Fig. 6: Biochemical reconstitution and structure NEDD1 WD β-propeller-Augmin complex.

A NEDD1 domain organization. N-terminal WD40 β-propeller domain (termed NEDD1-WD β-propeller). C-terminal helical domain (Note: Only WD β-propeller domain successfully purified). B Complex formation analysis: AUG1,2,3,4,5,6,7,8 binds NEDD1-WD β-propeller and while AUG1,3,4,5 shows no binding (See Fig. S12C–H). C SEC analysis of complex formation. Red: AUG1,2,3,4,5,6,7,8 alone. Blue: AUG1,2,3,4,5,6,7,8 with NEDD1-WD β-propeller. Green: NEDD1-WD alone. D Biochemical validation of SEC analysis in (C) by SDS-PAGE showing co-migration of NEDD1-WD with AUG1,2,3,4,5,6,7,8 and comparison with AUG1,2,3,4,5,6,7,8 alone (N = 2). E Cryo-EM 2D-class average comparison NEDD1-WD β-propeller binding to Augmin. Top: AUG1,2,3,4,5,6,7,8 V-junction-stem with β-barrel density. Bottom: AUG1,2,3,4,5,6,7,8-NEDD1-WD β-propeller complex. F Structural characterization of complex. Left: 12 Å Cryo-EM segmented reconstruction with fitted models. Right: Ribbon representation of complex. G 7.3-Å model showing β-barrel density bound to V-junction stem. Note the marked conformational change. H Comparative analysis of V-junction. Top: Segmented maps of complex vs. AUG1,2,3,4,5,6,7,8 alone. Bottom: Atomic models of NEDD1-WD and β-barrel regions. I NEDD1-WD-β propeller interface analysis. Right: Surface views of Augmin interaction site. Left: Conservation analysis across species.