NUCLEAR-PROTEIN IMPORT - RAN-GTP DISSOCIATES THE KARYOPHERIN-ALPHA-BETA HETERODIMER BY DISPLACING ALPHA FROM AN OVERLAPPING BINDING-SITE ONBETA

The alpha subunit of the karyopherin heterodimer functions in recognit ion of the protein import substrate and the beta subunit serves to doc k the trimeric complex to one of many sites on nuclear pore complex fi bers. The small GTPase Ran and the Ran interactive protein, p10, funct ion in the release of the docked complex. Repeated cycles of docking a nd release are thought to concentrate the transport substrate for subs equent diffusion into the nucleus. Ran-GTP dissociates the karyopherin heterodimer and forms a stoichiometric complex with Ran-GTP. Here we report the mapping of karyopherin beta's binding sites both for Ran-GT P and for karyopherin alpha. We discovered that karyopherin beta's bin ding site for Ran-GTP shows a striking sequence similarity to the cyto plasmic Ran-GTP binding protein, RanBP1. Moreover, rye found that Ran- GTP and karyopherin alpha bind to overlapping sites on karyopherin bet a. Having a higher affinity to the overlapping site, Ran-GTP displaces karyopherin alpha and binds to karyopherin beta. Competition for over lapping binding sites may be the mechanism by which GTP bound forms of other small GTPases function in corresponding dissociation-associatio n reactions. We also mapped Ran's binding site for karyopherin beta to a cluster of basic residues analogous to those previously shown to co nstitute karyopherin alpha's binding site to karyopherin beta.