Production of d-glucaric acid with phosphoglucose isomerase-deficient Saccharomyces cerevisiae

Mervi Toivari (Corresponding Author), Maija Leena Vehkomäki, Laura Ruohonen, Merja Penttilä, Marilyn G. Wiebe

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Abstract

d-Glucaric acid is a potential biobased platform chemical. Previously mainly Escherichia coli, but also the yeast Saccharomyces cerevisiae, and Pichia pastoris, have been engineered for conversion of d-glucose to d-glucaric acid via myo-inositol. One reason for low yields from the yeast strains is the strong flux towards glycolysis. Thus, to decrease the flux of d-glucose to biomass, and to increase d-glucaric acid yield, the four step d-glucaric acid pathway was introduced into a phosphoglucose isomerase deficient (Pgi1p-deficient) Saccharomyces cerevisiae strain. High d-glucose concentrations are toxic to the Pgi1p-deficient strains, so various feeding strategies and use of polymeric substrates were studied. Uniformly labelled 13C-glucose confirmed conversion of d-glucose to d-glucaric acid. In batch bioreactor cultures with pulsed d-fructose and ethanol provision 1.3 g d-glucaric acid L−1 was produced. The d-glucaric acid titer (0.71 g d-glucaric acid L−1) was lower in nitrogen limited conditions, but the yield, 0.23 g d-glucaric acid [g d-glucose consumed]−1, was among the highest that has so far been reported from yeast. Accumulation of myo-inositol indicated that myo-inositol oxygenase activity was limiting, and that there would be potential to even higher yield. The Pgi1p-deficiency in S. cerevisiae provides an approach that in combination with other reported modifications and bioprocess strategies would promote the development of high yield d-glucaric acid yeast strains.

Original languageEnglish
Pages (from-to)69-83
Number of pages15
JournalBiotechnology Letters
Volume46
DOIs
Publication statusPublished - Feb 2024
MoE publication typeA1 Journal article-refereed

Funding

Open Access funding provided by Technical Research Centre of Finland. This study was financially supported by the Academy of Finland through the grants Centre of Excellence in White Biotechnology—Green Chemistry (grant 118573), and SA SBio (grant 310191), and by the European Commission through the Seventh Framework Programme (FP7/2007-2013) under grant agreement N° FP7-241566 BIOCORE. This study was financially supported by the Academy of Finland through the grants Centre of Excellence in White Biotechnology—Green Chemistry (grant 118573), and SA SBio (grant 310191), and by the European Commission through the Seventh Framework Programme (FP7/2007-2013) under grant agreement N° FP7-241566 BIOCORE which are gratefully acknowledged. We thank Toni Paasikallio for assistance with fermentations, and Tuulikki Seppänen-Laakso, Kaarina Viljanen and Matti Hölttä for assistance with GC-MS measurements.

Keywords

  • Glucarate
  • Glucaric acid
  • Metabolic engineering
  • Myo-inositol
  • Phosphoglucose isomerase
  • Saccharomyces cerevisiae

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