Engineered Chlamydomonas reinhardtii Strains for Enhanced Astaxanthin Production

Microalgae have evolved a diverse carotenoid profile, enabling efficient light harvesting and photoprotection. Previous studies have demonstrated the feasibility of genome editing in the green algal model species Chlamydomonas reinhardtii, leading to significant modifications in carotenoid accumulation. By overexpressing a fully redesigned beta-carotene ketolase (bkt), the metabolic pathway of C. reinhardtii was successfully redirected toward astaxanthin biosynthesis, a high-value ketocarotenoid with exceptional antioxidant properties, naturally found in only a few microalgal species. In this study, a tailor-made double knockout targeting lycopene epsilon-cyclase (LCYE) and zeaxanthin epoxidase (ZEP) was introduced as a background for bkt expression to ensure higher substrate availability for bkt enzyme. The increased zeaxanthin availability resulted in a 2-fold increase in ketocarotenoid accumulation compared to the previously engineered bkt1 or bkt5 strain in the UVM4 background. Specifically, the best Delta zl-bkt-expressing lines reached 2.84 mg/L under low light and 2.58 mg/L under high light, compared to 1.74 mg/L and 1.26 mg/L, respectively, in UVM4-bkt strains. These findings highlight the potential of rationally designed microalgal host strains, developed through genome editing, for biotechnological applications and high-value compound production.