Nonnatural biosynthetic pathway for 2-hydroxylated xanthophylls with C₅₀-carotenoid backbone

Carotenoids are structurally diverse pigments with various important biological functions. There has been a large interest in the search for novel carotenoid structures, since only a slight structural changes can result in a drastic difference in their biological functions. Carotenoid-modifying enzymes show remarkable substrate promiscuity, allowing rapid access to a vast set of novel carotenoids by combinatorial biosynthesis. We previously constructed a nonnatural carotenoid biosynthetic pathway in Escherichia coli that can produce C₅₀ carotenoids having a longer chain than their natural C₄₀ counterparts. In this study, a carotenoid 2,2'-hydroxylase (crtG) from Brevundimonas sp. SD212 was coexpressed together with our laboratory-engineered C₅₀-zeaxanthin and C₅₀-astaxanthin biosynthetic pathways. We identified six novel nonnatural C₅₀-xanthophylls, namely, C₅₀-nostoxanthin, C₅₀-caloxanthin, C₅₀-adonixanthin, C₅₀-4-ketonostoxanthin, C₅₀-2-hydroxyastaxanthin, and C₅₀-2,2'-dihydroxyastaxanthin.