Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.487918
Title: Structural studies of large protein complexes using 3D cryo-electron microscopy and single particle analysis
Author: Young, Anna Louise
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 2007
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Abstract:
The research presented in this thesis involved cryo-electron microscopy and single particle analysis of two different complexes: clathrin cage assemblies and a GroEL single ring mutant. Clathrin is a cytoplasmic protein with important roles, not only in clathrin-mediated endocytosis, but also many other vital processes within the cell. GroEL, an 800 kDa bacterial chaperonin, is a double ring complex that requires both rings in order to perform its function in facilitating protein folding in the cell. Understanding how a GroEL single ring mutant is able to function may help to reveal the essential components of the allosteric pathway of native GroEL. The aims of the clathrin project were to identify the location of Hsc70 in a clathrin-auxilin lattice in order to determine how clathrin may be disassembled in vivo. The resulting 3D structure revealed Hsc70 beneath the clathrin vertex and showed significant alterations in the path of the clathrin triskelion leg, providing important insights into the uncoating process. In negative stain, two maps of functional GroEL single rings were obtained in the presence and absence of ATP. When the atomic structure ofa GroEL subunit was fitted to the apo single ring, an excellent fit of all domains was obtained with the exception of helix M. This is highly significant since helix M mediates salt bridge switching between the apical and equatorial domains during the ATPase cycle. In the ATP-bound state multiple conformations were observed, which may represent pre-hydrolysis structural intermediates. Functional single rings were also successfully imaged in ice and preliminary structures obtained.
Supervisor: Not available Sponsor: Not available
Qualification Name: University of Warwick, 2007 Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.487918  DOI: Not available
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