Fig. 2
From: Phagocytosis is mediated by two-dimensional assemblies of the F-BAR protein GAS7
Crystal structures of GAS7 and the membrane-binding sites. a Ribbon diagrams of the crystal structure of the isolated GAS7 F-BAR domain and the GAS7-specific N-terminal region including FFL1 (red) and FFL2 (orange). Secondary structural elements are indicated. R326 on helix α2b contacts N177 on FFL1 as well as D207 on FFL2. The K208, K209, Q212, K370 and R374 residues are also indicated. b Crystal lattice of the isolated F-BAR domain. The asymmetric unit of the crystal contains two F-BAR dimers, which both form the FFO by the crystal packing. The N-surface, which is perpendicular to the dimer axes, is indicated with the illustration of FFL1 and FFL2. c The hypothetical position of the WW domain of GAS7b on the FFO. The two WW domains of the F-BAR dimer can be localized to one side of the FFO. d Superimposition of two FFOs composed of different F-BAR dimers in the asymmetric unit of the crystal (cyan and magenta). The contact sites between the F-BAR domains are indicated. e Surfaces of the FFO. (left) Electrostatic surface potentials of FFOs composed of three F-BAR dimers; blue and red indicate positive and negative charges, respectively. (Right) In the same orientation as in d. The point mutations with defects in the F-BAR domain assembly (blue), and those without defects (cyan) are shown according to Supplementary Fig. 5. FFL1 (red), FFL2 (orange), Q212 (cyan), K208/K209 (cyan) and K370/R374 (blue) are also indicated. f–l GAS7b and its ΔFFL1, K370E/R374E, Q212R, ΔFFL2, K209E, K208A/K209A and D207R mutants were examined for their binding to the liposomes by liposome co-sedimentation assays. The presence of proteins in the pellet (ppt) indicates membrane binding. sup: supernatant. The liposomes were of the bovine Folch fraction (g–i, k), the PC, PE and PS lipids at a ratio of 20, 20 and 60 (f, j, l), and the PC, PE, PS and PIP3 lipids at a ratio of 40, 40, 20 and 5 (f). m Cross-linking of GAS7b and the D207R mutant treated with the BS(PEG)5 (PEGylated bis(sulfosuccinimidyl)suberate) cross-linker, in the presence or absence of PS liposomes
