초록
<P><B>Background</B></P><P>2,3-Butanediol is a chemical compound of increasing interest due to its wide applications. It can be synthesized via mixed acid fermentation of pathogenic bacteria such as <I>Enterobacter aerogenes</I> and <I>Klebsiella oxytoca.</I> The non-pathogenic <I>Saccharomyces cerevisiae</I> possesses three different 2,3-butanediol biosynthetic pathways, but produces minute amount of 2,3-butanediol. Hence, we attempted to engineer <I>S. cerevisiae</I> strain to enhance 2,3-butanediol production.</P><P><B>Results</B></P><P>We first identified gene deletion strategy by performing <I>in silico</I> genome-scale metabolic analysis. Based on the best <I>in silico</I> strategy, in which disruption of alcohol dehydrogenase (ADH) pathway is required, we then constructed gene deletion mutant strains and performed batch cultivation of the strains. Deletion of three ADH genes, <I>ADH1, ADH3</I> and <I>ADH5,</I> increased 2,3-butanediol production by 55-fold under microaerobic condition. However, overproduction of glycerol was observed in this triple deletion strain. Additional rational design to reduce glycerol production by <I>GPD2</I> deletion altered the carbon fluxes back to ethanol and significantly reduced 2,3-butanediol production. Deletion of <I>ALD6</I> reduced acetate production in strains lacking major ADH isozymes, but it did not favor 2,3-butanediol production. Finally, we introduced 2,3-butanediol biosynthetic pathway from <I>Bacillus subtilis</I> and <I>E. aerogenes</I> to the engineered strain and successfully increased titer and yield. Highest 2,3-butanediol titer (2.29 g·l<SUP>-1</SUP>) and yield (0.113 g·g<SUP>-1</SUP>) were achieved by <I>Δadh1</I> Δ<I>adh3</I> Δ<I>adh5</I> strain under anaerobic condition.</P><P><B>Conclusions</B></P><P>With the aid of <I>in silico</I> metabolic engineering, we have successfully designed and constructed <I>S. cerevisiae</I> strains with improved 2,3-butanediol production.</P>