Fig. 4: Scaffolding of diverse functional sites with RFdiffusion.

a, RFdiffusion outperforms other methods across 25 benchmark motif-scaffolding problems collected from six recent publications (Supplementary Table 9). In silico success is defined as AF2 r.m.s.d. to design model less than 2 Å, AF2 r.m.s.d. to the native functional motif less than 1 Å and AF2 pAE less than five. One hundred designs were generated per problem, with no previous optimization on the benchmark set (some optimization was necessary for Hallucination). Supplementary Table 10 presents full results. In silico success rates on the problems are correlated between the methods, and RFdiffusion can still struggle on challenging problems in which all methods have low success. b, Four examples of designs in which RFdiffusion significantly outperforms existing methods. Teal, native motif; colours, AF2 prediction of a design. Metrics (r.m.s.d. AF2 versus design/versus native motif (Å), AF2 pAE): 5TRV long, 1.17/0.57; 4.73; 6E6R long, 0.89/0.27, 4.56; 7MRX long, 0.84/0.82 4.32; 5TPN, 0.59/0.49 3.77. c, RFdiffusion can scaffold the p53 helix that binds MDM2 (left) and makes extra contacts with the target (right, average 31% increased surface area. Design was p53_design_89). Designs were generated with an RFdiffusion model fine-tuned on complexes. d, BLI measurements indicate high-affinity binding to MDM2 (p53_design_89, 0.7 nM; p53_design_53, 0.5 nM); the native affinity is 600 nM (ref. ⁴²). e, Out of 95 designs, 55 showed binding to MDM2 (more than 50% of maximum response). Thirty-two of these were monomeric (Supplementary Fig. 10h). f, After fine-tuning (Supplementary Methods), RFdiffusion can scaffold enzyme active sites. An oxidoreductase example (EC1) is shown (PDB 1A4I); catalytic site (teal); RFdiffusion output (grey, model; colours, AF2 prediction); zoom of active site. AF2 versus design backbone r.m.s.d. 0.88 Å, AF2 versus design motif backbone r.m.s.d. 0.53 Å, AF2 versus design motif full-atom r.m.s.d. 1.05 Å, AF2 pAE 4.47. g, In silico success rates on active sites derived from EC1-5 (AF2 Motif r.m.s.d. versus native: backbone less than 1 Å, backbone and sidechain atoms less than 1.5 Å, r.m.s.d. AF2 versus design less than 2 Å, AF2 pAE less than 5).