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-fold-higher levels of intracellular heme.

A new YNB medium containing very low concentrations of alkali metal cations has been developed to carry out experiments to study potassium homoeostas

Ergothioneine is a naturally occurring antioxidant that has shown potential in ameliorating neurodegenerative and cardiovascular diseases. In this study, we investigated the potential of the Crabtree‐negative, oleaginous yeast Yarrowia lipolytica as an alternative host for ergothioneine production. We expressed the biosynthetic enzymes EGT1 from Neurospora crassa and EGT2 from Claviceps purpurea to obtain 158 mg·L⁻¹ of ergothioneine in small‐scale cultivation, with an additional copy of each gene improving the titer to 205 mg·L⁻¹. The effect of phosphate limitation on ergothioneine production was studied, and finally, a phosphate‐limited fed‐batch fermentation in 1 L bioreactors yielded 1.63 ± 0.04 g·L⁻¹ ergothioneine in 220 h, corresponding to an overall volumetric productivity of 7.41 mg·L⁻¹·h⁻¹, showing that Y. lipolytica is a promising host for ergothioneine production.