Quantitative Structural Analysis of Importin-β Flexibility: Paradigm for Solenoid Protein Structures

Abstract

The structure of solenoid proteins facilitates a higher degree of flexibility than most folded proteins. In importin-β, a nuclear import factor built from 19 tandem HEAT repeats, flexibility plays a crucial role in allowing interactions with a range of different partners. We present a comprehensive analysis of importin-β flexibility based on a number of different approaches. We determined the crystal structure of unliganded Saccharomyces cerevisiae importin-β (Kap95) to allow a quantitative comparison with importin-β bound to different partners. Complementary mutagenesis, small angle X-ray scattering and molecular dynamics studies suggest that the protein samples several conformations in solution. The analyses suggest the flexibility of the solenoid is generated by cumulative small movements along its length. Importin-β illustrates how solenoid proteins can orchestrate protein interactions in many cellular pathways.

HEAT repeats – versatile arrays of amphiphilic helices working in crowded environments?

“…The overall conformation of the solenoid (i.e. its diameter, curvature and pitch) varies from protein to protein, and is also affected by the interactions formed with their binding partners (Conti et al, 2006;Lee et al, 2000;Forwood et al, 2010). For example, structural comparison of the karyopherin importin β with and without the cargo has revealed substantial differences in the curvature of the solenoid (Cingolani et al, 1999(Cingolani et al, , 2000.…”

“…2). The structural flexibility of HEAT repeats has been directly characterized by spectroscopic approaches (Tsytlonok et al, 2013) as well as by small angle X-ray scattering (Forwood et al, 2010). Molecular dynamics simulations have also demonstrated that, when external forces are applied at the ends of the molecule, the HEAT repeats exhibit unique elastic properties similar to a Hookean spring, whereby the extension is proportional to the tension applied (Grinthal et al, 2010;Kappel et al, 2010).…”

Structure‐based optimization of designed Armadillo‐repeat proteins

“…2(a) Q eliminate surface exposed hydrophobic residues on the accessible interface, and mutation D 23 P introduces a helixbreaking residue at the C-terminus of helix H2.…”

Nucleoporin Nup50 Stabilizes Closed Conformation of Armadillo repeat 10 in Importin α5

“…The ARM core generates a cargo-binding surface ideal to function as a static NLS-binding groove (46). This is very different from the HEAT-repeated architecture of ␤-karyopherins that form superhelical solenoids of remarkable structural flexibility (7,8,44). The results presented in this paper, together with previous structural (18) and biochemical (20) reports on importin ␣5 (Fig.…”