Ran-GTP interacts strongly with importin-ÃŸ, and this interaction promotes the release of the importin-'-nuclear localization signal cargo from importin-ÃŸ. Ran-GDP also interacts with importin-ÃŸ, but this interaction is 4 orders of magnitude weaker than the Ran-GTPÂ·importin-ÃŸ interaction. Here we use the yeast complement of nuclear import proteins to show that the interaction between Ran-GDP and importin-ÃŸ promotes the dissociation of GDP from Ran. The release of GDP from the Ran-GDP-importin-ÃŸ complex stabilizes the complex, which cannot be dissociated by importin-'. Although Ran has a higher affinity for GDP compared with GTP, Ran in complex with importin-ÃŸ has a higher affinity for GTP. This feature is responsible for the generation of Ran-GTP from Ran-GDP by importin-ÃŸ. Ran-binding protein-1 (RanBP1) activates this reaction by forming a trimeric complex with Ran-GDP and importin-ÃŸ. Importin-' inhibits the GDP exchange reaction by sequestering importin-ÃŸ, whereas RanBP1 restores the GDP nucleotide exchange by importin-ÃŸ by forming a tetrameric complex with importin-ÃŸ, Ran, and importin-'. The exchange is also inhibited by nuclear-transport factor-2 (NTF2). We suggest a mechanism for nuclear import, additional to the established RCC1 (Ran-guanine exchange factor)-dependent pathway that incorporates these results.