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Structural organisation of the human kinesin-12 Kif15
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Hussain, Hamdi (2018) Structural organisation of the human kinesin-12 Kif15. PhD thesis, University of Warwick.
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Official URL: http://webcat.warwick.ac.uk/record=b3253832~S15
Abstract
Kinesin-12, Kif15 is a molecular motor involved in bipolar spindle assembly. Kif15 function is regulated through autoinhibition of its C-terminal tail and binding to the microtubule-associated protein Tpx2. Previous studies have reported Kif15 to function as a tetramer as well as a dimer. In this study, a cross-linking mass spectrometry (XL-MS) protocol and analysis workflow was developed to study the structural organisation of Kif15. Using XL-MS studies, it was found that Kif15 adopts a parallel tetramer conformation, which is autoinhibited by its C-terminal leucine zipper. Next, we show that this autoinhibited conformation is stabilised by the binding partner Tpx2. We also show that there is a shift in the binding interface between Kif15 and Tpx2 when microtubules are present and absent. In the presence of microtubules, Tpx2 mainly binds to the leucine-zipper of the Kif15 motor, whereas in the absence of the microtubules, this binding is exclusively localised to the fourth coiled-coil. We also reveal that Tpx2 adopts a dimeric conformation at physiological ionic strength. Finally, to understand the function of Kif15 in-vivo, we have developed putative Kif15 knock-out cell lines and developed a cross-linking protocol to cross-link Kif15 in cells.
| Item Type: | Thesis or Dissertation (PhD) | ||||
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| Subjects: | Q Science > QP Physiology | ||||
| Library of Congress Subject Headings (LCSH): | Kinesin -- Structure, Oligomers, Proteins -- Crosslinking, Mass spectrometry | ||||
| Official Date: | August 2018 | ||||
| Dates: |
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| Institution: | University of Warwick | ||||
| Theses Department: | Molecular Organisation and Assembly in Cells | ||||
| Thesis Type: | PhD | ||||
| Publication Status: | Unpublished | ||||
| Supervisor(s)/Advisor: | McAinsh, Andrew D. ; Jones, Alexandra M. | ||||
| Sponsors: | University of Warwick. Molecular Organisation and Assembly in Cells ; Engineering and Physical Sciences Research Council | ||||
| Format of File: | |||||
| Extent: | xiii, 155 leaves : illustrations, charts | ||||
| Language: | eng |
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