Impact of liquid and solid-state cultures on hemoglobin production and oxidative state in Saccharomyces cerevisiae

Abstract

Recombinant human hemoglobin gained attention due to its potential use as a blood-free oxygen carrier substitute. To enhance human hemoglobin production in Saccharomyces cerevisiae, various genetic engineering strategies have been employed, including: increasing intracellular heme levels, minimizing heme and protein degradation pathways, and co-expressing the α-hemoglobin stabilizing protein (AHSP). Solid-state culture (SSC) may enhance hemoglobin production by increasing heme biosynthesis, as it relates to intracellular oxygen availability. A comparative analysis of heme and hemoglobin production was conducted between liquid culture (LC) and SSC using the S. cerevisiae AHSP strain. While both systems exhibited comparable heme and hemoglobin yields per cell, a significant 18 % increase in biomass was observed in SSC. The expression of the aerobic master gene HAP1 remained consistent between both systems, however, CYC1 (regulated by HAP1) was two-fold overexpressed in SSC, indicating higher oxygen transference and possibly more efficient electron transport. Several antioxidant genes were downregulated in the SSC, suggesting that LC may be more susceptible to electron leakage during oxidative phosphorylation, potentially due to the lower expression of CYC1. It is proposed that high expression of antioxidant genes in LC inhibits biomass production due to the metabolic burden of maintaining redox homeostasis. These differences between LC and SSC may explain the suitability of SSC as a platform for recombinant protein production.