Abstract

We describe the crystal structure of a complex between importin-β residues 1–442 (Ib442) and five FxFG nucleoporin repeats from Nsp1p. Nucleoporin FxFG cores bind on the convex face of Ib442 to a primary site between the A helices of HEAT repeats 5 and 6, and to a secondary site between HEAT repeats 6 and 7. Mutations at importin-β Ile178 in the primary FxFG binding site reduce both binding and nuclear protein import, providing direct evidence for the functional significance of the importin-β–FxFG interaction. The FxFG binding sites on importin-β do not overlap with the RanGTP binding site. Instead, RanGTP may release importin-β from FxFG nucleoporins by generating a conformational change that alters the structure of the FxFG binding site.