Hussain, Hamdi (2018) Structural organisation of the human kinesin-12 Kif15. PhD thesis, University of Warwick.

PDF WRAP_Theses_Hussain_2018.pdf - Submitted Version Embargoed item. Restricted access to Repository staff only until 5 September 2020. Contact author directly, specifying your specific needs. - Requires a PDF viewer. Download (62Mb)

Request Changes to record.

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)
Subjects:	Q Science > QP Physiology
Library of Congress Subject Headings (LCSH):	Kinesin -- Structure, Oligomers, Proteins -- Crosslinking, Mass spectrometry
Official Date:	August 2018
Dates:	Date Event August 2018 Submitted
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Publication Status:	Unpublished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:	pdf
Extent:	xiii, 155 leaves : illustrations, charts
Language:	eng

Request changes or add full text files to a record

Repository staff actions (login required)

View Item