Cryo-EM Tomographic Analysis of a Universal Influenza Virus Vaccine Candidate

Abstract

Influenza viruses cause hundreds of millions of infections every year, and vaccines currently in use need to be reformulated annually due to the rapid mutation potential of these viruses. A universal influenza vaccine would alleviate the need for the yearly vaccine. Chimeric hemagglutinin (cHA) proteins are currently in development as potential universal influenza virus vaccines based on the idea that when the immune system is sequentially exposed to HA proteins that express the same stalk, but different head domains, the resulting antibody response will be directed toward the conserved stalk domain instead of the rapidly mutating head domain. This vaccination strategy increases the body's production of broadly neutralizing antibodies. Due to the conformational nature of stalk epitopes, it is necessary to ensure that these epitopes remain correctly folded in the chimeric proteins, yet no prior structural information about cHA proteins exists. We used cryo-electron tomography to determine structures of a virus-bound cHA, cH5/1, comprising an H1 stalk and an H5 head domain. Compared to native H1 or H5 HA, the cHA shows a more open head domain with clear separation between HA monomers. The cHA structure also shows a dramatic rotation between the stalk and head domains compared to the native structures. Viruses expressing cH5/1 HA also had a significantly higher density of HA on the viral surface than those expressing native HA. Despite these dramatic differences, cH5/1 retains its antibody-binding function with both head and stalk-targeting antibodies, suggesting a surprising plasticity in HA protein structure that may benefit future vaccine design.