US-Switzerland Collaborative Research on Cryo-Electron Microscopy

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This award supports Dr. Edward Egelman of Yale University to collaborate in research with Dr. Jacques Dubochet of the Center for Electron Microscopy of the University of Lausanne, Switzerland. They are collaborating in a study of the RecA protein, the most intensively studied enzyme of general genetic recombination; their approach will also contribute substantially to improving techniques for the structural analysis of macromolecules in general. Dr. Egelman is a biophysicist who studies disordered and flexible macromolecules, specifically the RecA protein and actin. Traditional methods of structure determination, such as protein crystallography, have been generally unsuccessful with these molecules. Dr. Egelman has been examining them using computer-based image analysis of electron micrographs; he is developing a special facility for this purpose with support from the Instrumentation and Instrument Development Program of NSF. His particular expertise lies in graphics and software development; he depends on other laboratories for the electron micrographs to be studied. Dr. Dubochet has pioneered methods of cryo-electron microscopy of fully hydrated biological specimens. Only a handful of laboratories in the world are working in that field, and Dr. Dubochet is considered the leading expert on its use in imaging macromolecules. He will prepare frozen hydrated electron micrographs of RecA and other dynamical helical systems, which will provide one of the best means for both developing and demonstrating the effectiveness of Dr. Egelman's image processing hardware and software. Their complementary expertise and facilities will accelerate their mutual research on RecA and other dynamical protein polymers, and both laboratories will benefit from a reciprocal transfer of specialized techniques. The electron microscopy of frozen, fully hydrated biological specimens has emerged as one of the most promising new developments in structural biology in the past several years. It offers one of the most powerful methods available for visualizing disordered and dynamic helical protein polymers. It produces images of specimens with little or no damage or distortion from dehydration, heavy metal staining or electron dose. Dr. Egelman is developing instruments and programs to allow greatly accelerated analysis of electron micrographs, in particular to extract information about polymer dynamics and function from static images. His techniques for computer-aided enhancement and reconstruc- tion are even more necessary for frozen, hydrated specimens than for conventional electron micrographs, due to the problems of weak contrast and artifacts introduced by the optics of the electron microscope.