1. Academic Validation
  2. Genetically programmed cell-based synthesis of non-natural peptide and depsipeptide macrocycles

Genetically programmed cell-based synthesis of non-natural peptide and depsipeptide macrocycles

  • Nat Chem. 2022 Dec 22. doi: 10.1038/s41557-022-01082-0.
Martin Spinck # 1 Carlos Piedrafita # 1 Wesley E Robertson 2 Thomas S Elliott 1 Daniele Cervettini 1 Daniel de la Torre 1 Jason W Chin 3
Affiliations

Affiliations

  • 1 Medical Research Council Laboratory of Molecular Biology, Cambridge, UK.
  • 2 Medical Research Council Laboratory of Molecular Biology, Cambridge, UK. [email protected]
  • 3 Medical Research Council Laboratory of Molecular Biology, Cambridge, UK. [email protected]
  • # Contributed equally.
Abstract

The direct genetically encoded cell-based synthesis of non-natural peptide and depsipeptide macrocycles is an outstanding challenge. Here we programme the encoded synthesis of 25 diverse non-natural macrocyclic Peptides, each containing two non-canonical Amino acids, in Syn61Δ3-derived cells; these cells contain a synthetic Escherichia coli genome in which the annotated occurrences of two sense codons and a stop codon, and the cognate transfer RNAs (tRNAs) and release factor that normally decode these codons, have been removed. We further demonstrate that pyrrolysyl-tRNA synthetase/tRNA pairs from distinct classes can be engineered to direct the co-translational incorporation of diverse alpha hydroxy acids, with both aliphatic and aromatic side chains. We define 49 engineered mutually orthogonal pairs that recognize distinct non-canonical Amino acids or alpha hydroxy acids and decode distinct codons. Finally, we combine our advances to programme Syn61Δ3-derived cells for the encoded synthesis of 12 diverse non-natural depsipeptide macrocycles, which contain two non-canonical side chains and either one or two ester bonds.

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