Accessing Diverse Pyridine-Based Macrocyclic Peptides by a Two-Site Recognition Pathway

Dinh T. Nguyen, Tung T. Le, Andrew J. Rice, Graham A. Hudson, Wilfred A. Van Der Donk, Douglas A. Mitchell

Research output: Contribution to journalArticlepeer-review


Macrocyclic peptides are sought-after molecular scaffolds for drug discovery, and new methods to access diverse libraries are of increasing interest. Here, we report the enzymatic synthesis of pyridine-based macrocyclic peptides (pyritides) from linear precursor peptides. Pyritides are a recently described class of ribosomally synthesized and post-translationally modified peptides (RiPPs) and are related to the long-known thiopeptide natural products. RiPP precursors typically contain an N-terminal leader region that is physically engaged by the biosynthetic proteins that catalyze modification of the C-terminal core region of the precursor peptide. We demonstrate that pyritide-forming enzymes recognize both the leader region and a C-terminal tripeptide motif, with each contributing to site-selective substrate modification. Substitutions in the core region were well-tolerated and facilitated the generation of a wide range of pyritide analogues, with variations in macrocycle sequence and size. A combination of the pyritide biosynthetic pathway with azole-forming enzymes was utilized to generate a thiazole-containing pyritide (historically known as a thiopeptide) with no similarity in sequence and macrocycle size to the naturally encoded pyritides. The broad substrate scope of the pyritide biosynthetic enzymes serves as a future platform for macrocyclic peptide lead discovery and optimization.

Original languageEnglish (US)
Pages (from-to)11263-11269
Number of pages7
JournalJournal of the American Chemical Society
Issue number25
StatePublished - Jun 29 2022

ASJC Scopus subject areas

  • General Chemistry
  • Biochemistry
  • Catalysis
  • Colloid and Surface Chemistry


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