Exploring the Influence of Domain Architecture on the Catalytic Function of Diterpene Synthases.

ABSTRACT: Terpenoid synthases catalyze isoprenoid cyclization reactions underlying the generation of more than 80,000 natural products. Such dramatic chemodiversity belies the fact that these enzymes generally consist of only three domain folds designated as ?, ?, and ?. Catalysis by class I terpenoid synthases occurs exclusively in the ? domain, which is found with ?, ??, ??, and ??? domain architectures. Here, we explore the influence of domain architecture on catalysis by taxadiene synthase from Taxus brevifolia (TbTS, ???), fusicoccadiene synthase from Phomopsis amygdali (PaFS, (??)6), and ophiobolin F synthase from Aspergillus clavatus (AcOS, ??). We show that the cyclization fidelity and catalytic efficiency of the ? domain of TbTS are severely compromised by deletion of the ?? domains; however, retention of the ? domain preserves significant cyclization fidelity. In PaFS, we previously demonstrated that one ? domain similarly influences catalysis by the other ? domain [ Chen , M. , Chou , W. K. W. , Toyomasu , T. , Cane , D. E. , and Christianson , D. W. ( 2016 ) ACS Chem. Biol. 11 , 889 - 899 ]. Here, we show that the hexameric quaternary structure of PaFS enables cluster channeling. We also show that the ? domains of PaFS and AcOS can be swapped so as to make functional chimeric ?? synthases. Notably, both cyclization fidelity and catalytic efficiency are altered in all chimeric synthases. Twelve newly formed and uncharacterized C20 diterpene products and three C25 sesterterpene products are generated by these chimeras. Thus, engineered ??? and ?? terpenoid cyclases promise to generate chemodiversity in the greater family of terpenoid natural products.

SUBMITTER: Pemberton TA 

PROVIDER: S-EPMC5474940 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature