Methods for identifying proteins amenable to crystallization
Lead Inventors: Eric Gouaux, Ph.D.Problem or Unmet Need:Although protein crystallography is a powerful tool in both basic research and drug development, the process of protein crystallization is still poorly understood. This is evidenced by a lack of knowledge about what makes a protein a good candidate for crystallization as opposed to a poor one. Current methods rely on generating large numbers of expression constructs, expressing and purifying the proteins on a large scale, and then characterizing them prior to crystallization. This process is both expensive and time consuming. Ideally, it would be possible to characterize protein constructs prior to large scale expression and purification, reducing both cost and time.This invention includes a method for evaluating proteins as candidates for crystallization experiments. The method is consists of performing size exclusion chromatography on the proteins of interest expressing a fluorescent tag. The quantity of protein only needs to be on the order of 10ng and does not need to be in a purified state. As such, the chromatography can be performed on the soluble fraction of total cell lysate. The chromatography is used to detect the fluorescent elution profile of the protein, which can then be characterized to determine likelihood of crystallization. Proteins likely to crystallize have elution profiles that consist of a single, substantially symmetrical peak.
Characterize proteins earlier in the crystallization process than current methodso Decrease costso Increase efficiency Yields information about a protein's chromatographic behavioro More predictive of how amenable the protein is to crystallization than the standard yes/no answer for induction and solubility
Identification of proteins conducive to crystallization for use ino Basic scientific researcho Structure-based drug discovery
This invention includes a method for evaluating proteins as candidates for crystallization experiments. The method is consists of performing size exclusion chromatography on the proteins of interest expressing a fluorescent tag. The quantity of pro...
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