Protonation of a neutral (S)-β-bisabolene intermediate is involved in (S)-β-macrocarpene formation by the maize sesquiterpene synthases TPS6 and TPS11

Terpene synthases are responsible for the large diversity of terpene carbon skeletons found in plants. The unique, carbocationic reaction mechanism of these enzymes can form multiple products from a single prenyl diphosphate substrate. Two maize genes were isolated that encode very similar sesquiterpene synthases, TPS6 and TPS11, which both produce beta-bisabolene, a common monocyclic sesquiterpene, and beta-macrocarpene, an uncommon bicyclic olefin. Investigation of the reaction mechanism showed that the formation of beta-macrocarpene proceeds via a neutral beta-bisabolene intermediate and requires reprotonation by a proton that may ultimately be abstracted from water. This reprotonation is dependent on the pH and the presence of a Mg2+ cofactor. Mutational analysis of the enzyme demonstrated that a highly conserved tyrosine residue in the active center of the enzymes is important for the protonation process. TPS6 and TPS11 are transcribed both in leaves and roots of maize, but the respective terpene products were only detected in roots. The expression in roots was up-regulated by herbivore damage to the leaves, suggesting a long distance signal transduction cascade between leaves and roots.