The study offered a picture of VP7 in isolation from the virus. To visualize the outer-coat proteins on an intact virus particle, Harrison partnered with Nikolaus Grigorieff, an HHMI investigator at Brandeis University and expert in an emerging technique called electron cryomicroscopy (cryo-EM).

In cryo-EM, scientists preserve a large macromolecular complex by freezing it rapidly in a bath of liquid ethane (see Toolbox). Once immobilized, particles can be imaged from every angle with a narrow beam of electrons. The thousands of images are then averaged into a high-resolution three-dimensional picture. The symmetrical nature of rotavirus helps, Grigorieff explains, because each image reflects what the structure looks like from multiple angles.

The result was a picture that showed at near atomic resolution—four angstroms—VP7 proteins encircling the spike-shaped VP4 proteins in an interlinked web across the surface of the virus.

Harrison, Grigorieff, and colleagues published the structure in the June 30, 2009, issue of Proceedings of the National Academy of Sciences. The VP7 proteins appear to hold VP4 in place until conditions—probably a drop in calcium concentration in the host cell vesicle surrounding the engulfed virus particle—prime the particle for infection. The VP7 proteins are then released, freeing VP4 to puncture the membrane of a target cell. The antibody crystallized with VP7 apparently prevents infection by clamping VP7 onto the virus particle and preventing VP4 from puncturing the cellular membrane.

Harrison now wishes to understand how, and where within the cell, the spike protein VP4 carries out this puncturing step. He plans to investigate it with x-ray crystallography, with cryo-EM, and using light microscopy to image virus particles entering living cells. “We're now poised to ask, how does VP4 actually do it?” he says. “We don't know the answer yet, but we know a lot about what it must involve, and we know the kinds of experiments we need to do next.”

	PAGE 2 OF 2	Back