PMC

Single-particle EM analysis of free ctCRM1. EM models of the compact (orange) as well as the extended conformation (blue) of free ctCRM1 are shown. The crystal structures of free ctCRM1 and CRM1 in complex with SPN1 and RanGTP are fitted to the envelope models of the EM structures. The position of the NES cleft is marked by a black arrowhead.

Crystal structure of free ctCRM1 (gray). The acidic loop (green), the C-terminal helix (red) and the NES cleft (blue) are highlighted. The HEAT repeats are numbered, and termini are labeled. (Lower) Rotated detail view of the interactions of residues from the C-terminal helix with the acidic loop and a patch of CRM1 formed by helices of H8 to H12. Hydrogen bonds and salt bridges are represented by dashed lines, and interacting residues are labeled.

MD simulations of WT and mutant free ctCRM1. Projections of WT simulations (A, cyan) and simulations with deleted C-terminal helix (B, red) onto the difference vector between the extended and compact structure constitute a measure of how much the protein changes into the compact conformation. (C) Projections onto the plane in the configurational space spanned by the extended, compact, and almost compact crystal structure show that, after deletion of the C-terminal helix, the system adopts the configuration of the almost compact structure (magenta square) rather than the compact conformation (orange square).

Model for cooperative CRM1 export complex assembly and disassembly showing its conformational variability and the important structural features in different states of the transport cycle. CRM1 is shown in the respective conformations and colored in gray with the acidic loop highlighted in green. The C-terminal helix of CRM1 is shown in red, and the NES binding cleft is represented by blue ovals. The PDB ID codes of the individual crystal structures used are indicated.

Comparison of CRM1 conformations in different crystal structures. Free ctCRM1 (A), CRM1–RanGTP–SPN1 (B), CRM1–SPN1 (C), and CRM1–RanGTP–RanBP1 (D) are shown. CRM1 is depicted as rainbow colored surface from N (blue) to C terminus (red), whereas the interacting proteins are shown as tube models (SPN1, purple; RanGTP, beige; RanBP1, red). The position of the NES cleft in structures lacking cargo is marked by a black arrowhead. The bars at the side of the individual structures indicate the dimension of the protein in the shown orientation.

Comparison of NES cleft conformations between free ctCRM1 (PDB ID code 4FGV; green) and CRM1 bound to SPN1 and RanGTP (PDB ID code 3GJX; red). The NES cleft is shown in cartoon mode (Left) with the centers of mass of the helices 11A and 12A represented by blue spheres and their distances indicated. A surface model (Center) illustrates the differences of the NES clefts between both structures. A superposition of the SPN1 NES from the ternary CRM1–RanGTP–SPN1 complex (Right) highlights the structural changes in the NES cleft, which are incompatible with NES binding in free CRM1.