Dereplication and De Novo Sequencing of Nonribosomal

Nonribosomal peptides (NRPs) are of great pharmacological importance, but there is currently no technology for high-throughput NRP dereplication and sequencing. We employ multi-stage mass spectrometry followed by spectral alignment algorithms for sequencing of cyclic NRPs. We also present an algorithm for comparative NRP dereplication that establishes similarities between newly isolated and previously identified similar but non-identical NRPs, substantially reducing dereplication efforts.. The classical protein synthesis pathway (translation of template mRNA) is not the only mechanism for cells to assemble amino acids into proteins/peptides. Nonribosomal peptide synthesis is performed by Nonribosomal Peptide Synthetases (NRPS) that represent both the mRNA-free template and building machinery for the peptide biosynthesis1. NRPS produce nonribosomal peptides (NRP) that are not directly inscribed in genomes and thus cannot be inferred with traditional DNA sequencing. NRPs are of great pharmacological importance since they were optimized by evolution for chemical defense and communication. Starting from penicillin, NRPs and other natural products have an unparalleled track record i