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Structural basis for photoswitching in fluorescent proteins brought into focus

EUGENE, Ore.University of Oregon scientists have identified molecular features that determine the light-emitting ability green fluorescent proteins, and by strategically inserting a single oxygen atom they were able to keep the lights turned off for up to 65 hours.

The findings, published online this week by the Proceedings of the National Academy of Sciences, likely are applicable to most photoswitchable fluorescent proteins, said S. James Remington, professor of physics and member of the UO Institute of Molecular Biology.

"This new model makes specific predictions and improves the qualities of the protein as a photo-switchable label," Remington said. "It gives us the first picture of how these molecules can be switched on and off. That allows us to design new variants to make the proteins more useful."

For more than a decade, fluorescent proteins first isolated in jellyfish and since found in a variety of colors from coral reef organisms revolutionized molecular biology, allowing scientists to use them as markers for genetic expression, to locate molecules and observe activity within cells.

The recent discovery of photoswitchable fluorescent proteins which can be manipulated with a laser has been a significant development for cellular research.

"Photoswitchable fluorescent proteins have tremendous advantages over passive proteins," Remington said. "You can label all molecules but using a laser under a microscope, you can activate only a small group of them. That lets you follow the motion of subsets of molecules. We wanted to understand the process, so that we can permanently switch them off and on or vary the time delay."

However, he said, the mechanism of photoswitching was unknown, and in many cases the proteins returned to their stable state randomly and spontaneously.

Using a combination of rational mutagenesis and directed evolution, UO doctoral student J. Nathan Henderson determi
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Contact: Jim Barlow
jebarlow@uoregon.edu
541-346-3481
University of Oregon
10-Apr-2007


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