Efficient Secretory Expression and Purification of Food-Grade Porcine Myoglobin in Komagataella phaffii

using the suitable host (X33), signal peptide (α-factor signal peptide), and modified constitutive promoter (G1 promoter). In addition, the fermentation conditions for MG production were optimized at shaking-flask level (BMGY medium with 40 mg/L of hemin, 30 °C) and at fermenter level (30% DO, feeding 150 mg/L of hemin), resulting in the highest titer of 285.42 mg/L MG in fed-batch fermentations. Furthermore, a purification method for food-grade MG was developed, which can obtain 0.22 mol of heme/mol of MG with 88.0% purity and 66.1% recovery rate.