| Popis: |
Enveloped viruses employ membrane fusion during cell penetration in order to deliver their genetic material across the cell boundary. Large conformational changes in the proteins embedded in the viral membrane play a fundamental role in the membrane fusion process. Despite the tremendously wide variety of membrane containing viruses, it appears they all contain membrane fusion protein machinery with a remarkably conserved mechanism of action. The purpose of the research in this dissertation was to experimentally observe real time dynamic membrane fusion using fluorescence microscopy. An in vitro assay fusing viral membranes to supported lipid bilayers, triggered by the exposure to low pH and visualized with fluorescent dye molecules, was developed to observe viral membrane fusion. Studies of influenza and Sindbis virus were conducted with this assay. The intermediate structures of membrane fusion are too small to be spatially resolved with an optical microscope due to the diffraction limit, however fluorescence dequenching of dye molecules inserted into the viral membrane/lipid membrane system can provide information about the dynamics of membrane fusion as well as providing insight into the sequence of conformational changes of the lipid bilayer. Effects of adding the dye labels to the viruses are also investigated. |
