Individual binding pockets of importin-β for FG-nucleoporins have different binding properties and different sensitivities to RanGTP
- 21 October 2008
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 105 (42), 16101-16106
- https://doi.org/10.1073/pnas.0802647105
Abstract
Importin-beta mediates protein transport across the nuclear envelope through the nuclear pore complex (NPC) by interacting with components of the NPC, called nucleoporins, and a small G protein, Ran. Although there is accumulated knowledge on the specific interaction between importin-beta and the Phe-Gly (FG) motif in the nucleoporins as well as the effect of RanGTP on this interaction, the molecular mechanism by which importin-beta shuttles across the nuclear envelope through the NPC is unknown. In this study, we focused on four binding pockets of importin-beta for the FG motifs and characterized the interaction using a single-molecule force-measurement technique with atomic-force microscopy. The results from a series of importin-beta mutants containing amino acid substitutions within the FG-binding pockets demonstrate that the individual FG-binding pockets have different affinities to FG-Nups (Nup62 and Nup153) and different sensitivities to RanGTP; the binding of RanGTP to the amino-terminal domain of importin-beta induces the conformational change of the entire molecule and reduces the affinity of some of the pockets but not others. These heterogeneous characteristics of the multiple FG-binding pockets may play an important role in the behavior of importin-beta within the NPC. Single-molecule force measurement using the entire molecule of an NPC from a Xenopus oocyte also implies that the reduction of the affinity by RanGTP really occurs at the nucleoplasmic side of the entire NPC.Keywords
This publication has 40 references indexed in Scilit:
- Natively Unfolded Nucleoporins Gate Protein Diffusion across the Nuclear Pore ComplexCell, 2007
- Insights into E3 ligase activity revealed by a SUMO–RanGAP1–Ubc9–Nup358 complexNature, 2005
- Importin β contains a COOH-terminal nucleoporin binding region important for nuclear transportThe Journal of cell biology, 2003
- Regulating Access to the GenomeCell, 2003
- GLFG and FxFG Nucleoporins Bind to Overlapping Sites on Importin-βJournal of Biological Chemistry, 2002
- Proteomic analysis of the mammalian nuclear pore complexThe Journal of cell biology, 2002
- Visualization of a Ran-GTP Gradient in Interphase and Mitotic Xenopus Egg ExtractsScience, 2002
- Proteomic Analysis of Nucleoporin Interacting ProteinsJournal of Biological Chemistry, 2001
- Structural Basis for the Interaction between FxFG Nucleoporin Repeats and Importin-β in Nuclear TraffickingCell, 2000
- Importin beta , transportin, RanBP5 and RanBP7 mediate nuclear import of ribosomal proteins in mammalian cellsThe EMBO Journal, 1998