One ring or two?: Determination of ring number in carotenoids by lycopene ε-cyclases

被引:164
作者
Cunningham, FX [1 ]
Gantt, E [1 ]
机构
[1] Univ Maryland, Dept Mol Genet & Cell Biol, College Pk, MD 20742 USA
关键词
D O I
10.1073/pnas.051618398
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Carotenoids in the photosynthetic membranes of plants typically contain two beta -rings (e.g., beta -carotene and zeaxanthin) or one epsilon- and one beta -ring (e.g., lutein). Carotenoids with two epsilon -rings are uncommon. We reported earlier that the Arabidopsis thaliana lycopene epsilon -cyclase (LCYe) adds one epsilon -ring to the symmetrical linear substrate lycopene, whereas the structurally related lycopene beta -cyclase (LCYb) adds two beta -rings. Here we describe a cDNA encoding LCYe in romaine lettuce (Lactuca sativa var. romaine), one of the few plant species known to accumulate substantial quantities of a carotenoid with two epsilon -rings: lactucaxanthin. The product of the lettuce cDNA, similar in sequence to the Arabidopsis LCYe (77% amino acid identity), efficiently converted lycopene into the bicyclic epsilon -carotene in a heterologous Escherichia coli system. Regions of the lettuce and Arabidopsis epsilon -cyclases involved in the determination of ring number were mapped by analysis of chimeric epsilon -cyclases constructed by using an inverse PCR approach. A single amino acid was found to act as a molecular switch: lettuce LCYe mutant H457L added only one epsilon -ring to lycopene, whereas the complementary Arabidopsis LCYe mutant, L448H, added two epsilon -rings. An R residue in this position also yields a bi-epsilon -cyclase for both the lettuce and Arabidopsis enzymes. Construction and analysis of chimera of related enzymes with differing catalytic activities provide an informative approach that may be of particular utility for studying membrane-associated enzymes that cannot easily be crystallized or modeled to existing crystal structures.
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页码:2905 / 2910
页数:6
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