Biosynthetic Analysis of Marine Cyanobacterial Pathways

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DESCRIPTION (provided by applicant): Marine cyanobacteria are extraordinarily rich in their production of biologically-active and structurally- unique natural products. A number of these secondary metabolites or their derivatives are lead compounds n drug development programs aimed at providing new therapies to treat cancer, bacterial infections, inflammatory responses, and in crop protection to kill harmful microbial pathogens and insects. Isolation and structural analysis of marine and terrestrial cyanobacterial natural products has provided access to an unusually large number of mixed non-ribosomal peptide synthetase/polyketide synthase (NRPS/PKS) systems. The corresponding metabolic systems are comprised of an intriguing set of complex multifunctional proteins that along with allied enzymes generate structurally complex molecules via a modular multi-step process. Over the past several years the Sherman and Gerwick laboratories have developed a complementary program to clone and characterize the biosynthetic pathways of novel cyanobacterial secondary metabolites that possess significant potential for biotechnological applications. A full understanding of the molecular mechanisms, catalytic activities, kinetic properties, and substrate specificities within cyanobacterial biosynthetic pathways is just beginning to unfold. The proposed research will build upon our studies of the curacin and jamaicamide metabolic systems, two distinct yet related pathways that are genetically characterized and poised for detailed biochemical studies. This detailed genetic and biochemical understanding will facilitate the design of new biosynthetic systems that harness the growing potential of cyanobacterial secondary metabolism. Despite considerable gains over the past few years, the full promise of cyanobacterial natural products to yield new lead compounds for development as useful Pharmaceuticals, will only be realized by closing a series of key gaps in knowledge and technology. Solving these challenges will require development and optimization of genetic and biochemical methods that allow us to 1) utilize unique secondary metabolite enzymes for creation of novel small molecules, 2) manipulate cyanobacterial natural product gene clusters to produce analog structures. The specific aims are: 1. To investigate biochemically unique aspects of the curacin (Cur), and jamaicamide (Jam) biosynthetic pathways including formation of the cyclopropane ring, cis-alkene formation, and termination in Cur, and chain initiation, vinyl chloride formation and termination in Jam. 2. Perform bioassays on new compounds resulting from Specific Aim 1 including evaluation for inhibition of tubulin polymerization and binding site specificity, biochemical assays relevant to cancer, and in house screens at U-M and SIO relevant to anti-microbial activity and neurotoxicity, respectively.