EXPRESSION OF ALEUTIAN MINK DISEASE PARVOVIRUS CAPSID PROTEINS IN DEFINED SEGMENTS - LOCALIZATION OF IMMUNOREACTIVE SITES AND NEUTRALIZING EPITOPES TO SPECIFIC REGIONS

The capsid proteins of the ADV-G isolate of Aleutian mink disease parv ovirus (ADV) were expressed in 10 nonoverlapping segments as fusions w ith maltose-binding protein in pMAL-C2 (pVP1, pVP2a through pVP2i). Th e constructs were designed to capture the VP1 unique sequence and the portions analogous to the four variable surface loops of canine parvov irus (CPV) in individual fragments (pVP2b, pVP2d, pVP2e, and pVPZg, re spectively). The panel of fusion proteins was immunoblotted with sera from mink infected with ADV. Seropositive mink infected with either AD V-TR, ADV-Utah, or ADV-Pullman reacted preferentially against certain segments, regardless of mink genotype or virus inoculum. The most cons istently immunoreactive regions were pVP2g, pVP2e, and pVP2f, the segm ents that encompassed the analogs of CPV surface loops 3 and 4, The VP I unique region was also consistently immunoreactive. These findings i ndicated that infected mink recognize linear epitopes that localized t o certain regions of the capsid protein sequence. The segment containi ng the hypervariable region (pVP2d), corresponding to CPV loop 2, was also expressed from ADV-Utah, An anti-ADV-G monoclonal antibody and a rabbit anti-ADV-G capsid antibody reacted exclusively with the ADV-G p VP2d segment but not,vith the corresponding segment from ADV-Utah. Min k infected with ADV-TR or ADV-Utah also preferentially reacted with th e pVP2d sequence characteristic of that virus. These results suggested that the loop 2 region may contain a type-specific linear epitope and that the epitope may also be specifically recognized by infected mink . Heterologous antisera were prepared against the VP1 unique region an d the four segments capturing the variable surface loops of CPV. The a ntisera against the proteins containing loop 3 or loop 4, as well as t he anticapsid antibody, neutralized ADV-G infectivity in vitro and bou nd to capsids in immune electron microscopy. These results suggested t hat regions of the ADV capsid proteins corresponding to surface loops 3 and 4 of CPV contain linear epitopes that are located on the externa l surface of the ADV capsid. Furthermore, these linear epitopes contai n neutralizing determinants. Computer comparisons with the CPV crystal : structure suggest that these sequences may be adjacent to the threef old axis of symmetry of the viral particle.