The major neutralizing antigenic site on herpes simplex virus glycoproteinD overlaps a receptor-binding domain

Herpes simplex virus (HSV) entry is dependent on the interaction of virion glycoprotein D (gD) with one of several cellular receptors. We previously s hell-ed that go binds specifically to two structurally dissimilar receptors , HveA and HveC. We have continued our studies by using (i) a panel of bacu lovirus-produced go molecules with various C-terminal truncations and (ii) a series of go mutants with nonoverlapping 3-amino-acid deletions between r esidues 222 and 254. Binding of the potent neutralizing monoclonal antibody (MAb) DL11 (group Ib) was unaffected in forms of gD containing residues 1 to 250 but was greatly diminished in molecules truncated at residue 240 or 234. Both receptor binding and blocking of HSV infection were also affected by these C-terminal truncations, gD-1(234t) bound weakly to both HveA and HveC as determined by enzyme-linked immunosorbent assay (ELISA) and failed to block infection. Interestingly, gD-1(240t) bound well to both receptors but blocked infection poorly, indicating that receptor binding as measured by ELISA is not the only go function required for blocking. Optical biosens or studies showed that while gD-1(240t) bound HveC with an affinity similar to that of gD-1(306t), the rates of complex formation and dissociation wer e significantly faster than for gD-1(306t). Complementation analysis showed that any 3-amino-acid deletion between residues 222 and 251 of go resulted in a nonfunctional protein. Among this set of proteins, three had lost DL1 1 reactivity (those with deletions between residues 222 and 230). One of th ese proteins (deletion 222-224) was expressed as a soluble form in the bacu lovirus system. This protein did not react with DL11, bound to both HveA an d HveC poorly as shown by ELISA and failed to block HSV infection. Since th is protein was bound by several other MAbs that recognize discontinuous epi topes, we conclude that residues 222 to 224 are critical for go function. W e propose that the potent virus-neutralizing activity of DL11 (and other gr oup Ib MAbs) likely reflects an overlap between its epitope and a receptor- binding domain of gD.