Extended Data Fig. 4: Comparison of binding affinities when changing repeat numbers from either binder or peptide side. and top five flexible backbone docks for the four-repeat LRP binder RPB_LRP2_R4–LRPx4.

a, Six-repeat versions of RPB_LRP2_R6 and RPB_PEW2_R6 had higher affinity for eight-repeat LRP and PEW peptides than four-repeat versions without any decrease in specificity in yeast surface display. Biotinylated repeat proteins (the six-repeat versions RPB_LRP2_R6 and RPB_PEW2_R6 and the four-repeat versions RPB_LRP2_R4 and RPB_PEW2_R4) were detected by SAPE, and the expression of the designed repeat peptide on yeast surface was monitored by FITC-conjugated anti-Myc antibody. Serial dilutions were tested for each binder, and the full tested concentration is labelled. b, Six-repeat IYP and PLP peptides had higher affinity for six-repeat versions of the cognate designed repeat proteins (RPB_IYP1_R6 and RPB_PLP1_R6) than four-repeat versions by bio-layer interferometry. The full tested concentration is labelled. The biotinylated target peptides were loaded onto the streptavidin (SA) biosensors, and incubated with designed binders in solution to measure association and dissociation. The dissociation rate was markedly increased when testing against the six-repeat peptides as compared to the four-repeat peptides, indicating a much tighter binding event. c, Top five complex PDBs for RPB_LRP2_R4–LRPx4 from the flexible docking generated ensemble. Green, pink and grey are the ones closest to the crystal structure (shown in yellow) with RMSD over the peptide and the binding residues ≈ 0.03 Å, whereas the cyan dock RMSD = 3.89 Å.