Specificity matters. An all-by-all assay showed strong diagonal binding - each binder recognized only its cognate phosphopeptide, with negligible cross-reactivity. (6/8)

Crystal structures of CD3ε and EGFR complexes matched design models within ~2 Å. The intended phosphate-binding motifs were reproduced with atomic accuracy, validating the approach. (5/8)

We tested four sites: CD3ε (TCR), EGFR pY1068 & pY1173, INSR pY1361. In each case, RFD2-MI produced compact proteins that selectively bound the phosphorylated peptide, confirmed by yeast display & BLI with comparable affinities to native phosphorylation binding domains. (4/8)

We built RFD2-MI, an all-atom diffusion model for molecular interfaces based on RFD2. It co-designs binder + peptide and uses 1D conditioning features like hotspots, secondary structure, and solvent exposure to steer phosphate-pocket formation. (3/8)

Phosphorylation on tyrosines control key pathways in immunity, cancer, and metabolism. For the first time, we can now design proteins that specifically recognize individual phosphotyrosines, even in disordered regions. (1/8)

Preprint: www.biorxiv.org/content/10.1...

I would also be very interested in find some alternative letters, for example a B and P for a new ‘I ❤️ Biophysics’ sticker. Currently its hard to read for some people even with the Howarth and Chroma alphabet. SARS-CoV-2 NSP13 provided an amazing heart btw!