Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.596489
Title: The role of FxFG-repeat nucleoporins in nuclear trafficking
Author: Bayliss, R. W. R.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2000
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Abstract:
Trafficking of macromolecules between the nucleus and cytoplasm through nuclear pore complexes (NPCs) is an essential process in eukaryotic cells. For example, proteins containing a nuclear localisation sequence (NLS) are imported into the nucleus, whereas proteins containing a nuclear export sequence (NES) are exported into the cytoplasm. Many nucleoporins, the proteins from which NPCs are constructed, contain FxFG-repeats, sequences based on a hydrophobic core (of sequence PhexPheGly, where x is usually a small residue) and a hydrophilic linker. Previous work has shown that carrier molecules, including importin-β and Nuclear Transport Factor 2 (NTF2), bind to FxFG-repeats, and addition of soluble FxFG-repeats inhibits nuclear trafficking processes. This Thesis characterises the interactions between FxFG nucleoporin repeats and both NTF2 and importin-β. Binding studies on a rationally-designed NTF2 mutant localise the FxFG binding site to near Trp7. Although I show that the interaction between FxFG repeats and NTF2 is weak, I also show that the interaction is essential for NTF2 to mediate the nuclear import of Ran in permeabilised cells. An X-ray crystal structure of importin-β (residues 1-442) in complex with FxFG repeats is presented which shows, for the first time, how FxFG-repeats interact with a carrier molecule. The crystal structure is used to produce mutants of full-length importin-β which are deficient in FxFG repeat binding, and yet retain binding to RanGTP and importin-α. These mutants are used to demonstrate that an interaction with FxFG repeats is essential for importin-β to mediate nuclear protein import of a substrate containing a classical NLS. A model for how RanGTP may release importin-β from FxFG repeats is proposed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.596489  DOI: Not available
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