The first immunoglobulin-like domain of HveC is sufficient to bind herpes simplex virus gD with full affinity, while the third domain is involved in oligomerization of HveC

The human herpesvirus entry mediator C (HVeC/PRR1) is a member of the immun oglobulin family used as a cellular receptor by the alphaherpesviruses herp es simplex virus (HSV), pseudorabies virus, and bovine herpesvirus type 1. We previously demonstrated direct binding of the purified HveC ectodomain t o purified HSV type 1 (HSV-1) and HSV-2 glycoprotein D (gD). Here, using a baculovirus expression system, we constructed and purified truncated forms of the receptor containing one [HveC(143t)], two [HveC(245t)], or all three immunoglobulin-like domains [HveC(346t)] of the extracellular region. All three constructs were equally able to compete with HveC(346t) for go bindin g. The variable domain bound to virions and blocked HSV infection as well a s HveC(346t). Thus, all of the binding to the receptor occurs within the fi rst immunoglobulin-like domain, or V-domain, of HveC. These data confirm an d extend those of Cocchi ct al. (F. Cocchi, M. Lopez, L. Menotti, M. Aoubal a, P. Dubreuil, and G. Campadelli-Fiume, Proc. Natl. Acad. Sci. USA 95:1570 0, 1998). Using biosensor analysis, we measured the affinity of binding of go from HSV strains KOS and rid1 to two forms of HveC. Soluble gDs from the KOS strain of HSV-I had the same affinity for HveC(346t) and HveC(143t). T he mutant gD(rid1t) had an increased affinity for HveC(346t) and HveC(143t) due to a faster rate of complex formation. Interestingly, we found that Hv eC(346t) was a tetramer in solution, whereas HveC(143t) and HveC(245t) form ed dimers, suggesting a role for the third immunoglobulin-like domain of Hv eC in oligomerization. In addition, the stoichiometry between gD and HveC a ppeared to be influenced by the level of HveC oligomerization.