This proposal requests funds to upgrade facilities for analysis of biological macromolecules, using X-ray diffraction, at the University of Texas. The equipment will be used in the research of two established investigators and two newly appointed assistant professors. It falls into three categories, A. X-ray diffraction, B. high speed computing, and C. molecular graphics. A. The diffraction equipment consists of a high brilliance rotating anode X-ray generator and a state of the art imaging plate (IP) area detector. The detector captures X-rays as metastable centers on a BaFBr:EuX-phosphor, which is read by subsequent laser activation. This type of detector has the spatial resolution to accommodate both single crystal protein diffraction and the low angle scattering patterns generated by the users of the equipment. B. A high speed, dual processor, computer with 4 Gbytes of disk capacity will be used largely for molecular model refinement. This machine will allow timely turnover of simulated annealing refinements using the program XPLOR. It will also be used for sophisticated analysis of low angle scattering patterns. C. A modern molecular graphics station will be acquired. This instrument will augment a heavily used molecular graphics capacity and may be required to replace a failing instrument currently in use. The research interests of the user group are quite varied, involving a number of biologically important systems under study by a range of methodologies. Proteins under study by single crystal analysis include: ribosome inhibiting proteins like ricin, PAP and ebulin; plant and fungal chitinases; the antifungal protein zeamatin; at least four structurally different amino acid decarboxylases including ornithine, arginine and histidine decarboxylases; several invertebrate hemoglobins; multienzyme complexes, like PDC; and the ribosomal RNA binding protein L9. In addition to single crystal studies, the equipment will be used in the low angle scattering ana lysis of polymerizable lipid assemblies. Systematic studies are underway to define the rules governing subassembly in the planes of lipid bilayers. The above projects tend to extend well beyond simply describing the initial X-ray structure. Each protein is refined in a crystallographic sense to the limits of it data in order to produce as accupate a molecular model as possible. Also, the binding of substrate analogs or other ligands is assessed where appropriate. Several of the research groups have vigorous site-directed mutagenesis programs which create a family of proteins whose X-ray analysis is important to describing their structure/function relationships. Finally, structure-based drug design is being developed. The design paradigm requires analysis and energy refinement of numerous intermediate structures. The equipment requested is of great importance to the continuation of established programs and to the expansion of new ones. The X-ray generator and IP detector, together with a cold temperature apparatus, are important for data collection on the many systems under study. Some of these crystal systems have large unit cells and require lengthy time periods or several crystals for complete data collection. Some projects involve a large number of mutant, inhibited, or liganded structures, and in all cases it is desirable to collect as high resolution data as possible. All of these considerations argue for increased data collecting capacity. Furthermore, research projects involving low angle scattering work on bilayer structures cannot be carried out, for technical reasons, without acquisition of the new equipment. The requested computing upgrade will allow modern structural refinement programs, such as XPLOR, to run expeditiously. It will also allow the most modern and powerful data analysis programs for single crystal and low angle scattering analysis to be executed with comparative ease. The need for this system is intensified by the apparent demise of the University managed supercomputer. With over a dozen students and other scientists using the facilities, there is a strain on current molecular graphics equipment. A new unit will allow proper analysis of the mass of structural work being done particularly if one key instrument, currently in place, fails.
