Impact of ionic liquid pretreated plant biomass on Saccharomyces cerevisiae growth and biofuel production

AbstractCellulosic plant biomass is a promising sustainable resource for generating alternative biofuels and biochemicals with microbial factories. But a remaining bottleneck is engineering microbes that are tolerant of toxins generated during biomass processing, because mechanisms of toxin defense are only beginning to emerge. Here, we exploited natural diversity in 165 Saccharomyces cerevisiae strains isolated from diverse geographical and ecological niches, to identify mechanisms of hydrolysate-toxin tolerance. We performed genome-wide association (GWA) analysis to identify genetic variants underlying toxin tolerance, and gene knockouts and allele-swap experiments to validate the involvement of implicated genes. In the process of this work, we uncovered a surprising difference in genetic architecture depending on strain background: in all but one case, knockout of implicated genes had a significant effect on toxin tolerance in one strain, but no significant effect in another strain. In fact, whether or not the gene was involved in tolerance in each strain background had a bigger contribution to strain-specific variation than allelic differences. Our results suggest a major difference in the underlying network of causal genes in different strains, suggesting that mechanisms of hydrolysate tolerance are very dependent on the genetic background. These results could have significant implications for interpreting GWA results and raise important considerations for engineering strategies for industrial strain improvement.Author summaryUnderstanding the genetic architecture of complex traits is important for elucidating the genotype-phenotype relationship. Many studies have sought genetic variants that underlie phenotypic variation across individuals, both to implicate causal variants and to inform on architecture. Here we used genome-wide association analysis to identify genes and processes involved in tolerance of toxins found in plant-biomass hydrolysate, an important substrate for sustainable biofuel production. We found substantial variation in whether or not individual genes were important for tolerance across genetic backgrounds. Whether or not a gene was important in a given strain background explained more variation than the alleleic differences in the gene. These results suggest substantial variation in gene contributions, and perhaps underlying mechanisms, of toxin tolerance.

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Effects of Ultrasound on Fermentation of Glucose to Ethanol by Saccharomyces cerevisiae

Fermentation ◽

10.3390/fermentation5010016 ◽

2019 ◽

Vol 5 (1) ◽

pp. 16 ◽

Cited By ~ 5

Author(s):

Luis Huezo ◽

Ajay Shah ◽

Frederick Michel

Keyword(s):

Saccharomyces Cerevisiae ◽

Mass Transfer ◽

Glucose Uptake ◽

Ethanol Production ◽

Ethanol Yield ◽

Biofuel Production ◽

Inhibitory Effects ◽

Negative Effects ◽

Intraparticle Mass Transfer ◽

Improve Mass Transfer

Previous studies have shown that pretreatment of corn slurries using ultrasound improves starch release and ethanol yield during biofuel production. However, studies on its effects on the mass transfer of substrates and products during fermentation have shown that it can have both beneficial and inhibitory effects. In this study, the effects of ultrasound on mass transfer limitations during fermentation were examined. Calculation of the external and intraparticle observable moduli under a range of conditions indicate that no external or intraparticle mass transfer limitations should exist for the mass transfer of glucose, ethanol, or carbon dioxide. Fermentations of glucose to ethanol using Saccharomyces cerevisiae were conducted at different ultrasound intensities to examine its effects on glucose uptake, ethanol production, and yeast population and viability. Four treatments were compared: direct ultrasound at intensities of 23 and 32 W/L, indirect ultrasound (1.4 W/L), and no-ultrasound. Direct and indirect ultrasound had negative effects on yeast performance and viability, and reduced the rates of glucose uptake and ethanol production. These results indicate that ultrasound during fermentation, at the levels applied, is inhibitory and not expected to improve mass transfer limitations.

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Engineering of a novel cellulose-adherent cellulolytic Saccharomyces cerevisiae for cellulosic biofuel production

Scientific Reports ◽

10.1038/srep24550 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 25

Author(s):

Zhuo Liu ◽

Shih-Hsin Ho ◽

Kengo Sasaki ◽

Riaan den Haan ◽

Kentaro Inokuma ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Biofuel Production ◽

Cellulosic Biofuel

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Different Environmental Stress for Enhanced Biofuel Production from Plant Biomass

Plant Stress Biology ◽

10.1201/9781003055358-2 ◽

2020 ◽

pp. 21-34

Author(s):

S. Joshi ◽

M. Choudhary ◽

N. Srivastava

Keyword(s):

Environmental Stress ◽

Plant Biomass ◽

Biofuel Production

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Transcriptomic Changes Induced by Deletion of Transcriptional Regulator GCR2 on Pentose Sugar Metabolism in Saccharomyces cerevisiae

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2021.654177 ◽

2021 ◽

Vol 9 ◽

Author(s):

Minhye Shin ◽

Heeyoung Park ◽

Sooah Kim ◽

Eun Joong Oh ◽

Deokyeol Jeong ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Xylose Fermentation ◽

Sugar Metabolism ◽

Pentose Phosphate ◽

Biofuel Production ◽

Rna Seq ◽

Loss Of Function ◽

Lignocellulosic Biofuel ◽

Pentose Sugar ◽

Transcriptomic Changes

Being a microbial host for lignocellulosic biofuel production, Saccharomyces cerevisiae needs to be engineered to express a heterologous xylose pathway; however, it has been challenging to optimize the engineered strain for efficient and rapid fermentation of xylose. Deletion of PHO13 (Δpho13) has been reported to be a crucial genetic perturbation in improving xylose fermentation. A confirmed mechanism of the Δpho13 effect on xylose fermentation is that the Δpho13 transcriptionally activates the genes in the non-oxidative pentose phosphate pathway (PPP). In the current study, we found a couple of engineered strains, of which phenotypes were not affected by Δpho13 (Δpho13-negative), among many others we examined. Genome resequencing of the Δpho13-negative strains revealed that a loss-of-function mutation in GCR2 was responsible for the phenotype. Gcr2 is a global transcriptional factor involved in glucose metabolism. The results of RNA-seq confirmed that the deletion of GCR2 (Δgcr2) led to the upregulation of PPP genes as well as downregulation of glycolytic genes, and changes were more significant under xylose conditions than those under glucose conditions. Although there was no synergistic effect between Δpho13 and Δgcr2 in improving xylose fermentation, these results suggested that GCR2 is a novel knockout target in improving lignocellulosic ethanol production.

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Screening and directed evolution of transporters to improve xylodextrin utilization in the yeast Saccharomyces cerevisiae

10.1101/166678 ◽

2017 ◽

Author(s):

Chenlu Zhang ◽

Ligia Acosta-Sampson ◽

Vivian Yaci Yu ◽

Jamie H. D. Cate

Keyword(s):

Saccharomyces Cerevisiae ◽

Directed Evolution ◽

Plant Biomass ◽

Carbon Sources ◽

Industrial Applications ◽

Yeast Saccharomyces Cerevisiae ◽

Cellulosic Biofuel ◽

Economic Production ◽

Report Analysis ◽

Selection Medium

AbstractThe economic production of cellulosic biofuel requires efficient and full utilization of all abundant carbohydrates naturally released from plant biomass by enzyme cocktails. Recently, we reconstituted the Neurospora crassa xylodextrin transport and consumption system in Saccharomyces cerevisiae, enabling growth of yeast on xylodextrins aerobically. However, the consumption rate of xylodextrin requires improvement for industrial applications, including consumption in anaerobic conditions. As a first step in this improvement, we report analysis of orthologues of the N. crassa transporters CDT-1 and CDT-2. Transporter ST16 from Trichoderma virens enables faster aerobic growth of S. cerevisiae on xylodextrins compared to CDT-2. ST16 is a xylodextrin-specific transporter, and the xylobiose transport activity of ST16 is not inhibited by cellobiose. Other transporters identified in the screen also enable growth on xylodextrins including xylotriose. Taken together, these results indicate that multiple transporters might prove useful to improve xylodextrin utilization in S. cerevisiae. Efforts to use directed evolution to improve ST16 from a chromosomally-integrated copy were not successful, due to background growth of yeast on other carbon sources present in the selection medium. Future experiments will require increasing the baseline growth rate of the yeast population on xylodextrins, to ensure that the selective pressure exerted on xylodextrin transport can lead to isolation of improved xylodextrin transporters.

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Efficient Conversion of Glycerol to Ethanol by an Engineered Saccharomyces cerevisiae Strain

Applied and Environmental Microbiology ◽

10.1128/aem.00268-21 ◽

2021 ◽

Author(s):

Sadat M. R. Khattab ◽

Takashi Watanabe

Keyword(s):

Saccharomyces Cerevisiae ◽

Metabolic Flux ◽

Triosephosphate Isomerase ◽

Plant Biomass ◽

Agricultural Waste ◽

Glycerol Dehydrogenase ◽

Cell Wall Polysaccharides ◽

Plant Oil ◽

Cell Factories ◽

Engineered Strain

Glycerol is an eco-friendly solvent that enhances plant biomass decomposition via glycerolysis in many pretreatment methods. Nonetheless, inefficient conversion of glycerol to ethanol by natural Saccharomyces cerevisiae limits its use in these processes. Here, we have developed an efficient glycerol-converting yeast strain by genetically modifying the oxidation of cytosolic nicotinamide adenine dinucleotide (NADH) by an O 2 -dependent dynamic shuttle and abolishing both glycerol phosphorylation and biosynthesis in S. cerevisiae D452-2 strain, as well as vigorous expression of whole genes in the DHA-pathway ( Candid utilis glycerol facilitator, Ogataea polymorpha glycerol dehydrogenase, endogenous dihydroxyacetone kinase, and triosephosphate isomerase). The engineered strain showed conversion efficiencies (CE) up to 0.49 g ethanol/g glycerol (98% of theoretical CE), with production rate >1 g/L −1 h −1 when glycerol was supplemented in a single fed-batch fermentation in a rich medium. Furthermore, the engineered strain converted a mixture of glycerol and glucose into bioethanol (>86 g/L) with 92.8% CE. To the best of our knowledge, this is the highest reported titer of bioethanol produced from glycerol and glucose. Notably, we developed a glycerol-utilizing transformant from parent strain, which cannot utilize glycerol as a sole carbon source. The developed strain converted glycerol to ethanol with a productivity of 0.44 g/L −1 h −1 on minimal medium under semi-aerobic conditions. Our findings will promote the utilization of glycerol in eco-friendly biorefineries and integrate bioethanol and plant-oil industries. IMPORTANCE With the development of efficient lignocellulosic biorefineries, glycerol has attracted attention as an eco-friendly biomass-derived solvent that can enhance the dissociation of lignin and cell wall polysaccharides during the pretreatment process. Co-conversion of glycerol with the sugars released from biomass after glycerolysis increases the resources for ethanol production and lowers the burden of component separation. However, low conversion efficiency from glycerol and sugars limits the industrial application of this process. Therefore, the generation of an efficient glycerol-fermenting yeast will promote the applicability of integrated biorefineries. Hence, metabolic flux control in yeast grown on glycerol will lead to the generation of cell factories that produce chemicals, which will boost biodiesel and bioethanol industries. Additionally, the use of glycerol-fermenting yeast will reduce global warming and generation of agricultural waste, leading to the establishment of a sustainable society.

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Application of Plant Biomass Conversion Products for Biofuel Production

IOP Conference Series Earth and Environmental Science ◽

10.1088/1755-1315/272/2/022100 ◽

2019 ◽

Vol 272 ◽

pp. 022100

Author(s):

I N Tyaglivaya ◽

I Y Zhukova ◽

E N Papina

Keyword(s):

Plant Biomass ◽

Biomass Conversion ◽

Biofuel Production

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Recent Advances on Feasible Strategies for Monoterpenoid Production in Saccharomyces cerevisiae

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2020.609800 ◽

2020 ◽

Vol 8 ◽

Author(s):

Qiyu Gao ◽

Luan Wang ◽

Maosen Zhang ◽

Yongjun Wei ◽

Wei Lin

Keyword(s):

Saccharomyces Cerevisiae ◽

Plant Biomass ◽

Extraction Methods ◽

Active Components ◽

Plant Essential Oils ◽

Microbial Cell Factories ◽

Cell Factories ◽

Plant Extraction ◽

Low Efficiency ◽

Metabolic Activities

Terpenoids are a large diverse group of natural products which play important roles in plant metabolic activities. Monoterpenoids are the main components of plant essential oils and the active components of some traditional Chinese medicinal herbs. Some monoterpenoids are widely used in medicine, cosmetics and other industries, and they are mainly obtained by plant biomass extraction methods. These plant extraction methods have some problems, such as low efficiency, unstable quality, and high cost. Moreover, the monoterpenoid production from plant cannot satisfy the growing monoterpenoids demand. The development of metabolic engineering, protein engineering and synthetic biology provides an opportunity to produce large amounts of monoterpenoids eco-friendly using microbial cell factories. This mini-review covers current monoterpenoids production using Saccharomyces cerevisiae. The monoterpenoids biosynthetic pathways, engineering of key monoterpenoids biosynthetic enzymes, and current monoterpenoids production using S. cerevisiae were summarized. In the future, metabolically engineered S. cerevisiae may provide one possible green and sustainable strategy for monoterpenoids supply.

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