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The Penicillium chr...
The Penicillium chrysogenum transporter PcAraT enables high-affinity, glucose-insensitive l-arabinose transport in Saccharomyces cerevisiae
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Bracher, J. M. (author)
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Verhoeven, M. D. (author)
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Wisselink, H. W. (author)
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Crimi, B. (author)
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Nijland, J. G. (author)
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Driessen, A. J. M. (author)
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Klaassen, P. (author)
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- Van Maris, Antonius J.A. (author)
- KTH,Industriell bioteknologi,Delft University of Technology, Netherlands
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Daran, J. -MG. (author)
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Pronk, J. T. (author)
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(creator_code:org_t)
- 2018-03-13
- 2018
- English.
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In: Biotechnology for Biofuels. - : BioMed Central. - 1754-6834. ; 11:1
- Related links:
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https://biotechnolog...
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
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- Background: l-Arabinose occurs at economically relevant levels in lignocellulosic hydrolysates. Its low-affinity uptake via the Saccharomyces cerevisiae Gal2 galactose transporter is inhibited by d-glucose. Especially at low concentrations of l-arabinose, uptake is an important rate-controlling step in the complete conversion of these feedstocks by engineered pentose-metabolizing S. cerevisiae strains. Results: Chemostat-based transcriptome analysis yielded 16 putative sugar transporter genes in the filamentous fungus Penicillium chrysogenum whose transcript levels were at least threefold higher in l-arabinose-limited cultures than in d-glucose-limited and ethanol-limited cultures. Of five genes, that encoded putative transport proteins and showed an over 30-fold higher transcript level in l-arabinose-grown cultures compared to d-glucose-grown cultures, only one (Pc20g01790) restored growth on l-arabinose upon expression in an engineered l-arabinose-fermenting S. cerevisiae strain in which the endogenous l-arabinose transporter, GAL2, had been deleted. Sugar transport assays indicated that this fungal transporter, designated as PcAraT, is a high-affinity (K m = 0.13 mM), high-specificity l-arabinose-proton symporter that does not transport d-xylose or d-glucose. An l-arabinose-metabolizing S. cerevisiae strain in which GAL2 was replaced by PcaraT showed 450-fold lower residual substrate concentrations in l-arabinose-limited chemostat cultures than a congenic strain in which l-arabinose import depended on Gal2 (4.2 × 10-3 and 1.8 g L-1, respectively). Inhibition of l-arabinose transport by the most abundant sugars in hydrolysates, d-glucose and d-xylose was far less pronounced than observed with Gal2. Expression of PcAraT in a hexose-phosphorylation-deficient, l-arabinose-metabolizing S. cerevisiae strain enabled growth in media supplemented with both 20 g L-1 l-arabinose and 20 g L-1 d-glucose, which completely inhibited growth of a congenic strain in the same condition that depended on l-arabinose transport via Gal2. Conclusion: Its high affinity and specificity for l-arabinose, combined with limited sensitivity to inhibition by d-glucose and d-xylose, make PcAraT a valuable transporter for application in metabolic engineering strategies aimed at engineering S. cerevisiae strains for efficient conversion of lignocellulosic hydrolysates.
Subject headings
- NATURVETENSKAP -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)
Keyword
- l-Arabinose transporter
- Metabolic engineering
- Penicillium
- Proton symport
- Second-generation bioethanol
- Sugar transport
- Transcriptome
- Yeast
Publication and Content Type
- ref (subject category)
- art (subject category)
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- By the author/editor
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Bracher, J. M.
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Verhoeven, M. D.
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Wisselink, H. W.
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Crimi, B.
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Nijland, J. G.
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Driessen, A. J. ...
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show more...
-
Klaassen, P.
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Van Maris, Anton ...
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Daran, J. -MG.
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Pronk, J. T.
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show less...
- About the subject
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- NATURAL SCIENCES
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NATURAL SCIENCES
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and Biological Scien ...
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and Biochemistry and ...
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Biotechnology fo ...
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Royal Institute of Technology