Engineering of for production of astaxanthin.

Astaxanthin is a red-colored carotenoid, used as food and feed additive. Astaxanthin is mainly produced by chemical synthesis, however, the process is expensive and synthetic astaxanthin is not approved for human consumption. In this study, we engineered the oleaginous yeast for production of astaxanthin by fermentation. First, we screened 12 different isolates for β-carotene production by introducing two genes for β-carotene biosynthesis: bi-functional phytoene synthase/lycopene cyclase () and phytoene desaturase () from the red yeast . The best strain produced 31.1 ± 0.5 mg/L β-carotene. Next, we optimized the activities of 3-hydroxy-3-methylglutaryl-coenzyme A reductase () and geranylgeranyl diphosphate synthase () in the best producing strain and obtained 453.9 ± 20.2 mg/L β-carotene. Additional downregulation of the competing squalene synthase increased the β-carotene titer to 797.1 ± 57.2 mg/L. Then we introduced β-carotene ketolase () from sp. N81106 and hydroxylase () from to convert β-carotene into astaxanthin. The constructed strain accumulated 10.4 ± 0.5 mg/L of astaxanthin but also accumulated astaxanthin biosynthesis intermediates, 5.7 ± 0.5 mg/L canthaxanthin, and 35.3 ± 1.8 mg/L echinenone. Finally, we optimized the copy numbers of and to obtain 3.5 mg/g DCW (54.6 mg/L) of astaxanthin in a microtiter plate cultivation. Our study for the first time reports engineering of for the production of astaxanthin. The high astaxanthin content and titer obtained even in a small-scale cultivation demonstrates a strong potential for -based fermentation process for astaxanthin production.