Fig. 3: ASH1L_PHD is a reader of H3K4me3.

a Schematic of ASH1L domain organization. b Overlays of ¹H,¹⁵N HSQC spectra of ASH1L_PHD collected in the absence (black) and presence of the indicated molar ratios of histone H3K4me3, H3K4me2, H3K4me1 or H3K4me0 (all aa 1-12 of H3) peptides. c Binding affinities of wildtype and mutated ASH1L_PHD for indicated histone H3 peptides, as measured by tryptophan fluorescence^a, MST^b or NMR^c. d Representative binding curve used to determine K_d values by tryptophan fluorescence. K_ds are calculated as mean values +/- S.D. from three independent experiments. e Representative binding curves used to determine K_d values by NMR. ¹H/¹⁵N Normalization equation is shown in NMR methods. K_ds are represented as mean values +/- S.D. The experiment was performed once. f A ribbon diagram of the crystal structure of ASH1L_PHD (wheat) in complex with H3K4me3 peptide (green sticks). Zinc ions are shown as gray spheres and hydrogen bonds are shown as dashed lines. The trimethylammonium binding cage residues of ASH11L_PHD are shown as sticks and colored orange. g Electrostatic surface potential of the ASH1L_PHD bound to H3K4me3 peptide (green sticks). Electrostatic potential ranging from positive;blue (+100 kT/e) to negative;red (−100 kT/e) generated with PyMol vacuum electrostatics. h Overlay of the crystal structures of the H3K4me3-bound and H3K4me2-bound ASH11L_PHD (also see Supplementary Fig. 5). The methyllysine binding cage surface is represented in mesh. The H3K4me3 and H3K4me2 peptides are green and yellow, respectively.