Engineering high‐gravity fermentations for ethanol production at elevated temperature with <i>Saccharomyces cerevisiae</i>

Caspeta, Luis; Coronel, J Hernández; de, Arturo Montes; Abarca, Eduardo; González, L. C. Suárez; Martı́nez, Alfredo

doi:10.1002/bit.27103

Cited by 38 publications

(23 citation statements)

References 32 publications

(96 reference statements)

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“…Gene functions discovered by our screens can be directly applied in yeast biotechnology to generate strains with enhanced heat resistance or high temperature growth, i.e. to facilitate ethanol production 104,105 . If HSRmodulatory roles are conserved in human cells, they can be evaluated as therapeutic targets for disease treatment 7,10 .…”

Section: Crispri Effect Magnitude Depends On Temperature and Phenotypementioning

confidence: 99%

Gene dosage screens in yeast reveal core signalling pathways controlling heat adaptation

Jann

Johansson²,

Smith

et al. 2020

Preprint

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Heat stress causes proteins to unfold and lose their function, jeopardizing essential cellular processes. To protect against heat and proteotoxic stress, cells mount a dedicated stress-protective programme, the so-called heat shock response (HSR). Our understanding of the mechanisms that regulate the HSR and their contributions to heat resistance and growth is incomplete. Here we employ CRISPRi/a to down- or upregulate protein kinases and transcription factors in S. cerevisiae. We measure gene functions by quantifying perturbation effects on HSR activity, thermotolerance, and cellular fitness at 23, 30 and 38°C. The integration of these phenotypes allowed us to identify core signalling pathways of heat adaptation and reveal novel functions for the high osmolarity glycerol, unfolded protein response and protein kinase A pathways in adjusting both thermotolerance and chaperone expression. We further provide evidence for unknown cross-talk of the HSR with the cell cycle-dependent kinase Cdc28, the primary regulator of cell cycle progression. Finally, we show that CRISPRi efficiency is temperature-dependent and that different phenotypes vary in their sensitivity to knock-down. In summary, our study quantifies regulatory gene functions in different aspects of heat adaptation and advances our understanding of how eukaryotic cells counteract proteotoxic and other heat-caused damage.

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Section: Crispri Effect Magnitude Depends On Temperature and Phenotypementioning

confidence: 99%

Gene dosage screens in yeast reveal core signalling pathways controlling heat adaptation

Jann

Johansson²,

Smith

et al. 2020

Preprint

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show abstract

“…The ethanol tolerance phenotype is complex and also influenced by external factors, mainly as temperature, in many studies, there is an inherent interest in the particularity of responses involving similar genes that thermal and ethanolic stress induce and some even demonstrate that tolerance to a factor is dependent on another (Fujita et al, 2006;Caspeta et al, 2019). In this sense, research is aimed at understanding the mechanisms of tolerance of these microorganisms in the face of thermal and ethanolic stress.…”

Section: Introductionmentioning

confidence: 99%

The effects of thermal and ethanolic stress in industrial strains of Saccharomyces cerevisiae

Mueller

Santos

Cardoso

2020

RSD

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Saccharomyces cerevisiae is exceptional microorganisms used in biotechnological processes, mainly in the ethanol production chain. Studies of the cellular responses of industrial yeasts under ethanolic and thermal stress in an association are still incipient. This study aimed to evaluate the action of thermal and ethanolic stress in industrial strains of Saccharomyces cerevisiae under different temperatures and concentrations of ethanol, to understand whether these factors influence ethanol production. For cytotoxicity and genotoxicity tests, yeasts were grown in 2% YPD medium incubated for 10 hours at 250 rpm. After growth, the samples were grown in sugarcane juice in concentrations of 5, 10 and 15% ethanol and incubated at 30 and 40 ºC. In Petri dishes containing the solid medium YPD 2% the yeasts were dripped and incubated for 72 hours the cytotoxic action was analyzed by cell growth and genotoxicity through the comet assay and ethanol production by gas chromatography. Cell growth occurred in all conditions, however, at 30 ºC there was inhibition in 10% (v v-1) of ethanol being potentiated in 15% (v v-1), at 40 ºC. The genotoxicity analysis showed an induction of DNA damage in yeasts, however, the FLE yeast was the one with the highest DNA damage index. The yeast Pedra-2 was more tolerant and produced more ethanol, showing to be a tolerant strain concerning the analyzed fermentative interferents.

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“…Adaptive laboratory evolution (ALE) is considered as a simple yet robust method to develop strain tolerance to any speci c adverse condition [17]. There are numerous studies on Saccharomyces cerevisiae and ethanol production by SSF using evolved strains tolerant to temperature [18,19], pH [20] and other inhibitors [21]. Some studies have attempted to evolve yeast strains for lactate production to leverage its property to tolerate lower pH [22].…”

Section: Introductionmentioning

confidence: 99%

Evolutionary Engineering of Lactobacillus Bulgaricus Reduces Enzyme Usage and Enhances Conversion of Lignocellulosics to D-lactic Acid by Simultaneous Saccharification and Fermentation

Sahoo

Naveen

et al. 2020

Preprint

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BackgroundSimultaneous saccharification and fermentation (SSF) of pre-treated lignocellulosics to biofuels and other platform chemicals has long been a promising alternative to separate hydrolysis and fermentation processes. However, the disparity between the optimum conditions (temperature, pH) for fermentation and enzyme hydrolysis leads to execution of the SSF process at sub-optimal conditions, which can affect the rate of hydrolysis and cellulose conversion. The fermentation conditions could be synchronized with hydrolysis optima by carrying out the SSF at a higher temperature, but this would require a thermo-tolerant organism. Economically viable production of platform chemicals from lignocellulosic biomass has long been stymied because of the significantly higher cost of hydrolytic enzymes. The major objective of this work is to develop an SSF strategy for D- lactic acid production by a thermo-tolerant organism, in which the enzyme loading could significantly be reduced without compromising on the overall conversion. ResultsA thermo-tolerant strain of Lactobacillus bulgaricuswas developed by adaptive laboratory evolution (ALE) which enabled the SSF to be performed at 45 °C with reduced enzyme usage.Despite the reduction of enzyme loading from 15 FPU/gbiomass to 5 FPU/gbiomass, we could still achieve ~8% higher cellulose to D-LA conversion in batch SSF, in comparison to the conversion by separate enzymatic hydrolysis and fermentation processes at 45 °C and pH 5.5. Extending the batch SSF to an SSF with pulse-feeding of 5% pre-treated biomass and 5 FPU/g-biomass, at12-hour intervals (36th h – 96th h), resulted in a titer of 108 g/L D-LA and 60% conversion of cellulose to D-LA.This is one among the highest reported D-LA titers achieved from lignocellulosic biomass.ConclusionsWe have demonstrated that the SSF strategy, in conjunction with evolutionary engineering, could drastically reduce enzyme requirement and be the way forward for economical production of platform chemicals from lignocellulosics. We have shown that fed-batch SSF processes, designed with multiple pulse-feedings of the pre-treated biomass and enzyme, can be an effective way of enhancing the product concentrations.

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Engineering high‐gravity fermentations for ethanol production at elevated temperature with Saccharomyces cerevisiae

Cited by 38 publications

References 32 publications

Gene dosage screens in yeast reveal core signalling pathways controlling heat adaptation

Gene dosage screens in yeast reveal core signalling pathways controlling heat adaptation

The effects of thermal and ethanolic stress in industrial strains of Saccharomyces cerevisiae

Evolutionary Engineering of Lactobacillus Bulgaricus Reduces Enzyme Usage and Enhances Conversion of Lignocellulosics to D-lactic Acid by Simultaneous Saccharification and Fermentation

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