Genome-scale modeling drives 70-fold improvement of intracellular heme production in Saccharomyces cerevisiae

Olena Ishchuk; Iván Domenzain Del Castillo Cerecer; Benjamín José Sánchez; Facundo Muniz; Jose Luis Martinez Ruiz; Jens B Nielsen; Dina Petranovic Nielsen

doi:10.1073/pnas.2108245119

Genome-scale modeling drives 70-fold improvement of intracellular heme production in Saccharomyces cerevisiae
Journal article, 2022

Heme is an oxygen carrier and a cofactor of both industrial enzymes and food additives. The intracellular level of free heme is low, which limits the synthesis of heme proteins. Therefore, increasing heme synthesis allows an increased production of heme proteins. Using the genome-scale metabolic model (GEM) Yeast8 for the yeast Saccharomyces cerevisiae, we identified fluxes potentially important to heme synthesis. With this model, in silico simulations highlighted 84 gene targets for balancing biomass and increasing heme production. Of those identified, 76 genes were individually deleted or overexpressed in experiments. Empirically, 40 genes individually increased heme production (up to threefold). Heme was increased by modifying target genes, which not only included the genes involved in heme biosynthesis, but also those involved in glycolysis, pyruvate, Fe-S clusters, glycine, and succinyl-coenzyme A (CoA) metabolism. Next, we developed an algorithmic method for predicting an optimal combination of these genes by using the enzyme-constrained extension of the Yeast8 model, ecYeast8. The computationally identified combination for enhanced heme production was evaluated using the heme ligand-binding biosensor (Heme-LBB). The positive targets were combined using CRISPR-Cas9 in the yeast strain (IMX581-HEM15-HEM14-HEM3- δshm1-HEM2-δhmx1-FET4-δgcv2-HEM1-δgcv1-HEM13), which produces 70-foldhigher levels of intracellular heme.

Genome-scale modeling

heme ligand-binding biosensor

metabolic engineering

heme

Saccharomyces cerevisiae