Structure Prediction and Synthesis of Pyridine-Based Macrocyclic Peptide Natural Products

Abstract

The structural and functional characterization of natural products is vastly outpaced by the bioinformatic identification of biosynthetic gene clusters (BGCs) that encode such molecules. Uniting our knowledge of bioinformatics and enzymology to predict and synthetically access natural products is an effective platform for investigating cryptic/silent BGCs. We report the identification, biosynthesis, and total synthesis of a minimalistic class of ribosomally synthesized and post-translationally modified peptides (RiPPs) with the responsible BGCs encoding a subset of enzymes known from thiopeptide biosynthesis. On the basis of the BGC content, these RiPPs were predicted to undergo enzymatic dehydration of serine followed by [4+2]-cycloaddition to produce a trisubstituted, pyridine-based macrocycle. These RiPPs, termed "pyritides", thus contain the same six-membered, nitrogenous heterocycle that defines the thiopeptide RiPP class but lack the ubiquitous thiazole/thiazoline heterocycles, suggesting that thiopeptides should be reclassified as a more elaborate subclass of the pyritides. One pyritide product was obtained using an 11-step synthesis, and the structure verified by an orthogonal chemoenzymatic route using the precursor peptide and cognate pyridine synthase. This work exemplifies complementary bioinformatics, enzymology, and synthesis to characterize a minimalistic yet structurally intriguing scaffold that, unlike most thiopeptides, lacks growth-suppressive activity toward Gram-positive bacteria.