'Vision is the art of seeing things invisible,' wrote master of fantasy, Jonathan Swift. His voyages to kingdoms of changing visual scales served to emphasize the potential insights to be gleaned from viewing the world through continuously fresh perspectives.
Weizmann Institute researchers have recently demonstrated this principle's applicability to the art of scientific discovery. A decision on their part to change an experimental focus resulted in their capturing the first ever time-resolved 'movie' demonstrating how molecules break apart when exposed to synchotron radiation. Their findings, published in the January 18th issue of the Proceedings of the National Academy of Science USA (PNAS) may pave the way to improved techniques for studying biological molecules, as well as to pharmacological measures for preventing high-dose radiation damage, a common cause of cancer and birth defects.
The Weizmann scientists participating in this study were Dr. Gitay Kryger, Dr. Michal Harel and Prof. Joel Sussman of the Structural Biology Department, together with Prof. Israel Silman of the Neurobiology Department. The team worked in close collaboration with Martin Weik, Maria Raves, Piet Gros, and Jan Kroon from Holland's Bijvoet Center for Biomolecular Research at Utrecht, as well as Raimond Ravelli and Sean McSweeney of the European Molecular Biology Laboratory Outstation at Grenoble, France.
'In science, it is quite common to seek answers to one question and find answers to entirely different ones,' says Dr. Kryger, providing the background to their discovery. 'While looking down one avenue, we were essentially sidetracked into an alley, with perhaps even broader applications.'
In the course of studying one of nature's most intriguing enzymes, acetylcholinesterase (AChE), which plays a pivotal role in brain function and memory, the team decided to examine its enzymatic reaction in 'real-time', using X-ray crystallography, based on exposing crystals of the enzyme to high intensity X-ray beams.
Since this enzyme's reaction proceeds within microseconds, the researchers hoped to record the process by taking an extremely rapid series of X-ray 'snapshots.' The results, in fact, indicated a time-dependent change. However, upon closer examination, the researchers realized that in their attempt to capture the enzymatic reaction, they had actually obtained the first-ever 3D recording of radiation-induced breakage of specific chemical bonds in the protein. 'The observation was stunning. While the time-series movie looks like a simulated animation of chemical processes, we knew that we were seeing a direct experimental observation - something that had never been seen before,' said Prof. Sussman.
Subsequent studies revealed that, contrary to the previously held belief, radiation damage could actually be highly specific. The team found that disulfide bonds (which often bridge protein polypeptide chains) and carboxyl acids (such as those found at the 'active site,' where enzymatic reactions are initiated) are particularly prone to radiation damage. They also found a cross-species similarity, suggesting a more general phenomenon. Highly similar results were obtained when working with AChE crystals derived from the Torpedo fish (one of the richest sources of this enzyme), humans, and the Drosophila fruit fly, as well as from a different enzyme entirely, hen egg white lysozyme.
These findings have direct implications for improving data collection using X-ray crystallography. This technology presents significant challenges. While providing a dramatic glimpse into heretofore inaccessible microscopic worlds, it simultaneously introduces radiation damage, thereby often destroying the experimental sample. The crystallographic community has traditionally walked a thin experimental line, trying to enhance information gathering via increased X-ray intensity, while mitigating radiation damage through various techniques, in particular cryo-crystallography (data collection at extremely cold temperatures).
According to Dr. Kryger, 'One of the most important 'take-home' lessons is that in the attempt to better understand biological molecules, less intensive radiation may provide more accurate results. The key is to avoid introducing inadvertent changes into experimental samples, such as those induced by radiation damage.'
However, radiation damage and its prevention is a central issue which 'spills over' beyond laboratory walls. Organisms are constantly exposed to potentially detrimental radiation, whether from sunlight or radioactive materials. Such radiation, at high levels, is a common cause of cancer and birth defects. According to Prof. Sussman, 'The ability to visualize the specific damages caused by radiation on a 'test-tube' level offers an important diagnostic tool for developing pharmacological means to protect against radiation damage. These measures could be applied on a conventional or emergency situation basis, such as that which followed the Chernobyl nuclear power plant failure.' The Weizmann Institute team plans to collaborate with its European counterparts in examining the anti-radiation potential of various substances.
This study was funded by the U.S. Army Medical and Materiel Command, The European Union 4th Framework Program in Biotechnology, the Kimmelman Center for Biomolecular Structure and Assembly (Rehovot, Israel), and the Dana Foundation. The generous support of Mrs. Tania Friedman is gratefully acknowledged. RBGR acknowledges support from the TMR Access to Large Scale Facilities to the EMBL Grenoble Outstation.
Prof. Israel Silman, a member of the Weizmann Institute's Neurobiology Department, holds the Bernstein-Mason Chair of Neurochemistry. Prof. Joel Sussman is a member of the Weizmann Institute Structural Biology Department and has a joint appointment at the Brookhaven National Laboratory in New York.
The Weizmann Institute of Science is a major scientific research graduate study located in Rehovot, Israel. Its 2,500 scientists, students and support staff are engaged in more than 1,000 research projects across the spectrum of contemporary science.
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