Figure 1.
The major metabolic pathway leading to the formation of fumaric acid and in silico carbon flux distribution in the central metabolism of S. cerevisiae during fumaric acid production on glucose.
a/b, “a” represent the flux of parent strain, “b” represent the flux of the mutant strain FMME-002ΔFUM1. Fluxes are shown relative to a glucose uptake rate of 100.
Figure 2.
Robustness analysis for the D-glucose uptake rate and growth rate.
Closed triangle represents the original model iND750, open triangle represents the modified model iND750.
Figure 3.
Fermentation profile for cell growth and product accumulation during aerobic batch culture.
Closed square represents the parent strain, open square represents the mutant strain FMME-002ΔFUM1. (a): growth, (b): residual glucose, (c): fumaric acid, and (d): ethanol. Values are presented as means of three independent experiments. Bars represent the standard deviation.
Table 1.
Flux analysis to select the gene amplification targets for enhanced fumaric acid production.
Figure 4.
Relative gene expression levels.
RoPYC and SFC1 in the mutant strain FMME-002ΔFUM1, FMME-002ΔFUM1+↑RoPYC and FMME-002ΔFUM1+↑RoPYC+↑SFC1, Relative transcription levels were normalized to the transcription level of the β-ACTIN gene, which was taken as 1. The presented values are averages of three independent experiments; the error bars indicate standard deviations.
Figure 5.
The effect of RoPYC over-expression on fermentation profile of engineered strain.
Closed triangle represents the control strain FMME-002ΔFUM1, open triangle represents the mutant strain FMME-002ΔFUM1+↑RoPYC. (a): growth, (b): fumaric acid, (c) : glucose.
Figure 6.
The effect of SFC1 and RoPYC co-expression on fermentation profile of engineered strain.
Closed square represents the control strain FMME-002ΔFUM1+↑RoPYC+ pY26TEF-GPD, open square represents the mutant strain FMME-002ΔFUM1+↑RoPYC+↑SFC1. (a): growth, (b): fumaric acid, (c): glucose.
Table 2.
Strains and plasmids used in this study.
Table 3.
Primers used in this study.
Table 4.
Primers used in the transcriptional analysis.