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Updated: Jul 2, 2026

Comparing the Affinity of GTPase-binding Proteins using Competition Assays
Published on: October 8, 2015
Kap95p binding induces the switch loops of RanGDP to adopt the GTP-bound conformation: implications for nuclear
Jade K Forwood1, Thierry G Lonhienne, Mary Marfori
1School of Biomedical Sciences, Charles Sturt University, Wagga Wagga 2650, Australia. jforwood@csu.edu.au
Abstract:
The asymmetric distribution of the nucleotide-bound state of Ran across the nuclear envelope is crucial for determining the directionality of nuclear transport. In the nucleus, Ran is primarily in the guanosine 5'-triphosphate (GTP)-bound state, whereas in the cytoplasm, Ran is primarily guanosine 5'-diphosphate (GDP)-bound. Conformational changes within the Ran switch I and switch II loops are thought to modulate its affinity for importin-beta. Here, we show that RanGDP and importin-beta form a stable complex with a micromolar dissociation constant. This complex can be dissociated by importin-beta binding partners such as importin-alpha. Surprisingly, the crystal structure of the Kap95p-RanGDP complex shows that Kap95p induces the switch I and II regions of RanGDP to adopt a conformation that resembles that of the GTP-bound form. The structure of the complex provides insights into the structural basis for the gradation of affinities regulating nuclear protein transport.
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