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Title: Integrative structural biology of viruses
Author: Duyvesteyn, Helen Mary Elizabeth
ISNI:       0000 0004 9351 1861
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2020
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Picornaviruses include significant infectious agents and are challenging structural targets owing to their pathogenicity. They share a common architecture, but subtle differences in their structure mediate tissue tropism, receptor binding and uncoating cues. This thesis focuses on members of the Enterovirus and Aphthovirus genera, and aims to contribute structural understanding of their lifecycle and the forces that underpin and modulate capsid stability to ultimately inspire novel therapeutics and vaccines. I take advantage of improvements in cryoEM which enable structural biologists to routinely solve complex structures at close to atomic resolution and resolve sample heterogeneity. EV-F3 is a non-pathogenic enterovirus. Presented crystallographic structures or EV-F3 highlight a potentially druggable site, and soaks with siallactose suggest a cellular receptor. Cryo-EM investigations show intermediate states, confirming that heating can cause transitions to altered particle states seen in other enteroviruses. Long-wavelength X-ray data indicate the presence of K+, but not Ca2+ ions within the capsid. I present structural characterisation of recombinant VLP vaccine candidates for poliovirus (PV) and foot-and-mouth disease virus (FMDV). For FMDV, I show a continuum of cryo-EM structures with a potential S-S bridge at the 2-fold interface and varied 2-fold structures. A pentamer subassembly is characterised and analysis of metal ion composition in FMDV VLPs gives no evidence for bound Ca2+ ions. Single-particle cryo-EM experiments of yeast-derived thermostabilised PV-1 and PV-3 VLPs show both C and D antigenic states to be present for PV-1 and demonstrate a predominance of D-antigenic states for PV-3. Finally, preliminary investigations into aphthovirus uncoating include attempts to express full length αvβ6 integrin receptor and a preliminary structure of FMDV bound to αvβ3 head-group, demonstrating VLP affinity for virus receptors. I demonstrate the potential of caged protons for time-resolved structural studies of FMDV uncoating and present a novel expanded particle structure of an aphthovirus.
Supervisor: Huiskonen, Juha ; Stuart, David ; Orville, Allen Sponsor: Wellcome Trust
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Structural biology ; Foot-and-mouth disease virus ; Transmission electron microscopy ; Enteroviruses ; Virus-like particles ; Aphthoviruses ; Poliovirus ; X-ray crystallography ; Picornaviruses ; Virology