Comparison of genome‐wide selection strategies to identify furfural tolerance genes in <i>Escherichia coli</i>

Glebes, Tirzah Y.; Sandoval, Nicholas R.; Gillis, Jacob H.; Gill, Ryan T.

doi:10.1002/bit.25325

Cited by 29 publications

(25 citation statements)

References 68 publications

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“…For the e cient fermentation of lignocellulosic hydrolysates, it is highly desired to improve strain tolerance to speci c stresses, such as the inhibitors inevitably released from the pretreatment of lignocellulose biomass and the high temperature employed in the bioprocess of simultaneous sacchari cation and ethanol fermentation (SSF) [3,4]. Studies on genetic analysis to stress tolerance have yielded several bene cial genetic traits associated with furfural or acetic acid tolerance [5][6][7]. The deletion or overexpression of some resistant structural genes has proven useful for improving the strain tolerance to a single inhibitor [8][9][10].…”

Section: Introductionmentioning

confidence: 99%

Engineering prokaryotic regulator IrrE to enhance stress tolerance in budding yeast

Wang

et al. 2020

Preprint

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Background: Stress tolerance is one of the important desired microbial traits for industrial bioprocess and global regulatory protein engineering is an efficient approach to improve strain tolerance. In our study, IrrE, a global regulatory protein from prokaryotic organism Deinococcus radiodurans, was engineered to confer yeast the improved tolerance to the inhibitors in lignocellulose hydrolysates or high temperature.Results: Three IrrE mutants were developed through directed evolution and the expression of these mutants could improve the yeast fermentation rate by 3- to 4-fold in the presence of multiple inhibitors. Subsequently, the tolerance to multiple inhibitors of single-site mutants based on the mutations from the variants was then evaluated, and eleven mutants including L65P, I103T, E119V, L160F, P162S, M169V, V204A, R244G, Base 824 Deletion, V299A and A300V were identified to be critical for the improved FAP tolerance. Further studies indicated that IrrE caused genome-wide transcriptional perturbation in yeast, and the mutant I24 led to the rapid growth of Saccharomyces cerevisiae mainly by regulating the transcription level of transcription activators/factors, protecting the intracellular environment and enhancing the antioxidant capacity under inhibitor environment, which reflected the plasticity of IrrE. Meanwhile, we observed that the expression of the wild-type or mutant IrrE could also protect Saccharomyces cerevisiae from the damage caused by thermal stress. The recombinant yeast strains were able to grow with glucose at 42 ºC.Conclusions: IrrE from Deinococcus radiodurans can be engineered as a tolerance-enhancer for Saccharomyces cerevisiae. Systematic research on the regulatory model and mechanism of a prokaryotic global regulatory factor IrrE to increase yeast tolerance provided valuable insights for the improvement of microbial tolerance to complex industrial stress conditions.

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Section: Introductionmentioning

confidence: 99%

Engineering prokaryotic regulator IrrE to enhance stress tolerance in budding yeast

Wang

et al. 2020

Preprint

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“…Furfural is a key aldehyde inhibitor which acts synergistically with other inhibitors and presents one of the most potent toxic combination with 5-HMF [9]. Different cellular components involved in maintaining physiological homeostasis have been involved in conferring tolerance against furfural [10,11]. Components of DNA repair [10,12], cell membrane and chaperones [11], and MDR e ux pumps [13] have been reported to confer tolerance against furfural.…”

Section: Introductionmentioning

confidence: 99%

Deletion of pgi gene in E. coli increases tolerance to furfural and 5-hydroxymethyl furfural in media containing glucose-xylose mixture

Jilani

Dev

Eqbal

et al. 2020

Preprint

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Background: Furfural and 5-hydroxymethyl furfural (5-HMF) are key furan inhibitors that are generated due to breakdown of lignocellulosic sugars at high temperature and acidic treatment conditions. Both furfural and 5-HMF act in a synergistic manner to inhibit microbial metabolism and resistance to both is a desirable characteristic for efficient conversion of lignocellulosic carbon to ethanol. Genetic manipulations targeted toward increasing cellular NADPH pools have successfully imparted tolerance against furfural and 5-HMF. In present study, deletion of pgi gene as a strategy to augment carbon flow through pentose phosphate pathway (PPP) was studied in ethanologenic Escherichia coli strain SSK101 to impart tolerance towards either furfural or 5-HMFor both inhibitors together.Results: A key gene of EMP pathway, pgi, was deleted in an ethanologenic E. coli strain SSK42 to yield strain SSK101. In presence of 1 g/L furfural in minimal AM1 media, the rate of biomass formation for strain SSK101 was up to 1.9-fold higher as compared to parent SSK42 strain, and it was able to clear furfural in half the time. Tolerance to inhibitor was associated with glucose as carbon source and not xylose, and the tolerance advantage of SSK101 was neutralized in LB media. Bioreactor studies were performed under binary stress of furfural and 5-HMF (1 g/L each) and different glucose concentrations in a glucose-xylose mixture with final sugar concentration of 5.5%, mimicking major components of dilute acid treated biomass hydrolysate. In the mixture having 6 g/L and 12 g/L glucose, SSK101 strain produced ~18 g/L and 20 g/L ethanol, respectively. Interestingly, the maximum ethanol productivity was better at lower glucose load with 0.46 g/(L.h) between 96-120 h, as compared to higher glucose load where it was 0.33 g/(L.h) between 144-168 h. Importantly, parent strain SSK42 did not exhibit significant metabolic activity under similar conditions of inhibitor load and sugar concentration.Conclusions: E. coli strain SSK101 with pgi deletion had enhanced tolerance against both furfural and 5-HMF, which was associated with presence of glucose in media. Strain SSK101 also had improved fermentation characteristics under both hyperosmotic as well as binary stress of furfural and 5-HMF in media containing glucose-xylose mixture.

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“…Many efforts have been made to develop furfural-resistant fermentative strains. For example, the pentose phosphate pathway, c-aminobutyric acid (GABA) shunt, cofactor interconversion, high osmolality glycerol signalling and DNA binding processes seem to be associated with growth improvement under furfural stress (Modig et al, 2002;Gorsich et al, 2006;Kim et al, 2012;Wang et al, 2013a;Glebes et al, 2015). Several genes under furfural stress, which are involved in stress tolerance (e.g.…”

Section: Introductionmentioning

confidence: 99%

Intracellular metabolite profiling of Saccharomyces cerevisiae evolved under furfural

Jung

Kim

Yang

et al. 2016

Microbial Biotechnology

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SummaryFurfural, one of the most common inhibitors in pre‐treatment hydrolysates, reduces the cell growth and ethanol production of yeast. Evolutionary engineering has been used as a selection scheme to obtain yeast strains that exhibit furfural tolerance. However, the response of Saccharomyces cerevisiae to furfural at the metabolite level during evolution remains unknown. In this study, evolutionary engineering and metabolomic analyses were applied to determine the effects of furfural on yeasts and their metabolic response to continuous exposure to furfural. After 50 serial transfers of cultures in the presence of furfural, the evolved strains acquired the ability to stably manage its physiological status under the furfural stress. A total of 98 metabolites were identified, and their abundance profiles implied that yeast metabolism was globally regulated. Under the furfural stress, stress‐protective molecules and cofactor‐related mechanisms were mainly induced in the parental strain. However, during evolution under the furfural stress, S. cerevisiae underwent global metabolic allocations to quickly overcome the stress, particularly by maintaining higher levels of metabolites related to energy generation, cofactor regeneration and recovery from cellular damage. Mapping the mechanisms of furfural tolerance conferred by evolutionary engineering in the present study will be led to rational design of metabolically engineered yeasts.

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Comparison of genome‐wide selection strategies to identify furfural tolerance genes in Escherichia coli

Cited by 29 publications

References 68 publications

Engineering prokaryotic regulator IrrE to enhance stress tolerance in budding yeast

Engineering prokaryotic regulator IrrE to enhance stress tolerance in budding yeast

Deletion of pgi gene in E. coli increases tolerance to furfural and 5-hydroxymethyl furfural in media containing glucose-xylose mixture

Intracellular metabolite profiling of Saccharomyces cerevisiae evolved under furfural

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