A carotenogenic gene cluster from Brevibacterium linens with novel lycopene cyclase genes involved in the synthesis of aromatic carotenoids

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Abstract
The carotenogenic (crt) gene cluster from Brevibacterium linens, a member of the commercially important group of coryneform bacteria, was cloned and identified. An expression library of B. linens genes was constructed and a fragment of the crt cluster was obtained by functional complementation of a colourless B. flavum mutant, screening transformed cells for production of a yellow pigment. Subsequent screening of a cosmid library resulted in the cloning of the wholecrt cluster from B. linens. All genes necessary for the synthesis of the aromatic carotenoid isorenieratene were identified on the basis of sequence homologies. In addition a novel type of lycopene cyclase was identified by complementation of a lycopene-accumulating B. flavum mutant. Two genes, named crtYc and crtYd, which code for polypeptides of 125 and 107 amino acids, respectively, are necessary to convert lycopene to β-carotene. The amino acid sequences of these polypeptides show no similarity to any of the known lycopene cyclases. This is the first example of a carotenoid biosynthetic conversion in which two different gene products are involved, probably forming a heterodimer.