Interfacing Polyketide Synthase and Non-Ribosomal Peptide Synthetase Pathways Through Exploitation of Hairpin Docking Domain Pairs

Polyketide synthases (PKSs) and non-ribosomal peptide synthetases (NRPSs) play a critical role in the biosynthesis of numerous biologically-active natural products. Acting as molecular assembly lines, these modular megasynth(et)ases construct metabolic products in a stepwise fashion from simple precursors. To maintain biosynthetic fidelity, the interactions between subunits of these enzymatic machines are mediated by docking domains (DDs); short polypeptides appended to the C- and N-termini of the subunits.
This project focusses on the action of Short Linear Motif (SLiM) -hairpin docking domain (HDD) pairs, one of a handful of docking domain types known to mediate subunit interactions in hybrid NRPS-PKS systems. These docking domains are attached to the C-terminus of PKS carrier proteins and the N-terminus of their interacting NRPS domain. Recent work in the Challis group has shown these DDs are involved in the biosynthesis of HDAC inhibitors, where they are hypothesised to mediate the interaction between PKS and NRPS components of the assembly line. A range of computational, chemical and biochemical approaches will be taken to explore the prevalence of SLiM-HDD pairs in hybrid PKS-NRPSs, and how their interactions can be exploited to generate novel biosynthetic pathways. Overproduction of relevant protein domains in E.coli will form the basis for biochemical assays to assess the interaction between recombinant PKS and NRPS components containing the docking domains. Biosynthetic engineering will be undertaken to fuse the PKS and NRPS components from separate pathways, enabling the de novo production of novel HDAC inhibitors.