA, Yohei OKUYAMA, Masaaki KUNOU, Hanae IZU, Yongqiang CHENG, Yanxiang GAO, Beizhong HAN
Abstract: The effect of ingestion of acetic acid bacteria on alcoholic liver injury was evaluated. C57BL/6J mice (8 weeks old, male, 22-27 g) were divided into three groups: the control group (without ethanol), ethanol group (ethanol 2.5 g/kg), and ethanol + acetic acid bacteria group (ethanol 2.5 g/kg + acetic acid bacteria 10 mg). The mice were given three times per day for 14 consecutive days in each group. Then the serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels and liver lipids of the mice were measured. As a result, AST and ALT levels, liver triglyceride and cholesterol levels were significantly higher in the ethanol group than those in the control group (P＜0.05). The ethanol + acetic acid bacteria group had significantly lower levels than those in the ethanol group(P＜0.05). These results suggested that the ingestion of acetic acid bacteria might alleviate alcoholic liver injury.
Source: China Brewing 37(8), 28-31 (2018) in Chinese

9. Discrimination of wine from grape cultivated in Japan, imported wine, and others by multi-elemental analysis

Author: Hideaki SHIMIZU, Fumikazu AKAMATSU, Aya KAMADA, Kazuya KOYAMA, Masaki OKUDA, Hisashi FUKUDA, Kazuhiro IWASHITA, Nami GOTO-YAMAMOTO
Abstract: Differences in mineral concentrations were examined among three types of wine in the Japanese market place: Japan wine, imported wine, and domestically produced wine mainly from foreign ingredients (DWF), where "Japan wine" has been recently defined by the National Tax Agency as domestically produced wine from grapes cultivated in Japan. The main objective of this study was to examine the possibility of controlling the authenticity of Japan wine. The concentrations of 18 minerals (Li, B, Na, Mg, Si, P, S, K, Ca, Mn, Co, Ni, Ga, Rb, Sr, Mo, Ba, and Pb) in 214 wine samples were determined by inductively coupled-plasma mass spectrometry (ICP-MS) and ICP-atomic emission spectrometry (ICP-AES). In general, Japan wine had a higher concentration of potassium and lower concentrations of eight elements (Li, B, Na, Si, S, Co, Sr, and Pb) as compared with the other two groups of wine. Linear discriminant analysis (LDA) models based on concentrations of the 18 minerals facilitated the identification of three wine groups: Japan wine, imported wine, and DWF with a 91.1% classification score and 87.9% prediction score. In addition, an LDA model for discrimination of wine from four domestic geographic origins (Yamanashi, Nagano, Hokkaido, and Yamagata Prefectures) using 18 elements gave a classification score of 93.1% and a prediction score of 76.4%. In summary, we have shown that an LDA model based on mineral concentrations is useful for distinguishing Japan wine from other wine groups, and can contribute to classification of the four main domestic wine-producing regions of Japan.
Source: Journal of Bioscience and Bioengineering, 125, 413-418 (2018)

10. Construction of sake yeast with low production of dimethyl trisulfide (DMTS) precursor by a self-cloning method

Author: Yuriko IKEDA, Atsuko ISOGAI, Yuta MORIYOSHI, Ryoko KANDA, Kazuhiro IWASHITA, Tsutomu FUJII
Abstract: Dimethyl trisulfide (DMTS) is the primary component responsible for "hineka", the stale aroma of Japanese sake. Deletion of the MRI1 or MDE1 gene of sake yeast, encoding 5'-methylthioribose-1-phosphate isomerase and 5'-methylthioribulose-1-phosphate dehydratase, respectively, has been reported to greatly reduce the amount of DMTS precursor (DMTS-P1) in sake and to suppress the formation of DMTS during storage. In this study, we constructed sake yeast strains lacking MRI1 gene function by a self-cloning method. Two methods were applied: in one, a stop codon was introduced in the MRI1 ORF by point mutation; in the other, the entire MRI1 ORF was deleted from the genome. In both methods, a plasmid vector containing drug-resistance and counter-selectable markers was used to introduce the mutation. We successfully obtained the strains, which did not contain the plasmid sequences, by both methods. Small-scale sake brewing tests using these SC strains (strains obtained by the self-cloning method) found that DMTS-P1 was hardly detected in sake brewed with SC strains, and DMTS production after sake storage was greatly reduced as compared with the parent strain. The components of brewed sake were almost the same between the SC and parent strains. These results suggest that SC strains can produce sake with higher flavor stability without changing the sake brewing properties.
Source: Journal of Bioscience and Bioengineering, 125, 419-424 (2018)

11. The delaying effect of alpha-glycerophosphocholine on senescence, transthyretin deposition, and osteoarthritis in senescence-accelerated mouse prone 8 mice.

Author: Kiminori MATSUBARA, Mayumi OKUDA, Sachi SHIBATA, Shigeru MIYAKI, Takeshi OHKUBO, Hanae IZU, Tsutomu FUJII
Abstract: Administration of alpha-glycerophosphocholine (GPC), a choline compound in food, is expected to contribute to human health. In this study, we evaluated its effect on aging in senescence-accelerated mouse prone 8 (SAMP8) mice. Male SAMP8 mice had free access to a commercial stock diet and drinking water with or without GPC (0.07 mg/ml). Mice in the GPC group had significantly lower total senescence grading score than that of the control group at 36 weeks of age. Administration of GPC decreased the deposition of transthyretin (TTR), an amyloidogenic protein, in the brain. Aggregated TTR activated microglia and led to neuroinflammation. Thus, GPC would protect the brain by reducing TTR deposition and preventing neuroinflammation. In a histological study of knee joints, it was found that SAMP8 mice administered GPC showed decreased joint degeneration. These results suggest that GPC delays the aging process and may be a useful compound in anti-aging functional food development.
Source: Bioscience, Biotechnology, and Biochemistry, 82(4), 47-653 (2018)

12. MAL73, a novel regulator of maltose fermentation, is functionally impaired by single nucleotide polymorphism in sake brewing yeast

Author: Takumi OHDATE , Fumihiko OMURA, Haruyo HATANAKA, Yan ZHOU, Masami TAKAGI, Tetsuya GOSHIMA, Takeshi AKAO, Eiichiro ONO
Abstract: For maltose fermentation, budding yeast Saccharomyces cerevisiae operates a mechanism that involves transporters (MALT), maltases (MALS) and regulators (MALR) collectively known as MAL genes. However, functional relevance of MAL genes during sake brewing process remains largely elusive, since sake yeast is cultured under glucose-rich condition achieved by the co-culture partner Aspergillus spp.. Here we isolated an ethyl methane sulfonate (EMS)-mutagenized sake yeast strain exhibiting enhanced maltose fermentation compared to the parental strain. The mutant carried a single nucleotide insertion that leads to the extension of the C-terminal region of a previously uncharacterized MALR gene YPR196W-2, which was renamed as MAL73. Introduction of the mutant allele MAL73L with extended C-terminal region into the parental or other sakeyeast strains enhanced the growth rate when fed with maltose as the sole carbon source. In contrast, disruption of endogenous MAL73 in the sake yeasts decreased the maltose fermentation ability of sake yeast, confirming that the original MAL73 functions as a MALR. Importantly, the MAL73L-expressing strain fermented more maltose in practical condition compared to the parental strain during sake brewing process. Our data show that MAL73(L) is a novel MALR gene that regulates maltose fermentation, and has been functionally attenuated in sake yeast by single nucleotide deletion during breeding history. Since the MAL73L-expressing strain showed enhanced ability of maltose fermentation, MAL73L might also be a valuable tool for enhancing maltose fermentation in yeast in general.
Source: PLoS ONE 13(6): e0198744 (2018)

13. Estimation method for enzyme digestibility of steamed rice grains for sake making using alkali / urea solubility

Author: Masaki OKUDA, Midori JOYO, Hisashi FUKUDA
Abstract: In the present study, we investigated the estimation method for the enzyme digestibility of steamed rice grains for sake making using alkali / urea solubility. Visual observation of the alkali / urea concentration solubilizing rice grains showed significant correlations with the DSC (= Differential Scanning Calorimeter) gelatinization temperature, the enzyme digestibility of steamed rice grains, and the retrogradation of steamed rice grains. The absorbance of iodine staining after soaking rice grains in urea solutions measured using a spectrophotometer also showed significant correlations with the enzyme digestibility of steamed rice grains and the retrogradation of steamed rice grains. Therefore, the enzyme digestibility of steamed rice grains for sake making can be estimated easily by the alkali / urea concentration solubilizing rice grains and the absorbance of iodine staining after soaking the rice grains in a urea solution. However, the polishing ratio and the moisture content of rice grains influenced the urea concentration solubilizing rice grains, although the existence of a white-core did not. In conclusion, these methods will be useful to estimate the enzyme digestibility of steamed rice grains for sake making, if the estimation standards according to the polishing ratio and moisture content of the rice grains can be determined.
Source: J. Brew. Soc. Japan, 113, 315-330 (2018) in Japanese

14. Evaluation of Candida easanensis JK8 β-glucosidase with potentially hydrolyse non-volatile glycosides of wine aroma precursors

Author: Thongekkaew J, Fujii T, Masaki K, Koyama K
Abstract: Important 'floral' aromas naturally occur in grapes predominantly as flavourless glycoconjugate precursors. Since these aroma compounds can be released by hydrolysis, different glycosidase enzymes can potentially contribute different aromas to wines. In this paper, the effects of crude and purified Candida easanensis JK8 β-glucosidases on wine aroma precursors of Muscat of Alexandria grape powder were investigated by GC/MS combined with stir bar sorptive extraction (SBSE). A total of 19 bound volatile compounds were identified, including phenols, terpenes, aldehyde, ester and alcohols. The concentrations of terpenes especially nerol and geraniol, and β-Damascenone, a C13-norisoprenoid, contributes flowery and slightly fruity aromas were signiflcantly increased in enzyme treated. These results suggest the potential application of this yeast β-glucosidase as an aroma-enhancing enzyme in winemaking.
Source: Natural Product Research, 1-5 (2018)

15. Characteristics of the volatile composition of Shochu in the IzuIslands

Author: Hisashi Fukuda, Osamu Yamada and Jinshun Han
Abstract: To identify characteristics of compounds of Shochu made in the Izu Islands, we analyzed 78 volatile compounds by head space solid phase micro extraction(SPME), and 6 volatile compounds by direct head space analysis. There was a significant difference in 44 volatile compounds of sweet potato shochu between that in the Izu Islands and the in Honkaku Shochu Contest. In a stepwise discriminant analysis procedure, all of samples were rightly classified by 4 volatile compounds (1-octen-3-ol, ethyl butyrate, Ethyl enanthate, and vanillin). Estimating the accuracy of the discriminant analysis by cross validation, the ratios of appropriate classification were 100%. For barley shochu in the Izu Islands and in the Honkaku Shochu Contest, there was a significant difference in 29 of 84 volatile compounds. A total of 67 of 68 samples were rightly classified by 4 volatile compounds (Nonanal, Ethyl acetate, Ethyl butyrate, and DMTS). When estimating the accuracy of the discriminant analysis, the ratios of appropriate classification were 98.5%. For blended shochu of sweet potato shochu and barley shochu in the Izu Islands, there was a significant difference in 42 volatile compounds between sweet potato shochu, barley shochu, and blended shochu. In a stepwise discriminant analysis procedure, 30 of 33 samples were rightly classified by 8 volatile compounds (Citronellol, Ethyl isobutyrate, Ethyl caproate, Isoamyl caproate, Ethyl caprate, Ethyl cinnamate, Diethyl succinate and Guaiacol).
Source: J. Brew. Soc. Japan, 114, 151-158 (2019) in Japanese

16. Analysis of 4-mercapto-4-methylpentan-2-one in sake presented at the Sake Contest

Author: Sachiko Iizuka, Atsuko Isogai, Ryoko Kanda, and Tsutomu Fujii
Abstract: Recently, "muscat-like", "litchi-like" or "citrus-like" odor characteristics have been indicated in sake presented at Sake Contests. As thiols were considered to contribute to these odor characteristics, we analyzed one of thiol compounds, 4-mercapto-4-methylpentan-2-one (4MMP), in sake presented at the Sake Contest in 2015. The content of 4MMP in 25 samples was 5～14 ng/L, which was higher than the perception threshold of 4MMP in sake (1.2 ng/L) previously reported. To study the influence of 4MMP on the odor property of sake, a sensory evaluation was performed with the addition of 4MMP to the sake. The 'sulfur' odor property was significantly enhanced by the addition of ≥ 8.0 ng/L of 4MMP. Though the addition of 16.0 ng/L of 4MMP significantly lowered the total apparaisal, it significantly enhanced 'fruity' odor properties different from the aroma of ethyl caproate and isoamyl acetate.
Source: J. Brew. Soc. Japan, 114, 93-101 (2019) in Japanese

17. Carbon Stable Isotopic Compositions of Citric Acid and Malic Acid in Japanese Apricot Liqueur Decrease as the Fruit Ripens

Author: Fumikazu Akamatsu, Yasuhisa Tsuchida, Takaaki Oe, Yuri Hisatsune, Yukari Igi, Tomokazu Hashiguchi, Tsutomu Fujii
Abstract: The carbon stable isotopic composition (δ¹³C) is often analyzed to quantify the addition of acidulants to Japanese apricot liqueur, but little is known about the variation in theδ¹³C values of the main organic acids arising from differences in the ripeness of Japanese apricots. We show that in Japanese apricot liqueur prepared using fruits at different stages of ripeness, theδ¹³C values of citric acid and malic acid ranged from -25.1‰ to -23.7‰ and from -22.3‰ to -19.7‰, respectively, and theδ¹³C values decreased as the fruit ripened. The averageδ¹³C value of citric acid from liqueurs was 0.7‰ higher than that from fresh fruits, whereas theδ¹³C values of malic acid showed no isotope discrimination. The variation inδ¹³C values of the main organic acids in Japanese apricot liqueurs will help detect acidulant addition and control authenticity.
Source: Food Chemistry, 277, 70-74 (2019)

18. The potential aroma and flavor compounds in Vitis sp. cv. Koshu and V. vinifera L. cv. Chardonnay under different environmental conditions

Author: Sharon Marie Bahena-Garrido, Tomoko Ohama, Yuka Suehiro, Yuko Hata, Atsuko Isogai, Kazuhiro Iwashita, Nami Goto-Yamamoto and Kazuya Koyama
Abstract: BACKGROUND

Koshu, a hybrid of Vitis vinifera L. and V. davidii Foex, is the most popular indigenous cultivar for wine production in Japan. However, little is known about the potential aroma compounds it contains and how environmental factors affect these. In this study, we obtained comprehensive profiles of the volatile (both glycosidically bound and free) and phenolic compounds that occur in koshu berries, and compared these with similar profiles for V. vinifera cv. chardonnay. We then compared the response of these two cultivars to bunch shading and the ripening-related phytohormone abscisic acid (ABA).

RESULTS

Koshu berries contained significantly higher concentrations of phenolic compounds, such as hydroxycinnamic acid derivatives, and some volatile phenols, such as 4-vinyl guaiacol and eugenol, than chardonnay berries, which are thought to contribute to the characteristics of koshu wine. In addition, koshu berries had a distinctly different terpenoid composition from chardonnay berries. Shading reduced the concentration of norisoprenoid in both cultivars, as well as several phenolic compounds, particularly their volatile derivatives in koshu berries. The exogenous application of ABA induced ripening and increased the concentrations of lipid derivatives, such as hexanol, octanol, 1-nonanol, and 1-octen-3-ol. Multivariate and discriminant analyses showed that the potential aroma and flavor compounds in the berries could be discriminated clearly based on cultivar and environmental cues, such as light exposure.

CONCLUSION

The unique secondary metabolite profiles of koshu and their different responses to environmental factors could be valuable for developing various types of koshu wines and new cultivars with improved quality and cultural characteristics.
Source: Journal of the science of food and agriculture, 99, 1926-1937 (2019)

19. Meiotic chromosomal recombination defect in sake yeasts

Author: Hitoshi Shimoi, Yuta Hanazumi, Natsuki Kawamura, Miwa Yamada, Shohei Shimizu, Taro Suzuki, Daisuke Watanabe, and Takeshi Akao
Abstract: Sake yeast strains are classified into Saccharomyces cerevisiae and have a heterothallic life cycle. This feature allows cross hybridization between two haploids to breed new strains with superior characteristics. However, cross hybridization of sake yeast is very difficult because only a few spores develop in a sporulation medium, and most of these spores do not germinate. We hypothesized that these features are attributable to chromosome recombination defect in meiosis, which leads to chromosome loss. To test this hypothesis, we examined meiotic recombination of sake yeast Kyokai no. 7 (K7) using the following three methods: (i) analysis of the segregation patterns of two heterozygous sites in the same chromosome in 100 haploid K7 strains; (ii) sequencing of the whole genomes of four haploid K7 strains and comparison of the bases derived from the heterozygosities; and (iii) construction of double heterozygous disruptants of CAN1 and URA3 on the chromosome V of K7 and the examination of the genotypes of haploids after sporulation. We could not detect any recombinant segregants in any of the experiments, which indicated defect in meiotic recombination in K7. Analyses after sporulation of the same double heterozygous disruptants of K6, K9, and K10 also indicated meiotic recombination defect in these strains. Although rapamycin treatment increased the sporulation efficiency of K7, it did not increase the meiotic recombination of the double heterozygous K7. Moreover, the spo13 disruptant of the K7 derivative produced two spore asci without meiotic recombination. These results suggest that sake yeasts have defects in meiotic recombination machinery.
Source: Journal of Bioscience and Bioengineering, 127, 190-196 (2019)

20. Nutrient signaling via the TORC1-greatwall-PP2A^B55δ pathway responsible for the high initial rates of alcoholic fermentation in sake yeast strains of Saccharomyces cerevisiae

Author

Daisuke Watanabe, Takuma Kajihara, Yukiko Sugimoto, Kenichi Takagi, Megumi Mizuno, Yan Zhou, Jiawen Chen, Kojiro Takeda, Hisashi Tatebe, Kazuhiro Shiozaki, Nobushige Nakazawa, Shingo Izawa, Takeshi Akao, Hitoshi Shimoi, Tatsuya Maeda, Hiroshi Takagi.

Abstract

Saccharomyces cerevisiae sake yeast strain Kyokai no. 7 (K7) and its relatives carry a homozygous loss-of-function mutation in the RIM15 gene, which encodes a Greatwall-family protein kinase. Disruption of RIM15 in non-sake yeast strains leads to improved alcoholic fermentation, indicating that the defect in Rim15p is associated with the enhanced fermentation performance of sake yeast cells. In order to understand how Rim15p mediates fermentation control, we here focused on target-of-rapamycin protein kinase complex 1 (TORC1) and protein phosphatase 2A with the ^B55δ regulatory subunit (PP2A^B55δ), complexes that are known to act upstream and downstream of Rim15p, respectively. Several lines of evidence, including our previous transcriptomic analysis data, suggested enhanced TORC1 signaling in sake yeast cells during sake fermentation. Fermentation tests of the TORC1-related mutants using a laboratory strain revealed that TORC1 signaling positively regulates the initial fermentation rate in a Rim15p-dependent manner. Deletion of the CDC55 gene encoding ^B55δ abolished the high fermentation performance of Rim15p-deficient laboratory yeast and sake yeast cells, indicating that PP2A^B55δ mediates the fermentation control by TORC1 and Rim15p. The TORC1-Greatwall-PP2A^B55δ pathway similarly affected the fermentation rate in the fission yeast Schizosaccharomyces pombe, strongly suggested that the evolutionarily conserved pathway governs alcoholic fermentation in yeasts.

It is likely that elevated PP2A^B55δ activity accounts for the high fermentation performance of sake yeast cells. Heterozygous loss-of-function mutations in CDC55 found in K7-related sake strains may indicate that the Rim15p-deficient phenotypes are disadvantageous to cell survival.

IMPORTANCE

The biochemical processes and enzymes responsible for glycolysis and alcoholic fermentation by the yeast S. cerevisiae have long been the subject of scientific research. Nevertheless, the factors determining fermentation performance in vivo are not fully understood. As a result, the industrial breeding of yeast strains has required empirical characterization of fermentation by screening numerous mutants through laborious fermentation tests. To establish a rational and efficient breeding strategy, key regulators of alcoholic fermentation need to be identified. In the present study, we focused on how sake yeast strains of S. cerevisiae have acquired high alcoholic fermentation performance. Our findings provide a rational molecular basis to design yeast strains with optimal fermentation performance for production of alcoholic beverages and bioethanol. In addition, as the evolutionarily conserved TORC1-Greatwall-PP2A^B55δ pathway plays a major role in the glycolytic control, our work may contribute to research on carbohydrate metabolism in higher eukaryotes.

Source

Applied and Environmental Microbiology, 85,e02083-18 (2019)