Introduction continued

For those sesquiterpene synthases that convert FPP to compounds of the eremophilane and eudesmane classes, including epi-aristolochene, aristolochene , vetispiradiene, and d-selinene, it is postulated that the 10-membered-ring germacrenes (isomers A, B, C or D) are the initial products of the cyclisation and subsequent elimination and rearrangement reactions. These neutral compounds are apparently not released from the active site of the sesquiterpene synthase but are re-protonated by the enzyme to form new cations, which undergo further cyclisations to yield the final products.

Evidence for cryptic germacrene A synthase activity within aristolochene synthase has been provided by David Cane et al. who observed the formation of dihydrogermacrene A from the substrate analogue dihydro-FPP. More recently, site-directed mutagenesis of tobacco epi-aristolochene synthase (TEAS) has given rise to a mutant protein that produced germacrene A, albeit at lower efficiency compared to the wild-type production of epi-aristolochene. There are many plant natural products that are derived from oxidative metabolism of germacrene A, indicating that sesquiterpene synthases that produce and release this thermally unstable 10-membered ring hydrocarbon are widespread.

Germacrene A synthase is an important biotechnological target involved in the biosynthesis of parthenolide, the active ingredient of feverfew, as well as in the formation of the bitter sesquiterpenoids of lettuce and chicory. Next we describe the cDNA cloning, functional expression in E.coli, and characterisation of this gene - (+)-(10R)-germacrene A synthase - from the goldenrod, Solidago canadensis.