Table 1.
Crystallographic data collection and refinement statistics for WHSC1L1 1054–1285 with PTD2 and SAM.
Fig 1.
Norleucine-containing peptides can inhibit WHSC1 and WHSC1L1 activity in vitro.
Representative peptide inhibitor biochemical dose-response curves for (A) WHSC1 941–1240 and (B) WHSC1L1 1054–1285. Error bars represent the standard deviation of three independent replicates. Resulting IC50 values are reported in Table 2.
Table 2.
Biochemical and biophysical peptide inhibitor potency values for WHSC1 941–1240 and WHSC1L1 1054–1285a.
Table 3.
Biochemical IC50 values for norleucine peptide inhibitors derived from the H4K44 and H3K36 sequencesa.
Table 4.
PTD2 biochemical IC50 values for a panel of HMT enzymes.
Fig 2.
Representative sensorgram for PTD2 binding to Avi-tagged WHSC1 941–1240 from single-cycle kinetic SPR measurements.
WHSC1 was immobilized on a streptavidin-coated chip and peptide inhibitor was co-injected with SAM utilizing a 3-fold, 5-point dilution series ending at a 20 μM top concentration. Data reported in Table 2 is presented as the standard deviation of three independent experiments.
Fig 3.
Isothermal titration calorimetry of PTD2 binding to WHSC1 941–1240.
Upper panel, calorimetric trace for ligand titration; lower panel, binding isotherm from calorimetric trace. WHSC1 concentration in the cell was 10 μM supplemented with 100 μM SAM. PTD2 concentration in the syringe was 100 μM.
Fig 4.
Representative SPR sensorgrams for PTD2 binding to Avi-tagged WHSC1 941–1240 in the absence or presence of SAM analogs.
WHSC1 was immobilized on a streptavidin-coated chip and peptide inhibitor was either injected in the absence of cofactor (left panel), co-injected with SAH (middle panel), or co-injected with SFG (right panel) utilizing a 2-fold, 10-point dilution series ending at a 100 μM top concentration.
Fig 5.
WHSC1L1 1054–1285 has a similar overall structure in relation to other NSD family proteins and can form a ternary complex with SAM and PTD2.
(A) Superposition of NSD family proteins (green/yellow = WHSC1L1-PTD2 (PDB code = 6CEN); cyan = WHSC1L1 (PDB code = 5UPD); magenta = WHSC1 (PDB code = 5LSU); purple = NSD1 (PDB code = 3OOI). All protein chains are shown as ribbons; SAM and PTD2 are depicted in stick representation. (B) Structure of WHSC1L1-PTD2-SAM ternary complex. Hydrogen bonds are indicated with dashed lines. (C) Superposition of WHSC1L1-PTD2 and SETD2-H3.3 K36M (grey; PDB code = 5JJY).