Browsing by Author "Buckner, Cienna"

Coxsackievirus A9 (CVA9) is an enterovirus in the family Picornaviridae. While most infections are asymptomatic, it can cause diseases ranging from mild to serious, especially in neonates where it is known to cause aseptic meningitis and sepsis. There are currently no available vaccines or specific antiviral therapies against coxsackievirus infections. CVA9 has a 30 nm icosahedrally-symmetric capsid comprised of four viral proteins, VP1-4. These four proteins associate together to form a single protomer which is repeated 60 times to make the complete capsid. VP1 forms the five-fold vertex, at the base of which is located a hydrophobic pocket. This pocket is typically occupied by the pocket factor, palmitate. Upon internalization in an endosome, the pocket factor disassociates from the pocket, leading to pocket collapse and subsequently capsid expansion and loss of VP4 and the viral genome. Antiviral therapeutics against CVA9 have focused on stabilizing the hydrophobic pocket to ultimately prevent genome release, but there are currently no approved treatments. Ongoing research has expanded the search to the N-phenyl benzamides class of compounds, which have demonstrated antiviral activity against CVA9. An in-situ docking analysis proposed the hydrophobic pocket as one of several potential binding sites. Here, I used cryogenic electron microscopy and single particle processing to determine the binding site of one such compound, CL301, validating one proposed binding site and the antiviral mechanism of action. A dataset of 24,208 particles was reconstructed resulting in a 2.6 Å resolution electron density map. Atomic modelling showed that an averaged density contributed by both CL301 and palmitate occupied the VP1 hydrophobic pocket. There is not sufficient space for both molecules to occupy the pocket at once, so there is a mixed population, which could be individual virions having all 60 pockets occupied with one or the other molecule, or only a fraction of pockets per virion being occupied. As there is little conformational difference between the two, the statistical analysis used to remove heterogeneity in the dataset was not sensitive enough. In conclusion, CL301 is effective through capsid stabilization preventing expansion and release of the viral genome. Future work should concentrate on improving the affinity of this class of compounds.