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The Penicillium chrysogenum transporter PcAraT enables high-affinity, glucose-insensitive l-arabinose transport in Saccharomyces cerevisiae
- Source :
- Biotechnology for Biofuels, Vol 11, Iss 1, Pp 1-16 (2018)
- Publication Year :
- 2018
- Publisher :
- BMC, 2018.
-
Abstract
- Abstract 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 × 10ā3 and 1.8 g Lā1, 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 Lā1 l-arabinose and 20 g Lā1 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.
Details
- Language :
- English
- ISSN :
- 17546834
- Volume :
- 11
- Issue :
- 1
- Database :
- Directory of Open Access Journals
- Journal :
- Biotechnology for Biofuels
- Publication Type :
- Academic Journal
- Accession number :
- edsdoj.1f4f9f36c7364f7b83c83e3e784cda77
- Document Type :
- article
- Full Text :
- https://doi.org/10.1186/s13068-018-1047-6