An engineered cryptic Hxt11 sugar transporter facilitates glucose-xylose co-consumption in Saccharomyces cerevisiae

Hyun Yong Shin, Jeroen G Nijland, Paul P de Waal, René M de Jong, Paul Klaassen, Arnold J M Driessen

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Abstract

BACKGROUND: The yeast Saccharomyces cerevisiae is unable to ferment pentose sugars like d-xylose. Through the introduction of the respective metabolic pathway, S. cerevisiae is able to ferment xylose but first utilizes d-glucose before the d-xylose can be transported and metabolized. Low affinity d-xylose uptake occurs through the endogenous hexose (Hxt) transporters. For a more robust sugar fermentation, co-consumption of d-glucose and d-xylose is desired as d-xylose fermentation is in particular prone to inhibition by compounds present in pretreated lignocellulosic feedstocks.

RESULTS: Evolutionary engineering of a d-xylose-fermenting S. cerevisiae strain lacking the major transporter HXT1-7 and GAL2 genes yielded a derivative that shows improved growth on xylose because of the expression of a normally cryptic HXT11 gene. Hxt11 also supported improved growth on d-xylose by the wild-type strain. Further selection for glucose-insensitive growth on d-xylose employing a quadruple hexokinase deletion yielded mutations at N366 of Hxt11 that reversed the transporter specificity for d-glucose into d-xylose while maintaining high d-xylose transport rates. The Hxt11 mutant enabled the efficient co-fermentation of xylose and glucose at industrially relevant sugar concentrations when expressed in a strain lacking the HXT1-7 and GAL2 genes.

CONCLUSIONS: Hxt11 is a cryptic sugar transporter of S. cerevisiae that previously has not been associated with effective d-xylose transport. Mutagenesis of Hxt11 yielded transporters that show a better affinity for d-xylose as compared to d-glucose while maintaining high transport rates. d-glucose and d-xylose co-consumption is due to a redistribution of the sugar transport flux while maintaining the total sugar conversion rate into ethanol. This method provides a single transporter solution for effective fermentation on lignocellulosic feedstocks.

Original languageEnglish
Number of pages13
JournalBiotechnology for Biofuels
Volume8
Issue number176
DOIs
Publication statusPublished - 2-Nov-2015

Keywords

  • Sugar transport
  • Directed evolution
  • Lignocellulose conversion
  • Yeast
  • PERFORMANCE
  • INHIBITION
  • YEAST HEXOSE TRANSPORTERS
  • CANDIDA-INTERMEDIA
  • ETHANOL-PRODUCTION
  • FERMENTATION
  • STRAIN
  • GENES
  • METABOLISM
  • LIGNOCELLULOSE

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