(293g) Synthetic Pathways for the Microbial Production of Isoprenoids, Polyketides, and Prenylated Aromatics

Prenylated aromatics (PAs) are natural products with a broad range of pharmaceutical applications. PAs are typically sourced from plants, which limits their production due to agricultural and geographic variations and costly extraction. Furthermore, their biosynthesis relies on a relatively small number of chemistries and metabolic pathways that have been shaped by evolution to fulfill physiological roles and which are inherently constrained by carbon, energy, and kinetic inefficiencies. To address these limitations, we have engineered E. coli-based platforms for the synthesis of PAs, as well as their prenyl donors and aromatic acceptors. Our initial efforts focused on using polyketide synthase and olivetolic acid cyclase from Cannabis sativa to produce olivetolic acid (ACS Synth Biol 7:1886, 2018), along with the engineering of prenyltransferases for the synthesis of cannabigerovarinic, cannabigerolic, and grifolic acids (Biotechnol Bioeng 116:1116, 2019). More recently, we have engineered new-to-nature pathways for isoprenoid and polyketide biosynthesis, which provide the prenyl donors and aromatic acceptors for PA production. For example, we designed, prototyped, and implemented a route to isoprenoid biosynthesis, termed the Isoprenoid Alcohol (IPA) pathway, capable of operating at high flux with improved energy efficiency (PNAS 116:12810, 2019). We accomplished this through a retro-biosynthetic approach that identified isoprenoid alcohols as the key intermediate and namesake metabolites for the IPA pathway. We also recently discovered that certain thiolases, which we have termed polyketoacyl-CoA thiolases (PKTs), catalyze polyketide backbone formation and offer a synthetic and efficient alternative to polyketide biosynthesis (Nature Catalysis 3, 593-603, 2020). We showed that PKTs can synthesize polyketide backbones for representative lactone, alkylresorcinolic acid, alkylresorcinol, hydroxybenzoic acid and methylphenol polyketide families, and elucidated the basic catalytic mechanism and structural features enabling this previously unknown activity. These new-to-nature pathways have the potential to change the decades-long paradigm for natural product biosynthesis, which had primarily leveraged native chemistries and metabolic pathways.