Monday, April 30, 2007

Metabolic engineering of Saccharomyces cerevisiae microbial cell factories for succinic acid production

Josť Manuel Otero, Jens Nielsen, and Lisbeth Olsson. BioCentrum-DTU, Technical University of Denmark, Building 223, Lyngby, Denmark

Saccharomyces cerevisiae is a robust industrial production platform for various therapeutic agents, high added-value chemicals, and commodities. Central carbon metabolism has been investigaed in detail and glycolytic flux is distributed across C1 (CO2,g), C2 (ethanol, acetate), and C3 (glycerol, pyruvate) products. For the S. cerevisiae CEN.PK113-7D strain cultivated under carbon-limited, aerobic, batch fermentations, the distribution of carbon across biomass, C1, C2, and C3 products is 18, 14, 54, and 9 C-mol/C-mol-glucose, respectively, with <5 C-mol/C-mol-glucose unaccounted for.

A class of added-value chemicals being targeted for biotechnology production is C4 organic acids. Succinic acid is a key building block molecule for further conversion to precursors such as tetrahydrofuran, 1,4-butanediol, and butyrolactone. Succinic acid has the potential to become a commodity chemical, with global demand presently exceeding $2 billion USD/year, and over 160 million kilograms/year petrochemically produced. There are several biomass platforms, all prokaryotic, for succinic acid production; however, S. cerevisiae offers distinct advantages.  It has been awarded GRAS status for use in human consumables, grows well at low pH significantly minimizing purification and acidification costs, and can utilize diverse feedstocks.

Here we describe the use of systems biology tools to drive C6 carbon flux to succinic acid by enhancement of the two native succinic acid producing pathways: the TCA and glyoxylate cycles.  S. cerevisiae does not naturally accumulate succinic acid; however, through the use of in silico metabolic predictions guiding targeted gene deletions and over-expression, mutants that overproduce succinic acid have been engineered and thoroughly characterised.