Flávia Silva-Sousa, Bruna Oliveira, Ricardo Franco-Duarte, Carole Camarasa, Maria João Sousa
Climate change and consumer preferences are driving innovation in winemaking, with a growing interest in non-Saccharomyces species. Among these, Torulaspora delbrueckii (Td) has gained recognition for its ability to reduce volatile acidity and enhance aromatic complexity in wine. However, knowledge regarding its phenotypic and genomic diversity impacting alcoholic fermentation remains limited. Aiming to elucidate the metabolic differences between Td and Saccharomyces cerevisiae (Sc) and the Td intraspecies diversity, we conducted a comprehensive metabolic characterization of fifteen Td strains. This analysis delved beyond standard fermentation parameters (kinetics and major metabolites production) to explore non-conventional aromas and establish genotype-phenotype links. Our findings confirmed that most Td strains produce less acetic acid and more succinate and glycerol than Sc. The overall aromatic profiles of Td strains differed from Sc, exhibiting higher levels of monoterpenes and higher alcohols, while producing less acetate esters, fatty acids, their corresponding ethyl esters, and lactones. Moreover, we identified the absence of genes responsible for specific aroma profiles, such as decreased ethyl esters production, as well as the absence of cell wall genes, which might negatively affect Td performance when compared to Sc. This work highlights the significant diversity within Td and underscores potential links between its genotype and phenotype.
{"title":"Bridging the Gap: linking Torulaspora delbrueckii Genotypes to Fermentation Phenotypes and Wine Aroma.","authors":"Flávia Silva-Sousa, Bruna Oliveira, Ricardo Franco-Duarte, Carole Camarasa, Maria João Sousa","doi":"10.1093/femsyr/foae034","DOIUrl":"https://doi.org/10.1093/femsyr/foae034","url":null,"abstract":"<p><p>Climate change and consumer preferences are driving innovation in winemaking, with a growing interest in non-Saccharomyces species. Among these, Torulaspora delbrueckii (Td) has gained recognition for its ability to reduce volatile acidity and enhance aromatic complexity in wine. However, knowledge regarding its phenotypic and genomic diversity impacting alcoholic fermentation remains limited. Aiming to elucidate the metabolic differences between Td and Saccharomyces cerevisiae (Sc) and the Td intraspecies diversity, we conducted a comprehensive metabolic characterization of fifteen Td strains. This analysis delved beyond standard fermentation parameters (kinetics and major metabolites production) to explore non-conventional aromas and establish genotype-phenotype links. Our findings confirmed that most Td strains produce less acetic acid and more succinate and glycerol than Sc. The overall aromatic profiles of Td strains differed from Sc, exhibiting higher levels of monoterpenes and higher alcohols, while producing less acetate esters, fatty acids, their corresponding ethyl esters, and lactones. Moreover, we identified the absence of genes responsible for specific aroma profiles, such as decreased ethyl esters production, as well as the absence of cell wall genes, which might negatively affect Td performance when compared to Sc. This work highlights the significant diversity within Td and underscores potential links between its genotype and phenotype.</p>","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142603870","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The yeast Yarrowia lipolytica can assimilate n-alkane as a carbon and energy source. To elucidate the significance of phosphatidylserine (PS) in the utilization of n-alkane in Y. lipolytica, we investigated the role of the Y. lipolytica ortholog (PSS1) of Saccharomyces cerevisiae PSS1/CHO1, which encodes a PS synthase. The PSS1 deletion mutant (pss1Δ) of Y. lipolytica could not grow on minimal medium in the absence of ethanolamine and choline but grew when either ethanolamine or choline was supplied to synthesize phosphatidylethanolamine and phosphatidylcholine. The pss1Δ strain exhibited severe growth defects on media containing n-alkanes even in the presence of ethanolamine and choline. In the pss1Δ strain, the transcription of ALK1, which encodes a primary cytochrome P450 that catalyzes the hydroxylation of n-alkanes in the endoplasmic reticulum, was upregulated by n-alkane as in the wild-type strain. However, the production of functional P450 was not detected, as indicated by the absence of reduced CO-difference spectra in the pss1Δ strain. PS was undetectable in the lipid extracts of the pss1Δ strain. These results underscore the critical role of PSS1 in the biosynthesis of PS, which is essential for the production of functional P450 enzymes involved in n-alkane hydroxylation in Y. lipolytica.
{"title":"Phosphatidylserine synthase plays a critical role in the utilization of n-alkanes in the yeast Yarrowia lipolytica","authors":"Katsuro Matsuse, Mariho Hara, Ryo Iwama, Hiroyuki Horiuchi, Ryouichi Fukuda","doi":"10.1093/femsyr/foae030","DOIUrl":"https://doi.org/10.1093/femsyr/foae030","url":null,"abstract":"The yeast Yarrowia lipolytica can assimilate n-alkane as a carbon and energy source. To elucidate the significance of phosphatidylserine (PS) in the utilization of n-alkane in Y. lipolytica, we investigated the role of the Y. lipolytica ortholog (PSS1) of Saccharomyces cerevisiae PSS1/CHO1, which encodes a PS synthase. The PSS1 deletion mutant (pss1Δ) of Y. lipolytica could not grow on minimal medium in the absence of ethanolamine and choline but grew when either ethanolamine or choline was supplied to synthesize phosphatidylethanolamine and phosphatidylcholine. The pss1Δ strain exhibited severe growth defects on media containing n-alkanes even in the presence of ethanolamine and choline. In the pss1Δ strain, the transcription of ALK1, which encodes a primary cytochrome P450 that catalyzes the hydroxylation of n-alkanes in the endoplasmic reticulum, was upregulated by n-alkane as in the wild-type strain. However, the production of functional P450 was not detected, as indicated by the absence of reduced CO-difference spectra in the pss1Δ strain. PS was undetectable in the lipid extracts of the pss1Δ strain. These results underscore the critical role of PSS1 in the biosynthesis of PS, which is essential for the production of functional P450 enzymes involved in n-alkane hydroxylation in Y. lipolytica.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142263867","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Yeast cell wall chitin has been shown to bind grape pathogenesis-related chitinases that are the primary cause of protein haze in wines suggesting that yeast cell walls may be applied for haze protection. Here we present a high throughput screen to identify yeast strains with high cell wall chitin using a reiterative enrichment strategy and Fluorescence-Activated Cell Sorting of cells labelled with either GFP-tagged chitinase or with Calcofluor White. To assess the validity of the strategy, we first used a pooled deletion strain library of Saccharomyces cerevisiae. The strategy enriched for deletion mutants with genes that had previously been described as having an impact on chitin levels. Genes that had not previously been linked to chitin biosynthesis or deposition were also identified. These genes are involved in cell wall maintenance and/or membrane trafficking functions. The strategy was then applied to a mutagenized population of a commercial wine yeast strain, Saccharomyces cerevisiae EC1118. Enriched mutant strains showed significantly higher cell wall chitin than the wild type and significantly reduced the activity of chitinases in synthetic model wine, suggesting that these strains may be able to reduce haze formation in wine.
{"title":"Isolation and characterisation of Saccharomyces cerevisiae mutants with increased cell wall chitin using fluorescence-activated cell sorting","authors":"Lesiba Tyrone Chuene, Thulile Ndlovu, Debra Rossouw, Rene Kathleen Naidoo-Blassoples, Florian Franz Bauer","doi":"10.1093/femsyr/foae028","DOIUrl":"https://doi.org/10.1093/femsyr/foae028","url":null,"abstract":"Yeast cell wall chitin has been shown to bind grape pathogenesis-related chitinases that are the primary cause of protein haze in wines suggesting that yeast cell walls may be applied for haze protection. Here we present a high throughput screen to identify yeast strains with high cell wall chitin using a reiterative enrichment strategy and Fluorescence-Activated Cell Sorting of cells labelled with either GFP-tagged chitinase or with Calcofluor White. To assess the validity of the strategy, we first used a pooled deletion strain library of Saccharomyces cerevisiae. The strategy enriched for deletion mutants with genes that had previously been described as having an impact on chitin levels. Genes that had not previously been linked to chitin biosynthesis or deposition were also identified. These genes are involved in cell wall maintenance and/or membrane trafficking functions. The strategy was then applied to a mutagenized population of a commercial wine yeast strain, Saccharomyces cerevisiae EC1118. Enriched mutant strains showed significantly higher cell wall chitin than the wild type and significantly reduced the activity of chitinases in synthetic model wine, suggesting that these strains may be able to reduce haze formation in wine.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142263868","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Scotch Whisky, a product of high importance to Scotland, has gained global approval for its distinctive qualities derived from the traditional production process which is defined in law. However, ongoing research continuously enhances Scotch Whisky production and is fostering a diversification of flavour profiles. To be classified as Scotch Whisky, the final spirit needs to retain the aroma and taste of “Scotch”. While each production step contributes significantly to whisky flavour—from malt preparation and mashing to fermentation, distillation, and maturation—the impact of yeast during fermentation is crucially important. Not only does the yeast convert the sugar to alcohol, it also produces important volatile compounds, for example esters and higher alcohols, that contribute to the final flavour profile of whisky. The yeast chosen for whisky fermentations can significantly influence whisky flavour, so the yeast strain employed is of high importance. This review explores the role of yeast in Scotch Whisky production and its influence on flavour diversification. Furthermore, an extensive examination of non-conventional yeasts employed in brewing and winemaking is undertaken to assess their potential suitability for adoption as Scotch Whisky yeast strains, followed by a review of methods for evaluating new yeast strains.
{"title":"The potential for scotch malt whisky flavour diversification by yeast","authors":"Martina Daute, Frances Jack, Graeme Walker","doi":"10.1093/femsyr/foae017","DOIUrl":"https://doi.org/10.1093/femsyr/foae017","url":null,"abstract":"Scotch Whisky, a product of high importance to Scotland, has gained global approval for its distinctive qualities derived from the traditional production process which is defined in law. However, ongoing research continuously enhances Scotch Whisky production and is fostering a diversification of flavour profiles. To be classified as Scotch Whisky, the final spirit needs to retain the aroma and taste of “Scotch”. While each production step contributes significantly to whisky flavour—from malt preparation and mashing to fermentation, distillation, and maturation—the impact of yeast during fermentation is crucially important. Not only does the yeast convert the sugar to alcohol, it also produces important volatile compounds, for example esters and higher alcohols, that contribute to the final flavour profile of whisky. The yeast chosen for whisky fermentations can significantly influence whisky flavour, so the yeast strain employed is of high importance. This review explores the role of yeast in Scotch Whisky production and its influence on flavour diversification. Furthermore, an extensive examination of non-conventional yeasts employed in brewing and winemaking is undertaken to assess their potential suitability for adoption as Scotch Whisky yeast strains, followed by a review of methods for evaluating new yeast strains.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140841263","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Maintenance of asymmetric ion concentrations across cellular membranes is crucial for proper yeast cellular function. Disruptions of these ionic gradients can significantly impact membrane electrochemical potential and the balance of other ions, particularly under stressful conditions such as exposure to acetic acid. This weak acid, ubiquitous to both yeast metabolism and industrial processes, is a major inhibitor of yeast cell growth in industrial settings and a key determinant of host colonization by pathogenic yeast. Acetic acid toxicity depends on medium composition, especially on the pH (H+ concentration), but also on other ions' concentrations. Regulation of ion fluxes is essential for effective yeast response and adaptation to acetic acid stress. However, the intricate interplay among ion balancing systems and stress response mechanisms still presents significant knowledge gaps. This review offers a comprehensive overview of the mechanisms governing ion homeostasis, including H+, K+, Zn2+, Fe2+/3+, and acetate, in the context of acetic acid toxicity, adaptation, and tolerance. While focus is given on Saccharomyces cerevisiae due to its extensive physiological characterization, insights are also provided for biotechnologically and clinically relevant yeast species whenever available.
{"title":"The role of ion homeostasis in adaptation and tolerance to acetic acid stress in yeasts.","authors":"Miguel Antunes, Isabel Sá-Correia","doi":"10.1093/femsyr/foae016","DOIUrl":"https://doi.org/10.1093/femsyr/foae016","url":null,"abstract":"Maintenance of asymmetric ion concentrations across cellular membranes is crucial for proper yeast cellular function. Disruptions of these ionic gradients can significantly impact membrane electrochemical potential and the balance of other ions, particularly under stressful conditions such as exposure to acetic acid. This weak acid, ubiquitous to both yeast metabolism and industrial processes, is a major inhibitor of yeast cell growth in industrial settings and a key determinant of host colonization by pathogenic yeast. Acetic acid toxicity depends on medium composition, especially on the pH (H+ concentration), but also on other ions' concentrations. Regulation of ion fluxes is essential for effective yeast response and adaptation to acetic acid stress. However, the intricate interplay among ion balancing systems and stress response mechanisms still presents significant knowledge gaps. This review offers a comprehensive overview of the mechanisms governing ion homeostasis, including H+, K+, Zn2+, Fe2+/3+, and acetate, in the context of acetic acid toxicity, adaptation, and tolerance. While focus is given on Saccharomyces cerevisiae due to its extensive physiological characterization, insights are also provided for biotechnologically and clinically relevant yeast species whenever available.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140665352","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Anaerobic alcoholic fermentation, particularly in high-sugar environments, presents metabolic challenges for yeasts. Crabtree-positive yeasts, including Saccharomyces cerevisiae, prefer fermentation even in the presence of oxygen. These yeasts rely on internal NAD+ recycling and extracellular assimilation of its precursor, nicotinic acid (vitamin B3), rather than de novo NAD+ production. Surprisingly, nicotinic acid assimilation is poorly characterised, even in S. cerevisiae. This study elucidated the timing of nicotinic acid uptake during grape juice-like fermentation and its impact on NAD(H) levels, the NAD+/NADH ratio, and metabolites produced. Complete uptake of extracellular nicotinic acid occurred pre-mid-exponential phase, thereafter small amounts of vitamin B3 were exported back into the medium. Suboptimal levels of nicotinic acid were correlated with slower fermentation and reduced biomass, disrupting redox balance and impeding NAD+ regeneration, thereby affecting metabolite production. Metabolic outcomes varied with nicotinic acid concentrations, linking NAD+ availability to fermentation efficiency. A model was proposed encompassing rapid nicotinic acid uptake, accumulation during cell proliferation, and recycling with limited vitamin B3 export. This research enhances the understanding of nicotinic acid uptake dynamics during grape juice-like fermentation. These insights contribute to advancing yeast metabolism research and have profound implications for the enhancement of biotechnological practices and the winemaking industry.
{"title":"Nicotinic acid availability impacts redox cofactor metabolism in Saccharomyces cerevisiae during alcoholic fermentation","authors":"James D Duncan, Mathabatha E Setati, Benoit Divol","doi":"10.1093/femsyr/foae015","DOIUrl":"https://doi.org/10.1093/femsyr/foae015","url":null,"abstract":"Anaerobic alcoholic fermentation, particularly in high-sugar environments, presents metabolic challenges for yeasts. Crabtree-positive yeasts, including Saccharomyces cerevisiae, prefer fermentation even in the presence of oxygen. These yeasts rely on internal NAD+ recycling and extracellular assimilation of its precursor, nicotinic acid (vitamin B3), rather than de novo NAD+ production. Surprisingly, nicotinic acid assimilation is poorly characterised, even in S. cerevisiae. This study elucidated the timing of nicotinic acid uptake during grape juice-like fermentation and its impact on NAD(H) levels, the NAD+/NADH ratio, and metabolites produced. Complete uptake of extracellular nicotinic acid occurred pre-mid-exponential phase, thereafter small amounts of vitamin B3 were exported back into the medium. Suboptimal levels of nicotinic acid were correlated with slower fermentation and reduced biomass, disrupting redox balance and impeding NAD+ regeneration, thereby affecting metabolite production. Metabolic outcomes varied with nicotinic acid concentrations, linking NAD+ availability to fermentation efficiency. A model was proposed encompassing rapid nicotinic acid uptake, accumulation during cell proliferation, and recycling with limited vitamin B3 export. This research enhances the understanding of nicotinic acid uptake dynamics during grape juice-like fermentation. These insights contribute to advancing yeast metabolism research and have profound implications for the enhancement of biotechnological practices and the winemaking industry.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140624160","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Marc‐André Lachance, Christopher Burke, Karen Nygard, Marc Courchesne, Alexander V Timoshenko
Although filamentous Ascomycetes may produce structures that are interpreted as male and female gametangia, ascomycetous yeasts are generally not considered to possess male and female sexes. In haplontic yeasts of the genus Metschnikowia, the sexual cycle begins with the fusion of two morphologically identical cells of complementary mating types. Soon after conjugation, a protuberance emerges from one of the conjugants, eventually maturing into an ascus. The originating cell can be regarded as an ascus mother cell, hence as female. We tested the hypothesis that the sexes, female or male, are determined by the mating types. There were good reasons to hypothesize further that mating type α cells are male. In a conceptually simple experiment, we observed the early stages of the mating reaction of mating types differentially labeled with fluorescent concanavalin A conjugates. Three large-spored Metschnikowia species, M. amazonensis, M. continentalis, and M. matae, were examined. In all three, the sexes were found to be independent of mating type, cautioning that the two terms should not be used interchangeably.
尽管丝状子囊菌可能产生被解释为雌雄配子器的结构，但一般认为子囊酵母菌不具有雌雄性别。在梅茨尼科维亚属（Metschnikowia）的单倍体酵母菌中，性周期始于两个形态相同的互补交配型细胞的融合。结合后不久，从其中一个结合体中出现一个突起，最终成熟为腹腔。起源细胞可被视为腹水母细胞，因此是雌性的。我们测试了由交配类型决定性别（雌性或雄性）的假设。我们有充分的理由进一步假设交配型α细胞是雄性的。在一个概念简单的实验中，我们观察了不同交配类型的交配反应的早期阶段，这些交配类型的交配反应是用荧光凝集素A共轭物标记的。我们考察了三个大孢子 Metschnikowia 物种：M. amazonensis、M. continentalis 和 M. matae。在所有这三个物种中，性别都与交配类型无关，因此这两个术语不能互换使用。
{"title":"Yeast sexes: mating types do not determine the sexes in Metschnikowia species.","authors":"Marc‐André Lachance, Christopher Burke, Karen Nygard, Marc Courchesne, Alexander V Timoshenko","doi":"10.1093/femsyr/foae014","DOIUrl":"https://doi.org/10.1093/femsyr/foae014","url":null,"abstract":"Although filamentous Ascomycetes may produce structures that are interpreted as male and female gametangia, ascomycetous yeasts are generally not considered to possess male and female sexes. In haplontic yeasts of the genus Metschnikowia, the sexual cycle begins with the fusion of two morphologically identical cells of complementary mating types. Soon after conjugation, a protuberance emerges from one of the conjugants, eventually maturing into an ascus. The originating cell can be regarded as an ascus mother cell, hence as female. We tested the hypothesis that the sexes, female or male, are determined by the mating types. There were good reasons to hypothesize further that mating type α cells are male. In a conceptually simple experiment, we observed the early stages of the mating reaction of mating types differentially labeled with fluorescent concanavalin A conjugates. Three large-spored Metschnikowia species, M. amazonensis, M. continentalis, and M. matae, were examined. In all three, the sexes were found to be independent of mating type, cautioning that the two terms should not be used interchangeably.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140694115","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Mekonnen M Demeke, Dannele Echemendia, Edgard Belo, María R Foulquié-Moreno, Johan M Thevelein
Major progress in developing Saccharomyces cerevisiae strains that utilize the pentose sugar xylose has been achieved. However, the high inhibitor content of lignocellulose hydrolysates still hinders efficient xylose fermentation, which remains a major obstacle for commercially viable second-generation bioethanol production. Further improvement of xylose utilization in inhibitor-rich lignocellulose hydrolysates remains highly challenging. In this work, we have developed a robust industrial S. cerevisiae strain able to efficiently ferment xylose in concentrated undetoxified lignocellulose hydrolysates. This was accomplished with novel multi-step evolutionary engineering. First, a tetraploid strain was generated and evolved in xylose-enriched pretreated spruce biomass. The best evolved strain was sporulated to obtain a genetically diverse diploid population. The diploid strains were then screened in industrially relevant conditions. The best performing strain, MDS130, showed superior fermentation performance in three different lignocellulose hydrolysates. In concentrated corncob hydrolysate, with initial cell density of 1 g DW/L, at 35°C, MDS130 completely co-consumed glucose and xylose, producing ± 7% v/v ethanol with a yield of 91% of the maximum theoretical value and an overall productivity of 1.22 g/L/h. MDS130 has been developed from previous industrial yeast strains without applying external mutagenesis, minimizing the risk of negative side-effects on other commercially important properties and maximizing its potential for industrial application.
在开发可利用戊糖木糖的酿酒酵母菌株方面取得了重大进展。然而，木质纤维素水解物中的高抑制剂含量仍然阻碍着木糖的高效发酵，这仍然是商业上可行的第二代生物乙醇生产的主要障碍。进一步提高木糖在富含抑制剂的木质纤维素水解物中的利用率仍然极具挑战性。在这项工作中，我们开发出了一种强健的工业 S. cerevisiae 菌株，能够在浓缩的未解毒木质纤维素水解物中高效发酵木糖。这是通过新颖的多步骤进化工程实现的。首先，生成四倍体菌株，并在富含木糖的预处理云杉生物质中进行进化。对进化出的最佳菌株进行孢子培养，以获得基因多样化的二倍体群体。然后在工业相关条件下对二倍体菌株进行筛选。表现最好的菌株 MDS130 在三种不同的木质纤维素水解物中都表现出卓越的发酵性能。在浓缩玉米芯水解物中，初始细胞密度为 1 g DW/L，在 35°C 温度下，MDS130 完全共消耗葡萄糖和木糖，产生 ± 7% v/v 的乙醇，产量为最大理论值的 91%，总生产率为 1.22 g/L/h。MDS130 是在以前的工业酵母菌株基础上开发出来的，没有进行外部诱变，从而最大限度地降低了对其他重要商业特性产生负面副作用的风险，并最大限度地提高了其工业应用潜力。
{"title":"Enhancing xylose fermentation capacity of engineered Saccharomyces cerevisiae by multi-step evolutionary engineering in inhibitor-rich lignocellulose hydrolysate","authors":"Mekonnen M Demeke, Dannele Echemendia, Edgard Belo, María R Foulquié-Moreno, Johan M Thevelein","doi":"10.1093/femsyr/foae013","DOIUrl":"https://doi.org/10.1093/femsyr/foae013","url":null,"abstract":"Major progress in developing Saccharomyces cerevisiae strains that utilize the pentose sugar xylose has been achieved. However, the high inhibitor content of lignocellulose hydrolysates still hinders efficient xylose fermentation, which remains a major obstacle for commercially viable second-generation bioethanol production. Further improvement of xylose utilization in inhibitor-rich lignocellulose hydrolysates remains highly challenging. In this work, we have developed a robust industrial S. cerevisiae strain able to efficiently ferment xylose in concentrated undetoxified lignocellulose hydrolysates. This was accomplished with novel multi-step evolutionary engineering. First, a tetraploid strain was generated and evolved in xylose-enriched pretreated spruce biomass. The best evolved strain was sporulated to obtain a genetically diverse diploid population. The diploid strains were then screened in industrially relevant conditions. The best performing strain, MDS130, showed superior fermentation performance in three different lignocellulose hydrolysates. In concentrated corncob hydrolysate, with initial cell density of 1 g DW/L, at 35°C, MDS130 completely co-consumed glucose and xylose, producing ± 7% v/v ethanol with a yield of 91% of the maximum theoretical value and an overall productivity of 1.22 g/L/h. MDS130 has been developed from previous industrial yeast strains without applying external mutagenesis, minimizing the risk of negative side-effects on other commercially important properties and maximizing its potential for industrial application.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140600887","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Lulu Zhang, Zhou Meng, Richard Calderone, Weida Liu, Xiaodong She, Dongmei Li
How mutations in mitochondrial electron transport chain (ETC) proteins impact the cell cycle of Candida albicans was investigated in this study. Using genetic null mutants targeting ETC complexes I (CI), III (CIII), and IV (CIV), the cell cycle stages (G0/G1, S-phase, and G2/M) were analyzed via fluorescence-activated cell sorting (FACS). Four CI null mutants exhibited distinct alterations, including extended S- phase, shortened G2/M population and a reduction in cells size exceeding 10 μM. Conversely, CIII mutants showed an increased population in G1/G0 phase. Among four CI mutants, ndh51Δ/Δ and goa1Δ/Δ displayed aberrant cell cycle patterns correlated with previously reported cAMP/PKA downregulation. Specifically, nuo1Δ/Δ and nuo2Δ/Δ mutants exhibited increased transcription of RIM15, a central hub linking cell cycle with nutrient-dependent TOR1 and cAMP/PKA pathways and Snf1 aging pathway. These findings suggest that suppression of TOR1 and cAMP/PKA pathways or enhanced Snf1 disrupts cell cycle progression, influencing cell longevity and growth among CI mutants. Overall, our study highlights the intricate interplay between mitochondrial ETC, cell cycle, and signaling pathways.
{"title":"Mitochondria complex I deficiency in Candida albicans arrests the cell cycle at S phase through suppressive TOR and PKA pathways","authors":"Lulu Zhang, Zhou Meng, Richard Calderone, Weida Liu, Xiaodong She, Dongmei Li","doi":"10.1093/femsyr/foae010","DOIUrl":"https://doi.org/10.1093/femsyr/foae010","url":null,"abstract":"How mutations in mitochondrial electron transport chain (ETC) proteins impact the cell cycle of Candida albicans was investigated in this study. Using genetic null mutants targeting ETC complexes I (CI), III (CIII), and IV (CIV), the cell cycle stages (G0/G1, S-phase, and G2/M) were analyzed via fluorescence-activated cell sorting (FACS). Four CI null mutants exhibited distinct alterations, including extended S- phase, shortened G2/M population and a reduction in cells size exceeding 10 μM. Conversely, CIII mutants showed an increased population in G1/G0 phase. Among four CI mutants, ndh51Δ/Δ and goa1Δ/Δ displayed aberrant cell cycle patterns correlated with previously reported cAMP/PKA downregulation. Specifically, nuo1Δ/Δ and nuo2Δ/Δ mutants exhibited increased transcription of RIM15, a central hub linking cell cycle with nutrient-dependent TOR1 and cAMP/PKA pathways and Snf1 aging pathway. These findings suggest that suppression of TOR1 and cAMP/PKA pathways or enhanced Snf1 disrupts cell cycle progression, influencing cell longevity and growth among CI mutants. Overall, our study highlights the intricate interplay between mitochondrial ETC, cell cycle, and signaling pathways.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140600389","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Toni Rendulić, Andreea Perpelea, Juan Paulo Ragas Ortiz, Margarida Casal, Elke Nevoigt
Previously, we reported an engineered Saccharomyces cerevisiae CEN.PK113-1A derivative able to produce succinic acid (SA) from glycerol with net CO2 fixation. Apart from an engineered glycerol utilization pathway, the strain was equipped with the reductive branch of the TCA cycle (rTCA) and a heterologous SA exporter. However, the results indicated that a significant amount of carbon still entered the CO2-releasing oxidative TCA cycle. The current study aimed to tune down the flux through the oxidative TCA cycle by targeting the mitochondrial uptake of pyruvate and cytosolic intermediates of the rTCA pathway, as well as the succinate dehydrogenase complex. Thus, we tested the effects of deletions of MPC1, MPC3, OAC1, DIC1, SFC1, and SDH1 on SA production. The highest improvement was achieved by the combined deletion of MPC3 and SDH1. The respective strain produced up to 45.5 g/L of SA, reached a maximum SA yield of 0.66 gSA/gglycerol, and accumulated the lowest amounts of byproducts. Based on the obtained data, we consider a further reduction of mitochondrial import of pyruvate and rTCA intermediates highly attractive. Moreover, the approaches presented in the current study might also be valuable for improving SA production when sugars (instead of glycerol) are the source of carbon.
此前，我们报道了一种工程化的酿酒酵母 CEN.PK113-1A 衍生物，它能够利用甘油生产琥珀酸（SA），并具有净二氧化碳固定能力。除了改造甘油利用途径外，该菌株还配备了 TCA 循环的还原分支（rTCA）和异源 SA 导出器。然而，研究结果表明，仍有大量碳进入释放二氧化碳的氧化 TCA 循环。目前的研究旨在通过靶向丙酮酸的线粒体摄取和 rTCA 途径的细胞膜中间产物以及琥珀酸脱氢酶复合物来降低通过氧化 TCA 循环的通量。因此，我们测试了缺失 MPC1、MPC3、OAC1、DIC1、SFC1 和 SDH1 对 SA 产量的影响。MPC3和SDH1的联合缺失对SA产量的提高最大。该菌株的 SA 产量高达 45.5 克/升，最大 SA 产量为 0.66 克/克甘油，副产物积累量最低。根据所获得的数据，我们认为进一步减少线粒体丙酮酸和 rTCA 中间产物的输入极具吸引力。此外，当糖（而不是甘油）作为碳源时，本研究提出的方法可能对提高 SA 的产量也很有价值。
{"title":"Mitochondrial membrane transporters as attractive targets for the fermentative production of succinic acid from glycerol in Saccharomyces cerevisiae","authors":"Toni Rendulić, Andreea Perpelea, Juan Paulo Ragas Ortiz, Margarida Casal, Elke Nevoigt","doi":"10.1093/femsyr/foae009","DOIUrl":"https://doi.org/10.1093/femsyr/foae009","url":null,"abstract":"Previously, we reported an engineered Saccharomyces cerevisiae CEN.PK113-1A derivative able to produce succinic acid (SA) from glycerol with net CO2 fixation. Apart from an engineered glycerol utilization pathway, the strain was equipped with the reductive branch of the TCA cycle (rTCA) and a heterologous SA exporter. However, the results indicated that a significant amount of carbon still entered the CO2-releasing oxidative TCA cycle. The current study aimed to tune down the flux through the oxidative TCA cycle by targeting the mitochondrial uptake of pyruvate and cytosolic intermediates of the rTCA pathway, as well as the succinate dehydrogenase complex. Thus, we tested the effects of deletions of MPC1, MPC3, OAC1, DIC1, SFC1, and SDH1 on SA production. The highest improvement was achieved by the combined deletion of MPC3 and SDH1. The respective strain produced up to 45.5 g/L of SA, reached a maximum SA yield of 0.66 gSA/gglycerol, and accumulated the lowest amounts of byproducts. Based on the obtained data, we consider a further reduction of mitochondrial import of pyruvate and rTCA intermediates highly attractive. Moreover, the approaches presented in the current study might also be valuable for improving SA production when sugars (instead of glycerol) are the source of carbon.","PeriodicalId":12290,"journal":{"name":"FEMS yeast research","volume":null,"pages":null},"PeriodicalIF":3.2,"publicationDate":"2024-04-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140600718","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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