ENGINEERED BIOSYNTHESIS OF NOVEL KETOLIDE COMPOUNDS

The ketolides, semisynthetic macrolides prepared from erythromycin, have recently emerged as promising drugs for the treatment of several clinically important antibiotic resistant bacteria, including the suspected gastric cancer agent Helicobacter pylori. The long term goal of this project is to generate a library of molecules based on the ketolide class of compounds using a novel technology developed by Kosan and others. This technology, termed combinatorial biosynthesis, allows one to alter and construct novel polyketide products by genetically manipulating the polyketide synthases (PKSs) which biosynthesize this rich source of pharmacologically active compounds. Specifically, the erythromycin PKS will be engineered to produce, in vivo, a ketolide intermediate which is currently derived from a complicated synthetic strategy. An important advantage to this approach is the ability to manipulate structural features of ketolides which are inaccessible through standard techniques. Ultimately, a number of structurally diverse ketolide intermediates will be produced with anticipated activity against H. pylori and other resistant organisms. Phase I will consist of engineering the erythromycin PKS to produce a ketolide intermediate. Two mutations will be introduced by an in-vivo recombination strategy, and the resulting PKS will be expressed ina specially constructed Streptomyces host/vector system.