Acquition of a microdiffractometer for SER-CAT

University Of Georgia, Athens GA

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Abstract

DESCRIPTION (provided by applicant): Aided by technological advances in molecular biology, synchrotron X-rays, crystal cooling, detectors, phasing methods and structural genomics (notably high-throughput methods related to all aspects of the structure determination process) there has been an explosive growth in protein structure determination. It is estimated that fully 80% of all new structures are determined using synchrotron X-rays and many facilities are upgrading their beamlines facilities to meet this demand and to offer new capabilities, such as data collection from micro crystals (crystals whose dimensions are less than 50 ?). The desire for microdiffraction capability reflects need to collect data on smaller and smaller crystals as the community expands its research into larger and more difficult systems. For example, membrane proteins are both difficult to produce and crystallize, thus obtaining large crystals for diffraction studies is very difficult. Again, in another area, the study of protein- protein hetero complexes, production and crystallization of the intact complex is difficult and large crystals of the complex are hard to obtain. In both cases many of the systems under study are potential therapeutic targets. This proposal is a direct reflection of the desire of the Southeast Regional Collaborative Access Team's (SER-CAT) membership -- 90 plus research groups from 26 institutions (academic, industry, private and government) -- for microdiffraction capabilities on its undulator beamline (22ID) at the Advanced Photon Source, as was discussed during the March 17, 2007 meeting of the SER-CAT Executive Board. Since a majority of these groups (~75 funded research projects) are carrying out NIH funded extramural research the impact to NIH research program should be significant. To meet the demands of maintaining a microcrystal in a microbeam during data collection, three areas must be addressed beam size and shape, crystal imaging and goniometry. The proposed MAATEL MD2 microdiffractometer addresses these needs. Beam size - the 100? 22ID X-ray beam is too large. It can be reduced using pinholes similar but SER-CAT lacks the user-friendly beam imaging and diagnostic tools of the MD2;Crystal imaging - the SER-CAT crystal imaging system cannot easily image microcrystals, making alignment and data collection on sub 50? crystals difficult if not impossible. The on-axis microscope of the MD2 coupled with on-axis illumination makes it much easier to find and align microcrystals. In addition, the precise knowledge of the X-ray beam's size, shape and location (derived from beam imaging) insures that the microcrystal is centered in the microbeam;Goniometey - the ~5-10? sphere of confusion of the Rosenbaum goniometer used at SER-CAT is too large to ensure that microcrystal remains in the microbeam as the crystal is rotated during data collection. The MD2 goniometer has a 2msphere of confusion and an air bearing goniometer. This combination ensures that a 10??crystal remains in the beam during data collection. SER-CAT believes that the MD2 represents the most cost-effective solution for providing microdiffraction capabilities to its members. If funded will result in increased productivity and cost-effectiveness of beamline operation for both micro and general diffraction experiments (including remote data collection). This in turn will more and better science being produced. PUBLIC HEALTH RELEVANCE: Knowledge of a protein's 3-Dimensional structure can be a great aid in understanding fundamental biological processes and in the design of therapeutic agents. However, to understand the detailed nature of protein-ligand (drug) or protein-protein interactions high-resolution X-ray diffraction data, usually collected using synchrotron X-rays is required. As the community expands its research into larger and more difficult systems both protein production and crystallization become problematic usually resulting in microcrystals that are too small for data collection even when using synchrotron X-rays. Funds are requested to purchase a MAATEL MD2 microdiffractometer that will allow data collection on these microcrystals and should significantly impact the productivity and cost- effectiveness of synchrotron based X-ray structural biology resulting in more and better science being produced.