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.
Keywords: Glucarate; Glucaric acid; Metabolic engineering; Myo-inositol; Phosphoglucose isomerase; Saccharomyces cerevisiae.
© 2023. The Author(s).