Structure-guided product determination of the bacterial type II diterpene synthase Tpn2.

A grand challenge in terpene synthase (TS) enzymology is the ability to predict function from protein sequence. Given the limited number of characterized bacterial TSs and significant sequence diversities between them and their eukaryotic counterparts, this is currently impossible. To contribute towards understanding the sequence-structure-function relationships of type II bacterial TSs, we determined the structure of the terpentedienyl diphosphate synthase Tpn2 from Kitasatospora sp. CB02891 by X-ray crystallography and made structure-guided mutants to probe its mechanism. Substitution of a glycine into a basic residue changed the product preference from the clerodane skeleton to a syn-labdane skeleton, resulting in the first syn-labdane identified from a bacterial TS. Understanding how a single residue can dictate the cyclization pattern in Tpn2, along with detailed bioinformatics analysis of bacterial type II TSs, sets the stage for the investigation of the functional scope of bacterial type II TSs and the discovery of novel bacterial terpenoids. Crystal structure elucidation of terpene synthases plays an important role to understand the sequence-structure-function relationships of this enzyme class. Here, the authors determine the crystal structure of bacterial terpene synthase Tpn2, probe the catalytic mechanism by structure-guided mutation, and change the product preference from a clerodane skeleton to a syn-labdane skeleton. [ABSTRACT FROM AUTHOR]