Rational design and assembly of synthetic trimodular polyketide synthases

Type I polyketide synthases (PKSs) consist of modules that add two-carbon units in polyketide backbones. Rearranging modules from different sources can yield novel enzymes that produce unnatural products, but the rules that govern module-module communication are still not well known. The construction and assay of hybrid bimodular units with synthetic PKS genes were recently reported. Here, we describe the rational design of trimodular PKSs by combining bimodular units. A cloning-expression system was developed to assemble and test 54 unnatural trimodular PKSs flanked by the loading module and the thioesterase from the erythromycin synthase. Remarkably, 96% of them produced the expected polyketide. The obtained results represent an important milestone toward the ultimate goal of making new bioactive polyketides by rational design. Additionally, these results show a path for the production of customized tetraketides by fermentation, which can be an important source of advanced intermediates to facilitate the synthesis of complex products.