This proposal requests support to purchase and install a 200 kV cryo-electron microscope (Cryo-EM). The Cryo-EM is a key component of the strategic research plan of the School of Medicine, involving faculty within the Departments of Microbiology, Medicine, the Center for AIDS Research, and the Center for Bio-defense. Cryo-EM permits analysis of complex structures without the artifacts that are introduced with staining protocols used in conventional EM. Using image reconstruction software, cryo-EM produces 3-D renderings of complex structures, including complete virus structures revealing protein/nucleic acid interactions involved in viral assembly and maturation. Several ongoing NIH-funded research projects will be dramatically enhanced by access to a 200 kV cryo-EM on the UAB campus. These include determination of the mechanisms underlying maturation of the bacteriophage P22 (P. Prevelige), analysis of interactions between an RNA viral genome and the viral capsid (A. Ball), assembly of the bluetongue virus core (P, Roy), and packaging of the VSV RNA nucleocapsid complex (M. Luo). Additional studies of assembly of bacteriophages P2 and P4 will be performed by new recruit T. Dokland. All these projects focus on mechanisms of assembly of multi-protein complexes and establishment of nucleic acid-protein interactions in these complexes. Acquiring a 200 kV microscope is a key step in plans to develop a consolidated electron microscopy facility within the Shelby Research Building that will also include a 300 kV cryo-EM, conventional I20 kV transmission EM and scanning EM. The director of the Cryo-EM facility will be Dr. Terje Dokland. Acquisition of this instrument will dramatically enhance the structural biology capabilities of UAB scientists who are poised to apply this technology to analysis of cellular organelles and macromolecular complexes. Data collected using this technology will support ongoing and high-priority initiatives in structural biology, virology, trans-membrane channels, bio-defense, and vaccine development. ? ? ?
