Efficient synthesis of high-active multi-subunit hemoglobins through balanced expression of globins and robust heme-supply in Saccharomyces cerevisiae

The biosynthesis of multi-subunit hemoglobins that can be used in artificial oxygen carriers, artificial foods and medical diagnostics has become a hot research issue. However, the imbalanced expression of different subunits and insufficient heme-supply limit the efficient production of multi-subunit hemoglobins. After the equalized synthesis of different globins through the regulation at transcriptional and translational levels, the improvement of heme-supply by relieving the feedback inhibition of heme on Hem1p and the enhancement of key rate-limiting enzymes in yeast, a robust platform for the expression of high-active hemoglobins was constructed. The synthesis of hemoglobins with high-activity were achieved, including human hemoglobin (174.2 mg/L, 61.7 % mol heme/mol Hb), porcine hemoglobin (203.9 mg/L, 72.1 % mol heme/mol Hb) and bovine hemoglobin (258.2 mg/L, 64.9 % mol heme/mol Hb). Therefore, the applied strategies can be broadly used in the production of other heteromultimeric and heme-binding proteins.