Background: Saccharomyces cerevisiae, engineered for l-lactic acid production from glucose and xylose, is a promising production host for lignocellulose-to-lactic acid processes. However, the two principal engineering strategies-pyruvate-to-lactic acid conversion with and without disruption of the competing pyruvate-to-ethanol pathway-have not yet resulted in strains that combine high lactic acid yields (YLA) and productivities (QLA) on both sugar substrates. Limitations seemingly arise from a dependency on the carbon source and the aeration conditions, but the underlying effects are poorly understood. We have recently presented two xylose-to-lactic acid converting strains, IBB14LA1 and IBB14LA1_5, which have the l-lactic acid dehydrogenase from Plasmodium falciparum (pfLDH) integrated at the pdc1 (pyruvate decarboxylase) locus. IBB14LA1_5 additionally has its pdc5 gene knocked out. In this study, the influence of carbon source and oxygen on Y LA and QLA in IBB14LA1 and IBB14LA1_5 was investigated. Results: In anaerobic fermentation IBB14LA1 showed a higher Y LA on xylose (0.27gg Xyl -1 ) than on glucose (0.18gg Glc -1 ). The ethanol yields (Y EtOH, 0.15gg Xyl -1 and 0.32gg Glc -1 ) followed an opposite trend. In IBB14LA1_5, the effect of the carbon source on Y LA was less pronounced (~0.80gg Xyl -1 , and 0.67gg Glc -1 ). Supply of oxygen accelerated glucose conversions significantly in IBB14LA1 (QLA from 0.38 to 0.81gL-1h-1) and IBB14LA1_5 (QLA from 0.05 to 1.77g L-1h-1) at constant Y LA (IBB14LA1 ~0.18gg Glc -1 ; IBB14LA1_5 ~0.68gg Glc -1 ). In aerobic xylose conversions, however, lactic acid production ceased completely in IBB14LA1 and decreased drastically in IBB14LA1_5 (Y LA aerobic≤0.25gg Xyl -1 and anaerobic ~0.80gg Xyl -1 ) at similar QLA (~0.04gL-1h-1). Switching from aerobic to microaerophilic conditions (pO2~2%) prevented lactic acid metabolization, observed for fully aerobic conditions, and increased QLA and Y LA up to 0.11gL-1h-1 and 0.38gg Xyl -1 , respectively. The pfLDH and PDC activities in IBB14LA1 were measured and shown to change drastically dependent on carbon source and oxygen. Conclusion: Evidence from conversion time courses together with results of activity measurements for pfLDH and PDC show that in IBB14LA1 the distribution of fluxes at the pyruvate branching point is carbon source and oxygen dependent. Comparison of the performance of strain IBB14LA1 and IBB14LA1_5 in conversions under different aeration conditions (aerobic, anaerobic, and microaerophilic) further suggest that xylose, unlike glucose, does not repress the respiratory response in both strains. This study proposes new genetic engineering targets for rendering genetically engineering S. cerevisiae better suited for lactic acid biorefineries.
ASJC Scopus subject areas
- !!Applied Microbiology and Biotechnology