Charging of tRNAs using ribozymes and selection of cyclic peptides containing thioethers

Methods Mol Biol. 2012;805:335-48. doi: 10.1007/978-1-61779-379-0_19.

Abstract

In vitro selection methods represent a powerful approach toward identifying high-affinity peptide ligands from highly diverse peptide libraries against a desired target. We herein describe a method for the display and selection of cyclic thioether peptide libraries. Reprogramming the initiation event from fMet to an N-chloroacetyl-amino acid by utilizing flexizyme to rapidly and efficiently prepare the aa-tRNA can be effectively used to initiate translation, upon which the thiol group of an inserted cysteine at the C terminus of the designed library spontaneously reacts to yield a nonreducible cyclic thioether peptide readily compatible with any in vitro display methods. Thus, cyclic peptides already in a nonreducible stable form can be selected directly against the target of interest.

MeSH terms

  • Peptide Library
  • Peptides, Cyclic / chemistry
  • Peptides, Cyclic / genetics*
  • Peptides, Cyclic / metabolism*
  • Polymerase Chain Reaction
  • RNA, Catalytic / genetics
  • RNA, Catalytic / metabolism*
  • RNA, Transfer / genetics*
  • Ribosomes / genetics
  • Sulfides / metabolism*

Substances

  • Peptide Library
  • Peptides, Cyclic
  • RNA, Catalytic
  • Sulfides
  • RNA, Transfer