Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.533597
Title: Structural analysis of adenovirus bound to blood coagulation factors that influence viral tropism
Author: Lauder, Rebecca Pink
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 2011
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Abstract:
Adenoviruses are currently the most commonly used vectors for clinical gene therapy trials. Of these, Ad5 is the most commonly used serotype. Upon intravenous delivery, the vectors are sequestered in the liver which reduces their efficacy. While the coxsackievirus and adenovirus receptor is responsible for in vitro cell entry, this pathway is not used in vivo. Blood coagulation factors have been implicated in mediating in vivo hepatic transduction, and it is therefore important to characterise the interaction between these and Ad5 in order to permit development of more efficacious and safer viral vectors. Ad35 is a rare human pathogen and presents less pre-existing immunity than Ad5 and other common serotypes. It has potential as a gene therapy vector as this may reduce side effects and toxicity linked to pre-existing immunity. Ad35 uses CD46 as a receptor instead of CAR which allows it to infect hepatocytes, however blood coagulation factor X has also been implicated in in vivo hepatic transduction. This interaction must therefore also be characterised. We used low dose cryo electron microscopy to collect images of Ad5, Ad5 bound to FX and to FIX, Ad35 and Ad35 bound to FX. These were used to generate three dimensional icosahedral reconstructions of Ad5 at 27Å, Ad5 bound to FX at 26Å and 14Å, Ad5 bound to FIX at 21Å, Ad35 at 35Å and Ad35 bound to FX at 30Å. High resolution structural data were fitted to the Ad5 and Ad5-FX reconstructions in order to model the interaction and identify the binding sites. The structural data presented show that FX binds to Ad5 hexon with a stoichiometry of 1/3. Fitting experiments showed that the Gla domain of FX corresponds with the density in the centre of the hexon trimer, with arms corresponding to the EGF domains extending from this and terminating in the globular serine protease domain. Furthermore FX binds to Ad35 hexon in a similar manner. The data also shows that FIX does not bind to Ad5 hexon, however we were unable to confirm it if instead binds to the fiber as suggested by biochemical studies, due to limitations in the reconstruction technique used.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.533597  DOI: Not available
Keywords: QR355 Virology
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