Glucose assimilation rate determines the partition of flux at pyruvate between lactic acid and ethanol in Saccharomyces cerevisiae

Abstract

Engineered Saccharomyces cerevisiae expressing a lactic acid dehydrogenase can metabolize pyruvate into lactic acid. However, three pyruvate decarboxylase (PDC) isozymes drive most carbon flux toward ethanol rather than lactic acid. Deletion of endogenous PDCs will eliminate ethanol production, but the resulting strain suffers from C2 auxotrophy and struggles to complete a fermentation. Engineered yeast assimilating xylose or cellobiose produce lactic acid rather than ethanol as a major product without the deletion of any PDC genes. We report here that sugar flux, but not sensing, contributes to the partition of flux at the pyruvate branch point in S. cerevisiae expressing the Rhizopus oryzae lactic acid dehydrogenase (LdhA). While the membrane glucose sensors Snf3 and Rgt2 did not play any direct role in the option of predominant product, the sugar assimilation rate was strongly correlated to the partition of flux at pyruvate: fast sugar assimilation favors ethanol production while slow sugar assimilation favors lactic acid. Applying this knowledge, we created an engineered yeast capable of simultaneously converting glucose and xylose into lactic acid, increasing lactic acid production to approximately 17 g L−1 from the 12 g L−1 observed during sequential consumption of sugars. This work elucidates the carbon source‐dependent effects on product selection in engineered yeast.

Construction of an alternative NADPH regeneration pathway improves ethanol production in Saccharomyces cerevisiae with xylose metabolic pathway

“…This means glucose competes with xylose for flux throughout both the PPP and EMP. It is theoretically feasible to improve glucose and xylose co-fermentation by limiting the metabolic flux competition of glucose relative to xylose [ 22 ]. One successful approach is that Miskovic L et al reduced the glucose metabolism rate by deleting the hexokinase gene HXK2 [ 23 ], which reduced the rate of glucose phosphorylation, and improved glucose-xylose co-utilization.…”