Oral Presentation The 47th Lorne Conference on Protein Structure and Function 2022

Viral metamorphoses: using cryo-EM to understand the maturation of flaviviruses (#5)

Joshua M Hardy 1 , Natalee D Newton 2 , Naphak Modhiran 2 , Connor AP Scott 2 , Laura J Vet 2 , Jessica J Harrison 2 , Hariprasad Venugopal 3 , Jessica Balk 4 , Renee J Traves 2 , Leon E Hugo 5 , Alberto A Amarilla 2 , Summa Bibby 2 , Paul R Young 2 , Roy A Hall 2 , Jody Hobson-Peters 2 , Daniel Watterson 2 , Fasseli Coulibaly 4
  1. Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia
  2. School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, QLD, Australia
  3. Ramaciotti Centre for Cryo Electron Microscopy, Monash University, Clayton, VIC, Australia
  4. Biomedicine Discovery Institute and Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia
  5. QIMR Berghofer Medical Research Institute, Herston, QLD, Australia

Flaviviruses are notorious for their ability to cause seasonal and sporadic epidemics of unpredictable scale and severity. Vaccines are limited and no targeted antivirals have been clinically approved. To move forward, deeper insights into flavivirus biology are required.

Flaviviruses initially assemble as immature particles which undergo conformational changes and furin cleavage in the Golgi to metamorphosise into their infectious mature form. We discovered that the insect-specific Binjari flavivirus (BinJV) produces mainly immature virions and we resolved the structure of the immature virion to 3.9 Å using cryo-electron microscopy (cryo-EM)1. This resolution allowed the first complete model of the immature spike (E+prM proteins) and revealed that maturation requires conformational changes to expose the furin cleavage site. Unexpectedly, our structures show that the E spike is supported by a central pillar of prM rather than being pulled by a peripheral prM as in previous models. The revised architecture defines a ‘collapse’ model of maturation likely to be conserved across flaviviruses.

To study the mature flavivirus virion, we used our chimeric flavivirus platform which enables production of virions with an outer shell of E and M proteins from pathogenic viruses around a BinJV core2. Our chimeric viruses of dengue serotype 2, were highly homogenous and enabled us to achieve a record resolution for cryo-EM of an enveloped virus – 2.5 Å. Reaching this level of detail allowed us to visualise two lipid-like molecules bound to the E protein. Sequence analysis of the binding sites indicated a high degree of conservation across medically-relevant flaviviruses and mutation of interacting residues abolished infectivity.

We hypothesise that these pocket factors have a role in stabilising the virus particle in the late stages of viral maturation. Together, these structures provide alternative avenues to target the stem regions of E and prM for the development of improved vaccines and new therapeutics.

  1. Newton, N. D., Hardy, J. M., Modhiran, N., Hugo, L. E., Amarilla, A. A., Bibby, S., Venugopal, H., Harrison, J. J., Traves, R. J., Hall, R. A., Hobson-Peters, J., Coulibaly, F. & Watterson, D. (2021) The structure of an infectious immature flavivirus redefines viral architecture and maturation. Sci Adv, 7(20).
  2. Hardy, J. M., Newton, N. D., Modhiran, N., Scott, C. A. P., Venugopal, H., Vet, L. J., Young, P. R., Hall, R. A., Hobson-Peters, J., Coulibaly, F. & Watterson, D. (2021) A unified route for flavivirus structures uncovers essential pocket factors conserved across pathogenic viruses. Nat Commun, 12(1), 3266.