Major Enhancements to the X-Ray Crystallographic Data Collection Equipment in the Krebs Institute

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An extensive programme of macromolecular structure analysis is underway in the Krebs Institute, Sheffield University Institute. Our programmes of work include: (1) Structural proteomics. Major BBSRC-funded projects that are providing numerous targets for crystallisation include a targeted structural genomics project for the determination of structures of identified essential gene products in Gram positive organisms (B. subtilis, S. aureus), and a ground-breaking programme on yeast protein complexes. (2) A wide-ranging portfolio of projects on nucleic acid binding proteins, including proteins involved in recombination, replication, transcription and translation. (3) Investigation of protein stability in extreme environments, including enzymes from hyperthermophiles, psychrophiles and halophiles. (4) Enzymology, including the molecular basis of substrate specificity and design of novel enzymes. (5) combating drug resistance in bacteria and parasites. (6) Aspects of pathogenicity, including toxin structures. All these projects, aided by our REI-funded protein production and crystallisation facilities, are producing a rapid flow of protein crystals for structural analysis. However our work is limited by the deficiencies in our current ageing data collection equipment. Our existing pair of Mar345 detectors remain highly effective, and are matched by a state-of-the-art microfocus x-ray generator optics combination. However, the detectors are mounted on old goniostats that lack versatility, are controlled by ageing computers, and crystals are cooled by increasingly unreliable and obsolete cryogenic systems. Major advantages of the new configuration include: (1) The ability to optimise data collection for challenging crystal problems by using the MarResearch Desktop Beamline to visualise very small crystals, to minimise crystal icing during long data collections, to collect high resolution data from crystals with long unit cells and to minimise x-ray background. (2) Efficient crystal cryo-cooling during data collection by replacement of our 7 year old, outmoded and increasingly unreliable, Oxford Cryosystems Cryostream 600 crystal coolers with state of the art 700 series crystal cryo-coolers, and essential part of any protein data collection facility. (3) Faster data processing, and hence more efficient screening of crystals, using a fast, modern, linux based PC data processing farm. The enhancements will provide an efficient data collection facility to allow the rapid prosecution of a range of structural projects that probe key problems in biology.