Fig. 3: De novo design of high-affinity macrocycle binders to GABARAP.
From: Accurate de novo design of high-affinity protein-binding macrocycles using deep learning
a, AfCycDesign predicted model for design GAB_D8 bound to GABARAP shown as surface, with hotspot residues highlighted in green. b, Affinity determination of GAB_D8 using SPR. SPR sensorgram from a nine-point single-cycle kinetics experiment (fivefold dilution, highest concentration: 20 µM). Experimental data are shown in orange and global fits are shown with black lines. The Kd is also shown on the plot. c, Superposition of chains E and F from the X-ray crystal structure of GAB_D8 bound to GABARAPL1 and the AfCycDesign model. d, AfCycDesign predicted model for design GAB_D23 bound to GABARAP shown as surface, with hotspot residues highlighted in green. e, Affinity determination of GAB_D23 using SPR. SPR sensorgram from a nine-point single-cycle kinetics experiment (fivefold dilution, highest concentration: 20 µM). Experimental data are shown in pink and global fits are shown with black lines. The Kd is also shown on the plot. f, Alignment of chains A and B from the X-ray crystal structure of GAB_D23 bound to GABARAP and the AfCycDesign model. g, Alignments of GAB_D8 and GAB_D23 macrocycle models to X-ray crystal structures show close matches. h, Comparison of GAB_D8 and GAB_D23 binding modes in the design models. i, Competitive AlphaScreen dose-response plot, IC50 from the average of three experiments. Donor and acceptor beads in the assay are bound to GABARAP and GABARAP-binding peptide K1, respectively.
