Functional characterization of CmCCD1, a carotenoid cleavage dioxygenase from melon

Carotenoids are nutritionally important tetraterpenoid pigments that upon oxidative cleavage give rise to apocarotenoid (norisoprene) aroma volatiles. beta-Carotene is the predominant pigment in orange-fleshed melon (Cucumis melo L.) varieties, reaching levels of up to 50 microg/gFW. Pale green and white cultivars have much lower levels (0-10 microg/gFW). In parallel, beta-ionone, the 9,10 cleavage product of beta-carotene, is present (12-33ng/gFW) in orange-fleshed melon varieties that accumulate beta-carotene, and in much lower levels (0-5 ng/gFW) in pale green and white fleshed varieties. A search for a gene putatively responsible for the cleavage of beta-carotene into beta-ionone was carried out in annotated melon fruit EST databases yielding a sequence (CmCCD1) highly similar (84%) to other plant carotenoid cleavage dioxygenase genes. To test its function, the clone was overexpressed in Escherichia coli strains previously engineered to produce different carotenoids. We show here that the CmCCD1 gene product cleaves carotenoids at positions 9,10 and 9',10', generating geranylacetone from phytoene; pseudoionone from lycopene; beta-ionone from beta-carotene, as well as alpha-ionone and pseudoionone from delta-carotene. CmCCD1 gene expression is upregulated upon fruit development both in orange, pale-green and white melon varieties, despite the lack of apocarotenoid volatiles in the later. Thus, the accumulation of beta-ionone in melon fruit is probably limited by the availability of carotenoid substrate.