Davies, Sarah Margaret Anne

2018-05-14T10:12:17Z

2018-05-14T10:12:17Z

1998

Membrane fusion, the merging of two initially distinct membranes to form one common lipid bilayer, is a fundamental mechanism of life. It occurs many times each day within every eukaryotic cell as part of essential daily homeostatic processes, as well as between individual cells, such as sperm and egg during fertilisation. The fusion mechanism is, however, also crucial to the development of many diseases. All enveloped viruses, and indeed many other obligate intracellular parasites, must fuse their own surrounding lipid bilayer with the membrane of their host's target cell in order to gain cell entry and thus the ability to replicate. These infections produce disease states, and possibly even death, in the host species

Despite the clear importance of fusion, the precise molecular events that occur during this process are still not known. Fusion proteins of viruses have recently become popular tools for use in fusion studies. More specifically, several viruses have known fusion peptides, the sections of these proteins which confer their fusogenic activity. This thesis examines the structure and function of the putative fusion peptide of the retrovirus Feline leukaemia virus, (FeLV), using a variety of mainly biophysical techniques.

The structural effects of the FeLV fusion peptide on lipid polymorphism were studied. Using differential scanning calorimetry, ³¹P nuclear magnetic resonance and time-resolved X-ray diffraction this peptide was found to induce changes in lipid conformation and motion similar to those of known fusogens: it favoured the formation of non-bilayer lipid conformations which have a relatively large negative curvature, namely the inverted hexagonal phase and isotropic lipid states. Moreover, using X-ray diffraction, a new lipid phase was observed in the presence of the FeLV peptide

Neutron diffraction studies revealed a change in the packing of lipid molecules within a bilayer and also possible thinning ofthe bilayer, both ofwhich were induced by interaction with the FeLV fusion peptide.

Fusogenic activity for this putative viral fusion peptide was demonstrated, using fusion assays, which measured the merging of lipid membranes in the presence ofthe FeLV fusion peptide.

These findings are discussed in the light ofthe current concepts ofthe fusion mechanism. They add support to two currently favoured theories of fusion: precession by a fusion peptide as a means of inducing the initial destabilisation of a bilayer, and the formation ofhighly bent, high energy lipid intermediates, such as the 'modified stalk', in the multistep fusion pathway.

Circular dichroism was employed to determine the secondary structure ofthe FeLV fusion peptide under a variety of experimental conditions. This peptide was observed to flip readily between a-helical and p sheet conformations. This suggests that structural plasticity may be an important dynamic property offusion peptides. Possible relationships between peptide structure and function are discussed

http://hdl.handle.net/1842/29724

The University of Edinburgh

Annexe Thesis Digitisation Project 2018 Block 18

Already catalogued

Characterisation of the feline leukaemia virus fusion peptide: implications for the fusion mechanism

Thesis or Dissertation

Doctoral

PhD Doctor of Philosophy