Journal of the American Chemical Society 2012-12-26

Probing the mechanism of 1,4-conjugate elimination reactions catalyzed by terpene synthases.

Juan A Faraldos, Veronica Gonzalez, Amang Li, Fanglei Yu, Mustafa Köksal, David W Christianson, Rudolf K Allemann

Index: J. Am. Chem. Soc. 134(51) , 20844-8, (2012)

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Abstract

The reaction mechanisms of (E)-β-farnesene synthase (EBFS) and isoprene synthase (ISPS), enzymes that catalyze a formal regiospecific 1,4-conjugate elimination of hydrogen diphosphate from (E,E)-farnesyl and dimethylallyl diphosphate (FDP and DMADP) to generate the semiochemicals (E)-β-farnesene and isoprene, respectively, were probed with substrate analogs and kinetic measurements. The results support stepwise reaction mechanisms through analogous enzyme-bound allylic cationic intermediates. For EBFS, we demonstrate that the elimination reaction can proceed via the enzyme-bound intermediate trans-nerolidyl diphosphate, while for ISPS the intermediacy of 2-methylbut-3-enyl 2-diphosphate can be inferred from the product outcome when deuterated DMADPs are used as substrates. Possible implications derived from the mechanistic details of the EBFS-catalyzed reaction for the evolution of sesquiterpene synthases are discussed.