Fig. 3: Structural and biophysical analysis of PEAK/CrkII^NSH3 interaction.

a SPR data summarizing measured steady-state binding affinity (K_D) of synthetic PEAK PRM peptides toward immobilized CrkII^NSH3 domain. b Depiction of the published CrkII^NSH3:Abl⁷⁵⁸ structure (PDB: 5IH2)⁴¹, showing key residues within the CrkII^NSH3 consensus motif in Abl⁷⁵⁸ (peptide shown in gold stick representation) crucial for high affinity binding to CrkII^NSH3 (main chain light blue; CrkII^NSH3 surface is colored by electrostatic surface potential calculated using UCSF Chimera v 1.16; blue = positive, white = hydrophobic, red = negative). Aligned is the sequence of PEAK3^54–66, showing key conserved residues of the CrkII^NSH3 motif. c, d Interaction studies with recombinant PEAK1^IDR1/PEAK2^IDR1 and CrkII^FL underscore the role of avidity for high affinity binding of CrkII^FL to PEAK dimers; c Incubation of PEAK2^IDR1 with CrkII^FL dimer at a 1:1.3 ratio results in a complex formation while (d) incubation with CrkII^FL monomer does not result in a complex, as confirmed by SDS-PAGE analysis of SEC eluted fractions (n = 3, independent experiments). Plots of the CrkII^FL dimer and monomer are shown in light blue; PEAK2^IDR1 in pink; and the complex of PEAK2^IDR1 dimer:CrkII^FL dimer in black (see Supplementary Fig. 3d for PEAK1^IDR1 dimer:CrkII^FL dimer complex).