| Polyketide metabolites are produced by diverse bacterial taxa, including soil-dwelling bacteria, cyanobacteria, and bacterial symbionts living within insects or marine invertebrates, and are generated by decarboxylative condensations of simple coenzyme A (CoA) building blocks. At present, polyketide natural products find clinical utility as antibiotics, antiparasitics, antifungals, anticancer drugs, and immunosuppressants. These comprehensive pharmacological activities provide continued motivation to unravel polyketide biosynthetic mechanisms to enable the discovery of novel compounds for the benefit of human health. This dissertational research explores the molecular basis for guiding ACP-mediated protein-protein interactions in three diverse pathways (pikromycin, bryostatin and germicidin).;Accessing new members of the ketolide class of macrolide antibiotics remains an important goal given the increasing prevalence of drug-resistant pathogens. As a naturally occurring ketolide, the pikromycin could serve as a scaffold to build a diverse set of polyketides. The Sherman laboratory has spent over a decade investigating the catalytic mechanisms of pikromycin biosynthesis by the modular PKS-containing pathway found in Streptomyces venezuelae. Here, we explore the protein-protein interactions of the large, multifunctional polypeptides at the PikAIII/PikAIV interface. The combination of structural characterization of the docking domains together with discrete docking domain affinity measurements supports a paradigm wherein the binding specificity that determines the linear arrangement of proteins in modular PKS systems is encoded in the small, terminal docking domains. Additionally, a model for the observed docking domain specificity across a matrix of interacting pairs from the pikromycin and erythromycin pathways is presented.;Secondly, we profiled the ACP binding and catalysis of BryR, the HMG-ACP synthase from an uncultured symbiont of Bugula neritina, a marine bryazoan. BryR functions to install beta-branches in bryostatin, a PKC modulator with both anti-cancer and neuroprotective activities. Lastly, we have explored a unique type III PKS, Gcs, that is capable of using both acyl-CoA and acyl-ACP starter units in the catalysis of pyrones. A structural model for Gcs is reported. Together with the growing understanding of protein-protein interactions in PKSs, the knowledge and mechanistic understanding of the pikromycin, bryostatin, germicidin and other complex metabolic systems will provide additional opportunities to engineer chemical diverse polyketides using rational approaches. |
