ZHOU_PFL_POLYKETIDES

"Genome mining in Streptomyces violaceusniger, a prolific antibiotic producer"

 Partecipanti

Mappa

 Word cloud

Esplora la "nuvola delle parole (Word Cloud) per avere un'idea di massima del progetto.

 Obiettivo del progetto (Objective)

'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.'

Introduzione (Teaser)

Synthetic biology deals with the re-design of natural biosynthetic pathways for the production of specific molecules. A European project followed this approach to produce natural compounds with bioactive potential.

Descrizione progetto (Article)

Streptomyces is a genus of bacteria capable of growing in diverse environments. Their most intriguing property is the production of biologically active secondary metabolites that can serve as anti-fungal and anticancer agents or antibiotics. Expression of these molecules endows Streptomyces bacteria with a competitive advantage, thereby improving their overall survival. As a result, Streptomyces could be exploited in biotechnology for the production of antibiotics or other biologically relevant molecules.

In this context, the EU-funded 'Genome mining in Streptomyces violaceusniger, a prolific antibiotic producer' (ZHOU_PFL_POLYKETIDES) project focused on a particular family of enzymes that are implicated in the biosynthesis of antibiotics in the bacterial species Streptomyces violaceusniger DSM4137. These multi-enzyme complexes are called polyketide synthases and catalyse the production of polyketide compounds such as erythromycin and doxycycline.

In this context, scientists isolated, cloned and characterised large biosynthetic gene clusters from Streptomyces violaceusniger DSM4137 for the production of various natural products. To further understand the association between biosynthetic pathways, they deleted certain genes of these clusters or altered regulatory genes, and observed the synthesis output. Also, through genetic engineering, they obtained mutants of the polyketide synthase cluster to maximise expression.

These efforts led to successful production of the anti-fungal compound azalomycin, the anti-cancer agent nigericin and the antibiotic elaiophylin. Considerable work also went into the enzymology of these reactions and the molecular mechanism underlying the chemical biosynthesis of these compounds.

Collectively, the activities of the study support the transfer of biosynthetic enzymes in biotechnologically relevant microorganisms to produce a specified target molecule. This approach overcomes a significant bottleneck in the synthetic biology of natural products.