Terpenoids are a large, structurally diverse class of medicinally relevant natural products. Despite their role as a longstanding source of pharmaceuticals, however, the chemical complexity and poor natural abundance of these molecules has slowed the development of new terpenoid-based drugs. Systematic modifications to terpenoid-synthesizing pathways can produce diverse sets of terpenoid scaffolds in genetically tractable microbial hosts, but complimentary methods for identifying molecules with important biochemical activities remain largely absent. In this presentation, I will describe an approach for evolving terpene synthases that produce molecules with elusive biochemical activities. My work focuses on a therapeutically relevant objective: the inhibition of protein tyrosine phosphatase 1B (PTP1B), a target for various metabolic diseases and cancers.  We encoded this objective into a strain of <i>Escherichia coli</i> and used that strain to evolve terpene synthases that produce PTP1B inhibitors. This method yielded enzyme variants with novel product profiles, enhanced titers, and/or reduced cellular toxicities. Their terpenoid products, which we characterized with detailed biophysical studies, appear to target an allosteric site on the C-terminus of PTP1B. This presentation frames a new approach for using engineered microbial hosts for the evolution of targeted, biologically active metabolites.

Directed evolution of terpene synthases for the production of phosphatase inhibitors

ACS Fall 2021

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