Obiettivo
Complex polyketides are microbial natural products (including clinically-useful medicines) whose chemical structures correlate with the sequence of the modular enzymes responsible for their biosynthesis. This unique paradigm means that the structure of the gene can be inferred from the chemical structure of the molecule, and (with less certainty) the reverse is true. Now, genome-scale sequencing of actinomycetes has revealed untapped chemical diversity in culturable streptomycetes: there are many biosynthetic clusters present but most are cryptic (silent). Expression of biosynthetic genes is now a severe bottleneck in trying to develop a genuine “synthetic biology” of natural products, in which PKSs are designed and fabricated to order to produce a specificed target molecule. We have sequenced and will study an extreme example of “one strain-many clusters”: a strain of Streptomyces violaceusniger which houses 11 clusters (of which seven silent and some are novel). We will: (1) use in-frame PKS gene deletion followed by chemical analysis of the resulting mutants, to test whether and how inactivation of one pathway affects the simultaneous production of others; (2) attempt to activate cryptic pathway genes by alterations in regulatory genes and by ribosome engineering, and isolate any new products; (3) clone whole PKS clusters in high-yielding (and sequenced) strains of, respectively, the monensin-producing Streptomyces cinnamonensis and the erythromycin-producing Saccharopolyspora erythraea for heterologous expression of polyketides in “pre-optimised” backgrounds; and (4) use genome reduction triggered by cre-loxP recombination, to radically simplify the secondary metabolite profile of all three strains and test the mutants for improved expression of cloned PKS genes.
Campo scientifico
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FP7-PEOPLE-2010-IIF
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Meccanismo di finanziamento
MC-IIF - International Incoming Fellowships (IIF)Coordinatore
CB2 1TN Cambridge
Regno Unito