Gorrec F, MRC Laboratory of Molecular Biology, Cambridge, UK
Technology developments, including innovative crystallization screens, are needed to obtain X-ray diffraction-quality crystals from increasingly challenging protein and other macromolecular samples. MORPHEUS crystallization screens are continuously developed to further enhance initial screening. MORPHEUS screens integrate small molecules frequently observed in the PDB to co-crystallize with proteins. These molecules are included to function as additives that act as protein stabilizers, crystal packing bridges, or any other role beneficial to protein crystallization.
Each MORPHEUS screen integrates 96 conditions, a minimal format for limiting amounts of sample. To limit the number of conditions employed, the potential ligands found in the PDB are combined into mixes. Other mixes of compounds are used, such as buffer systems and precipitant mixes that also act as convenient and effective cryoprotectants. The different types of compound mixes are combined using a fixed ratio to generate 3-D grid screens. The preparation of a screen and the optimization of conditions are amenable to automation.
A multitude of test crystallization screens are used against novel and challenging samples produced at the LMB before making choices about the formulation. The main goal is to produce exclusive crystallization hits that were not observed in other screens. MORPHEUS screens, I and II, are now used routinely in many laboratories, while MORPHEUS III is being developed. Before describing the developments related to the MORPHEUS screens, I will briefly present theoretical and pragmatic aspects of macromolecular crystallography that were taken into consideration during the early stages of MORPHEUS development. In the last section I will show how to optimize conditions following the 4-corner method.
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