The recent re-emergence and outbreak of monkeypox virus Mpox has once again made poxviruses a public health threat and highlighted important gaps in core knowledge about poxviruses. Now, a research team at the Institute of Science and Technology Austria (ISTA) has unveiled the mystery of the core structure of poxviruses by combining various cryo-EM techniques with molecular modeling.
These research results were published in the journal Nature Structural & Molecular Biology and will help future therapeutic research targeting the core of poxviruses.
The variola virus is the most notorious poxvirus and one of the deadliest viruses to humans. It caused havoc by causing smallpox, which was not eradicated until 1980. Successful eradication was possible thanks to an extensive vaccination campaign using another vaccinia virus. The re-emergence and outbreak of monkeypox virus in 2022-2023 once again reminds us that viruses will always find ways to return to the forefront of public health threats. Importantly, this highlights fundamental questions about poxviruses that remain unanswered to this day.
One of the fundamental questions is at the heart of the matter: "We know that for a poxvirus to be infectious, its viral core must be formed correctly. But what is this poxvirus core made of, and how do its components fit together and function?" asked Florian Schur, the study's corresponding author and an assistant professor at the International Association for Science and Technology.
Schur and his team have now found the missing link: a protein called A10. Interestingly, A10 is a protein common to all clinically relevant poxviruses.
In addition, the researchers also found that A10 is one of the major components of the poxvirus core. This knowledge could aid future research into therapies targeting the core of poxviruses.
The viral core is one of the factors common to all infectious poxviruses. "Previous experiments in virology, biochemistry and genetics proposed several candidate core proteins for poxviruses, but no experimentally derived structures were available," said Julia Datler, co-first author of the study and a doctoral student at the International Institute of Science and Technology. "
Therefore, the research team first used the now famous artificial intelligence-based molecular modeling tool AlphaFold to perform computational predictions on the main candidate core protein model. At the same time, Datler laid the biochemical and structural foundation for the project, drawing on her virology background and the Shure group's primary expertise: cryo-electron microscopy, or cryo-EM for short.
"We combined state-of-the-art cryo-electron microscopy techniques with AlphaFold molecular modeling technology. This gave us the first detailed view of the poxvirus core - the 'safe' or 'bioreactor' inside the virus that encapsulates the viral genome and releases it into infected cells," said Schur.
"It was a bit of a gamble, but we ended up finding the right combination of techniques to study this complex problem," said postdoc Jesse Hansen, co-first author of the study.
Researchers at the International Institute of Science and Technology studied "live" Vaccinia virus mature viruses and purified poxvirus cores from every possible angle.
"We combined 'classic' single-particle cryo-electron microscopy, cryo-electron tomography, subimage averaging and AlphaFold analysis to obtain a holistic view of the poxvirus core," said Datler. "Using cryo-electron tomography, researchers can acquire images while gradually tilting the sample to reconstruct the three-dimensional volume of a biological sample as large as the entire virus."
"It's like doing a CT scan of a virus. Cryo-electron tomography is our laboratory's 'specialty' and allows us to obtain nanometer-scale resolution of the entire virus, its core and interior."
In addition, the researchers were able to fit the AlphaFold model into the observed shapes like a puzzle and identify the molecules that make up the core of the poxvirus. Among them, candidate core protein A10 stands out and becomes one of the main components. "We found that A10 defines key structural elements of the poxvirus core," Datler said.
These findings are an important resource for interpreting the structural and virological data generated over the past few decades.
The path to these discoveries was not easy. "We need to find our own path from the beginning," Datler said.
Datler used her expertise in biochemistry, virology, and structural biology to isolate, propagate, and purify Vaccinia virus samples and develop protocols for purifying intact viral cores while optimizing structural studies of these samples. "Structurally, these viral cores are extremely difficult to study. But luckily, our perseverance and optimism paid off."
ISTA researchers are convinced that their findings could provide a knowledge platform for future therapies targeting the core of poxviruses.
"For example, we could consider using drugs to prevent viral core assembly or even break down and release viral DNA during infection," Schur concluded. "Ultimately, the basic viral research done here will allow us to better prepare for possible future viral outbreaks."
Compiled source: ScitechDaily