Efficient Expression of Epidermal Growth Factor in Komagataella phaffii and Its Bioactivity Evaluation in Cell Culture.

Epidermal growth factor (EGF) is an important growth factor for low-serum cultured meat production, yet efficient recombinant expression remains challenging. Here, an integrated engineering strategy combining signal peptide optimization, transcription factor overexpression, and gene dosage amplification was established to enhance recombinant EGF production in Komagataella phaffii. Truncation of amino acids 57-70 in the α-factor signal peptide increased EGF secretion efficiency from 69.8% to 95.3% and yield from 21.86 to 36.07 mg/L. Mit1 overexpression increased the yield to 42.44 mg/L, while sequential gene dosage amplification enhanced EGF yield to 280.42 mg/L in shake-flask culture, representing a 12.8-fold increase. The engineered strain was evaluated in a 5 L bioreactor, achieving 370.05 mg/L EGF. Purified recombinant EGF exhibited biological activity by promoting porcine skeletal muscle satellite cell proliferation under low-serum conditions. This study provides an efficient strategy for recombinant EGF production in K. phaffii and supports its application in cultured meat development.