Defining the mechanism by which glycoprotein D triggers HSV fusion and entry

Herpes simplex virus entry into cells requires four essential glycoproteins. Three of the glycoproteins, gB and gH/gL, function in the fusion of the cellular and viral membranes. The fourth glycoprotein, gD, binds to one of several cellular receptors. Receptor-gD binding is the first interaction that is essential for virus entry. However, what remained unclear is how gD-receptor binding initiated fusion by gB and gH/gL. In this dissertation I have identified a conformational change in the C-terminus of the gD ectodomain that is critical for HSV entry. The resolution of the full gD ectodomain, including the previously unsolved C-terminus, predicted an interaction between the core and C-terminus of the gD ectodomain. The receptor binding sites were obscured by the position of the C-terminus in the crystal structure and thus would need to be displaced to permit receptor binding. I tested this hypothesis by introducing a series of engineered disulfide bonds between residues of the C-terminus and residues of the core that were designed to limit the extent of the conformational change. Using this panel of mutants, I showed that the conformational change between the core and C-terminus was critical for triggering fusion and entry. These mutants broadly implicated a portion of gD, either on the core, the C-terminus or both as being critical for triggering fusion. The region responsible for triggering fusion was further defined by the identification of neutralizing antibodies that did not interfere with gD-receptor binding. Therefore, these antibodies were inhibiting an essential function of gD that was unrelated to receptor binding. The epitope for at least one of these antibodies mapped to the core of gD. Further, when the C-terminus and the core of gD interacted, this epitope was obscured. Together, my data suggest that the binding of gD to a receptor does indeed displace the C-terminus of the gD ectodomain. This leads to exposure of a “fusion triggering” domain, present on the gD core, that is obscured by the native conformation of gD. I suggest that this newly exposed triggering domain then interacts with and activates gB and/or gH/gL to fuse the viral and cellular membranes