Fig. 2: A SAXS-derived ab initio envelope for the heterotetrameric Cep63 (424–541)•Cep152 (1205–1295) complex and the cooperative formation of its hexadecameric form.

a A schematic and the SAXS envelope for the heterotetrameric Cep63 (424–541)•Cep152 (1205–1295) complex are shown with the embedded crystal structure of Cep63 (502–541)•Cep152 (1205–1250) (PDB: 6CSU; red and blue helices). Averaged χ² (the difference between actual and expected data) and normalized spatial discrepancy (NSD) values calculated from 32 independent DAMMIN reconstructions and Rc (cross-sectional radius) calculated from low q Guinier fit are shown. Various physical parameters of the complex calculated from its respective SAXS curve are provided (table, right). ^a,b,c,d, MWs determined from the SAXS data using four different methods (see Methods for details). Raw data are provided in Supplementary Fig. 2a, b. A 3D-rendered envelope is provided as Supplementary Movie 1. b SEC of Cep63 (424–541)•Cep152 (1205–1295) performed at the indicated pH. Black arrow, the heterotetrameric complex; blue arrows, higher-MW complexes. c Sedimentation velocity c(s) profiles for the Cep63 (424–541)•Cep152 (1205–1295) complex under the indicated conditions. Colored arrows, faster-sedimenting, higher-MW species detected under the respective concentrations. Samples were analyzed in 3 mm pathlength cells. d Sedimentation equilibrium absorbance data collected for the same complex in (c) at 7,000 (blue), 11,000 (red), and 20,000 (green) rpm at the loading concentrations indicated. Data collected at pH 5.5 were analyzed globally in terms of a tetramer–octamer–hexadecamer reversible self-association model. For clarity, only every third experimental data point is shown. Best fits are represented by a solid line through the experimental points, and the combined residuals are shown above the plots. The table (bottom) shows concentrations for the tetramer, octamer, and hexadecamer (in tetramer units) calculated based on the best-fit reversible self-association model. e Histograms showing the particle distribution of the same complex in (c) as a function of molecular weight. The y-axis (Incidence) denotes the number of particles. The yellow histogram with the red dotted line was generated by reanalyzing the data after discarding a third of the particles, as described previously⁷⁷.