A Fence-like Coat for the Nuclear Pore Membrane Erik W. Debler, Yingli Ma, Hyuk-Soo Seo, Kuo-Chiang Hsia, Thomas R. Noriega, Günter Blobel, André Hoelz  Molecular Cell  Volume 32, Issue 6, Pages 815-826 (December 2008) DOI: 10.1016/j.molcel.2008.12.001 Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 1 Overview of the Seh1•Nup85 Structure (A) Schematic representation of the heptameric complex and the approximate localization of its seven nups (Lutzmann et al., 2002). (B) Domain structures of Seh1 and Nup85. For Seh1, the WD40 repeats (orange) and the numbering relative to yeast Seh1 are indicated. For Nup85, the unstructured N-terminal region (gray), the domain invasion motif (DIM) (purple), the α-helical solenoid domain (blue), and the C-terminal α-helical region (light pink) are indicated. The numbering is relative to yeast Nup85. The bars above the domain structures mark the crystallized fragments. (C) Ribbon representation of the Seh1•Nup85 hetero-octamer, showing Seh1 in yellow and orange and Nup85 in green and blue. A 90° rotated view is shown on the right. The three pseudo-2-fold axes (black ovals) and the overall dimensions are indicated. The Seh1•Nup85 hetero-octamer forms a slightly bent rod. (D) Schematic representation of the Seh1•Nup85 hetero-octamer. Magenta lines indicate interaction surfaces. Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 2 Detailed Structural Analysis of Nup85 and Seh1 (A) A ribbon representation of the Nup85 structure is shown in rainbow colors along the polypeptide chain from the N to the C terminus. The N-terminal domain invasion motif (DIM), the α-helical solenoid domain, and their secondary structure elements are indicated. (B) The structure of the Seh1•Nup85 heterodimer. The Nup85DIM (magenta), the Nup85 α-helical solenoid domain (blue), the Nup85 αQ-αR connector (red), the Seh1 β propeller (yellow), the disordered Seh1 5CD loop (gray dots), and the Seh1 2CD loop (orange) are indicated; a 90° rotated view is shown on the right. Dotted lines represent disordered regions. (C) Schematic representation of the Seh1•Nup85 interaction. The Seh1 2CD loop, the Nup85 αQ-αR connector, and the DIM region are highlighted in orange, red, and pink, respectively. (D) The β propeller domain of Seh1 in complex with the Nup85DIM. Seh1 is shown in yellow, and the six blades are indicated. The Nup85DIM contributes one strand to blade 6 and three strands to blade 7, completing the β propeller. (E) Schematic representation of the Seh1 β propeller and its interaction with the Nup85DIM. Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 3 Surface Properties of the Seh1•Nup85 Heterodimer (A) Surface rendition of the Seh1•Nup85 complex. The surface is colored according to the proteins (Seh1, yellow; Nup85, blue) and their participation in various interactions: with Seh1 of the adjacent heterodimer, orange; with Nup85 of the adjacent heterodimer, green; and with Seh1 of the adjacent heterotetramer, purple. (B) Nup85 is colored according to sequence conservation, from 40% similarity (yellow) to 100% identity (red). (C) Surface rendition of Nup85, colored according to the electrostatic potential, from red (−15 kBT/e) to blue (+15 kBT/e). Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 4 Interfaces in the Seh1•Nup85 Hetero-Octamer (A) A large surface area of ∼5100 Å2 is buried upon heterotetramer formation and extends over almost the entire length of a Seh1•Nup85 heterodimer. α helices that mediate heterotetramer formation are indicated. The Nup85 αQ-αR connector segments (red) are located at the center and contribute many contacts to the interface. (B) The Seh1•Seh1 dimerization interface located at the center of the Seh1•Nup85 hetero-octamer is significantly smaller than the interface in (A). The two interacting Seh1 β propeller domains and the two adjacent Nup85 molecules are colored yellow and blue, respectively. The locations of the pseudo-2-fold axes of symmetry that run through both interfaces are indicated (black ovals). Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 5 Flexibility of the Seh1•Nup85 Hetero-Octamer (A) The hetero-octamers of crystal forms 1 and 2 are related by an ∼35° hinge motion around the center of the hetero-octamer. On the right, a schematic of the two conformations is shown, where Seh1 and Nup85 are displayed as balls and cylinders, respectively. (B) Crystal form 3 harbors a heterododecamer in the asymmetric unit (small ribbon representation). The two interfaces between the heterotetramers are identical. In the upper two neighboring heterotetramers of crystal form 3 (boxed in the heterododecamer), one heterotetramer is rotated by ∼80° around its long axis with respect to crystal form 1. For clarity, a stripe of black lines marks the relative orientations of the heterotetramers. The alignment of the different structures was based on the lower heterotetramer of crystal form 1. Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions

Figure 6 Comparison of Seh1•Nup85 and Sec13•Nup145C (A) Comparison of the Seh1•Nup85 (left) and the Sec13•Nup145C (middle) heterodimers. Superimposition is based on the β propellers (right). (B) Comparison of the Seh1•Nup85 (first and second panel) and the Sec13•Nup145C hetero-octamers (third and fourth panel). Molecular Cell 2008 32, 815-826DOI: (10.1016/j.molcel.2008.12.001) Copyright © 2008 Elsevier Inc. Terms and Conditions