Flexizyme-Enabled Benchtop Biosynthesis of Thiopeptides

J Am Chem Soc. 2019 Jan 16;141(2):758-762. doi: 10.1021/jacs.8b11521. Epub 2019 Jan 8.


Thiopeptides are natural antibiotics that are fashioned from short peptides by multiple layers of post-translational modification. Their biosynthesis, in particular the pyridine synthases that form the macrocyclic antibiotic core, has attracted intensive research but is complicated by the challenges of reconstituting multiple-pathway enzymes. By combining select RiPP enzymes with cell free expression and flexizyme-based codon reprogramming, we have developed a benchtop biosynthesis of thiopeptide scaffolds. This strategy side-steps several challenges related to the investigation of thiopeptide enzymes and allows access to analytical quantities of new thiopeptide analogs. We further demonstrate that this strategy can be used to validate the activity of new pyridine synthases without the need to reconstitute the cognate prior pathway enzymes.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Anti-Bacterial Agents / chemical synthesis*
  • Aptamers, Nucleotide / chemistry*
  • Peptides, Cyclic / chemical synthesis*
  • Proof of Concept Study
  • RNA, Catalytic / chemistry*
  • Sequence Alignment
  • Thiazoles / chemical synthesis*


  • Anti-Bacterial Agents
  • Aptamers, Nucleotide
  • Peptides, Cyclic
  • RNA, Catalytic
  • Thiazoles