Alleviating glucose repression and enhancing respiratory capacity to increase itaconic acid production

The Crabtree effect products ethanol and acetic acid can be used for itaconic acid (IA) production in Saccharomyces cerevisiae. However, both the IA synthesis and oxidative phosphorylation pathways were hampered by glucose repression when glucose was used as the substrate. This study aimed to improve IA titer by increasing gene expressions related to glucose derepression without impairing yeast growth on glucose. Engineering the acetyl-CoA synthesis pathway increased the titer of IA to 257 mg/L in a urea-based medium. Instead of entire pathway overexpression, we found that some signaling pathways regulating glucose repression were effective targets to improve IA production and respiratory capacity. As a consequence of the reduced inhibition, IA titer was further increased by knocking out a negative regulator of the mitochondrial retrograde signaling MKS1. SNF1/MIG1 signaling was disturbed by deleting the hexokinase HXK2 or an endoplasmic reticulum membrane protein GSF2. The shaking results showed that XYY286 (BY4741, HO::cadA, Y::Dz.ada, 208a::Mt.acs, Δhxk2, pRS415-cadA, pRS423-aac2) accumulated 535 mg/L IA in 168 h in the YSCGLU medium. qRT-PCR results verified that deletion of MKS1 or HXK2 upregulated the gene expressions of the IA synthesis and respiratory pathways during the growth on glucose.