High-throughput crystallization-to-structure pipeline

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In the APCR-group, we developed a high-throughput crystallization-to-structure pipeline utilizing a variety of robotics, software and elemental technologies. The first 2 years of this effort focused on developing an automated environment for processing a huge number of samples in crystallization experiments, diffraction experiments and X- ray data analyses. Our automated environment enabled novice users in protein crystallography to perform structure determination. Utilizing the automated facilities, we spent the latter 3 years producing most of the research results, including the development of various elementaltechnologies for difficult target proteins. During the course of the Protein 3000 project, the present pipeline produced 307 PDB structures containing 169 structures for functional analysis, such as complexes and mutants (Fig. 1), indicat- ing the pipeline made a significant contribution to the large-scale determination of protein crystal structures. As shown in Table 1, this pipeline works most effectively in the analysis of mutant structures and in ligand screening. Thus use of the present system will be focused on not only elucidating the structure-function relationship of proteins but also on developing new technologies in protein engi- neering toward the design of useful proteins on the basis of the large number of mutant structures. One of the most important tasks in structural genomics is to develop a high- throughput protocol for recalcitrant targets such as those from mammalian membrane proteins. The current pipeline should be continuously re-evaluated and updated to improve both the throughput and the ability to process important but difficult target proteins.

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