ABSTRACT
We report the metabolomics-driven genome mining of a new cyclic-guanidino incorporating non-ribosomal peptide synthetase (NRPS) gene cluster and full structure elucidation of its associated hexapeptide product, faulknamycin. Structural studies unveiled that this natural product contained the previously unknown (R,S)-stereoisomer of capreomycidine, d-capreomycidine. Furthermore, heterologous expression of the identified gene cluster successfully reproduces faulknamycin production without an observed homologue of VioD, the pyridoxal phosphate (PLP)-dependent enzyme found in all previous l-capreomycidine biosynthesis. An alternative NRPS-dependent pathway for d-capreomycidine biosynthesis is proposed.
Subject(s)
Arginine/analogs & derivatives , Multigene Family , Streptomyces/genetics , Arginine/genetics , Arginine/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Biosynthetic Pathways , Genomics , Metabolomics , Peptide Synthases/genetics , Peptide Synthases/metabolism , Streptomyces/metabolismABSTRACT
Genome mining has become a key technology to exploit natural product diversity. Although initially performed on a single-genome basis, the process is now being scaled up to mine entire genera, strain collections and microbiomes. However, no bioinformatic framework is currently available for effectively analyzing datasets of this size and complexity. In the present study, a streamlined computational workflow is provided, consisting of two new software tools: the 'biosynthetic gene similarity clustering and prospecting engine' (BiG-SCAPE), which facilitates fast and interactive sequence similarity network analysis of biosynthetic gene clusters and gene cluster families; and the 'core analysis of syntenic orthologues to prioritize natural product gene clusters' (CORASON), which elucidates phylogenetic relationships within and across these families. BiG-SCAPE is validated by correlating its output to metabolomic data across 363 actinobacterial strains and the discovery potential of CORASON is demonstrated by comprehensively mapping biosynthetic diversity across a range of detoxin/rimosamide-related gene cluster families, culminating in the characterization of seven detoxin analogues.
Subject(s)
Actinobacteria/genetics , Biosynthetic Pathways/genetics , Computational Biology/methods , Genome, Bacterial , Algorithms , Biological Products , Cluster Analysis , Data Mining/methods , Genomics , Metabolomics , Microbiota , Multigene Family , Phylogeny , Reproducibility of Results , SoftwareABSTRACT
Natural products (NPs) are a rich source of medicines, but traditional discovery methods are often unsuccessful due to high rates of rediscovery. Genetic approaches for NP discovery are promising, but progress has been slow due to the difficulty of identifying unique biosynthetic gene clusters (BGCs) and poor gene expression. We previously developed the metabologenomics method, which combines genomic and metabolomic data to discover new NPs and their BGCs. Here, we utilize metabologenomics in combination with molecular networking to discover a novel class of NPs, the tyrobetaines: nonribosomal peptides with an unusual trimethylammonium tyrosine residue. The BGC for this unusual class of compounds was identified using metabologenomics and computational structure prediction data. Heterologous expression confirmed the BGC and suggests an unusual mechanism for trimethylammonium formation. Overall, the discovery of the tyrobetaines shows the great potential of metabologenomics combined with molecular networking and computational structure prediction for identifying interesting biosynthetic reactions and novel NPs.
