A Facile Method for Determining Lanthipeptide Stereochemistry

Abstract

ABSTRACTLanthipeptides are a large group of natural products that belong to the ribosomally synthesized and post-translationally modified peptides (RiPPs). Lanthipeptides contain lanthionine and methyllanthionine bis amino acids that have varying stereochemistry. The stereochemistry of new lanthipeptides is often not determined because current methods require equipment that is not standard in most laboratories. In this study, we developed a facile, efficient, and user-friendly method for detecting lanthipeptide stereochemistry utilizing advanced Marfey’s analysis. Under optimized conditions, 0.05 mg peptide is sufficient to characterize the stereochemistry of five (methyl)lanthionines of different stereochemistry using a simple liquid chromatography set-up, which is a much lower detection limit than current methods. In addition, we describe methods to readily access standards of the three different methyllanthionine stereoisomers and the two different lanthionine stereoisomers that have been reported in known lanthipeptides. The developed workflow uses commonly used non-chiral column system and offers a scalable platform to assist antimicrobial discovery. We illustrate its utility with an example of a lanthipeptide discovered by genome mining.