Secretory and metabolic engineering of squalene in Yarrowia lipolytica

This short communication systematically explored ATP-binding cassette (ABC) transporters and developed a carrier protein-mediated metabolite trafficking system to engineer Yarrowia lipolytica for secretion of squalene. Specially, the squalene biosynthesis pathway was overexpressed to improve squalene production. Subsequently, a series of ABC transporters and oxysterol-binding proteins (OSH) were screened, with the combination of SNQ2 and OSH3 showing the most significant improvement in squalene secretion. Additionally, we designed a carrier proteinmediated metabolite trafficking system by fusing the binding domain of OSH3 with a secretion signal peptide. This approach, combined with the optimization of signal peptides, increased squalene secretion to 144.67 mg/L, which represented a 17-fold improvement over the initial strain. This system also demonstrated versatility by facilitating the extracellular export of beta-carotene. In fed-batch cultivation, the engineered strain secreted 3.43 g/L of squalene extracellularly, accounting for 27.2 % of the total production. It represented the highest secretion level reported to date. This study establishes an efficient secretion platform for the transport of membrane-impermeable squalene in Y. lipolytica, providing valuable insights and strategies for the microbial production of terpenoids.