We report a synthetic methodology for the installation of methyllysine mimics on cysteine-containing peptides and bacteriophage peptide libraries. Strategies that allow for diversity and high throughput screening of PTM-containing peptides are critical for successfully targeting the many methyllysine reader proteins that are misregulated in cancer and disease. We have de- veloped conditions for alkylation of cysteine containing peptides with (2-haloethyl) amines, providing products that closely mimic methyllysine residues. Extensive optimization on C7C peptide phage constructs allowed for the successful installation of Kme3 mimics in 60–70% yields to create post-translational ε-Lys-N-methylated peptide phage libraries. Optimized reaction conditions between 2-bromo-N,N,N-trimethylethaninium bromide and commercially available PhD C7C library produce >2 × 1011 phage parti- cles and libraries of ~2 × 108 diversity in which each peptide sequence contains the Kme3 mimic. This process adds a new fragment into readily available genetically encoded libraries and opens new avenues for high throughput screening that may give rise to new ligands for a variety of methyllysine reader proteins.
Installation of cysteine-derived methyllysine mimics on phage-dis- played peptide libraries: optimization of reaction conditions for conversion and phage viability