Researchers team reveals molecular structures developed a 3-D model based on the 1918 H1 pandemic influenza virus. Virus-like particles (VLPs) are protein-based structures that mimic viruses and bind to antibodies. Because VLPs are not infectious, they show considerable promise as vaccine platforms for many viral diseases, including influenza. The study was published in Scientific Reports, could benefit VLP vaccine projects, targeting a range of viruses from HIV to Ebola and SARS coronavirus.
Researchers had produced VLPs for 1918 H1 influenza that successfully protected animals from different influenza viruses. The NIAID group prepared hundreds of such VLP samples and analyzed their structure with a technique called cryo-electron microscopy, which quick-freezes samples with glass-like clarity.
They then sliced through those VLP 3-D structures like slicing through a loaf of bread to analyze their internal structure, using computers to document the size and placement of key molecules. After averaging all their data, the group then created a 3-D 1918 influenza VLP model.
Image analysis of VLP surface spikes
HA spikes were observed on the surfaces of VLPs. To address the questions of HA conformation and epitope display of HA on VLPs, 2D-image analysis of cryo-EM images was performed along with immunoassays with antibodies to probe for epitope integrity and accessibility.
The internal component of VLPs
Interestingly, tomography and subsequent computational segmentation of a VLP indicated multiple structural components. Tomography allowed the internal components of VLPs to be visualized and analyzed. There was an HA-membrane layer and observed density inside corresponding to internal components.
The scientists found that about 90 percent of the VLPs are hemagglutinin (HA) proteins (by weight) found on the VLP surface. In contrast, HAs comprise less than half of the viral proteins of natural influenza viruses. The number and location of HA molecules may influence the efficacy of VLP vaccines, influencing the binding of antibodies to specific epitopes on the HA protein. Those antibodies can similarly bind live influenza viruses, preventing them from infecting cells.
Electron microscopy
Negative-staining electron microscopy of VLPs was similar to that reported previously for other samples, except that VLPs were stained with 1.5% phosphotungstic acid. Images were collected on a Tecnai 12 electron microscopy with LaB6 illumination operating at 100 kV at nominal magnifications of 52,000x and 110,000x.
The research group, in NIAID's Laboratory of Infectious Diseases, is continuing its work by comparing its VLP data to data from other natural influenza viruses. They believe the more that is understood about the molecular organization of influenza VLPs; the better scientists will be able to develop effective seasonal and universal influenza vaccines.
