UC San Diego

Carrier proteins (CPs) are essential proteins for many biosynthetic pathways responsible for coordinating the biosynthesis of natural products. However, the CP must be posttranslationally modified by a 4’-phosphopantetheinyl transferase (PPTase) prior to participation in biosynthesis. This essential covalent modification uniquely positions the PPTase as a potential drug target for pathogenic bacteria upon structural understanding of the PPTase. Our lab has solved two PPTase crystal structures from Mycobacterium species, providing important structural information regarding these drug targets for the development of anti-mycobacterial therapeutics. Our lab has also explored the poorly understood eukaryotic PPTases and CPs, using Arabidopsis thaliana as a model organism. Since CP-dependent natural product biosynthesis in plants is organelle specific, the PPTase must access all organelles in order to activate biosynthesis. Since processivity of these interacting proteins still remains unclear, our lab has developed chemical biological tools to elucidate the nature of these interactions. We have also applied chemical biological and chemical genetics tools to study non-ribosomal peptide synthetases (NRPSs) by focusing on BpsA, an NRPS from Streptomyces lavendulae. This model synthase will not only provide a basis for chemical genetic characterization of NRPSs, but will pave the way for elucidating NRPS protein-protein interactions.

Our research has also investigated CP-independent pathways. Previously, the Noel lab has made great strides in studying type III polyketide synthases (T3PKS), which produce a wide variety of plant metabolites from CoA thioester precursors. We investigated the effects of mutations in 2-pyrone synthase, a T3PKS that makes triacetic acid lactone (TAL) from acetyl CoA and malonyl CoA. Mutants were assayed in vitro for stability and activity, and for their in vivo TAL production when heterologously expressed in yeast. This study has uncovered exploitable enzymatic and organismal properties to increase heterologous production of valuable chemical products.

Additionally, we investigated aromatic prenyltransferases (aPTases), which add prenyl groups onto aromatic molecules, and their activity against a variety of aromatic natural products in order to discover the underlying enzyme properties that control substrate specificity. Knowledge of these properties will enable fine-tuning of aPTase activity to specifically modify molecules with prenyl groups for high-level production of natural prenylated molecules and new, unexplored analogs. This body of work contributes valuable knowledge to the field of natural product biosynthesis, with the duel aims of discovering and characterizing potential therapeutic targets in bacteria and modifying natural product pathways to produce valuable molecules through enzyme engineering.