High level secretion of cellobiohydrolases by Saccharomyces cerevisiae

Marja Ilmén, Riaan Den Haan, Elena Brevnova, John McBride, Erin Wiswall, Allan Froehlich, Anu Koivula, Sanni P. Voutilainen, Matti Siika-Aho, Danil C. La Grange, Naomi Thorngren, Simon Ahlgren, Mark Mellon, Kristen Deleault, Vineet Rajgarhia, Willem H. Van Zyl, Merja Penttilä (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

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Abstract

Background: The main technological impediment to widespread utilization of lignocellulose for the production of fuels and chemicals is the lack of low-cost technologies to overcome its recalcitrance. Organisms that hydrolyze lignocellulose and produce a valuable product such as ethanol at a high rate and titer could significantly reduce the costs of biomass conversion technologies, and will allow separate conversion steps to be combined in a consolidated bioprocess (CBP). Development of Saccharomyces cerevisiae for CBP requires the high level secretion of cellulases, particularly cellobiohydrolases. Results: We expressed various cellobiohydrolases to identify enzymes that were efficiently secreted by S. cerevisiae. For enhanced cellulose hydrolysis, we engineered bimodular derivatives of a well secreted enzyme that naturally lacks the carbohydrate-binding module, and constructed strains expressing combinations of cbh1 and cbh2 genes. Though there was significant variability in the enzyme levels produced, up to approximately 0.3 g/L CBH1 and approximately 1 g/L CBH2 could be produced in high cell density fermentations. Furthermore, we could show activation of the unfolded protein response as a result of cellobiohydrolase production. Finally, we report fermentation of microcrystalline cellulose (Avicel) to ethanol by CBH-producing S. cerevisiae strains with the addition of beta-glucosidase. Conclusions: Gene or protein specific features and compatibility with the host are important for efficient cellobiohydrolase secretion in yeast. The present work demonstrated that production of both CBH1 and CBH2 could be improved to levels where the barrier to CBH sufficiency in the hydrolysis of cellulose was overcome.

Original languageEnglish
Article number30
JournalBiotechnology for Biofuels
Volume4
DOIs
Publication statusPublished - 14 Sep 2011
MoE publication typeA1 Journal article-refereed

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Cellulose 1,4-beta-Cellobiosidase
secretion
Yeast
Saccharomyces cerevisiae
cellulose
Cellulose
enzyme
fermentation
Enzymes
ethanol
hydrolysis
Fermentation
Hydrolysis
Ethanol
protein
gene
Genes
Technology
Proteins
Unfolded Protein Response

Keywords

  • biofuels
  • cellulolytic yeast
  • UPR

Cite this

Ilmén, Marja ; Den Haan, Riaan ; Brevnova, Elena ; McBride, John ; Wiswall, Erin ; Froehlich, Allan ; Koivula, Anu ; Voutilainen, Sanni P. ; Siika-Aho, Matti ; La Grange, Danil C. ; Thorngren, Naomi ; Ahlgren, Simon ; Mellon, Mark ; Deleault, Kristen ; Rajgarhia, Vineet ; Van Zyl, Willem H. ; Penttilä, Merja. / High level secretion of cellobiohydrolases by Saccharomyces cerevisiae. In: Biotechnology for Biofuels. 2011 ; Vol. 4.
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Ilmén, M, Den Haan, R, Brevnova, E, McBride, J, Wiswall, E, Froehlich, A, Koivula, A, Voutilainen, SP, Siika-Aho, M, La Grange, DC, Thorngren, N, Ahlgren, S, Mellon, M, Deleault, K, Rajgarhia, V, Van Zyl, WH & Penttilä, M 2011, 'High level secretion of cellobiohydrolases by Saccharomyces cerevisiae', Biotechnology for Biofuels, vol. 4, 30. https://doi.org/10.1186/1754-6834-4-30

High level secretion of cellobiohydrolases by Saccharomyces cerevisiae. / Ilmén, Marja; Den Haan, Riaan; Brevnova, Elena; McBride, John; Wiswall, Erin; Froehlich, Allan; Koivula, Anu; Voutilainen, Sanni P.; Siika-Aho, Matti; La Grange, Danil C.; Thorngren, Naomi; Ahlgren, Simon; Mellon, Mark; Deleault, Kristen; Rajgarhia, Vineet; Van Zyl, Willem H.; Penttilä, Merja (Corresponding Author).

In: Biotechnology for Biofuels, Vol. 4, 30, 14.09.2011.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - High level secretion of cellobiohydrolases by Saccharomyces cerevisiae

AU - Ilmén, Marja

AU - Den Haan, Riaan

AU - Brevnova, Elena

AU - McBride, John

AU - Wiswall, Erin

AU - Froehlich, Allan

AU - Koivula, Anu

AU - Voutilainen, Sanni P.

AU - Siika-Aho, Matti

AU - La Grange, Danil C.

AU - Thorngren, Naomi

AU - Ahlgren, Simon

AU - Mellon, Mark

AU - Deleault, Kristen

AU - Rajgarhia, Vineet

AU - Van Zyl, Willem H.

AU - Penttilä, Merja

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PY - 2011/9/14

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N2 - Background: The main technological impediment to widespread utilization of lignocellulose for the production of fuels and chemicals is the lack of low-cost technologies to overcome its recalcitrance. Organisms that hydrolyze lignocellulose and produce a valuable product such as ethanol at a high rate and titer could significantly reduce the costs of biomass conversion technologies, and will allow separate conversion steps to be combined in a consolidated bioprocess (CBP). Development of Saccharomyces cerevisiae for CBP requires the high level secretion of cellulases, particularly cellobiohydrolases. Results: We expressed various cellobiohydrolases to identify enzymes that were efficiently secreted by S. cerevisiae. For enhanced cellulose hydrolysis, we engineered bimodular derivatives of a well secreted enzyme that naturally lacks the carbohydrate-binding module, and constructed strains expressing combinations of cbh1 and cbh2 genes. Though there was significant variability in the enzyme levels produced, up to approximately 0.3 g/L CBH1 and approximately 1 g/L CBH2 could be produced in high cell density fermentations. Furthermore, we could show activation of the unfolded protein response as a result of cellobiohydrolase production. Finally, we report fermentation of microcrystalline cellulose (Avicel) to ethanol by CBH-producing S. cerevisiae strains with the addition of beta-glucosidase. Conclusions: Gene or protein specific features and compatibility with the host are important for efficient cellobiohydrolase secretion in yeast. The present work demonstrated that production of both CBH1 and CBH2 could be improved to levels where the barrier to CBH sufficiency in the hydrolysis of cellulose was overcome.

AB - Background: The main technological impediment to widespread utilization of lignocellulose for the production of fuels and chemicals is the lack of low-cost technologies to overcome its recalcitrance. Organisms that hydrolyze lignocellulose and produce a valuable product such as ethanol at a high rate and titer could significantly reduce the costs of biomass conversion technologies, and will allow separate conversion steps to be combined in a consolidated bioprocess (CBP). Development of Saccharomyces cerevisiae for CBP requires the high level secretion of cellulases, particularly cellobiohydrolases. Results: We expressed various cellobiohydrolases to identify enzymes that were efficiently secreted by S. cerevisiae. For enhanced cellulose hydrolysis, we engineered bimodular derivatives of a well secreted enzyme that naturally lacks the carbohydrate-binding module, and constructed strains expressing combinations of cbh1 and cbh2 genes. Though there was significant variability in the enzyme levels produced, up to approximately 0.3 g/L CBH1 and approximately 1 g/L CBH2 could be produced in high cell density fermentations. Furthermore, we could show activation of the unfolded protein response as a result of cellobiohydrolase production. Finally, we report fermentation of microcrystalline cellulose (Avicel) to ethanol by CBH-producing S. cerevisiae strains with the addition of beta-glucosidase. Conclusions: Gene or protein specific features and compatibility with the host are important for efficient cellobiohydrolase secretion in yeast. The present work demonstrated that production of both CBH1 and CBH2 could be improved to levels where the barrier to CBH sufficiency in the hydrolysis of cellulose was overcome.

KW - biofuels

KW - cellulolytic yeast

KW - UPR

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U2 - 10.1186/1754-6834-4-30

DO - 10.1186/1754-6834-4-30

M3 - Article

AN - SCOPUS:80052569487

VL - 4

JO - Biotechnology for Biofuels

JF - Biotechnology for Biofuels

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