Chemical Control in the Battle against Fidelity in Promiscuous Natural Product Biosynthesis: The Case of Trichodiene Synthase

Abstract

Terpene cyclases catalyze the highly stereospecific molding of polyisoprenes into terpenes, which are precursors to most known natural compounds. The isoprenoids are formed via intricate chemical cascades employing rich, yet highly erratic, carbocation chemistry. It is currently not well understood how these biocatalysts achieve chemical control. Here, we illustrate the catalytic control exerted by trichodiene synthase, and in particular, we discover two features that could be general catalytic tools adopted by other terpenoid cyclases. First, to avoid formation of byproducts, the enzyme raises the energy of bisabolyl carbocation, which is a general mechanistic branching point in many sesquiterpene cyclases, resulting in an essentially concerted cyclization cascade. Second, we identify a sulfur–carbocation dative bonding interaction that anchors the bisabolyl cation in a reactive conformation, avoiding tumbling and premature deprotonation. Specifically, Met73 acts as a chameleon, shifting from an initial sulfur–π interaction in the Michaelis complex to a sulfur–carbocation complex during catalysis.

Comment on “Substrate Folding Modes in Trichodiene Synthase: A Determinant of Chemo- and Stereoselectivity”

“…1 incorrectly quoted several direct statements and structures that are plainly contrary to those reported in Ref. 2 ; unfortunately, the incorrect statements were used as evidence to justify their erroneous findings discussed above. In Ref.…”

“…These errors pertain to the claim of an “optimal” binding mode of the substrate farnesyl diphosphate (FPP), which, in fact, is inconsistent with experimental data, and numerous puzzling and plainly incorrect statements on our work that was published earlier in this Journal. 2 Importantly, the findings and key reaction steps appear to directly contradict the reaction pathway established by NMR isotope-labeling experiments.…”

“…2 In that study, we addressed numerous issues pertinent to the chemical control exerted by TDS to avoid by-product formation from a sequence of reactive carbocation intermediates, and highlighted several general catalytic principles that are potentially applicable to other terpenoid synthases. TDS has been extensively studied experimentally, 5–8 providing an unprecedented amount of data for judicious comparison and validation of theory.…”

“…There are multiple possible reaction channels leading to unwanted by-products, and it is necessary to use accurate quantum chemistry methods to treat the facile bond breaking and bond forming steps. 2, 11–15 To overcome these difficulties, we have developed and reported in reference 2 a systematic strategy to treat these biosynthetic processes computationally. 14, 16–17 …”

“…1 In that work, the authors followed essentially the same procedures, including exactly the same level of theory described in Ref. 2 , but started by docking the large, open structure of FPP directly into the active site. Then, the authors tried to delineate how this initial fold determines chemical specificity.…”