RiPPs from the Gut · Functional Exploration of Biosynthetic Dark Matter in the Human Gut Microbiome

Functional Exploration of Biosynthetic Dark Matter in the Human Gut Microbiome

The number of microbial cells in the human body is projected to be in the trillions, amounting to a 10-fold excess of microbial to human cells. Most of the microbial cells in the body are located in the gut, which harbors a great diversity of bacterial species. Functional studies of gut bacteria are generally challenging, since most are as-yet unculturable, fastidious growers or genetically intractable. Nonetheless, metagenomic sequencing of gut samples has generated an abundance of data, and “sequence-gazing” insights have provided interesting clues into microbial metabolism. The interactions within bacterial communities and between bacteria and host are often mediated by the molecular language of secondary metabolites, also termed natural products (NPs). Understanding the secondary metabolism of microbiome bacteria can elucidate their ecological roles within the microbiome and their contribution to human health. The molecular machinery required to synthesize secondary metabolites is encoded in bacterial genomes by autonomous units called biosynthetic gene clusters (BGCs). Remarkably, very little is known about the small molecule products of these BGCs or about why bacteria are producing them The great complexity, diversity and density of the gut microbiome results in a rich environment for the discovery and functional study of molecules that mediate interactions between members of the community and the host. To address the lack of structural and functional knowledge of the secondary metabolism of gut bacteria, our project used genome mining and synthetic biology to study the function of novel NPs from bacteria from human gut microbiome. In Work Package 1 (WP1), we explored the biosynthetic dark matter of human gut bacteria through genome mining approaches for the discovery of spliceotide and origamin RiPP natural products. And in Work Package 2 (WP2), we explored the function of spliceotide and origamin natural products and possible applications therapeutics. We uncovered new biosynthetic gene clusters encoding for spliceotides from gut bacteria. While we were unable to obtain functional data for these systems, we were uncovered the biological activity of 4 other spliceotide products. All products showed protease inhibition activity and, remarkably, two spliceotides reproducibly demonstrated low-nanomolar activity against Human Neutrophil Elastase. We also discovered origamin BGCs from gut microbes. We had previously demonstrated the activities for two enzymes in origamin biosynthesis in E. coli, but were not able to reproduce those results in origamins from gut bacteria. Nonetheless, we expanded our search of origamin clusters with the goal of isolating a final origamin product by obtaining activity of all key biosynthetic enzymes. We identified a new type of origamin BGC and were able to obtain activity for all three tailoring enzymes which produced one of the most extensively modified RiPPs known to date.

Текст от CORDIS, на английски · Данни: CORDIS, © Европейски съюз