Extended Data Fig. 2: Structural characterization of the RING1B-BMI1f interaction with RB-2.
From: Small-molecule inhibitors targeting Polycomb repressive complex 1 RING domain
a, Superposition of the crystal structures of RING1B-BMI1f (colored gray) and RING1B-BMI1 complex (PDB 2CKL; RING1B is colored in magenta and BMI1 is blue). Positions of N- and C-termini are shown. b, Crystal structure determined for RING1B-BMI1f cocrystalized with RB-2. Electron density is contoured at 1 σ (blue) and selected residues are labeled. c, Crystal structure of RING1B-BMI1f cocrystalized with RB-2 shown in surface representation. Opened site exposing hydrophobic side chains of L80 and L100 (both in pale green) is shown. Surrounding positively charged residues are colored in pale blue. d, Analysis of chemical shift perturbations determined for 120 μM DCN RING1B-BMI1f upon binding with 500 μM RB-2. ΔHN has been calculated as \(\sqrt {({\Delta}\delta _{HN}^2 + 0.1 \ast {\Delta}\delta _N^2)}\) in ppm (top) and ΔCO is a difference in CO chemical shifts in ppm (bottom). e, Strips from 3D 1H-13C HSQC-NOESY spectra for 290 μM ILV 2H,13C,15N RING1B-BMI1f (black) and 120 μM ILV 2H,13C,15N RING1B-BMI1f with 500 μM RB-2 (blue). Assignment indicates NOEs between RING1B-BMI1f protein and RB-2). f, Labeling of the RB-2 protons. g, 20 lowest energy conformers of RING1B-BMI1f with bound RB-2. RING1B residues are in pale green and BMI1 are in gray. RB-2 is shown with magenta carbons. h, Binding of RB-2 to the wild-type RING1B-BMI1 and RING1B(L94A)-BMI1f point mutant. 1H-15N HSQC spectra for 60 μM 15N RING1B-BMI1f or 60 μM 15N RING1B(L94A)-BMI1f (shown in red) are titrated with 100 μM RB-2 (shown in blue).
