Studies of canine parvovirus capsid: Assembly, disassembly and antibody neutralization

The virus capsid functions to protect the viral genome during passage between cells and hosts, which demands the capsid be a stable structure. The capsid also functions to deliver and release the viral genome correctly during infection, which requires that the capsid be ready to disassemble. Viral capsids are one of the main antibody's targets for host protection. Canine parvovirus (CPV) is one of the simplest animal virus, and its structure has been determined to atomic resolution. This makes CPV a good model in which to study the processes of capsid assembly and disassembly, the interactions of the capsid with antibodies, and the mechanisms of antibody neutralization, all of which are the objectives of this study.; Assembly studies revealed that CPV empty capsids assembled rapidly followed by a slower formation of CPV full (DNA-containing) capsids. Trimers of VP2 were formed as unstable, transient intermediates during assembly. Capsid nuclear localization was related to assembly. Mutations examined in this study which prevented capsid assembly disrupted the intermediate formation and also resulted in the protein being distributed throughout the cell. VP2 protein was also expressed in insect cells using recombinant baculoviruses. Those baculovirus-expressed VP2 molecules assembled inefficiently, and the proteins and capsids accumulated in the cytoplasm. The VP2 from the unassembled VP2 molecules and from capsids recovered from the insect cells differed in isoelectric point.; In vitro disassembly of capsids by pH or urea treatment released dimers, trimers, and pentamers of capsid proteins from empty capsids, while only trimers and pentamers were released from full capsids. Treatments with milder conditions exposed the 3'-end of the viral genome and the N-terminus of VP1 protein while the capsids remained essentially intact. The release of 3'-end of the viral genome occurred independently from the exposure of the N-terminus of VP1 protein.; Two single-chain variable domain antibodies (scFv) were prepared from two neutralizing monoclonal antibodies (Mab). One of those was examined to define its interactions with the CPV capsid by mutagenesis analysis, and some of the direct contact residues of the Mab were confirmed. The mechanisms of neutralization of CPV infectivity by scFv or Mab were analyzed, and some of the neutralization occurred through virion aggregation, but the scFv or Mab also interfered with the virus intracellular infection process.