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https://hdl.handle.net/20.500.14094/90005042
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2024-04-27
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90005042 (fulltext)
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メタデータID
90005042
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open access
出版タイプ
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タイトル
A pyruvate carbon flux tugging strategy for increasing 2,3-butanediol production and reducing ethanol subgeneration in the yeast Saccharomyces cerevisiae
著者
Ishii, Jun ; Morita, Keisuke ; Ida, Kengo ; Kato, Hiroko ; Kinoshita, Shohei ; Hataya, Shoko ; Shimizu, Hiroshi ; Kondo, Akihiko ; Matsuda, Fumio
著者ID
A1290
研究者ID
1000040512546
KUID
https://kuid-rm-web.ofc.kobe-u.ac.jp/search/detail?systemId=c06bf6c66d97f49e520e17560c007669
著者名
Ishii, Jun
石井, 純
イシイ, ジュン
所属機関名
先端バイオ工学研究センター
著者名
Morita, Keisuke
著者名
Ida, Kengo
著者名
Kato, Hiroko
著者名
Kinoshita, Shohei
著者名
Hataya, Shoko
著者名
Shimizu, Hiroshi
著者ID
A1715
研究者ID
1000040205547
KUID
https://kuid-rm-web.ofc.kobe-u.ac.jp/search/detail?systemId=a324eb4a1b052e53520e17560c007669
著者名
Kondo, Akihiko
近藤, 昭彦
コンドウ, アキヒコ
所属機関名
科学技術イノベーション研究科
著者名
Matsuda, Fumio
収録物名
Biotechnology for Biofuels
巻(号)
11
ページ
180-180
出版者
BMC
刊行日
2018-06-26
公開日
2018-07-18
抄録
Background: The yeast Saccharomyces cerevisiae is a promising host cell for producing a wide range of chemicals. However, attempts to metabolically engineer Crabtree-positive S. cerevisiae invariably face a common issue: how to reduce dominant ethanol production. Here, we propose a yeast metabolic engineering strategy for decreasing ethanol subgeneration involving tugging the carbon flux at an important hub branching point (e.g., pyruvate). Tugging flux at a central glycolytic overflow metabolism point arising from high glycolytic activity may substantially increase higher alcohol production in S. cerevisiae. We validated this possibility by testing 2,3-butanediol (2,3-BDO) production, which is routed via pyruvate as the important hub compound. Results: By searching for high-activity acetolactate synthase (ALS) enzymes that catalyze the important first-step reaction in 2,3-BDO biosynthesis, and tuning several fermentation conditions, we demonstrated that a stronger pyruvate pulling effect (tugging of pyruvate carbon flux) is very effective for increasing 2,3-BDO production and reducing ethanol subgeneration by S. cerevisiae. To further confirm the validity of the pyruvate carbon flux tugging strategy, we constructed an evolved pyruvate decarboxylase (PDC)-deficient yeast (PDCΔ) strain that lacked three isozymes of PDC. In parallel with re-sequencing to identify genomic mutations, liquid chromatography-tandem mass spectrometry analysis of intermediate metabolites revealed significant accumulation of pyruvate and NADH in the evolved PDCΔ strain. Harnessing the high-activity ALS and additional downstream enzymes in the evolved PDCΔ strain resulted in a high yield of 2,3-BDO (a maximum of 0.41 g g(-1) glucose consumed) and no ethanol subgeneration, thereby confirming the utility of our strategy. Using this engineered strain, we demonstrated a high 2,3-BDO titer (81.0 g L-1) in a fed-batch fermentation using a high concentration of glucose as the sole carbon source. Conclusions: We demonstrated that the pyruvate carbon flux tugging strategy is very effective for increasing 2,3-BDO production and decreasing ethanol subgeneration in Crabtree-positive S. cerevisiae. High activity of the common first-step enzyme for the conversion of pyruvate, which links to both the TCA cycle and amino acid biosynthesis, is likely important for the production of various chemicals by S. cerevisiae.
キーワード
Pyruvate flux
Crabtree effect
Acetolactate synthase
Ethanol subgeneration
2,3-Butanediol production
Pyruvate decarboxylase (PDC) deficient
カテゴリ
科学技術イノベーション研究科
先端バイオ工学研究センター
学術雑誌論文
権利
© The Author(s) 2018.
This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
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資源タイプ
journal article
言語
English (英語)
eISSN
1754-6834
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関連情報
DOI
https://doi.org/10.1186/s13068-018-1176-y
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