RFdiffusion
Upload your target structure, mark the residues you want gripped, and get back brand-new binders, each carrying a real AlphaFold2 confidence score against your target. A pilot run takes roughly 15 to 30 min.
What it is for
You have a target structure and a patch of its surface you want gripped, and you want brand-new binders of whatever shape works — the general-purpose starting point for de novo design. Every candidate comes back with a real AlphaFold2 confidence score against your own target. For antibody or nanobody formats use RFantibody or IgGM instead.
Generates brand-new protein backbones — 3D shapes with no sequence yet — by starting from noise and denoising toward something that fits the patch of your target you named. Ranomics then puts a sequence on each backbone with ProteinMPNN and refolds the pair with AlphaFold2 multimer, so every candidate that reaches you carries a confidence score measured against your own target rather than an idealised one. This is the general-purpose starting point for de novo binder design. RFdiffusion, Watson et al., Nature 2023.
When it fits:
- You want general-purpose de novo binders, with confidence numbers that came from your actual target.
- Your target is an ordinary protein epitope with no sugars or modified residues.
- You want the binder's length and shape left open rather than locked to an antibody scaffold.
Inputs
You will need:
- Target structure (
.pdb/.cif). - Chain ID of the target.
- At least one hotspot residue.
Each run uses a preset that sets the scale and scope:
- Your target, ~30 min start to first results
- Real RFdiffusion run against your uploaded target PDB with AF2 multimer validation. Pick 1 to 1000 candidates. Start with a small batch (4 designs, ~30 min) to confirm your target and hotspots, then scale to 100+ once the small batch looks reasonable. Results emailed when complete; A100-80GB.
Parameters you set on the form:
- Hotspot residues
- Comma-separated target-chain residues the binder should contact during diffusion.
- Binder length (min/max)
- How long the new binder should be. Each design draws its own length from this window. 55 to 65 is a compact single-domain binder and the right first try on a small, fairly flat patch; 90 to 150 suits a broad face or one sitting in a groove a short binder cannot reach across. Longer binders are harder to express at the bench, so lengthen only if nothing scores.
- Number of designs
- How many candidates to generate. Each passes ProteinMPNN sequence design and AF2 multimer scoring.
Typical runtime:
- pilot
- 15 to 30 min
How to read the results
Ranked candidates with ipTM, pLDDT, i_pAE, and downloadable PDBs. Aim for at least 1 in 5 with ipTM ≥ 0.65 on a tractable target before committing to a full pilot.
Where a tool reports them, the scores mean:
- ipTM
- Predicted confidence in the contact between two chains, on a 0 to 1 scale. Higher is better: > 0.75 strong; > 0.65 acceptable. Individual tools set their own pass bar a little either side of that — this guide's own results summary above states this tool's. On a multi-chain target the number may cover the target's own chain–chain interface as well as the binder's, so read the per-tool note on the results table before comparing designs on it.
- pLDDT
- Per-residue confidence in the predicted fold. Higher means the model is more sure of that part of the structure.
- i_pAE and pAE
- Predicted alignment error, at the interface (i_pAE) or across the whole structure (pAE). Lower is better.
References
Watson, J. L., Juergens, D., Bennett, N. R., et al. "De novo design of protein structure and function with RFdiffusion." Nature 620, 1089 to 1100 (2023). Composite pipeline: RFdiffusion backbones, ProteinMPNN sequences, and AF2 multimer validation.