Individual Virus-like Particles Research Articles

Budding Pathway in the Templated Assembly of Viruslike Particles

A new pathway for the assembly of viral capsid protein around inorganic nanoparticle cores was observed by time-course light scattering and cryo-electron tomography. Gold nanoparticles with an average diameter of 11.3 nm have been used as a template for the assembly of Brome mosaic virus (BMV) capsid protein at different concentrations. At least at low protein concentrations the kinetic features of the scattering and extinction measurements are consistent with the initial rapid formation of large nanoparticle-protein clusters, which subsequently separate into individual viruslike particles (VLPs). The occurrence of multiparticle clusters at short times after mixing nanoparticles and proteins was confirmed by cryo-EM. Cryo-electron tomography of the multiparticle clusters yielded an average surface-to-surface interparticle distance of ∼7.5 nm, equivalent to ∼1.5 times the thickness of a protein shell. We propose a scenario in which VLP generation may take place through monomer exchange between aggregated particles with defect-ridden or incomplete shells, leading to the formation of stable icosahedral shells, which eventually bud off the aggregate. Together with results from previous works, the findings highlight the astonishing versatility of plant virus capsid protein assembly. This previously unknown mechanism for VLP formation has features that may have relevance for the crowded environment characterizing virus factories in the cell.

Antigenicity and immunogenicity of novel chimeric hepatitis B surface antigen particles with exposed hepatitis C virus epitopes.

The small envelope protein of hepatitis B virus (HBsAg-S) can self-assemble into highly organized virus like particles (VLPs) and induce an effective immune response. In this study, a restriction enzyme site was engineered into the cDNA of HBsAg-S at a position corresponding to the exposed site within the hydrophilic a determinant region (amino acid [aa] 127-128) to create a novel HBsAg vaccine vector allowing surface orientation of the inserted sequence. We inserted sequences of various lengths from hypervariable region 1 (HVR1) of the hepatitis C virus (HCV) E2 protein containing immunodominant epitopes and demonstrated secretion of the recombinant HBsAg VLPs from transfected mammalian cells. A number of different recombinant proteins were synthesized, and HBsAg VLPs containing inserts up to 36 aa were secreted with an efficiency similar to that of wild-type HBsAg. The HVR1 region exposed on the particles retained an antigenic structure similar to that recognized immunologically during natural infection. VLPs containing epitopes from either HCV-1a or -1b strains were produced that induced strain-specific antibody responses in immunized mice. Injection of a combination of these VLPs induced antibodies against both HVR1 epitopes that resulted in higher titers than were achieved by vaccination with the individual VLPs, suggesting a synergistic effect. This may lead to the development of recombinant particles which are able to induce a broad anti-HCV immune response against the HCV quasispecies or other quasispecies-like infectious agents.