초록
<P>Fumaric acid (FA) is a promising biomass-derived building-block chemical. Bio-based FA production from renewable feedstock is a promising and sustainable alternative to petroleum-based chemical synthesis. Here we report on FA production by direct fermentation using metabolically engineered <I>Saccharomyces cerevisiae</I> with the aid of <I>in silico</I> analysis of a genome-scale metabolic model. First, <I>FUM1</I> was selected as the target gene on the basis of extensive literature mining. Flux balance analysis (FBA) revealed that <I>FUM1</I> deletion can lead to FA production and slightly lower growth of <I>S. cerevisiae</I>. The engineered <I>S. cerevisiae</I> strain obtained by deleting <I>FUM1</I> can produce FA up to a concentration of 610±31 mg L<SUP>–1</SUP> without any apparent change in growth in fed-batch culture. FT-IR and <SUP>1</SUP>H and <SUP>13</SUP>C NMR spectra confirmed that FA was synthesized by the engineered <I>S. cerevisiae</I> strain. FBA identified pyruvate carboxylase as one of the factors limiting higher FA production. When the <I>RoPYC</I> gene was introduced, <I>S. cerevisiae</I> produced 1134±48 mg L<SUP>–1</SUP> FA. Furthermore, the final engineered <I>S. cerevisiae</I> strain was able to produce 1675±52 mg L<SUP>–1</SUP> FA in batch culture when the <I>SFC1</I> gene encoding a succinate–fumarate transporter was introduced. These results demonstrate that the model shows great predictive capability for metabolic engineering. Moreover, FA production in <I>S. cerevisiae</I> can be efficiently developed with the aid of <I>in silico</I> metabolic engineering.</P>