title: Transport and metabolism of fumaric acid in Saccharomyces cerevisiae in aerobic glucose-limited chemostat culture.
abstract: Currently, research is being focused on the industrial-scale production of fumaric acid and other relevant organic acids from renewable feedstocks via fermentation, preferably at low pH for better product recovery. However, at low pH a large fraction of the extracellular acid is present in the undissociated form, which is lipophilic and can diffuse into the cell. There have been no studies done on the impact of high extracellular concentrations of fumaric acid under aerobic conditions in S. cerevisiae, which is a relevant issue to study for industrial-scale production. In this work we studied the uptake and metabolism of fumaric acid in S. cerevisiae in glucose-limited chemostat cultures at a cultivation pH of 3.0 (pH < pK). Steady states were achieved with different extracellular levels of fumaric acid, obtained by adding different amounts of fumaric acid to the feed medium. The experiments were carried out with the wild-type S. cerevisiae CEN.PK 113-7D and an engineered S. cerevisiae ADIS 244 expressing a heterologous dicarboxylic acid transporter (DCT-02) from Aspergillus niger, to examine whether it would be capable of exporting fumaric acid. We observed that fumaric acid entered the cells most likely via passive diffusion of the undissociated form. Approximately two-thirds of the fumaric acid in the feed was metabolized together with glucose. From metabolic flux analysis, an increased ATP dissipation was observed only at high intracellular concentrations of fumarate, possibly due to the export of fumarate via an ABC transporter. The implications of our results for the industrial-scale production of fumaric acid are discussed.
authors.affiliation: Department of Biotechnology, Delft University of Technology, The Netherlands.
authors.forename: Mihir V
authors.initials: MV
authors.lastname: Shah
authors.forename: Oscar
authors.initials: O
authors.lastname: van Mastrigt
authors.affiliation: Department of Biotechnology, Delft University of Technology, The Netherlands.
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Heijnen
authors.affiliation: Department of Biotechnology, Delft University of Technology, The Netherlands.
authors.forename: Walter M
authors.initials: WM
authors.lastname: van Gulik
mesh_terms: D000332:Aerobiosis; D000821:Animal Feed / Q000032:analysis; D000900:Anti-Bacterial Agents / Q000378:metabolism; D001234:Aspergillus niger / Q000737:chemistry; D018533:Biomass; D019149:Bioreactors; D002462:Cell Membrane / Q000378:metabolism; D029382:Dicarboxylic Acid Transporters / Q000235:genetics / Q000378:metabolism; D003998:Dicarboxylic Acids / Q000378:metabolism; D005650:Fumarates / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008293:Malates / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D010539:Permeability; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D019802:Succinic Acid / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D029382:Dicarboxylic Acid Transporters; D003998:Dicarboxylic Acids; D005650:Fumarates; D008293:Malates; C030298:malic acid; C032005:fumaric acid; D019802:Succinic Acid; D005947:Glucose; D010100:Oxygen
keywords: fumaric acid; futile cycling; metabolism; permeability coefficient; saccharomyces cerevisiae; transport mechanism