Secondary metabolites produced by the human gut microbiota are potentially relevant to human health, but identification of biosynthetic gene clusters in this environment has far outpaced the discovery of their small molecule products. Isolation-based natural product discovery in commensal human gut microbes is difficult, as secondary metabolite biosynthetic gene clusters from these organisms tend to remain “silent” under standard laboratory culture conditions. To circumvent this challenge, we targeted a biosynthetic gene cluster from the prominent human gut commensal <i>Ruminococcus bromii</i> for natural product discovery using an isolation-independent approach. This gene cluster contains a single-polypeptide non-ribosomal peptide synthetase (NRPS) and is predicted to produce an <i>N-</i>acylated dipeptide aldehyde, member of a known class of protease inhibitors. We used in vitro biochemical assays with purified NRPS enzymes to identify key building blocks and verified our structural prediction by performing a total in vitro biosynthetic reconstitution and total synthesis of the peptide aldehyde natural product, ruminopeptin. Further studies of ruminopeptin’s biological activity, including protease inhibition assays, support the hypothesis that this metabolite mediates microbe-microbe interactions in the human gut.

Discovery of a small molecule protease inhibitor from an abundant human gut commensal microbe

254th National Meeting (2017)

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