L-Lactic acid production from glucose and xylose with engineered strains of Saccharomyces cerevisiae: aeration and carbon source influence yields and productivities.

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 (Y) and productivities (Q) 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 and Q in IBB14LA1 and IBB14LA1_5 was investigated.

RESULTS

In anaerobic fermentation IBB14LA1 showed a higher Y on xylose (0.27 g g ) than on glucose (0.18 g g ). The ethanol yields (Y, 0.15 g g and 0.32 g g ) followed an opposite trend. In IBB14LA1_5, the effect of the carbon source on Y was less pronounced (~ 0.80 g g , and 0.67 g g ). Supply of oxygen accelerated glucose conversions significantly in IBB14LA1 (Q from 0.38 to 0.81 g L h) and IBB14LA1_5 (Q from 0.05 to 1.77 g L h) at constant Y (IBB14LA1 ~ 0.18 g g ; IBB14LA1_5 ~ 0.68 g g ). In aerobic xylose conversions, however, lactic acid production ceased completely in IBB14LA1 and decreased drastically in IBB14LA1_5 (Y aerobic ≤ 0.25 g g and anaerobic ~ 0.80 g g ) at similar Q (~ 0.04 g L h). Switching from aerobic to microaerophilic conditions (pO ~ 2%) prevented lactic acid metabolization, observed for fully aerobic conditions, and increased Q and Y up to 0.11 g L h and 0.38 g g , 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.