초록
<P><B>Abstract</B></P> <P>Metabolic engineering strategies are often devised to redirect precursor flux in an engineered pathway. To develop a new strategy, the effect of overexpression of the squalene biosynthesis (SB) pathway and squalene overproduction on the ethanol production (EP) and post-squalene biosynthesis (PB) pathways was determined in <I>Saccharomyces cerevisiae</I>. Through overexpression of the <I>HMG1, IDI1, ERG20</I> and <I>ERG9</I> genes of the SB pathway, production of squalene increased 10-fold in the M1EG strain compared to the wild-type strain (WT) [(34mg/L), without terbinafine, an inhibitor of squalene monooxygenase, and 35.02-fold (119.08mg/L) with terbinafine]. However, due to overexpression of the SB pathway and squalene overproduction, production of ethanol and functionality of the EP and PB pathways were synergistically downregulated by 51.61%, 95.86% and 81.79%, respectively, in the M1EG strain compared to the WT strain. Overexpression of the entire SB pathway also enhanced production of squalene by 76.12-fold (304.49mg/L) and synergistically downregulated production of ethanol and functionality of the EP and PB pathways by 66.13%, 97.02% and 87.56% in the FOH-2 strain compared to the WT strain, respectively. The EP and PB pathways were strongly downregulated in the FOH-2 strain compared to the M1EG strain because the FOH-2 strain overexpresses the entire SB pathway and produces more squalene than the M1EG strain.</P> <P>These data suggest that overexpression of the SB pathway and squalene overproduction downregulate the EP and PB pathways in engineered strains. Therefore, we speculate that a cryptic regulation mechanism may downregulate these pathways, and characterization of such a mechanism may enable us to divert the precursor flux from the EP and PB pathways to the squalene biosynthesis pathway.</P> <P><B>Highlights</B></P> <P> <UL> <LI> <I>S. cerevisiae</I> was engineered with SB pathway for squalene overproduction. </LI> <LI> Squalene overproduction effect on the EP and PB biosynthesis pathways was evaluated. </LI> <LI> EP pathway was downregulated by squalene overproduction. </LI> <LI> Expression of PB pathway was also downregulated by squalene overproduction. </LI> <LI> Higher metabolic burden resulted in stronger downregulation of EP and PB pathways. </LI> </UL> </P>