SARS-CoV-2 virus is the causative agent of COVID-19. Here we demonstrate that non-infectious SARS-CoV-2 virus like particles (VLPs) can be assembled by co-expressing the viral proteins S, M and E in mammalian cells. The assembled SARS-CoV-2 VLPs possess S protein spikes on particle exterior, making them ideal for vaccine development. The particles range in shape from spherical to elongated with a characteristic size of 129 ± 32 nm. We further show that SARS-CoV-2 VLPs dried in ambient conditions can retain their structural integrity upon repeated scans with Atomic Force Microscopy up to a peak force of 1 nN.

Immature HIV virions harbor a lattice of Gag molecules with significant ordering in CA-NTD, CA-CTD and SP1 regions. This ordering plays a major role during HIV maturation. To test the condition in which the Gag lattice forms in vivo, we assembled virus like particles (VLPs) by expressing only HIV Gag in mammalian cells. Here we show that these VLPs incorporate a similar number of Gag molecules compared to immature HIV virions. However, within these VLPs, Gag molecules diffuse with a pseudo-diffusion rate of 10 nm²/s, this pseudo-diffusion is abrogated in the presence of melittin and is sensitive to mutations within the SP1 region. Using cryotomography, we show that unlike immature HIV virions, in the Gag lattice of VLPs the CA-CTD and SP1 regions are significantly less ordered. Our observations suggest that within immature HIV virions, other viral factors in addition to Gag, contribute to ordering in the CA-CTD and SP1 regions.

HIV is a lentivirus characterized by its cone shaped mature core. Visualization and structural examination of HIV requires the purification of virions to high concentrations. The yield and integrity of these virions are crucial for ensuring a uniform representation of all viral particles in subsequent analyses. In this study, we present a method for the purification of HIV virions which minimizes the forces applied to virions while maximizing the efficiency of collection. This method, which relies on virion sedimentation simulations, allows us to capture between 1000 and 5000 HIV virions released from individual HEK293 cells after transfection with the NL4.3 HIV backbone. We utilized this approach to investigate HIV core formation from several constructs: pNL4-3(RT:D₁₈₅A&D₁₈₆A) with an inactive reverse transcriptase, NL4.3(IN: V₁₆₅A&R₁₆₆A) with a type-II integrase mutation, and NL4.3(Ψ: Δ(105–278)&Δ(301–332)) featuring an edited Ψ packaging signal. Notably, virions from NL4.3(Ψ: Δ(105–278)&Δ(301–332)) displayed a mixed population, comprising immature virions, empty cores, and cores with detectable internal density. Conversely, virions derived from NL4.3(IN: V₁₆₅A&R₁₆₆A) exhibited a type II integrase mutant phenotype characterized by empty cores and RNP density localized around the cores, consistent with previous studies. In contrast, virions released from pNL4-3(RT:D₁₈₅A&D₁₈₆A) displayed mature cores containing detectable RNP density. We suggest that the sedimentation simulations developed in this study can facilitate the characterization of enveloped viruses.