In the past fifteen years it has become increasingly clear that atomic structural information via x-ray crystallography has accelerated and enhanced our understanding of the fundamental molecular mechanisms behind disease processes. For the majority of challenging aqueous and membrane proteins, the ability to rapidly determine three-dimensional structures has become more and more dependent on our ability to produce diffraction-quality three dimensional crystals of the target protein. This proposal addresses a critical need of the structural biology community at the University of Alabama at Birmingham (UAB): the availability of high throughput crystal imaging systems that can accommodate crystallization at the two most common temperatures, 20 ?C and 4 ?C. The UAB structural biology program includes 11 faculty-level crystallographers who conduct structural studies on macromolecules associated with a diverse scope of biological research areas, all of which involve medically relevant proteins and proteins involved in complicated biological processes. Some of the current projects include (1) structural studies on prokaryotic and eukaryotic membrane proteins, (2) bacterial adherence proteins, (3) viral capsid formation proteins, (4) proteins involved in the integrin signaling pathway, (5) vesicular stomatitis virus (VSV) nucleocapsid (6) heat shock proteins, (7) cytokines and (8) several protein structure-based discovery projects that involve developing inhibitors for protein targets associated with diseases such as cancer, cystic fibrosis, anthrax, plague, tularemia, viral influenza and tuberculosis. Each of these projects stands to benefit significantly from the proposed crystal imaging/analysis instrumentation which is expected to have a major impact on the progress of several NIH-funded research projects. Many of these projects involve highly labile/temperature sensitive proteins requiring 4 ?C for purification and crystallization. In other cases (i.e. protein-protein complexes, protein-inhibitor complexes) crystals often display high mosaicity and/or poor diffracting resolution. However, many other projects previously crystallized, as well as many not yet crystallized, require trials at multiple temperatures since temperature is an important variabl in crystallization screening. For many of these challenging projects, the rate limiting step is the production of high-quality crystals. The addition of 4 ?C and 20 ?C high-throughput crystal imaging/analysis systems is requested to address the needs of eleven faculty level crystallographers and their pre-and postdoctoral level students. In addition, the requested systems will be incorporated into the Comprehensive Cancer Center's X-Ray Shared Facility thereby supporting any faculty member and/or students at the University of Alabama at Birmingham. The system's ability to remotely monitor a large number of crystallization experiments will enable utilization of this capability for several of the UAB faculty and students located in other buildings on campus.
