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Natural product pathway discovery through activity–guided single–cell genomics
Date
March 20, 2022
We have developed a new strategy to identify natural product biosynthetic pathways from complex microbial ecosystems that combines chemical biology tools with single cell sequencing. This methodology leverages a fluorescent in situ enzyme assay that targets carrier proteins common to polyketide (PKS) and non–ribosomal peptide synthetases (NRPS). By applying fluorescence–activated cell sorting to a treated microbiome, microbes with active secondary metabolic capabilities can be enriched and subjected to single–cell genomics. As an example, we demonstrated the genetic basis for biosynthetic diversity in complex marine organisms like tunacates. Here, the enzyme–active cells revealed a member of marine Oceanospirillales harboring a novel NRPS gene cluster with high similarity to phylogenetically distant marine and terrestrial bacteria. Interestingly, this synthase belongs to a larger class of siderophore biosynthetic gene clusters commonly associated with pestilence and disease. This strategy demonstrates activity–guided single–cell genomics as a tool to guide novel biosynthetic discovery.
Activity–guided microbial single–cell genomics of Ciona intestinalis
In the current study of polyketide and non-ribosomal peptide biosynthesis, a major goal has been the structural elucidation of the megasynthases that produce them. We have been developing chemical biology tools to study these enzyme systems, with a focus on dual site-selective crosslinking probes…
Carrier protein-dependent biosynthetic pathways produce a diverse range of primary and secondary metabolites, many of which manifest in our daily lives as therapeutics and antibiotics…
_Mycobacterium tuberculosis _(_Mtb_) is the causative agent of the disease Tuberculosis (TB), and it employs the type II fatty acid synthase (FAS-II) metabolic pathway…
Fatty acids are prevalent in all walks of life and are produced by fatty acid synthases (FASs) in which an acyl carrier protein (ACP) tethers a growing acyl chain as it is elongated and modified by a series of partner proteins…