High-level secretory production of leghemoglobin in Pichia pastoris through enhanced globin expression and heme biosynthesis.

Soy leghemoglobin is a key food additive that imparts meaty flavor and color to meat analogs. Here, a Pichia pastoris strain capable of high-yield secretory production of functional leghemoglobin was developed through gene dosage optimization and heme pathway consolidation. First, multi-copy integration of LegH expression cassette was achieved via both post-transformational vector amplification and CRISPR/Cas9 mediated genome editing methods.