Enhanced heme production in industrial Saccharomyces cerevisiae through metabolic engineering

This study aimed to develop an efficient Saccharomyces cerevisiae-based cell factory for heme production. The industrial strain S. cerevisiae KCCM 12638, a starter strain used in American whisky production, was selected for its naturally high heme concentration, and its medium composition was optimized. Next, CRISPR/Cas9-based genome editing was employed to enhance carbon flux through the heme biosynthetic pathway by overexpressing HEM2, HEM3, HEM12, and HEM13 genes, which encode key enzymes involved in heme synthesis. Additionally, the HMX1 gene, which encodes heme oxygenase 1, was inactivated to prevent heme degradation. The resulting ΔHMX1_H2/3/12/13 strain achieved a heme titer of 9 mg/L in batch fermentation, a 1.7-fold improvement over the wild-type KCCM 12638 strain. In a glucose-limited fed-batch fermentation, the ΔHMX1_H2/3/12/13 strain produced 67 mg/L heme. This study demonstrates that engineering industrial yeast strains can significantly enhance heme production, with promising applications in food, pharmaceuticals, and bioenergy.