Co-expression of TAL1 and ADH1 in recombinant xylose-fermenting Saccharomyces cerevisiae improves ethanol production from lignocellulosic hydrolysates in the presence of furfural

doi:10.1016/j.jbiosc.2013.07.007

Journal of Bioscience and Bioengineering

Volume 117, Issue 2, February 2014, Pages 165-169

https://doi.org/10.1016/j.jbiosc.2013.07.007 Get rights and content

Lignocellulosic biomass dedicated to bioethanol production usually contains pentoses and inhibitory compounds such as furfural that are not well tolerated by Saccharomyces cerevisiae. Thus, S. cerevisiae strains with the capability of utilizing both glucose and xylose in the presence of inhibitors such as furfural are very important in industrial ethanol production. Under the synergistic conditions of transaldolase (TAL) and alcohol dehydrogenase (ADH) overexpression, S. cerevisiae MT8-1X/TAL–ADH was able to produce 1.3-fold and 2.3-fold more ethanol in the presence of 70 mM furfural than a TAL-expressing strain and a control strain, respectively. We also tested the strains' ability by mimicking industrial ethanol production from hemicellulosic hydrolysate containing fermentation inhibitors, and ethanol production was further improved by 16% when using MT8-1X/TAL–ADH compared to the control strain. Transcript analysis further revealed that besides the pentose phosphate pathway genes TKL1 and TAL1, ADH7 was also upregulated in response to furfural stress, which resulted in higher ethanol production compared to the TAL-expressing strain. The improved capability of our modified strain was based on its capacity to more quickly reduce furfural in situ resulting in higher ethanol production. The co-expression of TAL/ADH genes is one crucial strategy to fully utilize undetoxified lignocellulosic hydrolysate, leading to cost-competitive ethanol production.

Section snippets

Microbial strains and media

Escherichia coli NovaBlue (Novagen, Inc., Madison, WI, USA) was used as the host strain for recombinant DNA manipulation. E. coli was grown in Luria–Bertani medium (10 g/L peptone, 5 g/L yeast extract, and 5 g/L sodium chloride) containing 100 mg/L ampicillin. S. cerevisiae strains were routinely cultivated at 30°C in synthetic medium [SD medium; 6.7 g/L yeast nitrogen base without amino acids (Difco Laboratories, Detroit, MI, USA), 20 g/L glucose] supplemented with appropriate amino acids and

Fermentation performance of TAL/ADH-co-expressing S. cerevisiae strains in the presence and absence of furfural

We determined the effects of furfural on ethanol fermentation with 50 g/L xylose as the sole carbon source in defined medium. Fig. 1 compares the fermentation performance of three strains: S. cerevisiae MT8-1X/404-405 (control strain), MT8-1X/TAL-405 (TAL-expressing strain) and MT8-1X/TAL–ADH (TAL/ADH-co-expressing strain) in the absence and presence of furfural. In the absence of furfural, 50 g/L xylose was fully consumed by MT8-1X/404-405 (Fig. 1A) and MT8-1X/TAL-405 within 48 h (Fig. 1B),

Discussion

Most biorefineries have been developed for the production of bioethanol from biomass, which requires utilization of xylose, but its hydrolysates also negatively affect microbial fermentation due to the presence of highly toxic compounds (21). We constructed a recombinant S. cerevisiae strain not only capable of utilizing xylose but also of producing higher amounts of ethanol in the presence of as high as 70 mM furfural. Previously, overexpression of an ADH-related gene reduced levels of toxic

Acknowledgments

This work has been supported through project P07015 by the New Energy and Industrial Technology Development Organization (NEDO) under the sponsorship of the Ministry of Economy, Trade, and Industry (METI) of Japan. We are also grateful to the Ministry of Higher Education of Malaysia and University Malaysia Perlis (UniMAP), Malaysia for providing a scholarship to Ku Syahidah Ku Ismail at Kobe University, Japan.

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Journal of Bioscience and Bioengineering

Section snippets

Microbial strains and media

Fermentation performance of TAL/ADH-co-expressing S. cerevisiae strains in the presence and absence of furfural

Discussion

Acknowledgments

References (23)

A comparative view of metabolite and substrate stress and tolerance in microbial bioprocessing: from biofuels and chemicals, to biocatalysis and bioremediation

Metab. Eng.

Evaluation of an adapted inhibitor-tolerant yeast strain for ethanol production from combined hydrolysate of softwood

Appl. Energy

Development of yeast cell factories for consolidated bioprocessing of lignocellulose to bioethanol through cell surface engineering

Biotechnol. Adv.

Deletion of the PHO13 gene in Saccharomyces cerevisiae improves ethanol production from lignocellulosic hydrolysate in the presence of acetic and formic acids, and furfural

Bioresour. Technol.

Overexpression of the yeast transcription activator Msn2 confers furfural resistance and increases the initial fermentation rate in ethanol production

J. Biosci. Bioeng.

Improvement of ethanol productivity during xylose and glucose co-fermentation by xylose-assimilating S. cerevisiae via expression of glucose transporter Sut1

Enzyme Microb. Technol.

Repeated-batch fermentation of lignocellulosic hydrolysate to ethanol using a hybrid Saccharomyces cerevisiae strain metabolically engineered for tolerance to acetic and formic acids

Bioresour. Technol.

Engineering inhibitor tolerance for the production of biorenewable fuels and chemicals

Curr. Opin. Chem. Eng.

Evolutionarily engineered ethanologenic yeast detoxifies lignocellulosic biomass conversion inhibitors by reprogrammed pathways

Mol. Genet. Genomics

Genomic adaptation of ethanologenic yeast to biomass conversion inhibitors

Appl. Microbiol. Biotechnol.

Identification of furfural as a key toxin in lignocellulosic hydrolysates and evolution of a tolerant yeast strain

Microb. Biotechnol.

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Genetic mapping of a bioethanol yeast strain reveals new targets for hydroxymethylfurfural- and thermotolerance

Understanding HMF inhibition on yeast growth coupled with ethanol production for the improvement of bio-based industrial processes

Valorisation of lignocellulosic biomass to value-added products: Paving the pathway towards low-carbon footprint