Infectious pancreatic necrosis virus: Identification of a VP3-containing ribonucleoprotein core structure and evidence for O-linked glycosylation of the capsid protein VP2

Virions of infectious pancreatic necrosis virus (IPNV) were completely disi ntegrated upon dialysis against salt-free buffers. Direct visualization of such preparations by electron microscopy revealed 5.0- to 6.5-nm-thick enta ngled filaments. By using a specific colloidal gold immunolabeling techniqu e, these structures were shown to contain the viral protein VP3. Isolation by sucrose gradient centrifugation of the filaments, followed by serologica l analysis, demonstrated that the entire VP3 content of the virion was reco vered together with the radiolabeled genomic material forming the unique th readlike ribonucleoprotein complexes. In a sensitive blotting assay, the ou ter capsid component of IPNV, i.e., the major structural protein VP2, was s hown to specifically bind lectins recognizing sugar moieties of N-acetylgal actosamine, mannose, and fucose. Three established metabolic inhibitors of N-linked glycosylation did not prevent addition of sugar residues to virion s, and enzymatic deglycosylation of isolated virions using N-glycosidase fa iled to remove sugar residues of VP2 recognized by lectins. However, gentle alkaline beta elimination clearly reduced the ability of lectins to recogn ize VP2. These results suggest that the glycosylation of VP2 is of the O-li nked type when IPNV is propagated in RTG-2 cells.