MODELING THE EFFECTS OF GENOME PACKAGING ON NAKED VIRUS

Despite remarkable advances in X-ray diffraction and cryo-electron microscopy enabling high-resolution visualization of viral capsids and virus-like particles (VLPs), a critical knowledge gap persists, understanding how viral genomes organize within their native capsids remains experimentally challenging. Computational modeling emerges as a powerful complementary approach to bridge this gap, offering unprecedented insights into complete virion architecture including genome-capsid interactions.

In this work, we present a multiscale, multistep protocol that explicitly models viral genome topology by integrating experimental structural data to guide genome confinement within the capsid. As a proof-of-concept, we focused on Circoviruses, the smallest pathogens able to infect mammals. Specifically, Porcine Circovirus type 2 (PCV2) that inflicts considerable harm to pig farms worldwide. A PCV2 structure reported on the PDB reveals 60 copies of the capsid proteins (CPs) in icosahedral symmetry with one bound tetranucleotide with sequence Py-Py-Pu-Pu (Py, pyrimidine; Pu, purine). Using this data, we modeled a new capsid with N-termini and generated ten genome topological models, classified into three categories based on genome arrangement within the capsid, ordered, intermediate, and disordered. Genome was generated using lattice polymer models and the SPQR RNA structural modeling technique. The virion models were relaxed through coarse-grained molecular dynamics simulations (CGMD) using SIRAH and Amber22.

CGMD simulations indicated that all systems gradually reached stability after 1 μs. The simulation showed highly compact virions, leaving only a few thousand water molecules in vacant areas after 2 μs. Despite all models conforming to sizes determined by AFM and TEM (18.0 nm +/- 2 nm compared to 19.4 nm +/- 3 nm), CGMD simulations reveal two distinct populations characterized by varying protein-DNA interaction energies. Disordered organizations had more instability compared to ordered ones and displayed positive potential energy at different temperatures (37°C against 67°C for ordered genomes). These findings demonstrate that genome packaging geometry critically influences virion stability and assembly pathways, suggesting the existence of local organizational rules governing complete virus architecture.