Synthetic rescue couples NADPH generation to metabolite overproduction in Saccharomyces cerevisiae

Engineering the redox cofactor metabolism is known to be a key challenge in developing a platform strain for biosynthesis of valuable products. Hence, general strategies for manipulation of co-factor metabolism in industrially relevant hosts are of significance. Here, we demonstrate an improvement in α-ketoglutarate (AKG) production in S. cerevisiae using a novel approach based on synthetic rescue. Here, we first perturb the cytosolic NADPH metabolism via deletion of glucose-6-phosphate dehydrogenase (ZWF1). In parallel, we used a strain design algorithm to identify strategies for further improvement in AKG production. Implementation of the identified genetic targets, including disruption of succinyl-CoA Ligase (LSC2) and constitutive expression of NADP+-specific isocitrate dehydrogenases (IDP1 and IDP2) resulted in more than 3 fold improvement in AKG production as compared to the wild type. Our results demonstrate this improvement is due to a synthetic rescue mechanism in which the metabolic flux was redirected towards AKG production through the manipulation of redox cofactors. Disrupting lsc2 in zwf1 mutant improved specific growth rate more than 15% as compared to the zwf1 mutant. In addition, our result suggests that cytosolic isocitrate dehydrogenase (IDP2) may be regulated by isocitrate pools. Together, these results suggest the ability to improve metabolite production via a model guided synthetic rescue mechanism in S. cerevisiae and the potential for using IDP2 expression as a generalized strategy to effectively meet NADPH requirements in engineered strains.