Defining the biosynthesis of ketocarotenoids in .

Carotenoids are important pigments in photosynthetic organisms where they play essential roles in photoreception and photoprotection. is a unicellular green alga that is able to accumulate high amounts of ketocarotenoids including astaxanthin, canthaxanthin and ketolutein when growing heterotrophically or mixotrophically with glucose as a carbon source. Here we elucidate the ketocarotenoid biosynthesis pathway in by analyzing five algal mutants. The mutants were shown to have a single nucleotide insertion or substitution in β-carotene ketolase () gene 1, which resulted in a lack of ketocarotenoid production in Cz-bkt1-1, and decreased ketocarotenoid content in the other four mutants. These mutants accumulated much higher amounts of non-ketocarotenoids (β-carotene, zeaxanthin and lutein). Interestingly, the Cz-bkt1-5 mutant synthesized 2-fold the ketolutein and only 1/30 of the canthaxanthin and astaxanthin as its parent strain, suggesting that the mutated BKT1 exhibits much higher activity in catalyzing lutein to ketolutein but lower activity in ketolating β-carotene and zeaxanthin. Mutant and WT 2 gene sequences did not differ. Taken together, we conclude that 1 is the key gene involved in ketocarotenoid biosynthesis in . Our study provides insight into the biosynthesis of ketocarotenoids in green algae. Furthermore, Cz-bkt1 mutants may serve as a natural source for the production of zeaxanthin, lutein, and β-carotene.