Engineering energetically efficient transport of dicarboxylic acids in yeast
            <i>Saccharomyces cerevisiae</i>

Darbani, Behrooz; Šťovíček, Vratislav; Hoek, Steven Axel van der; Borodina, Irina

doi:10.1073/pnas.1900287116

Cited by 65 publications

(54 citation statements)

References 49 publications

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“…The structure of TMDs and NBDs determine the specificity and efficiency of ABC transporter, which can be redesigned to improve the substrate specificity and transport activity, respectively. From the perspective of cellular economics, the energy-independent or less energy-dependent transporter may be the desired protein for product efflux [43]. In this sense, Snq2p may be the most suitable endogenous transporter for modification in future work because it can secret more β-carotene when consuming the same amount of ATP.…”

Section: Acetate Supplementation Enhances the β-Carotene Secretion Inmentioning

confidence: 99%

Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae

Lin

Cheng

et al. 2020

Biotechnol Biofuels

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Background Product toxicity is one of the bottlenecks for microbial production of biofuels, and transporter-mediated biofuel secretion offers a promising strategy to solve this problem. As a robust microbial host for industrial-scale production of biofuels, Saccharomyces cerevisiae contains a powerful transport system to export a wide range of toxic compounds to sustain survival. The aim of this study is to improve the secretion and production of the hydrophobic product (β-carotene) by harnessing endogenous ABC transporters combined with physiological engineering in S. cerevisiae. Results Substrate inducibility is a prominent characteristic of most endogenous transporters. Through comparative proteomic analysis and transcriptional confirmation, we identified five potential ABC transporters (Pdr5p, Pdr10p, Snq2p, Yor1p, and Yol075cp) for β-carotene efflux. The accumulation of β-carotene also affects cell physiology in various aspects, including energy metabolism, mitochondrial translation, lipid metabolism, ergosterol biosynthetic process, and cell wall synthesis. Here, we adopted an inducible GAL promoter to overexpress candidate transporters and enhanced the secretion and intracellular production of β-carotene, in which Snq2p showed the best performance (a 4.04-fold and a 1.33-fold increase compared with its parental strain YBX-01, respectively). To further promote efflux capacity, two strategies of increasing ATP supply and improving membrane fluidity were following adopted. A 5.80-fold increase of β-carotene secretion and a 1.71-fold increase of the intracellular β-carotene production were consequently achieved in the engineered strain YBX-20 compared with the parental strain YBX-01. Conclusions Overall, our results showcase that engineering endogenous plasma membrane ABC transporters is a promising approach for hydrophobic product efflux in S. cerevisiae. We also highlight the importance of improving cell physiology to enhance the efficiency of ABC transporters, especially energy status and cell membrane properties.

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Section: Acetate Supplementation Enhances the β-Carotene Secretion Inmentioning

confidence: 99%

Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae

Lin

Cheng

et al. 2020

Biotechnol Biofuels

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“…This transporter has been proven to increase malic acid titer significantly in S. cerevisiae (Zelle et al, 2008). We also recently showed that SpMae1 transporter is likely a SLAC1 voltage-dependent anion channel and is thus very energetically efficient (Darbani et al, 2019). Hence, we expressed transporter gene SpMae1 in Δ ACH1 ST8218, to obtain strain ST8413.…”

Section: Resultsmentioning

confidence: 89%

Engineering Oleaginous Yeast as the Host for Fermentative Succinic Acid Production From Glucose

Babaei

Kildegaard

Niaei

et al. 2019

Front. Bioeng. Biotechnol.

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Oleaginous yeast Yarrowia lipolytica is a prospective host for production of succinic acid. The interruption of tricarboxylic acid cycle through succinate dehydrogenase gene (SDH) deletion was reported to result in strains incapable of glucose utilization and this ability had to be restored by chemical mutation or long adaptive laboratory evolution. In this study, a succinate producing strain of Y. lipolytica was engineered by truncating the promoter of SDH1 gene, which resulted in 77% reduction in SDH activity but did not impair the ability of the strain to grow on glucose. The flux toward succinic acid was further improved by overexpressing the genes in the glyoxylate pathway and the oxidative TCA branch, and expressing phosphoenolpyruvate carboxykinase from Actinobacillus succinogenes. A short adaptation on glucose reduced the lag phase of the strain and increased its tolerance to high glucose concentrations. The resulting strain produced 7.8 ± 0.0 g/L succinic acid with a yield of 0.105 g/g glucose in shake flasks without pH control, while mannitol (11.8 ± 0.8 g/L) was the main by-product. Further investigations showed that mannitol accumulation was caused by low pH stress and buffering the fermentation medium eliminated mannitol formation. In a fed-batch bioreactor in mineral medium at pH 5, at which point according to Ka values of succinic acid, the major fraction of product was in acidic form rather than dissociated form, the strain produced 35.3 ± 1.5 g/L succinic acid with 0.26 ± 0.00 g/g glucose yield.

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“…In closing, the use of proton-dependent glucose import by B. bruxellensis may appear expensive from an energetics perspective when compared to the other members of the SLC2 family identified as facilitators (Darbani, Kell and Borodina 2018; Marques et al 2018; Darbani et al 2019). However, this property gives this yeast a competitive edge in nutrient poor environments.…”

Section: Resultsmentioning

confidence: 99%

Identification and characterisation of two high-affinity glucose transporters from the spoilage yeastBrettanomyces bruxellensis

Tiukova

Møller-Hansen

Belew

et al. 2019

FEMS Microbiology Letters

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The yeast Brettanomyces bruxellensis (syn. Dekkera bruxellensis) is an emerging and undesirable contaminant in industrial low-sugar ethanol fermentations that employ the yeast Saccharomyces cerevisiae. High-affinity glucose import in B. bruxellensis has been proposed to be the mechanism by which this yeast can outcompete S. cerevisiae. The present study describes the characterization of two B. bruxellensis genes (BHT1 and BHT3) believed to encode putative high-affinity glucose transporters. In vitro-generated transcripts of both genes as well as the S. cerevisiae HXT7 high-affinity glucose transporter were injected into Xenopus laevis oocytes and subsequent glucose uptake rates were assayed using 14C-labelled glucose. At 0.1 mM glucose, Bht1p was shown to transport glucose five times faster than Hxt7p. pH affected the rate of glucose transport by Bht1p and Bht3p, indicating an active glucose transport mechanism that involves proton symport. These results suggest a possible role for BHT1 and BHT3 in the competitive ability of B. bruxellensis.

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Engineering energetically efficient transport of dicarboxylic acids in yeast Saccharomyces cerevisiae

Cited by 65 publications

References 49 publications

Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae

Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae

Engineering Oleaginous Yeast as the Host for Fermentative Succinic Acid Production From Glucose

Identification and characterisation of two high-affinity glucose transporters from the spoilage yeastBrettanomyces bruxellensis

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