FEMS yeast research最新文献

ErCas12a is a class 2 type V CRISPR-Cas nuclease isolated from Eubacterium rectale with attractive fundamental characteristics, such as RNA self-processing capability, and lacks reach-through royalties typical for Cas nucleases. This study aims to develop a ErCas12a-mediated genome editing tool applicable in the model yeast Saccharomyces cerevisiae. The optimal design parameters for ErCas12a editing in S. cerevisiae were defined as a 21-nt spacer flanked by 19 nt direct repeats expressed from either RNApolII or III promoters, achieving near 100% editing efficiencies in commonly targeted genomic locations. To be able to transfer the ErCas12a genome editing tool to different strain lineages, a transportable platform plasmid was constructed and evaluated for its genome editing efficiency. Using an identical crRNA expression design, the transportable ErCas12a genome editing tool showed lower efficiency when targeting the ADE2 gene. In contrast to genomic Ercas12a expression, episomal expression of Ercas12a decreases maximum specific growth rate on glucose, indicating ErCas12a toxicity at high expression levels. Moreover, ErCas12a processed a multispacer crRNA array using the RNA self-processing capability, which allowed for simultaneous editing of multiple chromosomal locations. ErCas12a is established as a valuable addition to the genetic toolbox for S. cerevisiae.

Ethnic Indian people have been domesticating beneficial microorganisms (bacteria, yeasts, and moulds) by their wisdom of ethno-microbiological knowledge for production of flavoured and socio-culturally preferred fermented foods and alcoholic beverages for more than 8000 years. The purpose of this review is to collate the available literatures of diversity of Saccharomyces and non-Saccharomyces species associated with Indian fermented foods and alcoholic beverages. A colossal diversity of enzyme- and alcohol-producing yeasts under the phylum Ascomycota has been reported from Indian fermented foods and alcoholic beverages. The distributions of yeast species show 13.5% of Saccharomyces cerevisiae and 86.5% of some non-Saccharomyces spp. in Indian fermented foods and alcoholic beverages, based on the reported literatures available till date. There is a research gap on prospect of yeasts research in India. Hence, we suggest that validation of traditional knowledge of domestication of functional yeasts needs to be studied to develop the functional genomics platforms for Saccharomyces and non-Saccharomyces spp. in Indian fermented foods and alcoholic beverages.

Saccharomyces pastorianus, which is responsible for the production of bottom-fermented lager beer, is a hybrid species that arose from the mating of the top-fermenting ale yeast Saccharomyces cerevisiae and the cold-tolerant Saccharomyces eubayanus around the start of the 17th century. Based on detailed analysis of Central European brewing records, we propose that the critical event for the hybridization was the introduction of top-fermenting S. cerevisiae into an environment where S. eubayanus was present, rather than the other way around. Bottom fermentation in parts of Bavaria preceded the proposed hybridization date by a couple of hundred years and we suggest that this was carried out by mixtures of yeasts, which may have included S. eubayanus. A plausible case can be made that the S. cerevisiae parent came either from the Schwarzach wheat brewery or the city of Einbeck, and the formation of S. pastorianus happened in the Munich Hofbräuhaus between 1602 and 1615 when both wheat beer and lager were brewed contemporaneously. We also describe how the distribution of strains from the Munich Spaten brewery, and the development by Hansen and Linder of methods for producing pure starter cultures, facilitated the global spread of the Bavarian S. pastorianus lineages.

In winemaking, the development of new fermentation strategies, such as the use of mixed starter cultures with Saccharomyces cerevisiae (Sc) yeast and non-Saccharomyces (NS) species, requires a better understanding of how yeasts interact, especially at the beginning of fermentation. Despite the growing knowledge on interactions between Sc and NS, few data are available on the interactions between different species of NS. It is furthermore still unclear whether interactions are primarily driven by generic differences between yeast species or whether individual strains are the evolutionarily relevant unit for biotic interactions. This study aimed at acquiring knowledge of the relevance of species and strain in the population dynamics of cocultures between five yeast species: Hanseniaspora uvarum, Lachancea thermotolerans, Starmerella bacillaris, Torulaspora delbrueckii and Sc. We performed cocultures between 15 strains in synthetic grape must and monitored growth in microplates. Both positive and negative interactions were identified. Based on an interaction index, our results showed that the population dynamics seemed mainly driven by the two species involved. Strain level was more relevant in modulating the strength of the interactions. This study provides fundamental insights into the microbial dynamics in early fermentation and contribute to the understanding of more complex consortia encompassing multiple yeasts trains.

Consolidated bioprocessing (CBP) of starch requires recombinant Saccharomyces cerevisiae strains that produce raw starch-degrading enzymes and ferment the resultant sugars to ethanol in a single step. In this study, the native S. cerevisiae COX4 and RPS25A promoter-proximal introns were evaluated for enhanced expression of amylase genes (ateA, temA or temG_Opt) under the control of an S. cerevisiae promoter (ENO1P, TEF1P, TDH3P, or HXT7P). The results showed that different promoters and promoter-intron combinations differentially affected recombinant amylase production: ENO1P-COX4i and TDH3P-RPS25Ai were the best promoters for AteA, followed closely by HXT7P. The latter was also the best promoter for TemA and TemG production, followed closely by TDH3P-RPS25Ai for both these enzymes. Introducing promoter-proximal introns increased amylase activity up to 62% in Y294[ENO-COX-AteA] and Y294[TDH3-RPS-TemA], a significant improvement relative to the intron-less promoters. Strains co-expressing both an α-amylase and glucoamylase genes yielded up to 56 g/L ethanol from 20% w/v raw starch, with a higher carbon conversion observed with strains co-expressing TDH3P-RPS25Ai-temG_Opt than HXT7P-temG_Opt. The study showed that promoter-proximal introns can enhance amylase activity in S. cerevisiae and suggest that these alternative cassettes may also be considered for expression in more efficient ethanol-producing industrial yeast strains for raw starch CBP.

D-xylose utilization by yeasts is an essential feature for improving second-generation ethanol production. However, industrial yeast strains are incapable of consuming D-xylose. Previous analyzes of D-xylose-consuming or fermenting yeast species reveal that the genomic features associated with this phenotype are complex and still not fully understood. Here we present a previously neglected yeast enzyme related to D-xylose metabolism, D-xylose dehydrogenase (XylDH), which is found in at least 105 yeast genomes. By analyzing the XylDH gene family, we brought evidence of gene evolution marked by purifying selection on codons and positive selection evidence in D-xylose-consuming and fermenting species, suggesting the importance of XylDH for D-xylose-related phenotypes in yeasts. Furthermore, although we found no putative metabolic pathway for XylDH in yeast genomes, namely the absence of three bacterial known pathways for this enzyme, we also provide its expression profile on D-xylose media following D-xylose reductase for two yeasts with publicly available transcriptomes. Based on these results, we suggest that XylDH plays an important role in D-xylose usage by yeasts, likely being involved in a cofactor regeneration system by reducing cofactor imbalance in the D-xylose reductase pathway.

The fruit-like aroma of two valine-derived volatiles, isobutanol and isobutyl acetate, has great impact on the flavour and taste of alcoholic beverages, including sake, a traditional Japanese alcoholic beverage. With the growing worldwide interest in sake, breeding of yeast strains with intracellular valine accumulation is a promising approach to meet a demand for sakes with a variety of flavour and taste by increasing the valine-derived aromas. We here isolated a valine-accumulating sake yeast mutant (K7-V7) and identified a novel amino acid substitution, Ala31Thr, on Ilv6, a regulatory subunit for acetohydroxy acid synthase. Expression of the Ala31Thr variant Ilv6 conferred valine accumulation on the laboratory yeast cells, leading to increased isobutanol production. Additionally, enzymatic analysis revealed that Ala31Thr substitution in Ilv6 decreased sensitivity to feedback inhibition by valine. This study demonstrated for the first time that an N-terminal arm conserved in the regulatory subunit of fungal acetohydroxy acid synthase is involved in the allosteric regulation by valine. Moreover, sake brewed with strain K7-V7 contained 1.5-fold higher levels of isobutanol and isobutyl acetate than sake brewed with the parental strain. Our findings will contribute to the brewing of distinctive sakes and the development of yeast strains with increased production of valine-derived compounds.

Yeasts are important microorganisms used in different fermentation processes. The cocoa beans must go through a correct fermentation process to obtain good-quality chocolate, which involves the action of yeasts and bacteria, and yeasts play a crucial role since they act in the first days of fermentation. In coffee, several studies have shown that the microbiota in the fruits is also a relevant factor. The fermentation process (regardless of the processing type) improves the beverage's quality. In this sense, studies using starter cultures in these two raw materials are important for better control of the process, and optimization of fermentation time, in addition to the improvement and diversification of volatile and non-volatile compounds produced by yeasts. Thus, this review discusses the importance and role of yeasts during fermentation, their metabolism, the produced compounds, and how yeast and the different chemical reactions help increase the quality of chocolate and coffee.

Yeasts undergo intensive metabolic changes during the early stages of fermentation. Previous reports suggest the early production of hydrogen sulfide (H2S) is associated with the release of a range of volatile sulfur compounds (VSCs), as well as the production of varietal thiol compounds 3-sulfanylhexan-1-ol (3SH) and 3-sulfanylhexyl acetate (3SHA) from six-carbon precursors, including (E)-hex-2-enal. In this study, we investigated the early H2S potential, VSCs/thiol output, and precursor metabolism of 11 commonly used laboratory and commercial Saccharomyces cerevisiae strains in chemically defined synthetic grape medium (SGM) within 12 h after inoculation. Considerable variability in early H2S potential was observed among the strains surveyed. Chemical profiling suggested that early H2S production correlates with the production of dimethyl disulfide, 2-mercaptoethanol, and diethyl sulfide, but not with 3SH or 3SHA. All strains were capable of metabolizing (E)-hex-2-enal, while the F15 strain showed significantly higher residue at 12 h. Early production of 3SH, but not 3SHA, can be detected in the presence of exogenous (E)-hex-2-enal and H2S. Therefore, the natural variability of early yeast H2S production contributes to the early output of selected VSCs, but the threshold of which is likely not high enough to contribute substantially to free varietal thiols in SGM.