4 research outputs found
Malleilactone, a Polyketide Synthase-Derived Virulence Factor Encoded by the Cryptic Secondary Metabolome of Burkholderia pseudomallei Group Pathogens
Sequenced bacterial genomes are routinely found to contain
gene
clusters that are predicted to encode metabolites not seen in fermentation-based
studies. Pseudomallei group <i>Burkholderia</i> are emerging
pathogens whose genomes are particularly rich in cryptic natural product
biosynthetic gene clusters. We systematically probed the influence
of the cryptic secondary metabolome on the virulence of these bacteria
and found that disruption of the MAL gene cluster, which is natively
silent in laboratory fermentation experiments and conserved across
this group of pathogens, attenuates virulence in animal models. Using
a promoter exchange strategy to activate the MAL cluster, we identified
malleilactone, a polyketide synthase-derived cytotoxic siderophore
encoded by this gene cluster. Small molecules targeting malleilactone
biosynthesis either alone or in conjunction with antibiotics could
prove useful as therapeutics to combat melioidosis and glanders
Targeted Metagenomics: Finding Rare Tryptophan Dimer Natural Products in the Environment
Natural
product discovery from environmental genomes (metagenomics)
has largely been limited to the screening of existing environmental
DNA (eDNA) libraries. Here, we have coupled a chemical-biogeographic
survey of chromopyrrolic acid synthase (CPAS) gene diversity with
targeted eDNA library production to more efficiently access rare tryptophan
dimer (TD) biosynthetic gene clusters. A combination of traditional
and synthetic biology-based heterologous expression efforts using
eDNA-derived gene clusters led to the production of hydroxysporine
(<b>1</b>) and reductasporine (<b>2</b>), two bioactive
TDs. As suggested by our phylogenetic analysis of CPAS genes, identified
in our survey of crude eDNA extracts, reductasporine (<b>2</b>) contains an unprecedented TD core structure: a pyrrolinium indolocarbazole
core that is likely key to its unusual bioactivity profile. This work
demonstrates the potential for the discovery of structurally rare
and biologically interesting natural products using targeted metagenomics,
where environmental samples are prescreened to identify the most phylogenetically
unique gene sequences and molecules associated with these genes are
accessed through targeted metagenomic library construction and heterologous
expression
Discovery and Synthetic Refactoring of Tryptophan Dimer Gene Clusters from the Environment
Here we investigate bacterial tryptophan
dimer (TD) biosynthesis
by probing environmental DNA (eDNA) libraries for chromopyrrolic acid
(CPA) synthase genes. Functional and bioinformatics analyses of TD
clusters indicate that CPA synthase gene sequences diverge in concert
with the functional output of their respective clusters, making this
gene a powerful tool for guiding the discovery of novel TDs from the
environment. Twelve unprecedented TD biosynthetic gene clusters that
can be arranged into five groups (A–E) based on their ability
to generate distinct TD core substructures were recovered from eDNA
libraries. Four of these groups contain clusters from both cultured
and culture independent studies, while the remaining group consists
entirely of eDNA-derived clusters. The complete synthetic refactoring
of a representative gene cluster from the latter eDNA specific group
led to the characterization of the erdasporines, cytotoxins with a
novel carboxy-indolocarbazole TD substructure. Analysis of CPA synthase
genes in crude eDNA suggests the presence of additional TD gene clusters
in soil environments
Antimicrobials Inspired by Nonribosomal Peptide Synthetase Gene Clusters
Bacterial culture broth extracts
have been the starting point for
the development of numerous therapeutics. However, only a small fraction
of bacterial biosynthetic diversity is accessible using this strategy.
Here, we apply a discovery approach that bypasses the culturing step
entirely by bioinformatically predicting small molecule structures
from the primary sequences of the biosynthetic gene clusters. These
structures are then chemically synthesized to give synthetic-bioinformatic
natural products (syn-BNPs). Using this approach, we screened syn-BNPs
inspired by nonribosomal peptide synthetases against microbial pathogens,
and discovered an antibiotic for which no resistance could be identified
and an antifungal agent with activity against diverse fungal pathogens