Basic importance: mechanistic molecular modeling of the <i>ent</i> -copalyl diphosphate synthase from <i>Arabidopsis thaliana</i> (AtCPS)

Terpene synthases/cyclases catalyze the formation of complex polycyclic natural products <i>via</i> mechanisms proceeding through carbocation intermediates. Their active sites are generally lined with aliphatic and aromatic residues that prevent untoward termination of the reaction <i>via</i> carbocation deprotonation or addition of water, enabling use of the relevant catalytic base, whose positioning is then crucial to product outcome. This has been emphasized by studies with the <i>ent</i>-copalyl pyrophosphate synthase from <i>Arabidopsis thaliana</i> (AtCPS), representative of plant class II diterpene cyclases more generally, as displacement of its (conserved) catalytic base leads to such alternative product outcomes. Here we characterize the mechanistic basis for the profound effects of these disruptions in AtCPS using the TerDockin computational approach, combining quantum chemical electronic structure calculations and docking of the relevant carbocation intermediates, to provide insight into the underlying enzymatic structure-function relationships. Our predictions help identify important bases in the wild type and mutant systems of AtCPS, which include trapped water and the conjugate base of the catalytic acid that initiates terpene cyclization.