Fig. 3: High-resolution NMR structure of SERF2 reveals that its disordered and dynamic N-terminal domain binds TERRA rG4.

a 20 best NMR ensemble model structures of SERF2. The average converged helical structure spanning residues 33–47 in SERF2 is shown as the helix in orange. b 1H-15N NMR assignment (red spectrum) of 100 µM human SERF2 mixed with 20 µM (yellow) and 50 µM (purple) TERRA23 rG4 at 4 °C. Spectral zooms on the right illustrate chemical shift changes in several N- and C-terminal residues at 2:1 protein: rG4 ratio. c The 1H-15N chemical shift perturbations (left y-axis) were calculated from (b) and plotted as color-shaded peaks for each assigned residue in SERF2 at increasing TERRA23 rG4 concentrations. The yellow and purple colors in the graph correlate to their corresponding spectrum shown in (b); unassigned peaks are denoted with an asterisk. The heteroNOE values of SERF2 in the absence of TERRA23 rG4 were plotted to highlight the dynamic regions in SERF2. d 15N relaxation rates R2/R1 demonstrating that a significant change in dynamics occurs for 200 µM SERF2 on its own (pink dots) and in the presence of 100 µM interacting TERRA23 rG4 (gray squares), as a function of residue number. Error bars represent the standard error from per-residue exponential fitting of peak intensities in NMRFAM-Sparky. e, f 2D analysis plots derived from analytical ultracentrifugation experiments for 4.7 µM TERRA12 rG4 without (e) or mixed with a 2-molar excess SERF2 (f). The partial concentration shown in color on the right y-axis represents the abundance of individual species in the sample solution. g, h Cartoon shows the top (g) and side-view (h) of SERF2 and TERRA rG4 complex. Note the quadrupole-like and planar interactions shown respectively as ellipses in (g) and vertical slabs in (h). Residues generating the quadrupole-like interactions and distorting TERRA rG4 structure (PDB ID: 2M18) are labeled, and hydrogen bonds are indicated with dashed lines. i EMSA gel-shift assay of 5 µM TERRA23 rG4 mixed with an equimolar amount of wild-type SERF2 and different lysine to alanine SERF2 mutants. Similar results were obtained in two independent repeated experiments.