Metabolic engineering of β-carotene biosynthesis in Yarrowia lipolytica

Objective Carotenoids, as potent antioxidant compounds, have gained extensive attention, especially in human health. In this study, the combination of CRISPR/Cas9 integration strategy and fermenter cultivation was utilized to obtain efficient β-carotene-producing Yarrowia lipolytica cell factories for potential industrial application. Results The introduction of the genes of Mucor circinelloides , encoding phytoene dehydrogenase ( carB ) and bifunctional phytoene synthase/lycopene cyclase ( carRP ), contributed to the heterologous production of β-carotene in Y. lipolytica XK2. Furthermore, β-carotene production was efficiently enhanced by increasing the copy numbers of the carB and carRP genes and overexpressing of GGS1 , ERG13 , and HMG , the genes related to the mevalonate (MVA) pathway. Thus, the optimized strain overexpressed a total of eight genes, including three copies of carRP , two copies of carB , and single copies of GGS1 , HMG , and ERG13 . As a consequence, strain Y. lipolytica XK19 accumulated approximately 408 mg/L β-carotene in shake flask cultures, a twenty-four-fold increase compared to the parental strain Y. lipolytica XK2. Conclusions 4.5 g/L β-carotene was obtained in a 5-L fermenter through a combination of genetic engineering and culture optimization, suggesting a great capacity and flexibility of Y. lipolytica in the production of carotenoids.