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Virus Studies Reunite Two Families Of Major Insect-Borne Viruses

April 9, 2001
Purdue University
Two families of viruses, which were previously thought to be related and later were considered unrelated, are celebrating a scientific family reunion of sorts.

WEST LAFAYETTE, Ind. – Two families of viruses, which were previously thought to be related and later were considered unrelated, are celebrating a scientific family reunion of sorts.

In a study published in today's (Friday, 4/6) issue of the scientific journal Cell, scientists at Purdue University show that a protein structure on the outside surface of an alphavirus is related to a protein found on the surface of a family of viruses called flaviviruses. A second study, published in the same issue by a research team in France, shows a similar likeness.

"What we have then is two seemingly unrelated groups of viruses that have now been related based on the way they build their shell, or cage," says Richard Kuhn, associate professor of biological sciences and co-principal investigator for the Purdue study. "Together these two families of viruses cause more than half of the of insect-borne viral diseases in humans and animals worldwide. By better understanding the structure of these viruses, we may be able to uncover the mechanisms that allow them to thrive and develop new strategies for their prevention and control."

The findings provide new insights on how these major families of insect-borne viruses, which together cause millions of cases of human illness each year, are structured and how they evolved to infect cells. The discovery may help scientists develop antiviral compounds and other strategies to target the diseases caused by these viruses, including yellow fever and viral encephalitis.

The protein, called E1, was found to lie flat on the surface of an alphavirus called Sindbis, forming an icosahedral scaffold similar to the arrangement found in flaviviruses.

"This similarity in shell-structure tells us that the mechanism by which they enter the cells is likely the same," Kuhn says. "It also suggests that there is an evolutionary relationship between these two groups of viruses. Perhaps a progenitor that was different than either of them right now."

Ironically, alphaviruses and flaviviruses, which share a number traits, were once lumped into a single family of viruses. Both virus types contain well-ordered, symmetrical shells covered by a fatty, lipid layer. And both types of viruses use ribonucleic acid, or RNA, to carry their genomes.

In addition, both alphaviruses and flaviviruses are transmitted to humans or animals by insects, particularly blood-sucking varieties, such as mosquitoes and ticks. Currently, more than 100 types of flaviviruses and alphaviruses have been identified, causing illnesses such as yellow fever, encephalitis, dengue and various neurological disorders.

But 10 years ago, when scientists sequenced the genomes of these viruses, they found the gene sequences were organized very differently, and concluded that the two virus groups were unrelated.

Now, by studying the structure of the proteins used to make up the Sindbis virus, it appears that at least part of the genetic heritage of these viruses must have had a common primordial ancestor, Kuhn says.

"Using protein structure as a basis for classifying the viruses is based on the fact that protein structures change much more slowly than do the amino acid sequences that make up the genomes, thus providing clues about the viruses' evolution," he says, noting that this concept was developed 25 years ago by Purdue researcher Michael Rossmann.

X-ray crystallography is a technique often used to study structures such as proteins and viruses in atomic detail. But the process works only if the substances can be made to form crystals. Crystals are used because the diffraction pattern from one single molecule could be insignificant, but the many individual, identical molecules in a crystal amplify the pattern. Diffraction patterns are created when an X-ray beam hits a crystal, causing the electrons surrounding each atom to bend the beam. Computers can then be used to interpret this pattern and reconstruct the positions of the atoms.

Enveloped viruses, such as alphaviruses and flaviviruses, are more difficult to study because their surface features make it difficult to grow crystals that diffract, Kuhn says.

"Previously, people have been able to obtain crystals for some of these viruses, but those crystals diffract only to 30 Angstrom, or 30 hundred-millionths of a centimeter, which is not usable for high-resolution studies," he says.

The recent findings at Purdue originated from efforts to crystallize an alphavirus called Sindbis virus. In a collaboration between three laboratories at Purdue and the laboratory of Dennis Brown at North Carolina State University, an alternative strategy was developed based on the production of modified Sindbis viruses. The Purdue group included Timothy Baker, professor of biological sciences and a specialist in electron microscopy; Michael Rossmann, Hanley Distinguished Professor of Biological Sciences and an expert in X-ray crystallography; and Kuhn, a specialist in virology.

Sindbis virus has an ordered structure, with proteins imbedded in the membrane in specific patterns. However, the virus surface also contains a series of sugar molecules, located at sites called "glycosylation sites," attached to large, protruding protein spikes.

The Purdue team developed modified varieties of the virus, genetically altered to eliminate the glycosylation sites, that allowed them to study general surface features of the virus.

Along the way, they realized they could use the mutant viruses to map the glycosylation sites, providing a rare opportunity to analyze the structures of the two major proteins involved, called E1 and E2.

The researchers developed the map by creating a virus with a mutation in only one of the glycosylation sites, and then gathered images of the structure using cryo-electron microscopy and three-dimensional image reconstruction techniques. These methods, which use hundreds of two-dimensional images to develop a three-dimensional view of the virus, allowed the group to determine the overall shape of virus and see the symmetrical arrangement of its components.

The group then subtracted this image data from images of a non-mutant version of the virus, leaving only the images and data for the glycosylation site. This process was then repeated for all four of the glycosylation sites on the virus.

In addition to identifying the sites on the virus where glycosylation occurs, the Purdue team discovered that the two major proteins involved played distinct roles in forming the surface features along these sites.

"It's been long thought that these two proteins come together to form this spike protrusion, running up and down the length of the spike," Kuhn says. "Our findings show that while the E2 protein extends up and down the length of the spike, the E1 protein lies along the surface of the virus."

The unusual placement of the E1 protein suggests that it plays a role in helping the virus fuse to a host cell membrane, similar to the role played by an E protein found in tick-borne encephalitis, a virus from the flavivirus family.

"We were struck by the unusual placement of the E1 protein," Kuhn says. "There are very few proteins that extend themselves in such a way that they lie flat on the cell surface membrane. For viruses, we knew of only one example, and that was a protein found in tick-borne encephalitis virus."

When the Purdue group saw the distribution of the glycosylation sites of the Sindbis virus, it immediately reminded them of the other protein, whose structure had been previously solved.

In parallel with the work they were doing, a group in France, headed by Felix Rey, was working on the structure of the E1 protein. They showed, using X-ray crystallography, that the structure of the E1 protein of an alphavirus was similar to the E protein in flaviviruses. Their findings were in agreement with the Purdue analysis of an actual alphavirus structure.

The study at Purdue was funded by grants from the National Institutes of Health.

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Materials provided by Purdue University. Note: Content may be edited for style and length.

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Purdue University. "Virus Studies Reunite Two Families Of Major Insect-Borne Viruses." ScienceDaily. ScienceDaily, 9 April 2001. <>.
Purdue University. (2001, April 9). Virus Studies Reunite Two Families Of Major Insect-Borne Viruses. ScienceDaily. Retrieved December 5, 2023 from
Purdue University. "Virus Studies Reunite Two Families Of Major Insect-Borne Viruses." ScienceDaily. (accessed December 5, 2023).

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