A crtA-related gene from Flavobacterium P99-3 encodes a novel carotenoid 2-hydroxylase involved in myxol biosynthesis.

A genomic DNA fragment with carotenogenic genes involved in myxol biosynthesis (3',4'-didehydro-1',2'-dihydro-beta,psi-carotene-3,1',2'-triol) was cloned from Flavobacterium P99-3. It contains a gene highly homologous to crtA from purple bacteria encoding there an acyclic carotenoid 2-ketolase. Since no ketolation step is involved in myxol biosynthesis, the function of crtA-OH from Flavobacterium was assigned by complementation in Escherichia coli engineered to synthesize demethylspheroidene and 1'-hydroxy-demethylspheroidene. Upon co-expression of crtA-OH, the formation of 2-hydroxy derivatives of both carotenoids assigns CrtA-OH as a novel carotenoid hydroxylase. The gene was used to re-construct myxol biosynthesis in E. coli successfully. Additionally, 1',2'-dihydroxytorulene and 1,2,1'-trihydroxy-3,4,3',4'-tetradehydrolycopene were obtained. Their generation demonstrates that a new class of 2-hydroxy carotenoids can now be pursued by genetic engineering in E. coli.