Synthase-Selective Exploration of a Tunicate Microbiome by Activity-Guided Single-Cell Genomics.

While thousands of environmental metagenomes have been mined for the presence of novel biosynthetic gene clusters, such computational predictions do not provide evidence of their biosynthetic functionality. Using fluorescent enzyme assay targeting carrier proteins common to polyketide (PKS) and nonribosomal peptide synthetases (NRPS), we applied fluorescence-activated cell sorting to tunicate microbiome to enrich for microbes with active secondary metabolic capabilities. Single-cell genomics uncovered the genetic basis for a wide biosynthetic diversity in the enzyme-active cells and revealed a member of marine 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 demonstrates activity-guided single-cell genomics as a tool to guide novel biosynthetic discovery.