Transcription-factor centric genome mining strategy for discovery of diverse unprecedented RiPP gene clusters

Abstract

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a rapidly emerging group of natural products with diverse biological activity. Most of their biosynthetic mechanisms are well studied and the "genome mining" strategy based on homology has led to the unearthing of many new ribosomal natural products, including lantipeptides, lasso peptides, cyanobactins. These precursor-centric or biosynthetic protein-centric genome mining strategies have encouraged the discovery of RiPPs natural products. However, a limitation of these strategies is that the newly identified natural products are similar to the known products and novel families of RiPP pathways were overlooked by these strategies. In this work, we applied a transcription-factor centric genome mining strategy and diverse unique crosslinked RiPP gene clusters were predicted in several sequenced microorganisms. Our research could significantly expand the category of biosynthetic pathways of RiPP natural products and predict new resources for novel RiPPs. 2

Mammalian Commensal Streptococci Utilize a Rare Family of Class VI Lanthipeptide Synthetases to Synthesize Miniature Lanthipeptide-type Ribosomal Peptide Natural Products

“…Next, in addition to the genetic surroundings, each identified Rgg-type regulator led us to the discovery of diverse RiPP gene clusters. 43 As expected, most of the identified QS-regulated RiPP gene clusters belonged to the Ras−RiPP family, which has been reported previously. 31,43 These results indicated that multiple Streptococci widely use radical SAM enzymes to synthesize natural products to facilitate specific microbial communication.…”

“…43 As expected, most of the identified QS-regulated RiPP gene clusters belonged to the Ras−RiPP family, which has been reported previously. 31,43 These results indicated that multiple Streptococci widely use radical SAM enzymes to synthesize natural products to facilitate specific microbial communication. In addition to the Ras−RiPPs, we turned our attention to other types of RiPPs.…”

“…To examine this hypothesis, we used the Rgg transcriptional regulator (WP_011681385.1) from the streptide BGC as an inquiry, and intensive position-specific iterative (PSI)-BLAST searches were carried out against the NCBI database of nonredundant protein sequences. Next, in addition to the genetic surroundings, each identified Rgg-type regulator led us to the discovery of diverse RiPP gene clusters . As expected, most of the identified QS-regulated RiPP gene clusters belonged to the Ras–RiPP family, which has been reported previously. , These results indicated that multiple Streptococci widely use radical SAM enzymes to synthesize natural products to facilitate specific microbial communication.…”

“…When the cell density reaches a threshold concentration, SHP, which is a hydrophobic peptide signal, will bind to the Rgg response regulator and trigger the expression of the streptide operon, which normally contains a short precursor peptide, processing enzymes, and a transporter . Such a conservative genetic context makes it easy to discover unprecedented RiPP systems and predict a convincing precursor candidate between the regulator and the processing enzymes. ,− …”

“…In summary, we herein report the discovery of an unprecedented class VI lanthipeptides that widely existed in mammalian commensal Streptococci and have been overlooked in the past. With a transcription-factor-centric genome mining strategy, 43 class VI lanthipeptide synthetases that have low similarity with previous systems have been identified and are frequently associated with diverse QS systems, indicating their specific role in nature. By performing thorough in vitro studies, we showed that these lanthipeptide synthetases specifically recognized a small substrate with a short leader peptide to guide the catalysis and only installed one lanthionine moiety onto its short precursor peptide by connecting a conserved TxxC region.…”