(671c) Discovery and Chararacterization of a Class IV Lanthipeptide with a Novel Ring Topology

Lanthipeptides, which is short for lanthionine-containing peptides, constitute a major family of ribosomally synthesized and post-translationally modified peptides (RiPPs). Based on the characteristics of lanthipeptide synthases, lanthipepides are classified into four subfamilies: Class I-Class IV. While over a hundred lanthipeptides have been discovered to date, only very few of them are Class IV lanthipeptides and they are structurally similar. Here, a new group of Class IV lanthipeptide biosynthetic pathways was identified through bioinformatics analysis. The lanthipeptide biosynthetic pathway from Streptomyces sp. NRRL S-1022 was refactored and heterologously expressed in Escherichia coli, which generated a lanthipeptide with two non-overlapped rings, a new topology that distinguishes this compound from known Class IV lanthipeptides. In order to further study the mechanism of ring formation, the LanL was heterologously expressed and purified for in vitro assays. Through analysis of the intermediates by MS/MS, it was found that there are multiple routes for the ring formation while the cyclase domain exhibits some preference on the C to N direction for cyclization. Overall, we have successfully identified a new group of Class IV lanthipeptide and studied its biosynthesis in vitro, which expands our knowledge of the Class IV lanthipeptide structures and biosynthesis.