ICS Award
Virus capsid nanoparticle assembly reactions follow a narrow path through a complex reaction landscape

For many viruses, capsids assemble to protect genetic material and dissociate to release their cargo. To understand these contradictory properties, we analyzed capsid assembly for Hepatitis B virus; an endemic pathogen with an icosahedral, 120-homodimer capsid. We used solution X-ray scattering to examine trapped and equilibrated assembly reactions. To fit experimental results, we generated a library of unique intermediates, selected by umbrella sampling of Monte Carlo simulations. The number of possible capsid intermediates is immense, ∼10³⁰, yet assembly reactions are rapid and completed with high fidelity. If the huge number of possible intermediates were actually present, maximum entropy analysis shows that assembly reactions would be blocked by an entropic barrier, resulting in incomplete nanoparticles. When an energetic term was applied to select the stable species that dominated the reaction mixture, we found that only a few hundred intermediates, mapping out a narrow path through the immense reaction landscape. The grand canonical free energy landscape for assembly, calibrated by our experimental results, suggest that there is a narrow range of energies that supports on-path assembly. If association energy is too weak or too strong progressively more intermediates will be entropically blocked, spilling into paths leading to dissociation or trapped incomplete nanoparticles, respectively.

We then followed the assembly process at different conditions using time-resolved X-ray scattering. We found that within the successful assembly window the reaction follows a two state model where no intermediates were accumulated along the reaction assembly-path. At the strong interaction limit the assembly timescale was much faster where fast depletion in subunits concentration was observed along with accumulation of small intermediates complexes. The results show that when the interaction strength is slightly above an optimal value kinetic traps may hinder the formation of the complete T=4 capsid. These results are relevant to many viruses, provide a basis for simplifying assembly models and identifying new targets for antiviral intervention.