The nuclear transport factor karyopherin β binds stoichiometrically to Ran-GTP and inhibits the Ran GTPase activating protein

Abstract

The heterodimeric karyopherin functions in targeting a nuclear localization sequence (NLS)-containing protein to the nuclear pore complex followed by Ran-GTP and p10-mediated translocation of the NLS protein into the nucleoplasm. It was shown recently that Ran-GTP dissociated the karyopherin heterodimer and, in doing so, associated with karyopherin β (Rexach, M., and Blobel, G. (1995) Cell 83, 683-692). We show here, using all recombinant yeast proteins expressed in Escherichia coli, that karyopherin β binds to Ran-GTP and inhibits GTP hydrolysis stimulated by RanGAP (the Ran- specific GTPase activating protein). Inhibition of RanGAP-stimulated GTP hydrolysis by karyopherin β was dependent on karyopherin β concentration relative to Ran-GTP. Complete inhibition of RanGAP was observed at karyopherin β concentrations that were equimolar to Ran-GTP. In gel filtration experiments, we found Ran-GTP and karyopherin β to form a stoichiometric complex. Ran-GDP bound only weakly to karyopherin β. We propose that stoichiometric complex formation between karyopherin β and Ran- GTP renders Ran-GTP inaccessible to RanGAP.