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Engineering furfural tolerance in Escherichia coli improves the fermentation of lignocellulosic sugars into renewable chemicals.

Proceedings of the National Academy of Sciences of the United States of America (2013-02-23)
Xuan Wang, Lorraine P Yomano, James Y Lee, Sean W York, Huabao Zheng, Michael T Mullinnix, K T Shanmugam, Lonnie O Ingram
ABSTRACT

Pretreatments such as dilute acid at elevated temperature are effective for the hydrolysis of pentose polymers in hemicellulose and also increase the access of enzymes to cellulose fibers. However, the fermentation of resulting syrups is hindered by minor reaction products such as furfural from pentose dehydration. To mitigate this problem, four genetic traits have been identified that increase furfural tolerance in ethanol-producing Escherichia coli LY180 (strain W derivative): increased expression of fucO, ucpA, or pntAB and deletion of yqhD. Plasmids and integrated strains were used to characterize epistatic interactions among traits and to identify the most effective combinations. Furfural resistance traits were subsequently integrated into the chromosome of LY180 to construct strain XW129 (LY180 ΔyqhD ackA::PyadC'fucO-ucpA) for ethanol. This same combination of traits was also constructed in succinate biocatalysts (Escherichia coli strain C derivatives) and found to increase furfural tolerance. Strains engineered for resistance to furfural were also more resistant to the mixture of inhibitors in hemicellulose hydrolysates, confirming the importance of furfural as an inhibitory component. With resistant biocatalysts, product yields (ethanol and succinate) from hemicellulose syrups were equal to control fermentations in laboratory media without inhibitors. The combination of genetic traits identified for the production of ethanol (strain W derivative) and succinate (strain C derivative) may prove useful for other renewable chemicals from lignocellulosic sugars.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Furfural, 99%
Sigma-Aldrich
Succinic acid, BioXtra, BioRenewable, ≥99.0%
Sigma-Aldrich
Succinic acid, ReagentPlus®, BioRenewable, ≥99.0%
Sigma-Aldrich
Succinic acid, ACS reagent, ≥99.0%
Sigma-Aldrich
Succinic acid, puriss. p.a., ACS reagent, ≥99.5% (T)
Sigma-Aldrich
Furfural, ≥98%, FCC, FG
Sigma-Aldrich
Furfural, natural, ≥98%, FCC, FG
Sigma-Aldrich
Succinic acid, BioReagent, BioRenewable, suitable for cell culture, suitable for insect cell culture
Sigma-Aldrich
Furfural, ACS reagent, 99%
Sigma-Aldrich
Succinic acid, purum p.a., ≥99.0% (T)
Supelco
Succinic acid, certified reference material, TraceCERT®