August 18, 2006

Microscope to give biological sciences group a new look

WEST LAFAYETTE, Ind. — Purdue University's structural biology group received a $2 million grant to purchase a state-of-the-art electron microscope that will enhance the study of complex biological specimens, including viruses.

"Electron cryomicroscopy, or cryo-EM, is an emerging field," said Richard Kuhn, department head of biological sciences. "This microscope has the potential of improving the resolution of images in part because it has the facility to cool specimens to liquid helium temperatures, thereby deceasing damage resulting from the electron beam. In an electron microscope, the electron beam takes the place of a light beam in a conventional microscope. However, the wavelength of electrons is far shorter than the wavelength of light, which makes an electron microscope suitable for 'seeing' objects almost in atomic resolution.

"The microscope will allow us to see far more detail. With what we have now, a protein appears as a sort of glob. With this new microscope, we should be able to see many more detailed features."

The grant was provided by the National Institute of Health's National Center for Research Resources. This will make it possible for the Purdue structural biology team to purchase an electron microscope equipped with a field emission gun, a Helium-cooled stage and an energy filter.

Michael Rossmann, Hanley Professor of Biological Sciences and principal investigator of the grant application, said the new microscope is a welcome addition because the instrument should make it possible to visualize detail that can then be augmented by crystallography. Crystallography is a powerful technique that permits the determination of the three-dimensional structure of individual protein or nucleic acid molecules at near-atomic detail.

However, crystallography requires that these biological molecules can be crystallized, just like the common salt crystals you see in the salt shaker on the dining room table. The process of crystallization is difficult and becomes more difficult the larger and more complex the object being studied, Rossmann said. That is where the modern electron microscope comes in, for it can determine the general organization of these bigger molecular complexes.

"Molecules must be crystallized in order to use this method," Rossmann said. "Not everything can be crystallized, especially complicated things like viruses. Fortunately, electron microscopy technology has come along and advanced rapidly to fill this gap."

Purdue is planning to build a new structural biology building that is scheduled to be completed in 2009. A major component of the new building will be the microscopy facilities that will keep the Purdue Structure Group at the forefront of studying the physical features of biological molecules such as viruses and membrane protein complexes.

"The state-of-the-art microscope allows us to place one of our pre-existing electron microscopes into a containment facility planned for the new building," Rossmann said. "Obviously, there would be limited access to such an area, and one of the older microscopes would have to be dedicated to restricted use within the facility. Having such a facility is very special and will support research into the study of serious human pathogens responsible for widespread disease."

The Department of Biological Sciences currently has three electron microscopes that use a nitrogen cooling stage capable of cooling specimens down to about 100 degrees above absolute zero (minus 173 degrees Celsius).

The new microscope, estimated to cost a total of $4 million, will be able to use liquid helium, which has a temperature of only 4 degrees above absolute zero. This offers better protection to specimens, said Wen Jiang, assistant professor of biological sciences and an internationally recognized electron microscopist.

"When electrons pass through a specimen, they cause radiation damage," he said. "To minimize this damage, we freeze the specimens at temperatures well below the freezing point of water. Helium-cooled specimens should, in general, last about one-and-a-half times longer than those cooled with nitrogen for the same electron dosage. An electron microscope is similar to a common camera. Longer exposure time allows for sharper, more detailed images. The reduction in radiation damage at lower temperatures will allow an increase in exposure time, which will give the final image more accuracy and less noise.

"This microscope is necessary for our group to remain in the top of the field. It will allow us to do as well or better than the major labs around the world.

"Electron microscopy and crystallography are complementary. The strength of cryoelectron microscopy is its ability to provide large-scale information. The strength of crystallography is its ability to provide information at high resolution. We use both to piece together a detailed, comprehensive picture."

Rossmann said, "No one technology replaces another. Light microscopes, like those used in high school science labs, are still very important and are used in research. It is a matter of seeing the different layers of structure and obtaining a complete understanding."

Jiang said the software used in conjunction with the microscope is equally as important as the microscope's construction.

"We work to create custom software to extract information from the noisy images and provide a high-resolution end result," he said. "The software is critical in pushing cryoelectron microscopy to higher resolution. We work with the information technology department at Purdue to provide the large amount of computing power required for these studies."

The microscope's energy filters provide advantages for tomographic studies. This is a technique for the study of large biological objects such as a whole cell. It contrasts with more developed techniques currently in vogue for biological objects in which each object is exactly the same, such as small common cold or West Nile viruses. Tomography requires highly automated computer-controlled data collection procedures before computing three-dimensional reconstructions of the object being studied. This method allows scientists to view virus-cell interaction, Rossmann said.

"This new equipment offers the possibility of doing things not possible before, and it allows us to maintain our position as leaders in this field," he said.

The National Center for Research Resources provides grants to fund cutting-edge equipment to advance biomedical research and increase knowledge of the underlying causes of human disease. Awarded to research institutions around the country, the one-time grants support the purchase of sophisticated instruments costing more than $750,000. In addition to Rossmann, Jiang and Kuhn, Jue Chen and Wei Zhang are co-investigators on this award. Paul Chipman will be overseeing the purchase and maintenance of the instrument.

Writer: Elizabeth K. Gardner, (765) 494-2081, ekgardner@purdue.edu

Sources: Richard Kuhn, (765) 494-4407, kuhnr@purdue.edu

Michael Rossmann, (765) 494-4911, mr@purdue.edu

Wen Jiang, (765) 496-8436, jiang12@purdue.edu

Purdue News Service: (765) 494-2096; purduenews@purdue.edu

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