Fig. 7: Oligomeric state and structural organization of EDC4 C-terminal coiled-coil.

a, b Mass photometry histograms for purified Ce EDC4 residues 520–843 reveal a single species at ~315 kDa at 25 nM (a), consistent with a tetramer, and successive, equally spaced peaks beginning at ~303 kDa when the concentration is increased to 300 nM (b), indicating higher-order oligomers that grow in multiples of the tetrameric unit. Representative traces from three independent data sets are shown. c Residues 520–843 form an extended parallel four-helix bundle. The proximal stalk (rainbow-colored from green to yellow) is continuous with the distal α-helical bundle (magenta and cyan). The crystallized segment analyzed in panels (d–f) is bracketed (520–552). d Top, the 33-residue peptide (520–552) with residues resolved in the crystal structure colored blue; the canonical coiled-coil heptad positions (a–g) are annotated below. Bottom, helical-wheel diagrams illustrate the packing of hydrophobic a/d residues (gray) within a parallel tetramer. e Two orthogonal views show the crystal structure of the segment of the parallel four-helix bundle; a/d layer residues are depicted as sticks. f Superposition of the crystal structure (blue) onto the corresponding region of the AlphaFold-Multimer model (rainbow) demonstrates near-perfect register and side-chain packing. g Representative micrograph of negatively stained MBP-tagged Ce EDC4(520–843) with boxed inset (arrows mark individual particles); scale bar, 50 nm. The experiment was repeated independently three times with similar results. h Selected 2D class averages (particle counts and estimated resolutions indicated; scale bar, 51 nm) reveal elongated rod-shaped molecules terminating in a globular MBP density, consistent with a long coiled coil capped by the distal bundle.