Scheme 1.

Overview of a stapled-peptide library screen. Linear peptides synthesized on resin are cyclized using 1^st generation Grubbs catalyst, and excess catalyst is removed by washing with tris(hydroxymethyl)phosphine. X₁, X₂ and X₃ comprise the variable region. The one-bead-one-compound library of stapled peptides is incubated with a mixture of biotinylated target protein and non-biotinylated competitor to identify selective binders. Cyclized peptides that bind the target are visualized using a fluorescence microscope and then decoded by MALDI-TOF mass spectrometry.