A carotenoid desaturase homolog from Staphylococcus aureus (CrtOx) was identified. When expressed in engineered E. coli cells synthesizing linear C30 carotenoids, polar carotenoid products were generated, identified as aldehyde and carboxylic acid C30 carotenoid derivatives. The major product in this engineered pathway is the fully desaturated C30 dialdehyde carotenoid 4,4′-diapolycopen-4,4′-dial. Very low carotenoid yields were observed when CrtOx was complemented with the C 40 carotenoid lycopene pathway. But extension of an in vitro evolved pathway of the fully desaturated 2,4,2′,4′-tetradehydrolycopene produced the structurally novel fully desaturated C40 dialdehyde carotenoid 2,4,2′,4′-tetradehydrolycopendial. Directed evolution of CrtOx by error-prone PCR resulted in a number of variants with higher activity on C40 carotenoid substrates and improved product profiles. These findings may provide new biosynthetic routes to highly polar carotenoids with unique spectral properties desirable for a number of industrial and pharmaceutical applications.
Bibliographical noteFunding Information:
The authors gratefully acknowledge support from the Defense Advanced Research Projects Agency (DARPA-BIOS N66001-02-1-8928) and the David and Lucile Packard Foundation (grant 2001-18996).