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

Mahsa Babaei, Kanchana Rueksomtawin Kildegaard, Aligholi Niaei, Maryam Hosseini, Sirous Ebrahimi, Suresh Sudarsan, Irini Angelidaki, Irina Borodina*

*Corresponding author for this work

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Abstract

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.
Original languageEnglish
Article number361
JournalFrontiers in Bioengineering and Biotechnology
Volume7
Number of pages14
ISSN2296-4185
DOIs
Publication statusPublished - 2019

Cite this

@article{4cb8b917acad41f096f5c341533134c8,
title = "Engineering Oleaginous Yeast as the Host for Fermentative Succinic Acid Production From Glucose",
abstract = "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.",
author = "Mahsa Babaei and Kildegaard, {Kanchana Rueksomtawin} and Aligholi Niaei and Maryam Hosseini and Sirous Ebrahimi and Suresh Sudarsan and Irini Angelidaki and Irina Borodina",
year = "2019",
doi = "10.3389/fbioe.2019.00361",
language = "English",
volume = "7",
journal = "Frontiers in Bioengineering and Biotechnology",
issn = "2296-4185",
publisher = "Frontiers Media",

}

Engineering Oleaginous Yeast as the Host for Fermentative Succinic Acid Production From Glucose. / Babaei, Mahsa; Kildegaard, Kanchana Rueksomtawin; Niaei, Aligholi; Hosseini, Maryam; Ebrahimi, Sirous; Sudarsan, Suresh; Angelidaki, Irini; Borodina, Irina.

In: Frontiers in Bioengineering and Biotechnology, Vol. 7, 361, 2019.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Engineering Oleaginous Yeast as the Host for Fermentative Succinic Acid Production From Glucose

AU - Babaei, Mahsa

AU - Kildegaard, Kanchana Rueksomtawin

AU - Niaei, Aligholi

AU - Hosseini, Maryam

AU - Ebrahimi, Sirous

AU - Sudarsan, Suresh

AU - Angelidaki, Irini

AU - Borodina, Irina

PY - 2019

Y1 - 2019

N2 - 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.

AB - 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.

U2 - 10.3389/fbioe.2019.00361

DO - 10.3389/fbioe.2019.00361

M3 - Journal article

C2 - 31828067

VL - 7

JO - Frontiers in Bioengineering and Biotechnology

JF - Frontiers in Bioengineering and Biotechnology

SN - 2296-4185

M1 - 361

ER -