The biotechnological potential of the al-2 gene from Neurospra crassa for the production of monocyclic keto hydroxy carotenoids

The al-2 cDNA from Neurospora crassa was cloned, expressed and functionally characterized. The enzyme comprised the two catalytic activities of a phytoene synthase and a lycopene cyclase. In contrast to most other lycopene cyclases, single cyclizations were preferentially catalyzed. This N. crassa enzyme is the first CrtYB-type monocyclic-acting lycopene cyclase. Therefore, this cDNA has been evaluated for the heterologous synthesis of monocyclic hydroxy-keto carotenoids by combination with other carotenogenic genes in Escherichia coli. Depending on the degree of desaturation, 4-keto derivatives of γ-carotene and torulene with additional 2-hydroxy, 3-hydroxy and/or 1′-HO groups were generated and the following asymmetrical carotenoids identified and quantitated: 3-HO-4-keto-γ-carotene, 2-HO-4-keto-γ-carotene, 4-keto-1′-HO-γ-carotene, 3,1′-(HO)2-4-keto-γ-carotene, 3-HO-4-keto-torulene and 2-HO-4-keto-torulene. Among them all the monocyclic γ-carotene derivatives with 9 conjugated double bonds were not found naturally before. Furthermore, 2-HO-4-keto-torulene with 10 conjugated double bonds is another novel carotenoid.