Microbial Astaxanthin Synthesis by <i>Komagataella phaffii</i> through Metabolic and Fermentation Engineering

Astaxanthin is a kind of carotenoid with a strong antioxidant ability, which has shown broad applications in the areas of healthcare, medicine, cosmetics, food additives, and aquaculture. With the increasing demand for natural products, the microbial production of astaxanthin has become a new hot spot. In this study, the astaxanthin synthesis pathway was first metabolically constructed in <i>Komagataella phaffii</i> (<i>K. phaffii</i>)(<i>Pichia pastoris</i>). By exploring the combination of different promoters, astaxanthin producers were obtained. Then, the key enzymes in the astaxanthin synthesis pathway were explored, and different enzyme assembly strategies and an increase in NADPH supply were used to improve the astaxanthin production. Furthermore, fermentation parameters including temperature, the concentration of the carbon source, nitrogen sources, metal ions, BHT (2,6-di-<i>tert</i>-butyl-4-methylphenol), and Tween-80 were comprehensively investigated for the microbial growth and astaxanthin synthesis. Finally, the astaxanthin production reached 716.13 mg/L by fed-batch fermentation in a 5 L bioreactor, which was the highest production of astaxanthin synthesized by <i>K. phaffii</i>.