Figure 2.

(a) Schematic structures of three G4 bicyclic peptide ligands elicited by phage display, exhibiting both common G4 small molecule binding motifs and hydrophobic amino acids. (b) Next-generation sequencing permits motif analysis of the selection process; the optimal amino acids at each position in the final selected library are identified. Enriched amino acid functionalities include positive charges (position 8) and aromatic rings (9) but also hydrophobic residues (5) and proline (3). (c) ckit-1 G4 melting temperature increase (ΔT_m) as measured by FRET melting. ΔT_m demonstrates progressive improvement of G4 bicyclic peptides from b-G4pep1 to b-G4pep3. At the same time, no significant ΔT_m is observed for a short double-stranded DNA. (d) Apparent K_d values measured by fluorescence quench equilibrium binding assay for b-G4pep1 and b-G4pep3; the quoted K_d is for ckit-1.