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Keywords:

  • yeast;
  • Saccharomyces cerevisiae;
  • l-glycerol 3-phosphate;
  • glycerol;
  • fermentation;
  • industrial biotechnology

Abstract

Interest in l-glycerol 3-phosphate (l-G3P) production via microbial fermentation is due to the compound's potential to replace the unstable substrate dihydroxyacetone phosphate (DHAP) in one-pot enzymatic carbohydrate syntheses. A Saccharomyces cerevisiae strain with deletions in both genes encoding specific l-G3Pases (GPP1 and GPP2) and multicopy overexpression of l-glycerol 3-phosphate dehydrogenase (GPD1) was studied via small-scale (100 mL) batch fermentations under quasi-anaerobic conditions. Intracellular accumulation of l-G3P reached extremely high levels (roughly 200 mM) but thereafter declined. Extracellular l-G3P was also detected and its concentration continuously increased throughout the fermentation, such that most of the total l-G3P was found outside the cells as fermentation concluded. Moreover, in spite of the complete elimination of specific l-G3Pase activity, the strain showed considerable glycerol formation suggesting unspecific dephosphorylation as a mechanism to relieve cells of intracellular l-G3P accumulation. Up-scaling the process employed fed-batch fermentation with repeated glucose feeding, plus an aerobic growth phase followed by an anaerobic product accumulation phase. This produced a final product titer of about 325 mg total l-G3P per liter of fermentation broth. Biotechnol. Bioeng. 2008;100: 497–505. © 2008 Wiley Periodicals, Inc.