Synthetic Biology Tools for Polyketide Engineering

This thesis discusses the advancement of synthetic biology tools for polyketide engineering, specifically in the construction of a genetic device for the biosynthesis of a fluorinated monomer and the construction of polyketide pathways in yeast followed by heterologous expression in Streptomyces. The first project prioritizes fluorinated polyketides due to the advantage of fluorine substitution in drug design as well as the difficulty of chemically synthesizing polyketides. Synthetic biology was used to assemble a genetic device for the biosynthesis of polyketide monomer fluoroethylmalonyl-CoA by linking chosen genes from Streptomyces cattleya and Salinispora tropica. Once the production of this genetic device is shown in a heterologous host, it will consequently be used to produce fluorinated polyketides. The second project also contributes to expanding synthetic biology tools for polyketide systems, specifically for the heterologous expression of the ~100-kb meridamycin biosynthetic gene cluster. This was a collaboration with the Cornish Lab at Columbia University, and our contribution to this project was the understanding that it is more suitable to use vectors based on the phage ΦC31 integration system (int, attP) rather than ΦBT1 for polyketide pathway construction and heterologous expression in Streptomyces. This was concluded after various transfers into Streptomyces using vectors that contained both integration systems, and comparing their overall conjugation efficiencies.