Methionine and Glycine Stabilize Mitochondrial Activity in Sake Yeast During Ethanol Fermentation

Dodatak aminokiselina u fermentacijsku podlogu utječe na rast i aktivnost kvasca, a zadržavanje aktivnosti mitohondrija tijekom alkoholnog vrenja je kritično za aktivnost pivskog kvasca. Međutim, mehanizam djelovanja aminokiselina u fermentacijskoj podlozi i njihov utjecaj na aktivnost mitohondrija pivskog kvasca tijekom vrenja još uvijek nije poznat. U ovom radu smo utvrdili da aminokiseline u fermentacijskoj podlozi, osobito metionin i glicin, stabiliziraju aktivnost mitohondrija kvasca tijekom proizvodnje pića sake. Pomoću mutanta atg32△, koji ima narušenu aktivnost mitohondrija, istražili smo aminokiseline koje povećavaju aktivnost mitohondrija kvasca tijekom vrenja. Identificirali smo metionin i glicin kao aminokiseline koje bi mogle povećati aktivnost mitohondrija kvasca tijekom proizvodnje pića sake. Da bismo to potvrdili, izmjerili smo količinu reaktivnih oblika kisika u kvascu nakon vrenja u podlozi s metioninom i glicinom. Kvasac koji je fermentirao u podlozi s metioninom i glicinom zadržao je relativno veliku količinu reaktivnih oblika kisika u usporedbi s kvascem koji je fermentirao u podlozi bez dodatka aminokiselina. Osim toga, stanice koje su fermentirale u podlozi s dodatkom metionina imale su različit metabolom od stanica koje su fermentirale u podlozi bez dodatka aminokiseline. Rezultati pokazuju da određene aminokiseline, poput metionina i glicina, stabiliziraju aktivnost mitohondrija kvasca tijekom proizvodnje pića sake i na taj način upravljaju aktivnošću kvasca.Addition of amino acids to fermentation media affects the growth and brewing profiles of yeast. In addition, retaining mitochondrial activity during fermentation is critical for the fermentation profiles of brewer’s yeasts. However, a concrete mechanism linking amino acids in fermentation media with mitochondrial activity during fermentation of brewer’s yeasts is yet unknown. Here, we report that amino acids in fermentation media, especially methionine (Met) and glycine (Gly), stabilize mitochondrial activity during fermentation of sake yeast. By utilizing atg32Δ mutant sake yeast, which shows deteriorated mitochondrial activity, we screened candidate amino acids that strengthened the mitochondrial activity of sake yeast during fermentation. We identified Met and Gly as candidate amino acids that fortify mitochondrial activity in sake yeast during fermentation. To confirm this biochemically, we measured reactive oxygen species (ROS) levels in sake yeast fermented with Met and Gly. Yeast cells supplemented with Met and Gly retained high ROS levels relative to the non-supplemented sake yeast. Moreover, Met-supplemented cells showed a metabolome distinct from that of non-supplemented cells. These results indicate that specific amino acids such as Met and Gly stabilize the mitochondrial activity of sake yeast during fermentation and thus manipulate brewing profiles of yeast

Saccharomyces arboricola and Its Hybrids’ Propensity for Sake Production: Interspecific Hybrids Reveal Increased Fermentation Abilities and a Mosaic Metabolic Profile

The use of interspecific hybrids during the industrial fermentation process has been well established, positioning the frontier of advancement in brewing to capitalize on the potential of Saccharomyces hybridization. Interspecific yeast hybrids used in modern monoculture inoculations benefit from a wide range of volatile metabolites that broaden the organoleptic complexity. This is the first report of sake brewing by Saccharomyces arboricola and its hybrids. S. arboricola x S. cerevisiae direct-mating generated cryotolerant interspecific hybrids which increased yields of ethanol and ethyl hexanoate compared to parental strains, important flavor attributes of fine Japanese ginjo sake rice wine. Hierarchical clustering heatmapping with principal component analysis for metabolic profiling was used in finding low levels of endogenous amino/organic acids clustered S. arboricola apart from the S. cerevisiae industrial strains. In sake fermentations, hybrid strains showed a mosaic profile of parental strains, while metabolic analysis suggested S. arboricola had a lower amino acid net uptake than S. cerevisiae. Additionally, this research found an increase in ethanolic fermentation from pyruvate and increased sulfur metabolism. Together, these results suggest S. arboricola is poised for in-depth metabolomic exploration in sake fermentation

「「国際社会観光」基礎教育体系の研究」成果報告 平成30（2018）～令和２（2020）年度和洋女子大学教育振興支援助成報告

　本報告は、2018～2020年度の研究の成果報告である。2020年度に新しく国際学部国際学科が設置されたことにより、国際社会観光についての基礎教育体系を精緻化し、教材の作成を行った。さらに観光PBL（Project Based Learning）教育方法を探求し、企業や自治体とのPBLの拡充と、プロジェクトの経験からよりよい教育方法を探るFDを行った。　本稿では、『キーワードで読む国際観光』（国際学部国際学科編の冊子）で取り上げた内容について、学科FDで情報交換した過去のPBLの経験から学んだことについて、そして昨年度の急なコロナ禍でも積極的に行ったオンラインPBLの内容について、プロジェクト参加メンバーで分担執筆した

Saccharomyces arboricola and Its Hybrids’ Propensity for Sake Production: Interspecific Hybrids Reveal Increased Fermentation Abilities and a Mosaic Metabolic Profile

The use of interspecific hybrids during the industrial fermentation process has been well established, positioning the frontier of advancement in brewing to capitalize on the potential of Saccharomyces hybridization. Interspecific yeast hybrids used in modern monoculture inoculations benefit from a wide range of volatile metabolites that broaden the organoleptic complexity. This is the first report of sake brewing by Saccharomyces arboricola and its hybrids. S. arboricola x S. cerevisiae direct-mating generated cryotolerant interspecific hybrids which increased yields of ethanol and ethyl hexanoate compared to parental strains, important flavor attributes of fine Japanese ginjo sake rice wine. Hierarchical clustering heatmapping with principal component analysis for metabolic profiling was used in finding low levels of endogenous amino/organic acids clustered S. arboricola apart from the S. cerevisiae industrial strains. In sake fermentations, hybrid strains showed a mosaic profile of parental strains, while metabolic analysis suggested S. arboricola had a lower amino acid net uptake than S. cerevisiae. Additionally, this research found an increase in ethanolic fermentation from pyruvate and increased sulfur metabolism. Together, these results suggest S. arboricola is poised for in-depth metabolomic exploration in sake fermentation

Chromosomal aneuploidy improves the brewing characteristics of sake yeast

The effect of chromosomal aneuploidy on the brewing characteristics of brewery yeasts has not been studied. Here we report that chromosomal aneuploidy in sake brewery yeast (Saccharomyces cerevisiae) leads to the development of favorable brewing characteristics. We found that pyruvate-underproducing sake yeast, which produces less off-flavor diacetyl, is aneuploid and trisomic for chromosomes XI and XIV. To confirm that this phenotype is due to aneuploidy, we obtained 45 haploids with various chromosomal additions and investigated their brewing profiles. A greater number of chromosomes correlated with a decrease in pyruvate production. Especially, sake yeast haploids with extra chromosomes in addition to chromosome XI produced less pyruvate than euploids. Mitochondrion-related metabolites and intracellular oxygen species in chromosome XI aneuploids were higher than those in euploids, and this effect was canceled in their "petite" strains, suggesting that an increase in chromosomes upregulated mitochondrial activity and decreased pyruvate levels. These findings suggested that an increase in chromosome number, including chromosome XI, in sake yeast haploids leads to pyruvate underproduction through the augmentation of mitochondrial activity. This is the first report proposing that aneuploidy in brewery yeasts improves their brewing profile

Identification of clinical factors related to antibody-mediated immune response to the subfornical organ

Objective : We recently reported cases of adipsic hypernatremia caused by autoantibodies against the subfornical organ in patients with hypothalamic-pituitary lesions. This study aimed to clarify the clinical features of newly identified patients with adipsic hypernatremia whose sera displayed immunoreactivity to the mouse subfornical organ. Design : Observational cohort study of patients diagnosed with adipsic hypernatremia in Japan, United States, and Europe. Methods : The study included 22 patients with adipsic hypernatremia but without overt structural changes in the hypothalamic-pituitary region and congenital disease. Antibody response to the mouse subfornical organ was determined using immunohistochemistry. The clinical characteristics were compared between the patients with positive and negative antibody responses. Results : Antibody response to the mouse subfornical organ was detected in the sera of 16 patients (72.7%, female/male ratio, 1:1, 12 pediatric and 4 adult patients). The prolactin levels at the time of diagnosis were significantly higher in patients with positive subfornical organ (SFO) immunoreactivity than in those with negative SFO immunoreactivity (58.9 ± 33.5 vs. 22.9 ± 13.9 ng/ml, p <.05). Hypothalamic disorders were found in 37.5% of the patients with positive SFO immunoreactivity. Moreover, six patients were diagnosed with rapid-onset obesity with hypothalamic dysfunction, hypoventilation, and autonomic dysregulation/neural tumor syndrome after the diagnosis of adipsic hypernatremia. Plasma renin activity levels were significantly higher in patients with serum immunoreactivity to the Nax channel. Conclusions : The patients with serum immunoreactivity to the SFO had higher prolactin levels and hypothalamic disorders compared to those without the immunoreactivity. The clinical characteristics of patients with serum immunoreactivity to the subfornical organ included higher prolactin levels and hypothalamic disorders, which were frequently associated with central hypothyroidism and the presence of retroperitoneal tumors