Modification of the Endoplasmic Reticulum to Enhance Ovalbumin Secretion in <i>Saccharomyces cerevisiae</i>

Ovalbumin (OVA) is a high-quality protein for humans. Modifying microorganisms to produce proteins offers a solution to potential food protein shortages. In this study, OVA was expressed in <i>Saccharomyces cerevisiae</i>. Initially, screening signal peptides led to extracellular OVA reaching 3.4 mg/L using the INU1 signal peptide. Coexpressing Kar2 and PDI increased OVA production to 5.1 mg/L. Optimizing the expression levels of regulators OPI1, INO2, and INO4 expanded the endoplasmic reticulum membrane, raising yield to 5.5 mg/L. Combining both strategies increased OVA production to 6.2 mg/L, 82% higher than control. This strategy also enhanced secretion of other proteins. Finally, fed-batch fermentation in a 3-L bioreactor significantly boosted OVA production to 116.3 mg/L. This study provides insights for the heterologous synthesis of other high-quality proteins for future food applications.