Strain, medium, and chemicals
All experiments were performed with an industrial yeast strain S.
cerevisiae 4126. Yeast extract peptone dextrose (YPD) medium composed
of 5 g/L yeast extract powder, 3 g/L peptone (Sangon Biotech, Shanghai)
and different levels of glucose (30, 120, 280 g/L) was used. YPD medium
containing 30 g/L and 120 g/L glucose for pre-cultures in flasks and
fermenters was sterilized by the usual method (121°C, 20 min), while the
YPD medium composed of 280 g/L glucose for continuous ethanol
fermentation was sterilized at 110°C for 15 min to minimize the Maillard
reaction between amino acids and reducing sugars. YPD medium containing
of 120 g/L and 280 g/L glucose was perceived as low gravity medium and
very high gravity (VHG) medium, respectively. The following metabolites:
GLC, G6P, F6P, FBP, 6PG, R5P, T6P, TRE, GAP, GOLP, 2+3PG, PEP, PYR, OAA,
CIT, AKG, SUC, FUM, MAL, ATP, ADP, AMP, NAD+, NADH,
NADP+, NADPH, AcCoA (refer to the list of
abbreviations) used as standards for LC-MS/MS analysis were purchased
from Sigma-Aldrich.